Fig. 2

High glucose led to cell senescence and endothelial cell dysfunction. (A) Representative images of SA-β-gal staining and (B) quantification of SA-β-gal positive HUVECs treated with normal glucose (5.5 mM d-glucose, C), high mannitol (15 mM mannitol, M), high glucose (15 mM d-glucose, HG). Scale bar = 20 μm. (C–F) The mRNA levels of (C) P53, (D) P16, (E) P21, and (F) Cyclin D1 of C, M, and HG groups measured by qRT-PCR. (G) Representative images of Western Blotting analysis and (H-K) semi-quantification of (H) p53, (I) p21, (J) p16, and (K) Cyclin D1 of C, M, HG groups. (L) Cell proliferation of C, M, HG groups assessed by CCK-8 assays. Optical density (OD) = 450 nm. (M) The representative images of wound healing assay of HUVECs. Scale bar = 100 μm. (N) The semi‐quantitative analysis of the distance of migration. (O, P) Representative micrographs of EdU staining (O) and quantification (P) of EdU positive HUVECs. Scale bar = 100 μm. The values are expressed as the mean ± SEM. Experiments were carried out in triplicate and repeated at least three times. Results in B-F, H-K and P were analyzed with one-way ANOVA with Dunnett’s multiple comparisons test. Results in L and N were analyzed with two-way ANOVA with Dunnett’s multiple comparisons test.