Juan Jin1,2, Diandian Wu1,3, Li Zhao1,2, Wenli Zou1,2, Wei Shen1,2, Qiudi Tu1,2, Qiang He1,2. 1. Department of Nephrology, Zhejiang Provincial People's Hospital Hangzhou 310014, Zhejiang, P. R. China. 2. People's Hospital of Hangzhou Medical College Hangzhou 310014, Zhejiang, P. R. China. 3. Bengbu Medical College Bengbu, Anhui, P. R. China.
Abstract
AIM: We aimed to assess the effect of autophagy and stromal interaction molecule 1 (STIM1) on podocyte epithelial-mesenchymal transition in diabetic nephropathy. METHODS: The sera of 8-week-old db/db and C57BL/KsJ rats were used to culture MPC5 cells. The experiment was divided into 4 groups: MPC5 + siRNA-Scr + 10% C57BL/KsJ (Group A), MPC5 + siRNA-STIM1 + 10% C57BL/KsJ (Group B), MPC5 + siRNA-Scr + 10% db/db (Group C), and MPC5 + siRNA-STIM1 + 10% db/db (Group D). Podocyte autophagy was evaluated via immunofluorescence staining for LC3II and P62, and via Western blotting for P62 and LC3 (LC3II/LC3I). Western blotting was also used to assess the expression of TRPC6, Orai1, Beclin-1, Bcl-2, Caspase3, E-cadherin, fibronectin, and α-SMA protein. Furthermore, podocyte apoptosis was assessed via flow cytometry. RESULTS: We found that, in podocytes cultured in the serum of diabetic nephrotic rats, the autophagy level decreased, whereas the apoptosis level increased, and EMT can be advanced. However, after silencing STIM1 with siRNA, a converse outcome was noted. Furthermore, in diabetic nephropathy rats, the up-regulated expression of podocyte STIM1 can activate TRPC6 and Orai1 channels, which results in Ca2+ entry. CONCLUSIONS: We found that, in podocytes cultured in the serum of diabetic nephrotic rats, the autophagy level increased, whereas the apoptosis level decreased, and EMT can be inhibited by silencing STIM1 with siRNA. IJCEP
AIM: We aimed to assess the effect of autophagy and stromal interaction molecule 1 (STIM1) on podocyte epithelial-mesenchymal transition in diabetic nephropathy. METHODS: The sera of 8-week-old db/db and C57BL/KsJ rats were used to culture MPC5 cells. The experiment was divided into 4 groups: MPC5 + siRNA-Scr + 10% C57BL/KsJ (Group A), MPC5 + siRNA-STIM1 + 10% C57BL/KsJ (Group B), MPC5 + siRNA-Scr + 10% db/db (Group C), and MPC5 + siRNA-STIM1 + 10% db/db (Group D). Podocyte autophagy was evaluated via immunofluorescence staining for LC3II and P62, and via Western blotting for P62 and LC3 (LC3II/LC3I). Western blotting was also used to assess the expression of TRPC6, Orai1, Beclin-1, Bcl-2, Caspase3, E-cadherin, fibronectin, and α-SMA protein. Furthermore, podocyte apoptosis was assessed via flow cytometry. RESULTS: We found that, in podocytes cultured in the serum of diabetic nephroticrats, the autophagy level decreased, whereas the apoptosis level increased, and EMT can be advanced. However, after silencing STIM1 with siRNA, a converse outcome was noted. Furthermore, in diabetic nephropathyrats, the up-regulated expression of podocyte STIM1 can activate TRPC6 and Orai1 channels, which results in Ca2+ entry. CONCLUSIONS: We found that, in podocytes cultured in the serum of diabetic nephroticrats, the autophagy level increased, whereas the apoptosis level decreased, and EMT can be inhibited by silencing STIM1 with siRNA. IJCEP