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Abstract Background Podocytes have become a crucial target for interventions in proteinuric kidney diseases. Many studies have reported that overexpression of transient receptor potential cation channel protein 6 (TRPC6) in podocyte injury upregulates intracellular Ca2+ influx and stimulates Ca2+-dependent protease calpain-1 signaling. The traditional Chinese drug, tetrandrine, a nonselective Ca2+ channel blocker, has long been used to treat chronic kidney disease. This research aimed to explore the possible mechanisms underlying the anti-proteinuric properties of tetrandrine. Methods We investigated the involvement of tetrandrine in Ca2+ dependent calpain-1 signaling in mouse podocytes and adriamycin-induced nephropathy rats. Cyclosporine A (CsA) and U73122 were used as positive controls. Cell viability, cytotoxicity, Ca2+ concentration, calpain activity, and mRNA and protein expression levels of calpain-1 signaling pathways were examined. The clinical and pathological changes were measured. Results Tetrandrine decreased intracellular Ca2+ influx in cultured TRPC6-overexpressing podocytes. In both in vitro and in vivo studies, the administration of tetrandrine downregulated calpain activity and the expression of calpain-1 and restored the expression of downstream Talin-1 and nephrin. Compared to CsA, tetrandrine treatment exhibited superior inhibitory effects on calpain activity and calpain-1 expression. Conclusions Tetrandrine has therapeutic potential in podocyte damage by blocking Ca2+-dependent activation of the calpain-1 signaling pathway. Tetrandrine reduced proteinuria, improved renal function, and alleviate renal pathological damage.

Background: Podocytes have become a crucial target for interventions in proteinuric kidney diseases. Many studies have reported that overexpression of transient receptor potential cation channel protein 6 (TRPC6) in podocyte injury upregulates intracellular Ca2+ influx and stimulates Ca2+-dependent protease calpain-1 signaling. The traditional Chinese drug, tetrandrine, a nonselective Ca2+ channel blocker, has long been used to treat chronic kidney disease. This research aimed to explore the possible mechanisms underlying the anti-proteinuric properties of tetrandrine. Methods: We investigated the involvement of tetrandrine in Ca2+ dependent calpain-1 signaling in mouse podocytes and adriamycin-induced nephropathy rats. Cyclosporine A (CsA) and U73122 were used as positive controls. Cell viability, cytotoxicity, Ca2+ concentration, calpain activity, and mRNA and protein expression levels of calpain-1 signaling pathways were examined. The clinical and pathological changes were measured. Results: Tetrandrine decreased intracellular Ca2+ influx in cultured TRPC6-overexpressing podocytes. In both in vitro and in vivo studies, the administration of tetrandrine downregulated calpain activity and the expression of calpain-1 and restored the expression of downstream Talin-1 and nephrin. Compared to CsA, tetrandrine treatment exhibited superior inhibitory effects on calpain activity and calpain-1 expression. Conclusions: Tetrandrine has therapeutic potential in podocyte damage by blocking Ca2+-dependent activation of the calpain-1 signaling pathway. Tetrandrine reduced proteinuria, improved renal function, and alleviate renal pathological damage. [ABSTRACT FROM AUTHOR]

Jin Yu, Caifeng Zhu, Jiazhen Yin, Dongrong Yu, Feng Wan, Xuanli Tang, Xue Jiang Department of Nephrology, Guangxing Hospital Affiliated to ZheJiang Chinese Medical University (Key Laboratory of Zhejiang Province, Management of Kidney Disease), Hangzhou 310007, People’s Republic of ChinaCorrespondence: Xue JiangDepartment of Nephrology, Guangxing Hospital Affiliated to ZheJiang Chinese Medical University (Key Laboratory of Zhejiang Province, Management of Kidney Disease), Hangzhou 310007, People’s Republic of ChinaEmail monica_jiang@163.comObjective: Podocyte damage is common in many renal diseases characterized by proteinuria. Transient receptor potential cation channel protein 6 (TRPC6) plays an important role in renal function through its regulation of intracellular Ca2+ influx and RhoA/ROCK pathways. Chinese herb Stephania tetrandra, with the main active component being tetrandrine, has been used for the treatment of various kidney diseases for several years and has shown a positive effect. This study aimed at investigating the effect and mechanism of tetrandrine in podocyte damage induced by high expression of TRPC6.Methods: Immortalized, differentiated murine podocytes, MPC5 were treated with valsartan (0– 800 μM) and tetrandrine (0– 40 μM) for 48 h. The maximum safe concentrations of valsartan and tetrandrine were selected using a cell viability assay. MPC5 podocytes stably expressing TRPC6 were constructed using a lentivirus packaging system, followed by treatment with valsartan, tetrandrine, and Y-27632 for 48 h and U73122 (10 μM) for 10 min. The RhoA/ROCK pathway and podocyte-specific proteins (nephrin and synaptopodin) levels were quantified. Podocyte apoptosis and intracellular Ca2+ concentration were measured.Results: Maximum safe concentrations of 100 μM valsartan and 10 μM tetrandrine showed no observable toxicity in podocytes. MPC5 podocytes stably expressing TRPC6 had higher intracellular Ca2+ influx, apoptotic percentages, and expression of RhoA/ROCK proteins, but lower expression of nephrin and synaptopodin proteins. U73122 treatment for 10 min did not inhibit TRPC6, but suppressed RhoA/ROCK protein. Y-27632 decreased ROCK1 expression, but did not influence the expression of TRPC6 protein. Both 100 μM valsartan and 10 μM tetrandrine for 48 h significantly inhibited intracellular Ca2+ influx, apoptosis, and RhoA/ROCK pathway, and increased nephrin and synaptopodin proteins in podocytes stably expressing TRPC6.Conclusion: Elevated TRPC6 expression can lead to podocyte injury by inducing intracellular Ca2+ influx and apoptosis of podocytes, and this effect may be mediated by activation of the RhoA/ROCK1 pathway. Tetrandrine can alleviate podocyte injury induced by TRPC6 expression through inhibition of the RhoA/ROCK pathway, suggesting a protective role in podocyte damage.Keywords: tetrandrine, transient receptor potential cation channel protein 6, podocyte, RhoA/ROCK pathway