Research Paper Volume 12, Issue 11 pp 10370—10380

Keap1-Nrf2 signaling activation by Bardoxolone-methyl ameliorates high glucose-induced oxidative injury in human umbilical vein endothelial cells

Figure 4. Keap1 silencing mimics BARD-induced cytoprotection in HG-stimulated HUVECs. The stable HUVECs with Keap1 shRNA lentiviral particles (“sh-Keap1”) were treated with or without Bardoxolone Methyl (BARD, at 50 nM) for applied time periods, control cells were transduced with the scramble control shRNA (“sh-C”), expression of listed genes was tested by qPCR and Western blotting analyses (A, C, D), with the relative ARE activity examined as well (B); Alternatively, cells were pretreated with BARD (50 nM) for 1h, followed by HG stimulation and cultured for 48h, cell viability (CCK-8 assay, E) and apoptosis (nuclear TUNEL staining assay, F) were tested. Expression of the listed proteins was quantified, normalizing to the indicated loading control protein (A and D). Error bars stand for mean ± standard deviation (SD, n=5). ##p<0.01. vs. “sh-C” cells. Each experiment was repeated five times to insure the consistency of experimental results.