Roxadustat attenuates heat stress-induced apoptosis and senescence in renal tubular epithelial cells by upregulating HIF-1α

Objective To investigate the effect and underlying mechanism of roxadustat on apoptosis and senescence of renal tubular epithelial cell line HK-2 induced by heat stress. Methods After HK-2 cells were treated with different concentrations of roxadustat (10, 20, 30, 40 and 50 μmol/L) for 24 h, CCK-8 assay was used to determine the optimal intervention concentration of roxadustat. HK-2 cells were divided into 4 groups (n=3): control group, roxadustat group (30 μmol/L, 24 h), heat-stress group (43 ℃, 2 h), and heat-stress+roxadustat group (30 μmol/L roxadustat treatmnet for 24 h followed by heat-stress 2 h). Cell viability was detected by CCK-8 assay. Expression of hypoxia-inducible factor-1α (HIF-1α), Cleaved Caspase-3, p16 and p21 at protein level was detected by Western blotting. Immunofluorescence assay was employed to observe the distribution of HIF-1α. β-galactosidase staining kit was utilized to detect SA-β-Gal activity. TUNEL staining was used to measure cell apoptosis. Results The highest cell viability was observed in the cells after 30 μmol/L roxadustat treatment. Heat stress resulted in a significant decrease in cell viability (P < 0.05), elevated protein levels of HIF-1α, Cleaved Caspase-3, p16 and p21(P < 0.05), enhanced SA-β-Gal activity (P < 0.05) and increased percentage of TUNEL-positive cells (P < 0.05) when compared with the cells in the control group. In comparison with the heat-stress group, the heat-stress+roxadustat group showed significant decrease in the protein levels of Cleaved Caspase-3, p16 and p21 (P < 0.05), reduced activity of SA-β-Gal [(65.44±5.00)% vs (77.15±2.61)%, P < 0.05] and decreased percentage of TUNEL-positive cells [(16.73±2.20)% vs (46.40±13.87)%, P < 0.05], but increase in cell viability [(86.33±4.51)% vs (66.33±8.50)%, P < 0.05] as well as HIF-1α protein expression (P < 0.05). Furthermore, immunofluorescence assay showed that HIF-1α was mainly distributed in the nucleus and perinucleus. Conclusion Roxadustat attenuates heat stress-induced apoptosis and senescence of renal tubular epithelial cells by upregulating HIF-1α.