Autophagy Regulation Mechanism Mediated By TRPC6 And Its Role In Renal Proximal Tubular Cell Injury Upon Oxidative Stress

Reactive oxygen species（ROS）are chemically reactive chemical species containing oxygen.Examples include peroxides,superoxide,hydroxyl radical.They are formed as a natural byproduct of the normal metabolism of oxygen,and also generated under various pathological conditions such as renal ischemia/reperfusion（I/R）injury.Overproduction of ROS causes oxidative stress and contributes to damages of renal proximal tubular cells（PTC）,which are the main cause of the pathogenesis of renal I/R injury.Autophagy is a dynamic process that removes long-lived proteins and damaged organelles via lysosome-mediated degradation,which has an antioxidant effect that relieves oxidative stress.The canonical transient receptor potential channel 6（TRPC6）,a non selective cation channel that allows passage of Ca²⁺,plays an important role in renal disease.Yet,the relationship between TRPC6 and autophagy,as well as their functions in renal oxidative stress injury,remains unclear.In this study,renal tubular segments were extracted from adult male WT or TRPC6knockout mice and primary PTC were cultured under sterile conditions.H₂O₂ or t-BOOH treated PTC were used to simulate damages caused by ROS in vitro and to explore the function of TRPC6 on ROS-induced autophagy and apoptosis of PTC.In this study,we found that oxidative stress triggered TRPC6-dependent Ca²⁺influx in PTC to inhibit autophagy,thereby rendering cells more susceptible to death.We also demonstrated that TRPC6 knockout(TRPC6^-/-)or inhibition by SAR7334,a TRPC6 selective inhibitor,increased autophagic flux and mitigated oxidative stress-induced apoptosis of PTC.The protective effects of TRPC6 ablation were prevented by autophagy inhibitors Chloroquine and Bafilomycin A1.Moreover,this study also shows that TRPC6 blockage promotes autophagic flux via inhibiting the PI3K/AKT/mTOR and ERK1/2 signaling pathways.This is the first evidence showing that TRPC6-mediated Ca²⁺influx plays a novel role in suppressing cytoprotective autophagy triggered by oxidative stress in PTC,and it may become a novel therapeutic target for the treatment of renal oxidative stress injury in the future.