BACKGROUND/AIMS: Although increased expression and gain function of transient receptor potential cation channel 6 (TRPC6) has been associated with the pathogenesis of some proteinuric glomerular diseases, it remains elusive how TRPC6 participates in the process of podocyte damage. METHODS: The potential signaling responsible for TRPC6 activation was investigated using immunoblot assays in an in vitro podocyte injury model induced by Adriamycin (ADR). Podocyte apoptosis was measured using FITC-conjugated Annexin V and Propidium Iodide staining. The channel activity of TRPC6 was assessed using the Ca2+ influx assay. RESULTS: Increase of TRPC6 expression was detected in ADR-treated podocytes, and TRPC6 knockdown significantly decreased ADR-induced podocytes apoptosis. Following ADR treatment, phospho-mTORSer2481 and phospho-AktSer473 was significantly increased in a time-dependent manner, whereas phospho-mTORSer2448 and phospho-p70S6KThr389 showed no change. ADR-induced apoptosis was prevented by ku0063794 (a dual mTOR complexes inhibitor), not by rapamycin (a specific mTORC1 inhibitor). Furthermore, nuclear translocation of NFκB/p65 was detected in ADR-treated podocytes, which was prevented by an Akt inhibitor triciribine. Of note, NFκB inhibitor PDTC prevented ADR-induced increase of TRPC6, and decreased ADR-induced apoptosis. We found that Akt activation and NFκB nuclear translocation was significantly inhibited by knockdown of mTORC2 protein Rictor, not by mTORC1 protein Raptor. In comparison with control, the Ca2+ influx was significantly increased in ADR-treated podocytes, which was remarkably prevented by TRPC6 knockdown. ADR-induced increase of TRPC6 channel activity was dramatically prevented by ku0063794, but not by rapamycin. Additionally, knockdown of Rictor, not Raptor, prevented ADR-induced increase of the Ca2+ influx. Moreover, the application of NFκB inhibitor PDTC also prevented the Ca2+ influx in ADR-treated podocytes. CONCLUSIONS: Our findings revealed that the mTORC2/Akt/NFκB pathway-mediated activation of TRPC6 participates in ADR-induced podocyte apoptosis.
BACKGROUND/AIMS: Although increased expression and gain function of transient receptor potential cation channel 6 (TRPC6) has been associated with the pathogenesis of some proteinuric glomerular diseases, it remains elusive how TRPC6 participates in the process of podocyte damage. METHODS: The potential signaling responsible for TRPC6 activation was investigated using immunoblot assays in an in vitro podocyte injury model induced by Adriamycin (ADR). Podocyte apoptosis was measured using FITC-conjugated Annexin V and Propidium Iodide staining. The channel activity of TRPC6 was assessed using the Ca2+ influx assay. RESULTS: Increase of TRPC6 expression was detected in ADR-treated podocytes, and TRPC6 knockdown significantly decreased ADR-induced podocytes apoptosis. Following ADR treatment, phospho-mTORSer2481 and phospho-AktSer473 was significantly increased in a time-dependent manner, whereas phospho-mTORSer2448 and phospho-p70S6KThr389 showed no change. ADR-induced apoptosis was prevented by ku0063794 (a dual mTOR complexes inhibitor), not by rapamycin (a specific mTORC1 inhibitor). Furthermore, nuclear translocation of NFκB/p65 was detected in ADR-treated podocytes, which was prevented by an Akt inhibitor triciribine. Of note, NFκB inhibitor PDTC prevented ADR-induced increase of TRPC6, and decreased ADR-induced apoptosis. We found that Akt activation and NFκB nuclear translocation was significantly inhibited by knockdown of mTORC2 protein Rictor, not by mTORC1 protein Raptor. In comparison with control, the Ca2+ influx was significantly increased in ADR-treated podocytes, which was remarkably prevented by TRPC6 knockdown. ADR-induced increase of TRPC6 channel activity was dramatically prevented by ku0063794, but not by rapamycin. Additionally, knockdown of Rictor, not Raptor, prevented ADR-induced increase of the Ca2+ influx. Moreover, the application of NFκB inhibitor PDTC also prevented the Ca2+ influx in ADR-treated podocytes. CONCLUSIONS: Our findings revealed that the mTORC2/Akt/NFκB pathway-mediated activation of TRPC6 participates in ADR-induced podocyte apoptosis.
Authors: Kunyu Shen; David W Johnson; David A Vesey; Michael A McGuckin; Glenda C Gobe Journal: Cell Stress Chaperones Date: 2017-09-27 Impact factor: 3.667
Authors: Akshaya Chandrasekaran; May Y Lee; Xuexin Zhang; Shaheen Hasan; Habben Desta; Scott A Tenenbaum; J Andrés Melendez Journal: Exp Biol Med (Maywood) Date: 2020-07-19
Authors: Daniel J Serie; Julia E Crook; Brian M Necela; Travis J Dockter; Xue Wang; Yan W Asmann; DeLisa Fairweather; Katelyn A Bruno; Gerardo Colon-Otero; Edith A Perez; E Aubrey Thompson; Nadine Norton Journal: Pharmacogenet Genomics Date: 2017-10 Impact factor: 2.089
Authors: Samira Musah; Akiko Mammoto; Thomas C Ferrante; Sauveur S F Jeanty; Mariko Hirano-Kobayashi; Tadanori Mammoto; Kristen Roberts; Seyoon Chung; Richard Novak; Miles Ingram; Tohid Fatanat-Didar; Sandeep Koshy; James C Weaver; George M Church; Donald E Ingber Journal: Nat Biomed Eng Date: 2017-05-10 Impact factor: 25.671