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Fig. 3 | Stem Cell Research & Therapy

Fig. 3

From: Development of a surrogate potency assay to determine the angiogenic activity of Stempeucel®, a pooled, ex-vivo expanded, allogeneic human bone marrow mesenchymal stromal cell product

Fig. 3

Activity of VEGF secreted from phBMMSCs is mediated by VEGFR2. a Proliferation of HUVECs using 50% CM from phBMMSCs and 50% CM with Axitinib at concentrations of 0.25 nM, 0.29 nM and 1.2 nM to block VEGFR2, VEGFR2 and VEGFR3, and VEGFR2, VEGFR3 and VEGFR1 respectively. Significant reduction in HUVEC proliferation was observed in the CM with all Axitinib treatment groups, ***p = 0.0001. EGM was used as a positive control for HUVEC proliferation. Analysis of VEGFR1 (b), VEGFR2 (c) and VEGFR3 (d) fold expression in HUVECs treated with phBMMSC CM and phBMMSC CM with anti-VEGF mAb by qRT-PCR. Significant reduction only in VEGFR2 expression was observed in the phBMMSC CM with anti-VEGF mAb group compared with the CM alone group, *p = 0.01. e Analysis of VEGF pathway-dependent markers PI3K and AKT1 in HUVECs cultured in the presence of phBMMSC CM by qRT-PCR; mRNA expression levels were normalized with housekeeping gene β-Actin as an internal standard and subsequently normalized to expression levels in HUVECs cultured in EGM as control. Data generated using two batches of phBMMSC CM, assayed in triplicates and presented as mean ± SEM. f Schematic diagram to depict the possible signal transduction pathway of VEGF secreted by phBMMSCs in HUVECs. SEMs shown as error bars. HUVEC Human umbilical vein endothelial cell, EGM endothelial growth medium, CM conditioned medium, VEGFR vascular endothelial growth factor receptor, mAb monoclonal antibody, Ang-1 angiopoietin-1, SDF-1 stromal-derived factor-1, HGF hepatocyte growth factor, IL interleukin, TGF-β transforming growth factor beta

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