Study On The Mechanism Of E2F1 Regulation On Autophagy In High Glucose-induced EMT In Renal Tubular Epithelial Cell And The Intervention Of Resveratrol

Objective: E2F1 has an effect on cell cycle?apoptosis and autophagy in tumour cells,but it is unclear that the regulation and mechanism of E2F1 on autophagy and EMT in renal tubular epithelial cells(NRK-52E)cultured with high-glucose.This study aims to explore whether resveratrol can promote autophagy by down-regulating E2F1 and further inhibit EMT in NRK-52 E,so as to explore a new effective target for DN prevention and treatment.Methods:(1)The diabetic nephropathy model was established,and the renal pathological changes were observed by HE,Masson staining.Immunohistochemical staining was performed toobserve the expression of E2F1 protein in renal tissue.(2)The protein expressions of E2F1,LC3,p62 and E-cadherin,?-SMA and Collagen IV were identified by western blot.We collected protein and RNA samples after 48 hours.Western blot and RT-q PCR were used to detect the protein and m RNA expressions of E2F1,LC3 and p62,E-cadherin,?-SMA and Collagen IV.(3)The Control and E2F1-sh RNA were respectively transfected in NG and HG groups,and after 48 h,we collected samples.Then we detected the protein and m RNA expressions of E2F1,LC3,p62,E-cadherin,?-SMA and Collagen IV by Western blot and RT-q PCR.(4)The NRK-52 E cells were added to resveratrol(Res)in NG and HG groups and we collected samples.Then we detected the protein expressions of E2F1,LC3,p62,E-cadherin,?-SMA and Collagen IV by Western blot.(5)The Vector and overexpressed plasmid E2F1 were transfected in NG and HG groups and then resveratrol was add to NG and HG groups for 48 h.We collected samples and detected the protein and m RNA expressions of E2F1,LC3,p62,E-cadherin,?-SMA and Collagen IV by western blot and RT-q PCR.Results:(1)Diabetic nephropathy was successfully replicated.The results of HE and Masson staining showed that fibrotic lesions were arisen in DN group.Immunohistochemical showed that: the cytoplasm of renal tubular epithelial cells in DN group showed higher expression of E2F1.(2)(1)Compared with NC group,the protein expressions of E2F1,p62,?-SMA and Collagen IV were significantly increased in DN group(P<0.05),and the protein expressions of LC3 and Ecadherin were significantly decreased(P<0.05).Compared with NG group,the expressions of E2F1,p62,?-SMA and Collagen IV was significantly increased in NRK-52 E cells in HG group(P<0.05),and the protein expressions of LC3 and E-cadherin were decreased(P<0.05).(2)In HG+E2F1-sh RNA group,the m RNA and protein expressions of E2F1,p62,?-SMA and Collagen IV were significantly decreased(P<0.05),the expressions of LC3 and Ecadherin were significantly increased(P<0.05).(3)(1)Compared with HG group,the protein expressions of E2F1,p62,?-SMA and Collagen IV in HG+Res group were significantly decreased(P<0.05),and the protein expressions of LC3 and E-cadherin were increased(P<0.05).(2)The protein expressions of E2F1,p62,?-SMA and Collagen IV in HG+Res+E2F1 group wereincreased compared with HG+Res group(P<0.05).Conclusions:(1)The expression of E2F1 is increased in diabetic nephropathy.(2)The expression of E2F1 is increased in HG group and E2F1 inhibits autophagy in renal tubular epithelial cells with high-glucose cultured and promotes EMT.(3)Resveratrol promotes autophagy by down-regulating E2F1 and inhibits the occurrence and development of EMT.