Study On The Molecular Mechanism Of Intervention By Gan-song Yin On Autophagy-apoptosis Of Renal Podocytes And Renal Tubular Epithelial Cells In Mice Induced By High Glucose

Objective To study the effect and related molecular mechanisms of intervention by Gan-song Yin(GSY)medicated serum on renal podocytes and renal tubular epithelial cells induced by high glucose.Methods The podocytes(MPC5)and renal tubular epithelial cells(TCMK-1)were cultured in vitro,modeled with glucose solution,and GSY-containing serum was used as an intervention drug.The experiment was designed as normal group(NOR),model group(MOD),and Gan-song Yin high dose group(H-GSY),Gan-song Yin medium dose group(M-GSY),Gan-song Yin low-dose group(L-GSY).After intervention with glucose and GSY,the rate of apoptosis,the level of reactive oxygen species(ROS)and mitochondrial membrane potential,which were measured on a flow cytometer.The incidence of autophagy was measured by the MDC method.Western Blot method was used to detect the expression of apoptosis-related proteins Bcl-2,Bax,caspase7 and caspase9,the expression of autophagy marker proteins LC3,P62,the protein expression of TSC1,m TOR,Atg5 in renal podocytes,and the protein expression of SGLT2 and SIRT1 in tubular epithelial cells.Results 1.The effect of GSY on MPC5 induced by high glucose: After intervention of GSY for 24 hours,compared with the model group,the proportion of cell apoptosis in GSY group gradually decreased(P<0.05),and the accumulation of cellular reactive oxygen species decreased(P<0.01),the level of intracellular mitochondrial membrane potential increased(P<0.01),intracellular autophagosomes increased(P<0.01),and change in the H-GSY group was the most obvious.Western Blot showed that,compared with the Model group,among apoptosis-related proteins,the expression of Bcl-2 in the GSY group increased(P<0.01),while the expression of Bax,caspase7 and caspase9 decreased(P<0.05);among the autophagy-related proteins,the expression of P62 and m TOR decreased(P<0.05),the expression of Atg5 and TSC1 increased(P<0.05).2.The effect of GSY on TCMK-1 induced by high glucose: After intervention of GSY,compared with the model group,the proportion of cell apoptosis decreased in GSY group(P<0.01),and the level of ROS decreased(P<0.01),the level of intracellular mitochondrial membrane potential increased(P<0.05),especially in the M-GSY group and H-GSY group(P<0.01),and intracellular autophagosomes increased(P<0.01).Western Blot showed that,compared with the Model group,the expression of Bcl-2 in GSY group increased(P<0.01),and the expression of Bax,caspase7 and caspase9 decreased(P<0.05).Among the autophagy-related proteins,the expression of LC3 and SIRT1 increased(P<0.05),the expression of SGLT2 and P62 decreased(P<0.05).Conclusion 1.GSY may has a protective effect on MPC5 and TCMK-1 cells induced by high glucose,which can reduce the level of ROS and cell apoptosis,increase the level of mitochondrial membrane potential.2.GSY can down-regulate m TOR,and up-regulate Atg5 and TSC1,and the mechanism may be to promote autophagy of MPC5 cells by activating TSC1-m TOR-Atg5 signal pathway.GSY can down-regulate SGLT2,up-regulate SIRT1,which may promote autophagy in TCMK-1 cells by activating SGLT2-SIRT1 signaling pathway and alleviate glomerular injury.