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UNIVERSITY OF HASANUDDIN

TNF- AND IL-10 LEVEL INRECURRENT APHTHOUS

STOMATITIS ULCER BY SWAB TECHNIC

Will submit to RDM&E USU Medan,

November 11-12 2011

This study aimed to examine validity of swab technic in cytokine level measurement

of RAS with promising result.


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TNF- AND IL-10 LEVEL INRECURRENT APHTHOUS

STOMATITIS ULCER BY SWAB TECHNIC

E.Marlina*, . Ha!i**, M. "#$ri*** Faculty of entistry! "niversity of #asanuddin! $akassar! South Sulawesi!

%&'()!ndonesia

** Faculty of entistry! "niversity of Airlangga! Surabaya! +ast ,ava! -&/'! ndonesia

0orresponding author 1 drg. +rni $arlina! Sp.2$! tel.13-'4&(5677'&/!/%'') #21

3-'67/6)))'68! e6mail1e.marlina9yahoo.co.id

A%$&ra'&

Recurrent Aphthous Stomatitis4RAS5 are among the most common problem

seen by clinician. The aetiology remain uncertain but possible predispotition factors

were systemic condition! hereditary! and autoimmun. Some study focused on cytokine

using invasive technic such as vena puncture for blood peripheral! and biopsy. :ut

others used swab mucosal to examine cytokine. This study aimed to examine validity

of swab technic in cytokine level measurement of RAS. The result by the paired T-

testshowed statistically significant level of T;F6< with p = &!&&- and >6& withp =

&!&( with each interval confidence %)?. Swab technic could use for cytokine

measurement for ulser and mucosal RAS patient.

In&r(!#'&i(n

Recurrent Aphthous Stomatitis4RAS5 are among the most common problem

seen by clinician. 4Field et al! '&&(@ reenberg! '&&/ 5. The aetiology remain

uncertain 4Field et al!'&&(@ Scully! '&&(@ RegeBy et al! '&&/5! but possible

predispotition factors were systemic condition! hereditary! and autoimmun. Some

study focused on T;F6 < and >6& as cytokine which plays a maCor role in

pathogenesis of RAS 4;atah! '&&(5. This studies use pheripheral blood mononuclear

cell as a cytokine source but this assumed that the cytokine production and

distribution not representative as local action rather than other stimulus. And it was an

invasive examination even though it was stabile 4:uno! %%7@ >in '&&)@ Damamoto!

%%(5. Some other study using exicional biopsy of RAS ulcer to compared cytokine

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with normal side. This technic representative enough for local production and

distribution even still invasive. 4;atah! '&&@ ;atah! '&&&5.

erber and partners 4'&&/5 study >6E gene polymorphisme using

vestibulum vaginal or buccal mucosal swab to examine cytokine roles in vulvar

vestibulitis syndrome. t could applied in RAS lesion because non invasive nature!

and because RAS ulcer is a superficial tissue necrosis with fibrinopurulent exudate

consisting of clotted fibrin! and numerous red blood cells forming haemorrhagic

focci. ;eutrophils and cellular debris cover the necrotic area. The epithelium is

infiltrated with variable numbers of intraepithelial lymhocytes and some neutrophils.

;eutrophils predominate in the immediate ulcerated area! although peripheral areas

surrounding the ulcer remain mononuclear in nature. 4>ehner %-%a@ $ills et al.

%7&@ Schroeder et al. %7/@ #yrinen6mmonen et al. %%5. ;eutrophil!

>ymphocytes! and fibrin clothing6contained pseudomembrane are sources for

cytokine. Swabbing this pseudomembrane could show T;F6< that found in burden

level! and low level of >6& as a basic examination to demonstrate the validity of

swabbing technic in cytokine level of RAS.

This study aimed to examine validity of swab technic in cytokine level

measurement of RAS.

Ma&)rial an! M)&(!)

All reccurent ulcer patients 4men5 age range from 7 to () visited to Gral

$edicine epartement of entistry Faculty of Airlangga "niversity asked to

participate in this study. 2articipant follow the anamnesis! clinical examination! and

full blood count to eliminate anemis condition! vitamin :' and folic acid deficiency.

2articipant with gastrointestinal or other systemic condition excluded. iagnosis of

RAS was made from clinical presentation which show discrete! painful! shallow!

recurrent ulcer! covered by a yellow6grey pseudomembrane and surrounded by an

erythenatous halo.

+ntry reHuirements were that the patient should have RA" ulser on non6

keratiniBed mucosal with at least / episode per year on average! RAS lesions studied

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were '6/ days old and located at simetrical sided such as the right buccal with left

buccal as normal mucosal. 2atients should have not systemic condition or other

immunodefisiensi! treated their current ulcer with any form of topical or systemic

medication.

The study protocolwas approved by The +thics 0ommittee of Faculty of

entistry! "niversity of Airlangga! Surabaya and informed cosent was obtained from

each participant of the study. Seven aphthous ulcers were swabs from 8 patients

RAS! all man 4mean age ' years old! ranging from % to '' years5. n addition!

specimens from the clinically unaffected area at the corresponding site opposite to the

ulcer were obtained from all patients to demonstrate cytokine diffrences between

clinically unaffected mucosa and RAS lesions. 0ottol roll sterile used to swab RAS

lesions and the opposite site! and cotton roll tip drawn on effendorf tube contain

$+$ and stored at 6'&o0 until +>SA tes done.

+>SA Iit Assay2ro 0at ;o. +T'&&6 used as Iit assay! and the result

measure in pikogram 4pH5. Gne Sample Iomlogorof Semirnov test used for sample

normality distribution. The means of the two groups were compared by using the

paired6T with considered statistically significant &!&) or less. S2SS /.) for windows

was used for all calculations.

R)$#l&

Iolmogorov Smirnov as normality distribution test used for each group and

showed normal distrbution withpJ &!88& for T;F6level of normal mucosal andp

= &!%%7 for T;F6ulser. >6& also showed normal distribution withp= &!/( for

>6& normal mucosal andp = &!7& for >6& ulcer.

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Ta%)l.TNF- an! IL-10 (n Recurrent Aphthous Stomatitis +RAS #l')r$ an!

n(ral #'($al RAS a&i)n$

S#%/)'& TNF-

IL-10

N #'($al +l #l')r +l N #'($al +l #l$)r +l

(& '%- 87 -)

' & '/& 77 -8

/ -' 7) 8- -)( 8& ')& 78 8

) ') '8& 77 8'

- & 7) %/ -)

8 & /7& ' %$ean 7/!7) 4S %&!-75 ')-!)8 4S -7!'5 &/ 4S (75 8( 4S %!-75

Table ! showed result of the paired T-test to compare T;F6 normal

mucosal dan T;F6 ulcers with statistically significant level of p = &!&&- and

interval confidence %)?. ifferent level of >6& normal mucosal and >6& RAS

ulcer showed significant level withp = &!&( and interval confidence %)?.

DISCUSSI2N

0ommonly! cytokine measurement using vena puncture technic for serum

specimen or both of Fine Needle Aspiration biopsy 4F;A:5 and exicional biopsy

technic 4>ewkowicB! '&&)@ ;atah!'&&&5. Gur knowledge and assay tecnology support

the measurement of cytokine for research and clinical approach 4#ill! '&&&@

GKorman! '&&75. Gne of this approach was swab technic using cotton strip sterile or

brush previously used for mucosal microorganism. This technic applied by a stroke of

mucosal / to ) times to carry microorganisme or cell exfoliation 4Spafford! '&&5.

This technic also used for cytokine gene polymorphisme on vaginal! gut and oral

mucosal 4reber et al! '&&)@ uimares! '&&85. Also for immunobloth analisi and

densitometry optica for cyclin biomarker expression change and as blood

examination alternative for them with contraindication or reCect to cytokine gene

polymorphisme for #odgkin >imfoma 4>an Hing! '&&-5. This approach also used to

collected servical cells and cervicovaginal mucous 4#ill! '&&&5! for urethral cytokine

examination 42ate! '&&5! cytokine of vaginal fluid 4agnew et al! '&&75! and the


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validity measuremen studied with comparing >6- cytokine concentration between

swab aplicator used by the professional dan by subCect and concluded that this technic

valid for both of them 4Faro et al! '&&-5.

Swab technic for mucosal organ could demonstrate cytokine level by

lymphosite! macrophage! and dendritic cell as cell source of cytokine which burden in

Mucosal associated lymphoid tissues 4$A>T5 covered oral mucosal.

n the current study we observed that swabs technic on oral mucosal could

show T;F6< and >6& level. Some sample show Bero value but it was a normality

fenomena because T;F6< commonly not found in epithelium. #owever this does not

mean that these cells are not producing and secreting this cytokine. t is ossible that

such cell types may contain insufficient T;F6< to be demonstrate by our assay

4;atah! '&&5.

T;F6< test in normal distribution with differences of T;F6< level between

normal mucosal and ulcers RAS significant atp = &!&'7. This value similiar to that

previously reported by ;atah!'&&/ and :uno %%7 which T;F6 significantly

increased in RAS. +levated levels of this cytokine related to the mechanism of

proinflamation cytokine as a respon for oral stimulus such as bacteri! minor trauma!chemical! or thermis as a triggering factor for RAS.

>6& level show expected value which decrease significantly withp= &!(8.

This previously reported by >ewkowicB '&&) studied this cytokine in 2:$0 of RAS

patients. This decrease possible because abnormality balance of cytokine profile as a

cause of RAS ulcers. And this decrease assumed as immune failure for suppress oral

inflamation reaction 4:uno!%%75.

n conclusion! swab technic could use to measure proinflamation and

antiinflamation cytokine especially T;F6and >6& level. Gur study need further to

continue as our result did not show specifity of this cytokine because of we do not

compare this result with 2:$0 or biopsy technic which relativly stabile and more

representative and with small sample siBe.

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AC3N2WLED4MENT

This research was supportd by South Sulawesi goverment and our gratefull to

2rof.Savage and ;atah S for the inspiring study and written. ;ot metion for $rs.

Rahayu at Tropical iseases 0entre of Airlangga "niversity that conduted all

laboratories procedurs.

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