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Evaluation of viability of rat cortical

neural cells on surface-modified

PLGA films

Min Sub Lee

The Graduate School

Yonsei University

The graduate program in Biomedical Engineering

Major of Biomaterials



PLGA films

A Master Thesis

Submitted to the Graduate Program in

Biomedical Engineering and the Graduate School of

Yonsei University in partial fulfillment of

the requirements for the degree of Master of Science in

Biomedical Engineering

Min Sub Lee

December 2003

감사의 글

생체공학이라는 새로운 학문의 터에 발을 들인 지 엊그제 같은데 벌써 졸업이라는 과

정만을 앞두게 되었습니다 물론 앞으로 나아갈 길의 끝은 아니지만 삶의 하나의 마디로서

대학원이라는 새로운 환경을 무사히 마칠 수 있기까지 물심양면으로 도와주신 많은 분들

덕분에 이렇게 미비하나마 하나의 결과물을 조심스레 보여드릴 수 있게 되었습니다

우선 학위과정 동안 학문의 가르침만이 아니라 인생의 조언을 해주신 박종철 선생님께

감사의 마음을 전하고 싶습니다 철없고 게으른 저를 따뜻하게 때로는 따끔하게 이끌어주

신 은혜 잊지 못할 것 같습니다 그리고 바쁘신 와중에도 부족한 저의 논문을 심사해주시

고 새로운 분야의 지식까지 소개해주신 표면과학연구센터의 이인섭 교수님과 성형외과학

교실의 나동균 선생님께도 깊은 감사드립니다

학위과정 동안 깊은 관심으로 보살펴주신 의학공학교실 서활 선생님 김덕원 선생님

김남현 선생님 유선국 선생님께 감사드립니다 언제나 맑은 미소로 대해주시고 스키도 가

르쳐 주신 세종대 이권용 선생님 실험에 필요한 PLGA를 무진장 제공해주신 인제대의 김

정구 선생님 식약청의 류규하 선생님께도 감사의 마음을 전합니다 그리고 실험에 대해서

많은 조언과 편의를 제공해주신 전자현미경실의 정동룡 선생님 일본의 Uzawa Masakazu

씨에게도 감사의 마음을 전합니다

실험실 생활을 하는 동안 제가 힘들어할 때마다 용기를 북돋워주신 믿음직한 동희형

많은 실험 기술과 학생으로서의 자세를 전해주신 봉주형 눈빛만으로 통하는 동욱형 언제

나 싱글벙글 유머만점 인기만점의 현숙누나 눈웃음이 매력적인 동영형 동기이자 식약청

의 실세 원선누나 부지런한 얼짱 자영이 싹싹한 현주 군대에서 고생하는 덜렁이 태윤이

힘든 일도 성실히 해준 학희 모두에게 고마움을 전합니다 대장님의 풍모를 갖춰가는 친절

한 시내누나 질문할 때마다 명쾌한 답을 주셨던 유식형 유쾌한 남자 종훈형 aseptic life

형범형 인도 미소녀 한희 누나 멋진 유석형 든든한 재민형 항상 웃는 아람누나 모두 함

께 있던 시간들을 잊지 못할 듯 합니다

세종대의 철인 상국이형과 노래방의 지배자 환이형에게도 감사의 마음을 전합니다 의

학공학교실의 젠틀맨 창용형 늘 편안히 대해주신 수찬형 저의 antibody이자 맑은 웃음의

기창형 술자리서 허심탄회하게 이야기할 수 있는 동기-재성형과 선희 과묵하지만 든든한

철이형께 감사드립니다 학사업무 및 랩 관련 서류들을 빈틈없이 챙겨주신 유정숙씨와 김

태화씨께도 감사합니다 임상센터에서 함께 일하면서 많은 것을 도와주셨던 학부 선배인

성재형 양띠 모임 경희 언혜 정형외과 김향 선생님 소아과 김설 선생님께도 고마움을 전

합니다

친누나 이상으로 챙겨주셨던 자인누나 정 많은 명아누나 상병형 도형형 용언님 보

영님 홍대를 함께 누비던 많은 분들이 생각납니다 그리고 무엇보다 즐거웠던 lt공중캠프gt

의 많은 분들께도 감사의 마음을 전하고 싶습니다 이장님 기영형 사당누나 정원형 시린

누나 만담콤비 촬스형과 벙구리형 물꼭누나 수테키 경민형 성훈형 성우형 형우 동희

가은 미선누나 우영 정우 지윤이 소현님 가을이 모두 제게 힘이 되어 주었습니다 행복

한 가장이 되신 일환형 업무에 바빠 선물도 안챙겨주는 미진씨 lt아락동gt의 용석형 상연

형도 고마웠습니다 명경지수의 많은 분들-잊지 못할 진이형님 호용형 은오형 종섭형 창

진형 광용 윤호 세곤 철중 경래 그리고 메신저에서 말벗이 되어준 생명공학과의 재휘누

나 대윤 희원 호선 형이에게도 감사의 마음을 전합니다 나름대로의 길을 찾아 맹진하고

있을 철환이와 태균이 자주 보진 못하지만 잘 챙겨주는 정현누나 모두에게 감사드립니다

그리고 이 자리에 제가 설 수 있기까지 누구보다 절 사랑해주신 아버지 어머니께 말

로는 일할도 표현치 못할 그 마음을 전하고 싶습니다 멀리 있어 자주 찾아뵙지도 못했지

만 저의 결정에 대해 무한한 신뢰를 보여주신 만큼 앞으로 그 보답을 할 수 있을지 아직

도 자신이 없습니다 무덤덤한 오빠를 꼼꼼히 챙겨준 하나뿐인 여동생 민정이에게도 정말

미안하고 고맙습니다

행복한 시지프를 꿈꾸며

2004년 1월

이 민 섭

- i -

CONTENTS

FIGURE LEGENDS ⅲ

TABLE LEGENDS ⅵ

ABBREVIATIONS ⅶ

ABSTRACT ⅷ

1 INTRODUCTION 1

11 Neural tissue engineering 1

12 Biodegradable poly(lactic-glycolic) acids (PLGA) 3

13 Extracellular matrix proteins 5

131 Laminin 7

132 Fibronectin 9

133 Collagen 9

14 Poly-D-lysine 10

15 Titanium dioxide coating 10

16 Objectives of this study 11

2 MATERIALS AND METHODS 12

21 Experimental procedure 12

22 Primary culture of rat cortical neural cells 14

23 Characterization of neural cells 16

231 Microscopy observation 16

232 Immunofluorescence observation 16

233 Scanning electron microscopy (SEM) observation 17

24 Preparation of PLGA film 18

25 ECM coating 19

26 TiO2 coating 20

261 X-ray photoelectron spectroscopy (XPS) analysis 22

- ii -

262 Water contact angle measurement 22

27 Cell viability assay 23

271 WST-8 assay 23

28 Statistical analysis 26

3 RESULTS 27

31 Characterization of rat cortical neural cells 27

311 Morphological observation of cultured cells 27

312 Immunofluorescence observation of cultured cells 27

32 Effects of ECM coated PLGA film to cortical neural cells 30

321 Assessment of cell viability on ECM coated PLGA film 30


323 SEM observation of cultured cells 31

33 Effects of TiO2 coated PLGA film to cortical neural cells 42

331 Surface composition of TiO2 coated PLGA film 42

332 Hydrophilicity of TiO2 coated PLGA film 42

333 Assessment of cell viability on TiO2 coated PLGA film 45



4 DISCUSSION 49

5 CONCLUSION 51

REFERENCES 52

국 문 요 약 59

- iii -

FIGURE LEGENDS

Figure 1 Structures of biodegradable polymer (PLA PGA PLGA) 4

Figure 2 Structural model of laminin Chains designated by Greek letters

domains by Roman numerals cys-rich rod domains in the short arms

are designated by symbols S the triple coiled-coil region (domain I

II) of the long arm by parallel straight lines In the β1 chain the α-

helical coiled-coil domains are interrupted by a small cys-rich domain

α Interchain disulfide bridges are indicated by thick bars Regions of

the molecule corresponding to fragments E1X 4 E8 and 3 are indi-

cated G1-G5 are domains within the terminal globule of the α1 chain

[32] 8

Figure 3 Experimental procedure of this study 13

Figure 4 Primary culture of rat cortical neural cells (A) Preparation of

embryos of E-16 SD rat (B) Separation of head and body (C)

Dissection of cerebrum without meninges (D) Micro-dissection of

cortex (E) Trituration of neural cells with pasteur pipet (F) Cultured

neural cells on PDL-LN coated coverslip (20X105 cellsml at 200X

magnification) 15

Figure 5 Structure of fabricated PLGA film coated with or without TiO2 18

Figure 6 Appearance and diagram of plasma equipment The working pressure

was maintained around 42 x 10-3 torr and the reactive gas of O2

was properly mixed with Ar for oxide deposition (Ar O2 = 50sccm

9sccm) To increase the hydrophilicity of surface TiO2 was deposited

on PLGA surface to the thickness of 25 nm by the reactive sputter

deposition under the temperature of 40 21

- iv -

Figure 7 Procedure of neural cell viability assay 24

Figure 8 Structures of WST-8 and WST-8 formazan 25

Figure 9 Phase contrast microscopy observation of cortical neural cells cultured

on coverslip coated with a mixture of PDL (50 μgml) and LN (100μ

gml) 28

Figure 10Immunofluorescence observation of cortical neural cells cultured on

coverslip coated with PDL+LN (50+100 μgml) at 200X magnification

(A) NF has a prominent somatodendritic localization and is present

within dendritic growth cones (green) (B) Neuron observed under

the filter for MAP-2 staining only (C) Double labelling of rat cortical

neural cells for NF and MAP-2 Sites of low MAP-2 staining in

dendritic growth correspond to locations of intense NF localization

In the cell body and some dendritic regions NF (green) and MAP-2

(red) colocalized as shown as yellow color 29

Figure 11Viability of rat cortical neural cells on PLGA films coated with

PDLECM after 4 and 8 days culture and mark indicate plt005

relative to non-coating condition at 4 days and 8 days culture

respectively The results shown are mean data from three

experiments and error bars indicate standard deviation 33


PLGA films coated with a mixture solution of PDL (10 μgml) and

LN (100 μgml) (A) and (B) were observed at 100X magnification

and (C) and (D) were observed at 400X magnification 34

Figure 13SEM photograph of cortical neural cells on PLGA films coated with

of without PDLLNFNCol and their mixture 35

Figure 14Surface composition of PLGA films coated with of without TiO2 43

Figure 15Hydrophilicity of uncoated PLGA film and TiO2 coated PLGA film

The contact angles were determined with direct optical image instead

- v -

of numerical values because the subtly warped thin PLGA film

during plasma treatment could not apply to contact angle analyzer

(A) control (B) 0 hr after coating (C) 12 hr after coating (D) 60 hr

after coating 44

Figure 16Viability of rat cortical neural cells on PLGA films with or without

coating TiO2 after 4 days culture mark indicate plt005 relative to

non-coating condition at 4 days The results shown are mean data

from three experiments and error bars indicate standard deviation

46


PLGA films coated with TiO2 after 4 days culture (A) and (B) were

observed at 100X magnification 47


of without TiO2 48

- vi -

TABLE LEGENDS

Table 1 Applied concentration ranges of ECM and poly-D-lysine 19

- vii -

ABBREVIATIONS

CNS Central nervous system

Col Collagen

ECM Extracellular matrix

FN Fibronectin

LN Laminin

MAP-2 Microtubule associated protein-2

NF Neurofilament

PBS Phosphate buffered saline

PDL Poly-D-lysine

PGA Poly glycolic acids

PLA Poly lactic acids

PLGA Poly (lactic glycolic) acids

SEM Scanning electron microscopy

XPS X-ray photoelectron spectroscopy

- viii -

ABSTRACT

The purpose of this study is to evaluate the viability of primary cultured

cortical neural cells from E 14 Sprague Dawley rat on surface modified PLGA

films

To characterize the primary cultured neural cells cultured cells were

analyzed the cellular morphology such as axon and dendrite outgrowth with

phase-contrast microscopy and identified with immunofluorescence observation

for NF and MAP-2 To modify the PLGA surface in biological aspect PLGA

films were coated with various concentrations and combinations of

poly-D-lysine(PDL) and extracellular matrix protein(ECM) such as laminin(LN)

fibronectin(FN) and collagen(Col) To modify the PLGA surface in

physicochemical aspect PLGA films were coated with TiO2 by plasma

magnetron sputtering method to increase the hydrophilicity of surface The

PLGA films used in this study were fabricated as non-porous to clarify the

interaction between ECM and PLGA surface After incubation for 4 and 8 days

the cultured neural cells on surface modified PLGA film were analyzed the cell

viability with CCK-8 assay and certified the cellular morphology and

differentiation with immunofluorescence and scanning electron microscopy(SEM)

observation

In cell viability test the cultured neural cells on PLGA coated with a

mixture of PDL and LN as well as PDL and FN increased 220 (plt005)

compared to those on uncoated PLGA With the exception of Col all kind of

coating condition enhanced the cell growth and proliferation proportion to

increase of coating density In TiO2 coating experiment the cell viability was

increased 20 compared to non-coating condition However the coating with

only PDL or TiO2 just enhanced the neural cells attachment ability and were

- ix -

not appropriate in cellular stationary growth and regular differentiation through

the immunofluorescence and SEM observation On the contrary cultured neural

cells on PLGA films coated with a mixture of PDL and LN as well as PDL

and FN indicated higher viability and stationary growth and regular

differentiation

Therefore these results suggest that this approaches could be helpful to

apply neural tissue engineering and to design an optimal in vitro culture

system for neural cells

Key Word Rat cerebral cortical neural cell Primary culture PLGA Cell

viability Extracellular matrix (ECM) Laminin Fibronectin Collagen TiO2

Plasma magnetron sputtering Hydrophilicity

- 1 -

1 INTRODUCTION

11 Neural tissue engineering

Every year thousands are disabled by neurological disease and injury

resulting in the loss of functioning neuronal circuits and regeneration failure

Several therapies offered significant promise for the restoration of neuronal

function including the use of growth factors to prevent cell death following

injury [1] stem cells to rebuild parts of the nervous system [2-3] and the use

of functional electrical stimulation to bypass CNS lesions [4-5] However

functional recovery following brain and spinal cord injuries and neuro-

degenerative diseases were likely to require the transplantation of exogenous

neural cells and tissues since the mammalian central nervous system had little

capacity for self-repair [6] Such attempts to replace lost or dysfunctional

neurons by means of cell or tissue transplantation or peripheral nerve grafting

have been intensely investigated for over a century [7] Biological interventions

for neural repair and motor recovery after brain and spinal cord injury may

involve strategies that replace cells or signaling molecules and stimulate the

regrowth of axons The fullest success of these interventions will depend upon

the functional incorporation of spared and new cells and their processes into

sensorimotor networks [8]

As an alternative to conventional grafting technologies a new approach is

being investigated which uses cell engineering-derived biomaterials to influence

function and differentiation of cultured or transplanted cells Neural tissue

engineering is an emerging field which derives from the combination of

various disciplines intracerebral grafting technologies advances in polymer

- 2 -

bioprocessing and surface chemistry that have been achieved during the last

decade and molecular mapping of cell-cell and cell-matrix interactions that

occur during development remodeling and regeneration of neural tissue The

ultimate goal of neural tissue engineering is to achieve appropriate biointer-

actions for a desired cell response [6]

In rebuilding damaged neuronal circuits in vivo it is not clear how much of

the normal anatomical architecture will have to be replaced to approximate

normal function Thus it is necessary to develope methods to promote neurite

extension toward potential targets and possibly to provoke some of these

neurons to migrate to specified locations for the reestablishment of the

neuronal circuitry Since the nervous system has an extremely complicated 3D

architecture in all of its anatomical subdivisions which 2D systems have been

considered not enough to replicate For example Hydrogel scaffoldings [9]

agarose gels [10] collagen gels [11] PLA porous scaffold [12] and PGA fibers

scaffold [3] were good candidates for building 3D substrate patterns for

neuronal culture However there are critical factors to consider when

immobilizing neural cells in 3D matrices including pore size and matrix

material properties such as gel density permeability and toxicity Therefore

non-porous 2D PLGA film was employed in vitro system to investigate how

extracellular cues accurately influence neuronal behavior and growth on

modified surface

- 3 -

12 Biodegradable Poly(Lactic-Glycolic)Acids (PLGA)

Tissue engineering has arisen to address the extreme shortage of tissues and

organs for transplantation and repair One of the most successful techniques

has been the seeding and culturing of cells on biodegradable scaffolds in vitro

followed by implantation in vivo [13-14] While matrices have been made from

a host of natural and synthetic materials there has been particular interest in

the biodegradable polymer of poly(glycolic acid) (PGA) poly(lactic acid) (PLA)

and their copolymers poly(lactic-co-glycolic acid) (PLGA) (figure 1) This

particular family of degradable esters is very attractive for tissue engineering

because the members are readily available and can be easily processed into a

variety of structures their degradation can be controlled through the ratio of

glycolic acid to lactic acid subunits and the polymers have been approved for

use in a number of application by FDA Furthermore recent research has

shown this family of polymers to be biocompatible in the brain and spinal

cord [15-16]

The first step in developing a polymer-cell construct is the identification of

the appropriate bulk and surface characteristics of the scaffold for the intended

application PGA PLA and PLGA all support the growth of neural stem cells

without surface modification However surface modification can further control

cellular behavior with respect to the surfaces and may afford an opportunity

to achieve more than simple adhesion controlled differentiation migration and

axonal guidance There has been a great deal of work in the field of bio-

materials with development of complex patterned surface structures but on

simple level the surfaces of the degradable polyesters may be altered by the

simple adsorption of peptide such as polylysine and proteins such as laminin

- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n

Poly(lactic acid) (PLA) Poly(glycolic acid) (PGA)

CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m

Poly(lactic-co-glycolic acid) (PLGA)

Figure 1 Structures of biodegradable polymer (PLA PGA PLGA)

- 5 -

13 Extracellular matrix proteins

Although cells are the key players in the formation of new tissue they

cannot function without the appropriate extracellular matrix (ECM) For many

cell types including neurons it has been demonstrated that cell signal is

influenced by multiple factors such as soluble survivalgrowth factors signals

from cell-cell interactions and perhaps most importantly cell-ECM signals [17]

The matrix influences the cells and cells in turn modify the matrix thus

creating a constantly changing microenvironment throughout the remodeling

process The localized microenvironment in which a tissue develops must

support structural as well as temporal changes to facilitate the formation of

final cellular organization This is mediated by specific types and concentrations

of ECM molecules that appear at defined times to direct tissue development

repair and healing The ECM components are constantly remodeled degraded

and re-synthesized locally to provide the appropriate type and concentration of

proteins with respect to time [18]

The survival differentiation and extension of processes by neurons during

these treatments require the interaction of cells with biomaterials and their

extracellular environment There is wide variety of cell-cell adhesion and ECM

molecules that effect developing and injured neurons These can serve as

potential tools to direct the growth of recovering neurons or transplanted

precursors [19] These adhesion molecules work in conjunction with growth

factors to modulate neuron adhesion migration survival neurite outgrowth

differentiation polarity and axon regeneration [20-23]

The other factor to consider is cell attachment to the matrix which is

necessary for neural cell culture In vivo neurons are surrounded by ECM and

like most cells require adhesion to extracellular matrix for survival since the

- 6 -

most cell types are anchorage-dependent [24] Neurons in the brain usually

attach to the ECM for a proper growth function and survival When the

cell-ECM contacts are lost neural cells may undergo apoptosis a physiological

form of programmed cell death Adhesion dependence permits cell growth and

differentiation when the cell is in its correct environment within the organism

The importance of ECM components for cell culture has been demonstrated to

regulate programmed cell death and to promote neurite extension in 3D

immobilization scaffolds [25-26] The importance of cell attachment has been

demonstrated in several studies where neural cells were immobilized in agarose

gels that had been modified with ECM proteins [26-27] Those studies showed

that neurite outgrowth was significantly enhanced in the presence of ECM

components which promote cell adhesion

However there are many variables for the development of these devices

including not only the material to be used but also the geometry and spatial

distribution To move toward their clinical application researcher need

improved knowledge of the interactions of ECM proteins with neurons and

glia Therefore the primary objective of this study was to investigate the role

of three ECM components (laminin collagen and fibronectin) on the survival

and differentiation of rat cortical neurons grown on biodegradable PLGA film

- 7 -

131 Laminin

Laminin (LN) is a large (Mr=850000) multi-domained cross-shaped glyco-

protein that is organized in the meshwork of basement membranes such as

those lining epithelia surrounding blood vessels and nerves and underlying

pial sheaths of the brain [28] It also occurs in the ECM in sites other than

basement membranes at early stages of development and is localized to

specific types of neurons in the central nervous system (CNS) during both

embryonic and adult stages Synthesized and secreted by cells into their

extracellular environment laminin in turn interacts with receptors at cell

surfaces an interaction that results in changes in the behavior of cells such as

attachment to a substrate migration and neurite outgrowth during embryonic

development and regeneration

The multi-domain structure of laminin means that specific domains are

associated with distinct functions such as cell attachment promotion of neurite

outgrowth and binding to other glycoproteins or proteoglycans Within these

domains specific fragments of polypeptide sequences as few as several amino

acids have been identified with specific biological activity For example two

different peptide sequences IKVAV and LQVQLSIR within the E8 domain on

the long arm function in cell attachment and promote neurite outgrowth

(figure 2) [29-30]

During peripheral nerve regeneration laminin plays a role in maintaining a

scaffold within the basement membrane along which regenerating axons grow

Lamininrsquos role in mammalian central nervous system injury in controversial

although other vertebrates that do exhibit central regeneration have laminin

associated with astrocytes in the regenerating regions [31]

- 8 -

[Beck K et al Structure and function of laminin anatomy of a multidomain

glycoprotein 1990]


domains by Roman numerals cys-rich rod domains in the short arms are

designated by symbols S the triple coiled-coil region (domain I II) of the long

arm by parallel straight lines In the β1 chain the α-helical coiled-coil domains

are interrupted by a small cys-rich domain α Interchain disulfide bridges are

indicated by thick bars Regions of the molecule corresponding to fragments

E1X 4 E8 and 3 are indicated G1-G5 are domains within the terminal

globule of the α1 chain [32]

- 9 -

132 Fibronectin

Fibronectin (FN) is involved in many cellular processes including tissue

repair embryogenesis blood clotting and cell migrationadhesion Fibronectin

sometimes serves as a general cell adhesion molecule by anchoring cells to

collagen or proteoglycan substrates FN also can serve to organize cellular

interaction with the ECM by binding to different components of the

extracellular matrix and to membrane-bound FN receptors on cell surfaces [33]

Fibronectin is not present in high levels in the adult animal however

fibronectin knockout animals demonstrate neural tube abnormalities that are

embryonic lethal [34] During peripheral nerve regeneration fibronectin plays a

role as neurite-promoting factors [35-36] Furthermore primary neural stem

cells injected into the traumatically injured mouse brain within a FN-based

scaffold (collagen IFN gel) showed increased survival and migration at 3

weeks relative to injection of cells alone [37]

133 Collagen

Collagen (Col) is major component of the ECM and facilitate integrate and

maintain the integrity of a wide variety of tissues It serves as structural

scaffolds of tissue architecture uniting cells and other biomolecules It also

known to promote cellular attachment and growth [38] When artificial

materials are inserted in our bodies they must have strong interaction with

collagen comprised in soft tissue Therefore the artificial materials whose

surface is modified with collagen should easily adhere to soft tissue Thus

various collagen-polymer composites have been applied for peripheral nerve

regeneration [39-40] Alignment of collagen and laminin gels within a silicone

- 10 -

tube increased the success and the quality of regeneration in long nerve gaps

The combination of an aligned matrix with embedded Schwann cells should be

considered in further steps for the development of an artificial nerve graft for

clinical application [41-42] For this reason collagen was coated onto PLGA

films to immobilize primary neural cells

14 Poly-D-lysine

Poly-D-lysine (PDL) is a synthetic molecule used as a thin coating to

enhance the attachment of cells to plastic and glass surfaces It has been used

to culture a wide variety of cell types including neurons

15 Titanium dioxide coating

The efficacy of different titanium dioxide materials on cell growth and

distribution has been studied [43] In this study in order to improve PLGA

surfacecells interaction PLGA surface was modified with TiO2 using

magnetron sputtering method A magnetron sputtering is the most widely

method used for vacuum thin film deposition The changes of their surface

properties have been characterized by means of contact angle measurement

X-ray photoelectron spectroscopy (XPS) For the assessment of cell viability

WST-8 assay and scanning electron microscopy (SEM) observation were carried

out

- 11 -

16 Objectives of this study

To approach toward understanding the interaction of neural cells with the

ECM this study concentrated to examine neural cells behavior (viability and

differentiation) on two-dimensional biodegradable PLGA environments that are

built step-by-step with biologically defined ECM molecules Since neural cells

are known to be strongly affected by the properties of culture substrates

[44-45] the possibility of improving the viability of neural cells was

investigated through PLGA culture with surface modification as coating with a

variety of substrates that have been widely used in neural cultures including

poly-D-lysine laminin fibronectin collagen Furthermore in order to improve

PLGA surfacecells interaction PLGA surface was modified with TiO2 using

magnetron sputtering method These modified PLGA surfaces were compared

with non-coated PLGA film for their effects on the viability and growth and

proliferation of rat cortical neural cells The effect of coating density was also

examined This approaches could be useful to design an optimal culture system

for producing neural transplants

- 12 -

2 MATERIALS AND METHODS

21 Experimental procedure

The purpose of this study was to compare the viability of rat cortical

neural cells on surface modified various PLGA films which was mainly

evaluated by neural cell attachment growth and differentiation in this work

Experimental procedure of this study was briefly represented as figure 3 First

of all rat cortical neural cells were primary cultured and characterized by

morphological and immunobichemical observation In the second place surface

modified PLGA films 6535 composite ratio were prepared by titanium

dioxide deposition and PDL-ECM molecules coating TiO2 deposited surface

was characterized with contact angle measurement and XPS For 4 and 8 day

incubation after neural cells seeding cultured cells viability was investigated

and compared with WST-8 assay and SEM observation

- 13 -

TiOTiO22 or ECM coatingor ECM coating

NonNon--porous PLGA filmporous PLGA film

Neural cells seedingNeural cells seeding

Contact angle

measurementXPS analysis Cell viability

assay Imuno-

fluorescence analysis

SEM analysis

PLGA filmPLGA 6535

Treament withChloroform

Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying

Figure 3 Experimental procedure of this study

- 14 -

22 Primary culture of rat cortical neural cells

Pregnant rats embryonic day 14~16 days were purchased from

Daehan-Biolink in Korea Dissociated rat cortical cells were prepared by

digestion of embryonic- day-14~16 rat (Sprague-Dawley) whole cortices as

described previously [46] The cerebral cortex was microdissected free of

meninges and dissociated in Hanks Balanced Salt Solution (Gibco BRL

Gaithersburg MD USA) containing 1 gl glucose (Sigma USA G-5767) 1 mM

pyruvate 1 mM NaHCO3 and 10 mM hepes and triturated with a

fire-polished pasteur pipet Dissociated cortical cells were grown in Neurobasal

medium with 2 wv B-27 supplement (both from Gibco) 05 mM L-glutamine

(Sigma G-5763) 25 μM glutamate 100 μgml streptomycin and 100 Uml

penicillin G In the case of immunofluorescence analysis 200 μl of a neural cell

suspension containing 10X105 cellsml was plated onto ECM or TiO2-coated

and uncoated PLGA film in 96 well plates (Falcon Becton dickinson labware

NJ USA) However in the cases of WST-8 assay and SEM observation the

density of cells was more dense as 20X106 cellsml The cells were incubated

at 37 degC in a humidified atmosphere of 5 CO2 for 4 and 8 days (figure 4)

Cell media was exchanged every 4 days with half volume

- 15 -


embryos of E-16 SD rat (B) Separation of head and body (C) Dissection of

cerebrum without meninges (D) Micro-dissection of cortex (E) Trituration of

neural cells with pasteur pipet (F) Cultured neural cells on PDL-LN coated

coverslip (20X105 cellsml at 200X magnification)

- 16 -

23 Characterization of neural cells

To characterize the cultured cells after 4 days in vitro cultured cells on

coverslip coated with poly-D-lysine (50 μgml) and laminin (100 μgml) were

observed with phase-contrast microscopy and fluorescence microscopy The

characterization of neural cells were verified based on the expression of MAP-2

and neurofilament

231 Microscopy observation

Cell morphology was monitored with a phase-contrast microscope (Olympus

Optical Co Tokyo Japan) Images of the cultures were captured with

Olympus DP-12 digital CCD camera

232 Immunofluorescence observation

To assess the development of cortical neurons on PLGA films double

immunostaining for axonal filament marker Neurofilament (145 kD) and for

dendritic marker MAP-2 was carried out in cultures MAP-2 is a

well-established marker for the identification of neuronal cell bodies and

dendrites [47] Neurofilament (NF) is the intermediate filaments of neurons and

their processes For immunocytochemistry cultures were fixed with 35

paraformaldehyde in 01 M phosphate buffer (pH 70) for 10~15 min at room

temperature and rinsed in PBS To permeate the cellular membranes cells on

PLGA films were exposed to 01 Triton X-100 (Sigma T-9284) for 10 min at

room temperature and rinsed in PBS twice Then the cells were incubated with

- 17 -

1 bovine serum albumin in PBS for 30 min at room temperature to block

non-specific antibody binding The cells were subsequently incubated with a

mixture of rabbit polyclonal anti-Neurofilament M(145 kD) (1200) (Chemicon

Temecula CA USA) and mouse monoclonal anti-MAP-2 (1200) (Chemicon

Temecula CA USA) was treated for 1 hr at room temperature The cells were

incubated for 40~60 min at room temperature with a mixture of fluorescein

anti-rabbit IgG and texas red anti-mouse IgG (1250) (Vector Laboratories

Burlingame CA USA) After rinses in PBS cultures were photographed using

fluorescent microscope (Olympus BX60 Olympus Optical Co Tokyo Japan)

233 Scanning electron microscopy (SEM) observation

The seeded neural celllsquos density and morphology on surface modified PLGA

films were characterized by scanning electron microscope (S-800 HITACHI

Tokyo Japan) SEM is one of the best way to characterize both the density

and nature of neural cells on opaque PLGA film With SEM one can see cell

attachment and spreading as well as process extension The films were washed

with 01 M PBS (pH 74) to remove unattached cells The cells were fixed with

25 glutaraldehyde solution overnight at 4 and dehydrated with a series

of increasing concentration of ethanol solution The films were vacuum dried

and coated by ultra-thin layer (300 Å) of goldpt in an ion sputter (E-1010

HITACHI Tokyo Japan) An image analyzer program (Escan 4000 Bum-Mi

Universe Co Ltd Ansan korea) was used to captured the images of cells and

modified surfaces

- 18 -

24 Preparation of PLGA film

The biodegradable PLGA thin film used in this study was fabricated as

non-porous 5 mm in diameter 05 mm in thickness and 6535 composite

ratios (figure 5) For accurate analysis about the properties of modified surface

PLGA films used in this study were fabricated as non-porous structure The

6535 lactideglycolide copolymer degrades in about 3 months [48] PLGA films

were sterilized in 70 ethanol for 2 hrs washed two times with PBS and

finally stored under sterile conditions To prevent the PLGA films from boating

in media-submerged 96 well plate autoclaved vacuum greese was previously

attached between PLGA film and well plate bottom The autoclaved vacuum

greese was confirmed to do not have any cytotoxicity in cell culture in vitro

(Data not shown)

AA BB A control PLGA B TiO2 coated PLGA

Figure 5 Structure of fabricated PLGA film coated with or without TiO2

- 19 -

25 ECM coating

In this study the three kinds of ECM were employed including collagen

(COL) (Type I atelocollagen from calf skin Seoul Korea) laminin (LN) (Sigma

USA) fibronectin (FN) (Sigma USA) and Poly-D-lysine (PDL) (Sigma USA)

The applied concentrations of each or combined ECM were PDL (01 1 10 μ

gml) COL (100 μgml) LN (100 μgml) FN (100 μgml) PDL+COL (01+100

10+100 μgml) PDL+LN (01+100 10+100 μgml) PDL+FN (01+100 10+100 μ

gml) (table 1) In previous study the effect of COL LN FN was not found

any difference in the range of concentration 10 to 100 μgml (Data not

shown) For ECM coatings Each PLGA film was placed in 96 well plates and

soaked in 50 μl of various concentration of ECM for 2 hr at 37 degC incubator

Then the supernatant of ECM on PLGA films were aspirated and washed 2

times with fresh PBS

Table 1 Applied concentration ranges of ECM and poly-D-lysine

Extracellular Matrix Proteins

LN (100 ) FN (100 ) Col (100 ) Non-coating

PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating

The PLGA films were treated on one side with TiO2 using magnetron

sputtering method in a plasma reactor (figure 6) Magnetron sputtering is most

widely used method for vacuum thin film deposition The films were placed in

the plasma reactor chamber which was evacuated to 10x10-5 Torr The chamber

was then filled with oxygen and argon The pressure was raised to the

processing pressure (42x10-3 Torr) Once the processing pressure was reached

the generator was activated at 11 kW for 20 min under 40˚C TiO2 was

deposited on the PLGA film to the thickness of 25 nm by the reactive sputter

deposition At the end of this exposure the PLGA films were stored in a

desiccator until they were used

- 21 -

(A) Plasma equipment (Outside) (B) Plasma equipment (Inside)

Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+

Optimal gas pressure

Ionization of targetelectrons

Deposition of atomsonto sample

Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+




(C) Plasma equipment diagram


was maintained around 42 x 10-3 torr and the reactive gas of O2 was properly

mixed with Ar for oxide deposition (Ar O2 = 50sccm 9sccm) To increase

the hydrophilicity of surface TiO2 was deposited on PLGA surface to the

thickness of 25 nm by the reactive sputter deposition under the temperature of

40

- 22 -

261 X-ray photoelectron spectroscopy (XPS) analysis

The surface chemical composition of the deposited layers was determined

using an X-ray photoelectron spectroscopy Samples were cleaned by ultrasonic

vibration in ethanol and air dried prior to analysis Wide scan spectra in the

12000 eV binding energy range wererecorded with a pass energy of 50 eV for

all samples Spectra were corrected for peak shifting due to sample charging

during data acquisition by setting the main component of the C 1s line to a

value generally accepted for carbon contamination (2850 eV)

262 Water contact angle measurement

To certify the increase of hydrophilicity of TiO2-coated PLGA films the

surfaces of PLGA with or without coating were characterized by static water

contact angle measurements using the sessile drop method Forthe sessile drop

measurement a water droplet of approximately 10 μl was placed on the dry

surfaces The contact angles were determined with direct optical images instead

of numerical values because the thin PLGA film had warped subtly during

plasma treatment At least 10 measurements of different water droplets on at

least three different locations were considered to determined the hydrophilicity

- 23 -

27 Cell viability assay

This study compared the number of viable neural cells and estimated their

proliferation activity on PLGA film with or without coating The number of

viable cells was quantified indirectly by WST-8 assay (Cell Counting Kit-8

Dojindo Lab Japan) To assess the outgrowth of nuerite and formation of

neural network the cultured cells were observed with immunofluorescence and

SEM observation (figure 7)

271 WST-8 assay

The Cell Counting Kit-8 contained WST-8 (2-(2-methoxy-4-nitrophenyl)-3-

(4-nitrophenyl)-5-(24-disulfophenyl)-2H-tetrazolium monosodium salt) a water-

soluble tetrazolium salt which is reduced by dehydrogenase activities of viable

cells to produce a yellow color formazan dye (figure 8) The total dehydro-

genase activity of viable cells in the medium can be determined by the

intensity of the yellow color It found that the numbers of viable neural

stemprogenitor cells to be directly proportional to the metabolic reaction

products obtained in the WST-8 [49]

After incubating the cells in 96 well plate at 37degC in 5 CO2 -95 air for 4

and 8 days the WST-8 assay were carried out according to the manufacturerrsquos

instructions Briefly 20μl of the Cell Counting Kit solution was applied to each

well in the assay plate and incubated for 4 hr at 37degC The absorbance was

measured at 570 nm by an ELISA reader (Spectramax 340 Molecular devices

Co Sunnyvale CA USA)

- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell

WST-8 solutionIsolation of cortexneural cells

Neural cell platingInsert each wellAdd WST-8 solution

Cell cultured onPLGA

SEM analysis

PLGA filmWST-8 assay

Immuno-fluorescenceanalysis

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Figure 7 Procedure of neural cell viability assay

- 25 -

Figure 8 Structures of WST-8 and WST-8 formazan

- 26 -

28 Statistical analysis

All results were analyzed using the Students t-test (Excel 2002 Microsoft

WA USA) and expressed as meanplusmnthe standard deviation A value of plt005

was accepted as statistically significant

- 27 -

3 RESULTS

31 Characterization of rat cortical neural cells

The cultures were characterized morphologically and immunochemically

311 Morphological observation of cultured cells

A phase contrast image of cortical neural cell on a glass coverslip coated

with poly-D-lysine and laminin on day 4 after cell plating was observed as

well established neural network (figure 9)

312 Immunofluorescence observation of cultured cells

A immunofluorescence observation of cortical neural cell on a glass

coverslip coated with poly-D-lysine and laminin on day 4 after cell plating was

showed cultured cells were MAP-2 and NF positive and constituted a neutire

organization (figure 10) Immunoblotting for specific dendritic and neuronal cell

bodys proteins verified their enrichment MAP-2 immunopositive cells were

prevalent in this cortical neuron culture system (figure 10-B) verifying the

predominance of neurons in this cell culture

- 28 -

X200

X400


on coverslip coated with a mixture of PDL (50 μgml) and LN (100 μgml)

- 29 -

(A) Neuro Filament (FITC) (B) MAP-2 (Texas Red)

(C) Dual image

Figure 10 Immunofluorescence observation of cortical neural cells cultured on

coverslip coated with PDL+LN (50+100 μgml) at 200X magnification (A) NF

has a prominent somatodendritic localization and is present within dendritic

growth cones (green) (B) Neuron observed under the filter for MAP-2 staining

only (C) Double labelling of rat cortical neural cells for NF and MAP-2 Sites

of low MAP-2 staining in dendritic growth correspond to locations of intense

NF localization In the cell body and some dendritic regions NF (green) and

MAP-2 (red) colocalized as shown as yellow color

- 30 -

32 Effects of ECM coated PLGA film to cortical neural

cells

321 Assessment of cell viability on ECM coated PLGA film

To investigate the interplay between the ECM and neural cells primary

cultured cortical neural cells from E 14 Sprague Dawley rats were plated onto

PLGA films coated with various substrates Figure 11 shows the results of the

WST-8 assay experiment of rat cortical neural cells cultured for 4 and 8 days

respectively on all kind of ECM coated PLGA films The amount of neural

cells on PLGA film are shown as percentage of control non-coated PLGA film

The cell attachment on various substrate was different in each culture duration

In 4 days culture PDL LN FN coating could not show any effects to neural

cells attachment on PLGA films However in 8 days culture and PDL LN FN

coating showed an opposite results in this case only FN could not increase

the viability of neural cell

To examine if further improvement could be attained at higher coating

density PLGA surfaces with PDL coated at 01 1 10 μgml and with a

PDL-ECM coated at 01 10 μgml were tested As shown in figure 11 only

PDL coating also increased the cell attachment and spreading in proportion to

the coating density Especially in 8 days culture number of attached cells

under PDL 10 μgml condition showed an increase of 65 over that of PDL

01 μgml condition Unexpected results for neuronal growth on untreated

surfaces of PLGA to the growth achieved with either PDL alone or in

combination with the ECM proteins LN FN Col Although PDL-LN substrates

on glass coverslips are routinely used to generate the maximum response for

neurons growing in culture as on PLGA films PDL-FN showed the highest

- 31 -

neural cell viability after 8 days in vitro

In the cases of mixing with PDL-LN PDL-FN PDL-col higher PDL density

promoted more increase of neural cell attachment and proliferation with the

exception of PDL-col treatment

322 Immunoflurescence observation of cultured cells

A immunofluorescence observation of cortical neural cell on a PLGA films

coated with poly-D-lysine and laminin on day 4 after cell plating was showed

cultured cells were MAP-2 and NF positive and constituted a neurite

organization (figure 12)

At low level magnification observation cultured neural cells were clustered

and constituted axon and dendrite outgrowth only in boundary of clusters

These phenomenon were caused by high density cell plating on limited space

At high level magnification observation however bipolar shaped neural cells

were detected on coated PLGA films

323 SEM observation of cultured cells

Figure 13 shows neural cells cultured for 4 days on PLGA films coated

with either PDL alone or in combination with the ECM proteins LN FN Col

The photographs of 4 days culture reflect the status of neural cell attachment

and spreading at 100X magnification as well as the conditions of neural cell

growth at 1000X magnification

In images of 100X magnification cultured neural cells aggregated and form

several clusters in PDL or ECM protein coated substrates On the other hand

cells on non-coated PLGA film were hardly detected and could not form a

- 32 -

cluster These results implicate the 2D PLGA hydrophobic surface is not

efficient to support neural cell attachment and adhesive molecules such as

PDL and ECM assist the cell attachment to hydrophobic surface even in

cell-cell adhesion

In images of higher magnification PLGA surface coating with mixtures of

PDL versus LN (figure 13H-I) or FN (figure 13 J-K) had a much higher ratio

of bipolar-shaped cells than others indicating their greater ability to promote

neural cell spreading than others In these conditions observed cells had

smooth round to oval somata and neurites that appeared uniform in diameter

and smooth in appearance The number of flat cells on LN and FN-coated

PLGA was even less than that on non-coated and col-coated PLGA showing

that neural cell attachment and differentiation was less efficient on non-coated

and Col-coated PLGA In these inadequate conditions observed cells had

rough swollen and vacuolated somata and irregular fragmented or beaded

neurites (1000X ~2000X magnification) Therefore compared with WST-8 assay

PDL itself roles just in initial phase cell attachment but not in cell proliferation

and differentiation and maintaining of proper cellular state However PDL

enhance the effect of LN and FN coating as well as intercellular adhesion

In morphologically observation with SEM images PDL and ECM proteins

effect on neurite outgrowth were concentration dependent In the cases of

mixing with higher dose of PDL versus LN or FN higher dose of PDL (10 μ

gml) enhances significantly more neurite outgrowth than lower dose of PDL

(01 μgml)

- 33 -

Neural cells on modified PLGA films (1X106 cellsml)

0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days

Coating density (microgml)


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days4 days8 days


Figure 11 Viability of rat cortical neural cells on PLGA films coated with

PDLECM after 4 and 8 days culture and mark indicate plt005 relative

to non-coating condition at 4 days and 8 days culture respectively The results

shown are mean data from three experiments and error bars indicate standard

deviation

- 34 -

(A) Neuro Filament (B) MAP-2

(C) Neuro Filament (D) MAP-2


PLGA films coated with a mixture solution of PDL (10 μgml) and LN (100 μ

gml) (A) and (B) were observed at 100X magnification and (C) and (D) were

observed at 400X magnification

- 35 -

(A) SEM of cortical neural cells on uncoated PLGA film

Figure 13 SEM photograph of cortical neural cells on PLGA films coated with

of without PDLLNFNCol and their mixture

- 36 -

(B) SEM of cortical neural cells on poly-D-lysine(01 μgml) coated PLGA film

(C) SEM of cortical neural cells on poly-D-lysine(1 μgml) coated PLGA film

(Figure 13 continued)

- 37 -

(D) SEM of cortical neural cells on poly-D-lysine(10 μgml) coated PLGA film

(E) SEM of cortical neural cells on laminin(100 μgml) coated PLGA film


- 38 -

(F) SEM of cortical neural cells on fibronectin(100 μgml) coated PLGA film

(G) SEM of cortical neural cells on collagen(100 μgml) coated PLGA film


- 39 -

(H) SEM of cortical neural cells on PDL(01 μgml)

and LN(100 μgml) coated PLGA film

(I) SEM of cortical neural cells on PDL(10 μgml)



- 40 -

(J) SEM of cortical neural cells on PDL(01 μgml)

and FN(100 μgml) coated PLGA film

(K) SEM of cortical neural cells on PDL(10 μgml)



- 41 -

(L) SEM of cortical neural cells on PDL(01 μgml)

and Col(100 μgml) coated PLGA film

(M) SEM of cortical neural cells on PDL(10 μgml)


- 42 -

33 Effects of TiO2 coated PLGA film to cortical neural

cells

331 Surface composition of TiO2 coated PLGA film

XPS analysis of the surface of uncoated PLGA and TiO2 coated PLGA

showed clear differences in the interstitial O content (figure 14) Analysis of

the O 1s high-resolution spectra revealed that on the TiO2 coated PLGA

surface oxygen was predominantly in the form of interstitial oxygen double the

amount present at the surface of the uncoated PLGA XPS analysis also

showed that TiO2 coated PLGA surface was oxidized by titanium On the other

hand the uncoated PLGA showed just the peak of C 1s line relatively

332 Hydrophilicity of TiO2 coated PLGA film

Titanium dioxide has received much attention for good hydrophilic

properties of its surface The water contact angle was measured on the

TiO2-coated films and uncoated films respectively It could be seen that the

surface hydrophilicity of the PLGA films was improved by TiO2 deposition

with magnetron sputtering method (figure 6)

It has been reported there were some defects that plasma treatment was

utilized as the sole method to modify the materials because the hydrophilicity

of materials would decrease with preserving time owing to the surface mobility

[50] In my study the stability of TiO2 coating on the PLGA films was

measured for 60 hr after coating and the TiO2-coated PLGA films maintained

their hydrophilicity for 60 hr (Figure 15)

- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

Figure 14 Surface composition of PLGA films coated with of without TiO2

- 44 -

AA BB

CC DD

Figure 15 Hydrophilicity of uncoated PLGA film and TiO2 coated PLGA film

The contact angles were determined with direct optical images instead of

numerical values because the subtly warped thin PLGA film during plasma

treatment could not apply to contact angle analyzer (A) control (B) 0 hr after

coating (C) 12 hr after coating (D) 60 hr after coating

- 45 -

333 Assessment of cell viability on TiO2 coated PLGA film

Rat cortical neural cells were deposited onto PLGA thin films with or

without TiO2 coating and cultured for 4 days The metabolic activity of cortical

neural cells was monitored after 4 days of incubation with WST-8 assay Cell

viability was higher on the TiO2 coated PLGA film than uncoated one (figure

12) Since hydrophilicity was supposed to support cell adhesion and

proliferation these results correlated well with the physical characteristics of

film surfaces



coated with TiO2 on day 4 after cell plating was showed cultured cells were

MAP-2 and NF positive and constituted a neurite organization (figure 17)

At 100X magnification observation cultured neural cells were clustered and

constituted axon and dendrite outgrowth only in boundary of clusters


In SEM photographs of cortical neural cells after 4 days of incubation the

cells were spread on both film surfaces as clustering to a large extent (Figure

18) But the higher number of clusters was attached to the modified surface

comparatively

- 46 -

Figure 16 Viability of rat cortical neural cells on PLGA films with or without


non-coating condition at 4 days The results shown are mean data from three

experiments and error bars indicate standard deviation

- 47 -

(A) Neuro Filament (FITC)

(B) MAP-2 (Texas Red)


PLGA films coated with TiO2 after 4 days culture (A) and (B) were observed

at 100X magnification

- 48 -

SEM of cortical neural cells on uncoated PLGA film

SEM of cortical neural cells on TiO2 coated PLGA film


of without TiO2

- 49 -

4 DISCUSSION

The purpose of this study is to evaluate the feasibility and usefulness of

surface modified PLGA with various ECM or titanium dioxide to apply neural

cell transplanting in neural tissue engineering fields This work is done because

other studies of primary cultured neurons against ECM proteins were only in

tissue culture plate Therefore this study is concentrated to clarify the effects of

various ECM molecules and their own concentration and TiO2 to coating for

cortical neuron cell extension on non-porous PLGA surface

Membranes with adequate permeation characteristics and excellent cell

adhesive properties are essential for the development of bioengineered tissues

and biohybrid organs [51] In this respect principally only a nanometer deep

region of the material is of importance as the cell-material interaction is

strongly influenced by the physicochemical properties of the surface Although

many efforts were already taken it is still not clear which properties may be

critical to the host responses [52] Several authors have already reported on

enhanced cell adhesion on hydrophilic surfaces [53-54] The presence of specific

functional groups [55-56] immobilized adhesive proteins [57-58] surface charge

[59] and morphology [60] were also found to be regulative factors

The results of neural cell attachment and spread may be explained by the

physicochemical properties of materials and the adsorption of the ECM

molecule There are two phases in the process of cell attachment The first is

nonspecific adsorption of cells on the materials mainly mediated by the

physicochemical interaction between cells and materials Material properties

such as hydrophilicity and positive surface charges contribute to this

physicochemical interaction Hydrophilicity could be obtained from the water

contact angles on the materials and the surface charges of all the materials

- 50 -

could be estimated from their components In this study PDL and TiO2

coating correspond to such hypothesis The second phase is the specific

adsorption of cells on the materials ECM molecules such as laminin and

fibronectin participate in the process of specific cell attachment and cell spread

After nonspecific adhesion which is mostly dependent on material properties

cells will secrete some ECM molecules which can adsorb onto the material

surface bind the integrins on the surface of normal neural cells and mediate

the specific interaction between cells and materials It observed that rat cortical

neural cells exhibited high growth potential on most of the ECM coating PLGA

films The only exception was observed with surface coated with collagen on

which cell proliferation was limited possibly due to insufficient cell attachment

It seems that laminin and fibronectin play an even more important role in

neural cell spreading than collagen It suggests that ECM adhesive proteins

play a more important role than material properties especially in cell spread

Cells receive important signals from ECM which play a critical role in cell

development and survival Due to the anchorage-dependence of neural cells for

growth and survival any disruption of cell attachment can lead to the

interruption of these signals causing stress to the cells and eventually leading

to their death Successful attachment is conducive to the later spreading

process Hence the results of the cell culture experiment may be explained

comprehensively by the material properties and the amount of adsorption of

ECM molecules on the materials

Functional rewiring of neuronal networks requires functional synaptic

formation Additionally functional neuronal networks immobilized in

biodegradable polymer scaffold have potential uses in applications ranging

from implantation for disease treatment [61] to providing active neuronal

networks for use in cell-based biosensors [62]

- 51 -

5 CONCLUSION

In this study the viability of primary cultured cortical neural cells from E

14 SD rat was evaluated on surface modified PLGA films The cultured cells

were preliminary characterized with phase-contrast microscopy and immunoflu-

orescence observation To modify the PLGA surface PLGA films were coated

with various concentrations and combinations of PDL and ECM or deposited

with TiO2 by magnetron sputtering method to increase the hydrophilicity of

surface After incubation for 4 and 8 days the cultured neural cells on surface

modified PLGA film were analyzed the cell viability with CCK-8 assay and

certified the cellular morphology and differentiation with immunofluorescence

and SEM observation












differentiation




- 52 -

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Pancrazio JJ Detection of physiologically active compounds using cell-based

biosensors Trends Biotechnol 200119304-309

- 59 -

국 문 요 약

표면개질된 PLGA 필름상에서 쥐 대뇌피질 신경세포의

생존률의 평가

연세대학교 대학원

생체공학협동과정

생체재료학 전공

이 민 섭

임신 14령의 쥐의 태아에서 대뇌피질 신경세포(rat cortical neural cell)를 초대

배양(Primary culture)하여 표면개질된 PLGA 필름상에서 생존률을 비교하 다

초대배양한 쥐 대뇌피질 신경세포의 확인은 위상차 현미경(Phase-contrast

microscopy)을 통해서 신경세포 특유의 형태 분석과 신경세포 특이 항체를 이용한

면역형광화학(Immunofluorescence)법으로 검증하 다 PLGA 필름은 각각 생물학

적 측면과 물리화학적 측면에서 개질되었다 생물학적 측면에서는 인공 세포부착

물질인 폴리라이신(poly-D-lysine)과 생체내 세포외기질(Extracellular matrix)에서

유래한 라미닌(laminin) 파이브로넥틴(fibronectin) 콜라겐(collagen)을 혼합별 농

도별로 PLGA 필름에 코팅하 고 물리화학적 측면에서는 재료 표면의 친수성을

증가시키기 위해 플라즈마를 이용한 TiO2 코팅을 시도하 다 이 연구에 사용된

PLGA 필름은 세포에 대한 표면개질의 향을 정확히 분석하기 위해서 비공성

(Non-porous) 표면을 가지도록 제조되었다

표면개질된 PLGA필름 상에서 4일 8일 동안 배양된 초대배양 신경세포는

WST-8 assay를 통해서 실제 생존률을 비교하고 면역형광화학법과 주사전자현미

경(SEM) 분석을 통해서 세포의 생장 상태 및 형태를 확인하 다

실제 실험 결과 초대배양된 쥐 신경세포는 폴리라이신과 라미닌 폴리라이신

과 파이브로넥틴을 각각 혼합 코팅한 PLGA 필름상에서 코팅하지 않은 PLGA 필

름상에서보다 통계학적으로 의미있는 (plt005) 220의 생존률 증가를 보여주었다

- 60 -

그리고 콜라겐을 제외한 모든 경우에서 코팅되는 농도에 비례해서 신경세포의 생

존률 또한 증가됨을 확인할 수 있었다 TiO2 코팅의 경우에는 대조군과 비교해서

20 가량의 생존률 증가를 확인하 다 그렇지만 면역형광화학법과 주사전자현미

경을 통한 분석 결과 폴라라이신 코팅과 TiO2 코팅은 단순히 뇌세포의 부착능력

만을 향상시킬 뿐 PLGA 필름 상에서 뇌세포의 안정화된 생장과 분화에는 적합

하지 않음이 밝혀졌다 반면 폴리라이신을 각각 라미닌이나 파이브로넥틴과 혼합

코팅한 PLGA 필름상에서 배양된 뇌세포는 높은 생존률을 보여줄 뿐만 아니라

안정화된 생장과 분화가 촉진되었음이 확인되었다

따라서 이러한 결과들은 실제로 손상된 뇌조직의 재생에 있어서 필요한 이식

용 세포의 대량 증식이나 직접 이식의 경우에 유용하게 활용될 수 있음을 제시하

고 있다

핵심 단어 대뇌피질 신경세포(Cerebral cortical neural cell) 초대배양(Primary

culture) PLGA 세포 생존률(Cell viability) 세포외기질(ECM) 라미닌(Laminin)

파이브로넥틴(Fibronectin) 콜라겐(Collagen) TiO2 친수성(Hydrophilicity)

CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS




32 Effects of ECM coated PLGA film to cortical neural cells




33 Effects of TiO_(2) coated PLGA film to cortical neural cells

331 Surface composition of TiO_(2) coated PLGA film

332 Hydrophilicity of TiO_(2) coated PLGA film

333 Assessment of cell viability on TiO_(2) coated PLGA film



4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약



PLGA films

A Master Thesis

Submitted to the Graduate Program in

Biomedical Engineering and the Graduate School of

Yonsei University in partial fulfillment of

the requirements for the degree of Master of Science in

Biomedical Engineering

Min Sub Lee

December 2003

감사의 글






























합니다

















2004년 1월

이 민 섭

- i -

CONTENTS

FIGURE LEGENDS ⅲ

TABLE LEGENDS ⅵ

ABBREVIATIONS ⅶ

ABSTRACT ⅷ

1 INTRODUCTION 1




131 Laminin 7

132 Fibronectin 9

133 Collagen 9

14 Poly-D-lysine 10











25 ECM coating 19

26 TiO2 coating 20


- ii -



271 WST-8 assay 23


3 RESULTS 27














4 DISCUSSION 49

5 CONCLUSION 51

REFERENCES 52

국 문 요 약 59

- iii -

FIGURE LEGENDS










[32] 8







magnification) 15








- iv -





gml) 28
























- v -




after coating 44





46





of without TiO2 48

- vi -

TABLE LEGENDS


- vii -

ABBREVIATIONS


Col Collagen


FN Fibronectin

LN Laminin


NF Neurofilament


PDL Poly-D-lysine






- viii -

ABSTRACT



films















observation








- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

감사의 글






























합니다

















2004년 1월

이 민 섭

- i -

CONTENTS

FIGURE LEGENDS ⅲ

TABLE LEGENDS ⅵ

ABBREVIATIONS ⅶ

ABSTRACT ⅷ

1 INTRODUCTION 1




131 Laminin 7

132 Fibronectin 9

133 Collagen 9

14 Poly-D-lysine 10











25 ECM coating 19

26 TiO2 coating 20


- ii -



271 WST-8 assay 23


3 RESULTS 27














4 DISCUSSION 49

5 CONCLUSION 51

REFERENCES 52

국 문 요 약 59

- iii -

FIGURE LEGENDS










[32] 8







magnification) 15








- iv -





gml) 28
























- v -




after coating 44





46





of without TiO2 48

- vi -

TABLE LEGENDS


- vii -

ABBREVIATIONS


Col Collagen


FN Fibronectin

LN Laminin


NF Neurofilament


PDL Poly-D-lysine






- viii -

ABSTRACT



films















observation








- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

















2004년 1월

이 민 섭

- i -

CONTENTS

FIGURE LEGENDS ⅲ

TABLE LEGENDS ⅵ

ABBREVIATIONS ⅶ

ABSTRACT ⅷ

1 INTRODUCTION 1




131 Laminin 7

132 Fibronectin 9

133 Collagen 9

14 Poly-D-lysine 10











25 ECM coating 19

26 TiO2 coating 20


- ii -



271 WST-8 assay 23


3 RESULTS 27














4 DISCUSSION 49

5 CONCLUSION 51

REFERENCES 52

국 문 요 약 59

- iii -

FIGURE LEGENDS










[32] 8







magnification) 15








- iv -





gml) 28
























- v -




after coating 44





46





of without TiO2 48

- vi -

TABLE LEGENDS


- vii -

ABBREVIATIONS


Col Collagen


FN Fibronectin

LN Laminin


NF Neurofilament


PDL Poly-D-lysine






- viii -

ABSTRACT



films















observation








- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- i -

CONTENTS

FIGURE LEGENDS ⅲ

TABLE LEGENDS ⅵ

ABBREVIATIONS ⅶ

ABSTRACT ⅷ

1 INTRODUCTION 1




131 Laminin 7

132 Fibronectin 9

133 Collagen 9

14 Poly-D-lysine 10











25 ECM coating 19

26 TiO2 coating 20


- ii -



271 WST-8 assay 23


3 RESULTS 27














4 DISCUSSION 49

5 CONCLUSION 51

REFERENCES 52

국 문 요 약 59

- iii -

FIGURE LEGENDS










[32] 8







magnification) 15








- iv -





gml) 28
























- v -




after coating 44





46





of without TiO2 48

- vi -

TABLE LEGENDS


- vii -

ABBREVIATIONS


Col Collagen


FN Fibronectin

LN Laminin


NF Neurofilament


PDL Poly-D-lysine






- viii -

ABSTRACT



films















observation








- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- ii -



271 WST-8 assay 23


3 RESULTS 27














4 DISCUSSION 49

5 CONCLUSION 51

REFERENCES 52

국 문 요 약 59

- iii -

FIGURE LEGENDS










[32] 8







magnification) 15








- iv -





gml) 28
























- v -




after coating 44





46





of without TiO2 48

- vi -

TABLE LEGENDS


- vii -

ABBREVIATIONS


Col Collagen


FN Fibronectin

LN Laminin


NF Neurofilament


PDL Poly-D-lysine






- viii -

ABSTRACT



films















observation








- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- iii -

FIGURE LEGENDS










[32] 8







magnification) 15








- iv -





gml) 28
























- v -




after coating 44





46





of without TiO2 48

- vi -

TABLE LEGENDS


- vii -

ABBREVIATIONS


Col Collagen


FN Fibronectin

LN Laminin


NF Neurofilament


PDL Poly-D-lysine






- viii -

ABSTRACT



films















observation








- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- iv -





gml) 28
























- v -




after coating 44





46





of without TiO2 48

- vi -

TABLE LEGENDS


- vii -

ABBREVIATIONS


Col Collagen


FN Fibronectin

LN Laminin


NF Neurofilament


PDL Poly-D-lysine






- viii -

ABSTRACT



films















observation








- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- v -




after coating 44





46





of without TiO2 48

- vi -

TABLE LEGENDS


- vii -

ABBREVIATIONS


Col Collagen


FN Fibronectin

LN Laminin


NF Neurofilament


PDL Poly-D-lysine






- viii -

ABSTRACT



films















observation








- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- vi -

TABLE LEGENDS


- vii -

ABBREVIATIONS


Col Collagen


FN Fibronectin

LN Laminin


NF Neurofilament


PDL Poly-D-lysine






- viii -

ABSTRACT



films















observation








- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- vii -

ABBREVIATIONS


Col Collagen


FN Fibronectin

LN Laminin


NF Neurofilament


PDL Poly-D-lysine






- viii -

ABSTRACT



films















observation








- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- viii -

ABSTRACT



films















observation








- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- ix -





differentiation







- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 1 -

1 INTRODUCTION
























- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 2 -





















modified surface

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 3 -















cord [15-16]











- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 4 -

CH C O

CH3

O

n

CH C O

CH3

O

n

CH2 C O

O

n

CH2 C O

O

n


CH C O

CH3

O

n

CH2 C O

O

m

CH C O

CH3

O

n

CH2 C O

O

m



- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 5 -



























- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 6 -




















- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 7 -

131 Laminin



















(figure 2) [29-30]






- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 8 -


glycoprotein 1990]









- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 9 -

132 Fibronectin














133 Collagen










- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 10 -






14 Poly-D-lysine













out

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 11 -

















- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 12 -














- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 13 -




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying




Contact angle


assay Imuno-


SEM analysis

PLGA filmPLGA 6535


Vacuum drying


- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 14 -



















- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 15 -






- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 16 -






and neurofilament
















- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 17 -






















modified surfaces

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 18 -












(Data not shown)



- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 19 -

25 ECM coating
















PDL (01 )

(10 )

(100 )

Non-coating

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 20 -

26 TiO2 coating











- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 21 -


Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




Sample

Target(Ti)T C

Plasma

Gas

ArO2

TMP

CathodeTarget

Ar+




CathodeTarget

Ar+




CathodeTarget

Ar+










40

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 22 -


















- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 23 -








271 WST-8 assay















- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 24 -

Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis



Incubatefor 4 days

Incubatefor 4 hr

2X105 cellswell




SEM analysis




- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 25 -


- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 26 -





- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 27 -

3 RESULTS















- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 28 -

X200

X400



- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 29 -


(C) Dual image









- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 30 -


cells
























- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 31 -
























- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 32 -




cell-cell adhesion



















(01 μgml)

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 33 -


0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200

2504 days8 days



0Wellplate

Non-coating

PDL01

PDL1

PDL10

LN100

FN100

Col100

PDL01+

LN100

PDL10+

LN100

Perc

enta

ge o

f con

trol

PDL01+

FN100

PDL10+

FN100

PDL01+

Col100

PDL10+

Col100

50

100

150

200







deviation

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 34 -







- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 35 -




- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 36 -




- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 37 -




- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 38 -




- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 39 -






- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 40 -






- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 41 -





- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 42 -


cells





















- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 43 -

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA

1200 1000 800 600 400 200 0

N(E

)E

Binding Energy(ev)

Titanium oxide

Control

O 1s

Ti2p3

Ti2p1C 1s

OKLL

OKLL

TiLMMTiLMM

NKLL

Ti3sTi3p

coated PLGA


- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 44 -

AA BB

CC DD






- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 45 -








film surfaces











comparatively

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 46 -





- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 47 -






- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 48 -




of without TiO2

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 49 -

4 DISCUSSION


























- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 50 -




























- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 51 -

5 CONCLUSION










and SEM observation












differentiation




- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 52 -

REFERENCES




Nature 2000407963-970
















200013655-659


20011011869-1879



2001221077-1084

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 53 -



2003340169-172




1033-1040



151-161











Physiol 198345581-600






A37-A48


- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 54 -










1445-1449





Eng 19995291-304



Mater Res 199840392-400






J Biol Chem 198926416174-16182





- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 55 -




Bio 19946648-655






Neck Surg 1990103509-518







283-295



Raton FL 1995627~647









- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 56 -







2385-91






















- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 57 -

2607-2614























19991472479-485





- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 58 -





Science 2002297178-181




- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 59 -

국 문 요 약


생존률의 평가




이 민 섭



















- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

- 60 -











고 있다




CONTENTS

FIGURE LEGENDS

TABLE LEGENDS

ABBREVIATIONS

ABSTRACT

1 INTRODUCTION




131 Laminin

132 Fibronectin

133 Collagen

14 Poly-D-lysine











25 ECM coating

26 TiO_(2) coating




271 WST-8 assay


3 RESULTS














4 DISCUSSION

5 CONCLUSION

REFERENCES

국문요약

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