Early Activate Autophagy Can Suppress The Podocyte Damage Induced By Aldosterone

The glomerular filtration barrier consists of the visceral glomerular epithelial cell,also known as the podocyte, the glomerular basement membrane (GBM) and theglomerular endothelial cell. The damage of glomerular filtration barrier is thepathophysiological basis of proteinuria. Recently, many researches suggested thatpodocytes, the terminally differentiated, post-mitotic cells, have lost their ability toproliferate. Loss of20%podocytes will cause mesangium expansion and a transient,mild proteinuria. Loss of20%～40%podocytes will lead to more serious mesangiumexpansion, lasting proteinuria, the glomerular capsule adhesion and FSGS. More than40%loss of podocytes will aggravate glomerulosclerosis and induce progressive lossof renal function culminating in end-stage renal disease (ESKD).Aldosterone (Aldo) is one of important biological active agents inrenin-angiotensin-aldosterone system (RAAS). It plays the biological roles throughthe classic gene action and the non-gene action. Much attention has focused on thenon-classical effect of Aldo, which is independent of renin-angiotensin-aldosteronesystem and can change the growth behavior and structure of cells withouthaemodynamics effect. In cell level, Aldo can induce inflammatory reaction,oxidative stress, fibrosis, mesangial cells proliferation and podocyte injury, and thencause renal injury.Autophagy is a multi-step degradation process, which degrades organelles andproteins to maintain the cellular homeostasis. Recently, with the founding ofautophagy related genes and the application of methods for monitoring autophagy, theimportant of autophagy has been increasingly recognized. When cells are subjected tothe external stress, autophagy may paly a protective role, but if long-term overstimulated, autophagy may induce programmed cell death. In the renal ischemia-reperfusion model and the injury of renal tubular epithelial cell induced bycisplatin or cyclopsorin A, or in the doxorubicin or purine induced podocyte injurymodel, autophagy plays a protective role.Ractive oxygen species ROS are defined as oxygen-containing, reactive chemicalspecies, including superoxide anions, hydroxyl radicals, hydrogen peroxide and so on.A main source of ROS is produced in the mitochondria (mitochondrial electron-tran-sport chain mETC) produce. In addition, NADPH oxdiase (NOX), peroxisom,lipoxygenase and cytochrome P450oxidase are involved in the production ofintracellular ROS. ROS, as a kind of signal molecules, regulates cell proliferation,differentiation, apoptosis and other important biological activities. Recent studiessuggest that ROS, especially the mitochondria ROS, as a signal moleculesparticipates in the regulation of autophagy.Objective: To investigate the role of autophagy in the aldosterone-induced apoptosisand the role of ROS in aldosterone-induced autophagy in cultured podocytes.Methods: Aldosterone-induced apoptosis in cultured mouse podocyte clones (MPC5)and were detected by Flow cytometry. The generation of ROS was detected byFluorescence microplate reader. After24h treatment with aldosterone, the existence ofapoptotic body and autophagosome were evaluated by electron microscopy. Theprotein expressions of LC3, caspase-3and nephrin were detected by Western Blot.The mRNAexpression of Beclin-1was detected by real-time PCR.Results: Aldosterone induced podocyte apoptosis and autophagy in a time-dependentmannner. After24h treatment, the ratio of apoptosis was increased by26.5%(P＜0.05) and the expression of nephrin was decreased by28%(P＜0.05), as comparedwith the control group. Aldosterone remarkably (P＜0.05) increased the expression ofBeclin-1at6h and promoted transforming LC3-I to LC3-II at12h. Compared with with aldosterone treatment, the apoptosis rate of podocyte was further increased by39%(P＜0.05), the expression of nephrin was further declined by19.5%（P＜0.05）and the activation of caspase-3was further increased by90%after3-MApretreatment.Aldosterone time-dependently induced ROS generation in podocytes. NACsignificantly inhibited the expression of Beclin-1and decreased transforming LC3-Ito LC3-II induced by aldosterone at24h treatment.Conclusion: Aldosterone induced autophagy and apoptosis in podocyte. Autophagyoccurred ealier (12h) than apoptosis (24h). The occurrence of autophagy inhibitedapoptosis. ROS participated in the inducing of autophagy by aldosterone. Therefore,the strategy targeting autophagy pathway may respresent a new way of glomerulardiseases treatment.