Aldosterone inhibits autophagy activation of mesangial cells and accelerates cell apoptosis under oxidative stress condition / 第二军医大学学报

ABSTRACT

Objective To investigate the effect of aldosterone on the activation of autophagy in mesangial cells. Methods Three classic techniques were used to detect the autophagy activity in mesangial cells with or without aldosterone treatment in this study. (1) Western blotting analysis was used to examine the expression of autophagy protein LC3, SQSTM1/P62 after human mesangial cell line (HMCL) was treated with different concentrations of aldosterone. (2) Confocal laser scanning microscope was used to observe the change of autophagy points after transforming eukaryotic expression vector GFP-LC3 into HMCL cells. (3) Transmission electron microscopy was used to observe the autophagic vacuoles of HMCL cells after treatment with aldosterone. Microscope and Western blotting analysis were used to observe the expression of apoptosis protein and poly(ADP-ribose) polymerase in mesangial cells under the conditions of oxidative stress (2.5×10-6mol/L H2O2 solution) and under the presence or absence of aldosterone. Results All the three methods confirmed that high physiological dose of aldosterone (10-7 mol/L) could inhibit mesangial cell autophagy activation (1) The autophagy marker LC3 I converting to LC3 II had a decrease of approximately 40% after 10-7 mol/L aldosterone stimulation for 12 h. (2) The numbers of autophagy point of mesangial cells induced by Earle's balanced salt solution (EBSS) and rapamycin were also reduced by 60% and 47% upon aldosterone treatment, respectively. (3) Aldosterone treatment significantly reduced starvation and rapamycin-induced vacuole formation in mesangial cells examined by transmission electron microscopy. The mesangial cells treated by aldosterone had a significantly increased apoptosis rate than the control group under oxidative stress condition with hydrogen peroxide stimulation (2.5×10-6 mol/L, P<0.05). Conclusion High physiological concentration of aldosterone shows an inhibition against the basic autophagy activation of mesangial cells and accelerates cell apoptosis under conditions of oxidative stress.