marker (%) normal epcs mean ± sm (%) mmd epcs mean ± sm (%) cd3426.9 ± 11.531.0 ± 6.8 kdr27.3 ±...
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Marker (%)Normal EPCs
Mean ± SM (%)
MMD EPCs Mean
± SM (%)
CD34 26.9 ± 11.5 31.0 ± 6.8
KDR 27.3 ± 5.52 22.4 ± 8.90
CD133 24.5 ± 4.64 12.6 ± 3.93
CD31 99.0 ± 0.9 98.8 ± 0.5
CD45 0.42 ± 1.23 1.53 ± 3.12
CD34 KDR CD133 CD31B
(Scale bar : 50 μm)
CD31 vWF
C D
(Scale bar : 100 μm)
Dil-Ac-LDL uptake
Supplemental Figure I
Normal MMD
(Scale bar : 100 μm)
A 21 days
28 days 35 days
7 days CD45
Normal MMD0
10
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50
Nu
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ell)
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β-actin
RALDH2
-
(-)con50 nM
siRNA(A)
50 nM
siRNA(B)
50 nM
(-) control siRNA
Non treated 10 nM
30 nM 50 nM
A B
C D
RALDH2 siRNA (A) (nM)
RALDH2 siRNA (B) (nM)
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Negative control siRNA (nM)
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GAPDH
RALDH2
-(-)con50 nM
siRNA(A)
5 nM
siRNA(A)
10 nM
siRNA(A)
30 nM
siRNA(A)
50 nM
GAPDH
RALDH2
-(-)con50 nM
siRNA(B)
5 nM
siRNA(B)
10 nM
siRNA(B)
30 nM
siRNA(B)
50 nM
(Scale bar : 50 μm)
Supplemental Figure II
Se-ries1
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1.6 N1N2N3N4
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55 KD
42 KD
GAPDH
RALDH2
-(-)con50 nM
siRNA(A)
50 nM
siRNA(B)
50 nM
A B
C
(Scale bar : 500 μm)
ATRA (-) ATRA 10 μM
Supplemental Figure III(-
) c
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iRN
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RA
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Via
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Nontreated
(-)controlsiRNA
RALDH2siRNA
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RALDH2siRNA
(B)
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n Fo
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Nontreated
(-)controlsiRNA
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RALDH2siRNA
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Nontreated
(-)controlsiRNA
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(B)
Supplemental Figure I. Phenotypic characterization of endothelial colony forming cells (ECFCs).(A) ECFCs show temporal morphological changes and colonies display a cobblestone morphology. (B) FACS analysis of CD34, KDR, CD133, CD31 and CD45. (C) DiI-labeled Ac-LDL uptake by ECFCs. (D) Immunofluorescence staining of ECFCs labeled with antibodies against CD31 and vWF. Supplemental Figure II. Retinaldehyde dehydrogenase 2 (RALDH2) knockdown by siRNA transfection in normal endothelial colony forming cells (ECFCs).(A) Visualization of efficient transfection of siRNA into normal ECFCs as green cells. (B) RT-PCR shows effective knockdown of RALDH2 by using two types of siRNA. (C) Similar results from real-time quantitative PCR, demonstrates that the expression of the RALDH2 mRNA is decreased after transfection with siRNA. (D) Gene knockdown was confirmed at the protein level. (N1: normal sample #1, N2: normal sample #2, N3: normal sample #3, N4: normal sample #4) Supplemental Figure III. Effective knockdown retinaldehyde dehydrogenase 2 (RALDH2) and the effect of all-trans retinoic acid (ATRA) treatment on tube formation in human umbilical vein endothelial cell (HUVEC).(A) Normal tube formation capabilities of HUVECs in vitro are inhibited after RALDH2 siRNA was trans-fected. The inhibited tube formation ability of HUVECs after RALDH2 knock down is partially restored after treatment with ATRA. (B) RT-PCR shows effective knock-down of RALDH2 by using 2 types of siRNA in HUVECs. Real-time quantitative PCR also confirms that the expression of RALDH2 mRNA is decreased after transfection with siRNA. (C) The viability and proliferation are not affected by transfection with RALDH2 siRNA.
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