Fig. 4.
aFGF activated Wnt/β-catenin signaling pathway to promote c-Myc expression and protect the endothelial function against HG + PA. (A) TEF/LEF-luciferase reporter activity in HUVECs. (B) Immunoblotting and sqRT-PCR analysis of c-Myc in HUVECs. HUVECs were cultured either in NG or HG + PA medium in the presence or absence of aFGF (100 ng/mL) for 72 h, MAN was served as the osmotic control for the HG + PA. For manipulation of Wnt/β-catenin pathway, IWR (5 μM) and ICG-001 (10 μM) was pretreated for 2 h before aFGF administration. (C) OCR was analysed using a Seahorse XF analyser. (D) ATP production in HUVEC. (E) Mitochondrial O2•− in HUVEC was measured by mitochondria targeted probe MitoSOX and UPLC after accumulation of O2•−-specific product 2-OH-Mito-E+. (F) mtROS of HUVECs was detected by MitoSOX staining assay, scale bars = 1000 μm, (G) Mitochondrial membrane potential was detected by TMRM fluorescence staining, scale bars = 5 μm, (H) TUNEL assay of HUVECs, scale bars = 100 μm, (I) Capillary-like tube formation of HUVECs, scale bars = 300 μm. All values displayed are means ± SEM of 6 independent experiments. #p < 0.05 vs. NG or MAN; *p < 0.05 vs. HG + PA; % p < 0.05 vs. HG + PA co-incubated with aFGF.