Gengting Dong1, Tingbo Chen1, Xuecong Ren1, Zhifeng Zhang1, Weixue Huang2, Liang Liu1, Pei Luo3, Hua Zhou4. 1. State Key Laboratory of Quality Research in Chinese Medicine, Macau University of Science and Technology, Avenida Wai Long, Taipa, Macau, China. 2. State Key Laboratory of Bio-organic and Natural Products Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai, China. 3. State Key Laboratory of Quality Research in Chinese Medicine, Macau University of Science and Technology, Avenida Wai Long, Taipa, Macau, China. Electronic address: pluo@must.edu.mo. 4. State Key Laboratory of Quality Research in Chinese Medicine, Macau University of Science and Technology, Avenida Wai Long, Taipa, Macau, China. Electronic address: hzhou@must.edu.mo.
Abstract
PURPOSE: Mitochondrial dysfunction is a prominent feature of ischemia heart disease but the underlying mechanism of dynamics (fusion/fission) is still unclear. Here we investigated a novel function and underlying mechanism of Rg1 on an in vitro cardiomyocyte model of hypoxia/reoxygenation (H/R). METHODS: Cellular cytotoxicity was evaluated by MTT, mitochondrial viable staining, and cardiac marker detection. Mitochondrial function was evaluated by ATP content measurement, MMP determination, ROS, OCR and ECAR assay. Mitochondrial dynamics was investigated by Live-cell imaging with time-lapse fluorescence microscopy and morphological features were evaluated by the high-content image analysis. Mitochondrial fusion and fission-related proteins, GDH were determined by Western blot, RT-PCR and immunofluorescence. RESULTS: Rg1 moderated GDH dysregulation and then protected against H/R-induced cellular damage and mitochondrial dysfunction in a dose-dependent manner. Rg1 significantly increased mitochondrial length, reduced the number of cells with fragmented mitochondria and up-regulated the MFN2 expression finally leading to preventing the imbalance of mitochondrial dynamics following H/R. Knock-down of MFN2 by specific siRNA completely abolished the ability of Rg1 to cell survival by H/R. CONCLUSION: Rg1 through modulation of GDH and MFN2 maintained mitochondrial dynamics that resulted in protection against H/R-induced cardiomyocyte injury. All these results put forward a new protective mechanism of Rg1 on the therapeutic potential in cardiac I/R disorders.
PURPOSE:Mitochondrial dysfunction is a prominent feature of ischemia heart disease but the underlying mechanism of dynamics (fusion/fission) is still unclear. Here we investigated a novel function and underlying mechanism of Rg1 on an in vitro cardiomyocyte model of hypoxia/reoxygenation (H/R). METHODS: Cellular cytotoxicity was evaluated by MTT, mitochondrial viable staining, and cardiac marker detection. Mitochondrial function was evaluated by ATP content measurement, MMP determination, ROS, OCR and ECAR assay. Mitochondrial dynamics was investigated by Live-cell imaging with time-lapse fluorescence microscopy and morphological features were evaluated by the high-content image analysis. Mitochondrial fusion and fission-related proteins, GDH were determined by Western blot, RT-PCR and immunofluorescence. RESULTS:Rg1 moderated GDH dysregulation and then protected against H/R-induced cellular damage and mitochondrial dysfunction in a dose-dependent manner. Rg1 significantly increased mitochondrial length, reduced the number of cells with fragmented mitochondria and up-regulated the MFN2 expression finally leading to preventing the imbalance of mitochondrial dynamics following H/R. Knock-down of MFN2 by specific siRNA completely abolished the ability of Rg1 to cell survival by H/R. CONCLUSION:Rg1 through modulation of GDH and MFN2 maintained mitochondrial dynamics that resulted in protection against H/R-induced cardiomyocyte injury. All these results put forward a new protective mechanism of Rg1 on the therapeutic potential in cardiac I/R disorders.
Authors: Declan McLaughlin; Youyou Zhao; Karla M O'Neill; Kevin S Edgar; Philip D Dunne; Anna M Kearney; David J Grieve; Barbara J McDermott Journal: Br J Pharmacol Date: 2017-04-22 Impact factor: 8.739