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Published byPhoebe Hodges Modified over 9 years ago
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Marker (%) Normal EPCs Mean ± SM (%) MMD EPCs Mean ± SM (%) CD3426.9 ± 11.531.0 ± 6.8 KDR27.3 ± 5.5222.4 ± 8.90 CD13324.5 ± 4.6412.6 ± 3.93 CD3199.0 ± 0.998.8 ± 0.5 CD450.42 ± 1.231.53 ± 3.12 CD34 KDRCD133CD31 B (Scale bar : 50 μm) CD31 vWF C D (Scale bar : 100 μm) Dil-Ac-LDL uptake Supplemental Figure I NormalMMD (Scale bar : 100 μm) A 21 days 28 days 35 days 7 days CD45 ***
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β-actin RALDH2 - (-)con 50 nM siRNA (A) 50 nM siRNA (B) 50 nM (-) control siRNA Non treated 10 nM 30 nM 50 nM AB C D RALDH2 siRNA (A) (nM) RALDH2 siRNA (B) (nM) - - Negative control siRNA (nM) - 50 - - - 5 - - 10 - - 30 - - 50 - - - 5 - - 10 - - 30 - - 50 GAPDH RALDH2 - (-)con 50 nM siRNA (A) 5 nM siRNA (A) 10 nM siRNA (A) 30 nM siRNA (A) 50 nM GAPDH RALDH2 - (-)con 50 nM siRNA (B) 5 nM siRNA (B) 10 nM siRNA (B) 30 nM siRNA (B) 50 nM (Scale bar : 50 μm) Supplemental Figure II 55 KD 42 KD
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GAPDH RALDH2 - (-)con 50 nM siRNA (A) 50 nM siRNA (B) 50 nM A B C (Scale bar : 500 μm) ATRA (-)ATRA 10 μM Supplemental Figure III (-) control siRNA RALDH2 siRNA(A) RALDH2 siRNA(B) Non treated (-)control siRNA RALDH2 siRNA (A) RALDH2 siRNA (B) Non treated (-)control siRNA RALDH2 siRNA (A) RALDH2 siRNA (B) Non treated (-)control siRNA RALDH2 siRNA (A) RALDH2 siRNA (B)
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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 transfected. 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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