Salvianolate inhibits reactive oxygen species production in H2O2-treated mouse cardiomyocytes in vitro via the TGFβ pathway
Abstract
Ai-hua FEI1, Qing CAO3, Shu-yan CHEN3, Hai-rong WANG1, Fei-long WANG1, Shu-ming PAN1, Zhao-fen LIN2, *
1Emergency Department, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200092, China; 2Emergency Department, Shanghai Changzheng Hospital, Second Military Medical University, Shanghai 200003, China; 3Department of Geriatrics, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200092, China
Aim: To investigate the effects of salvianolate, a water-soluble active compound from Salvia miltiorrhiza Bunge, on reactive oxygen species (ROS) production in mouse cardiomyocytes in vitro.
Methods: Primary ventricular cardiomyocytes were prepared from neonatal mouse. The cell viability was determined using MTT assay. Culture medium for each treatment was collected for measuring the levels of NO, iNOS, total antioxidant capacity (TAOC) and transforming growth factor β1 (TGFβ1). TGFβ1 and Smad2/3 expression in the cells was detected with Western blotting.
Results: H2O2 (1.25 mmol/L) did not significantly affect the cell viability, whereas the high concentration of salvianolate (5 g/L) alone dramatically suppressed the cell viability. Treatment of the cells with H2O2 (1.25 mmol/L) markedly increased ROS and iNOS production, and decreased the levels of NO, TAOC and TGFβ1 in the culture medium. Furthermore, the H2O2 treatment significantly increased TGFβ1 and Smad2/3 expression in the cells. Addition of salvianolate (0.05, 0.1, and 0.5 g/L) concentration-dependently reversed the H2O2-induced alterations in the culture medium; addition of salvianolate (0.05 g/L) reversed the H2O2-induced increases of TGFβ1 and Smad2/3 expression in the cells. Blockage of TGFβ1 with its antibody (1 mg/L) abolished the above mentioned effects of salvianolate.
Conclusion: Salvianolate inhibits ROS and iNOS production and increases TAOC and NO levels in H2O2-treated cardiomyocytes in vitro via downregulation of Smad2/3 and TGFβ1 expression. High concentration of salvianolate causes cytotoxicity in mouse cardiomyocytes.
Keywords: salvianolate; Salvia miltiorrhiza Bunge; cardiac fibrosis; cardiomyocytes; reactive oxygen species; NO; iNOS; TGFβ1; Smad2/3
* To whom correspondence should be addressed.
E-mail linzhaofen@sina.com
Received 2012-08-14 Accepted 2012-12-24
Keywords:
1Emergency Department, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200092, China; 2Emergency Department, Shanghai Changzheng Hospital, Second Military Medical University, Shanghai 200003, China; 3Department of Geriatrics, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200092, China
Aim: To investigate the effects of salvianolate, a water-soluble active compound from Salvia miltiorrhiza Bunge, on reactive oxygen species (ROS) production in mouse cardiomyocytes in vitro.
Methods: Primary ventricular cardiomyocytes were prepared from neonatal mouse. The cell viability was determined using MTT assay. Culture medium for each treatment was collected for measuring the levels of NO, iNOS, total antioxidant capacity (TAOC) and transforming growth factor β1 (TGFβ1). TGFβ1 and Smad2/3 expression in the cells was detected with Western blotting.
Results: H2O2 (1.25 mmol/L) did not significantly affect the cell viability, whereas the high concentration of salvianolate (5 g/L) alone dramatically suppressed the cell viability. Treatment of the cells with H2O2 (1.25 mmol/L) markedly increased ROS and iNOS production, and decreased the levels of NO, TAOC and TGFβ1 in the culture medium. Furthermore, the H2O2 treatment significantly increased TGFβ1 and Smad2/3 expression in the cells. Addition of salvianolate (0.05, 0.1, and 0.5 g/L) concentration-dependently reversed the H2O2-induced alterations in the culture medium; addition of salvianolate (0.05 g/L) reversed the H2O2-induced increases of TGFβ1 and Smad2/3 expression in the cells. Blockage of TGFβ1 with its antibody (1 mg/L) abolished the above mentioned effects of salvianolate.
Conclusion: Salvianolate inhibits ROS and iNOS production and increases TAOC and NO levels in H2O2-treated cardiomyocytes in vitro via downregulation of Smad2/3 and TGFβ1 expression. High concentration of salvianolate causes cytotoxicity in mouse cardiomyocytes.
Keywords: salvianolate; Salvia miltiorrhiza Bunge; cardiac fibrosis; cardiomyocytes; reactive oxygen species; NO; iNOS; TGFβ1; Smad2/3
* To whom correspondence should be addressed.
E-mail linzhaofen@sina.com
Received 2012-08-14 Accepted 2012-12-24