ABSTRACT
Objective: To explore the protective mechanism of resvertrol (RSV) suppressing the rapid electrical stimulation incurred oxidative stress injury in neonatal rats cardiomyocytes. Methods: The neonatal rats cardiac ifbroblasts and myocytes were isolated by double enzyme digestion and differential adhesion method, and the cardiomyocytes were randomly divided into 5 groups:①Control (CTR) group,②Rapid electrical stimulation (RES) group,③RES+APO group, cells were pretreated with NADPH oxidase inhibitor APO,④RES+RSV group,⑤RES+AIP group, cells were pretreated with CaMKII inhibitor AIP. In order to conifrm whether RSV protection was via MsrA-OX-CaMKⅡpathway, the cells were further divided into another 3 groups:①DMSO control group,②RSV+DMSO group,③RES+ RSV+DMSO group. The best dose of RSV was measured with Kit-8 by cardiomyocytes surviving condition, the optimal electrical stimulation time was detected with ELISA by Ang II level in conditioned medium. The level of reactive oxygen species (ROS) in cardiomyocytes was detected by flow cytometry, the protein expressions of MsrA, Nox4, Nox2, P22phox, OX-CaMK II and apoptosis related cleaved caspase-3 were observed by Western blot analysis.Results:①Compared with CTR group, RES group showed increased AngII secretion with increased protein expressions of Nox4, Nox2, P22phox, OX-CaMK II and cleaved caspase-3.②Compared with RES group, the RES+APO, RES+RSV, RES+AIP groups had decreased ROS level, the ROS was even lower in RES+RSV group.③Compared with RES group, the RES+APO, RES+RSV groups presented decreased protein expressions of Nox4, Nox2, P22phox, OX-CaMK II and cleaved caspase-3, while RES+AIP group only had decreased Nox2, OX-CaMK II and cleaved caspase-3.④Compared with DMSO control group, RES+ RSV+DMSO group had the lower level of cleaved caspase-3 expression. Conclusion: RSV has protective effect on rapid electrical stimulation incurred oxidative stress injury in neonatal rats cardiomyocytes, which might be via NADPH oxidase with the increased MsrA expression .
ABSTRACT
Objective: To explore the effect of atorvastatin (Atv) on high glucose-induced oxidative stress injury in human umbilical vein endothelial cells (HUVECs) by SIRT1/NADPH oxidase pathway with the possible mechanisms. Methods: HUVECs were cultured in low glucose medium and then divided into 6 experimental groups:①Normal group,②Osmotic pressure control group,③High glucose (HG) group,④HG+Atv (0.1, 1.0, 10.0)μmol/L group,⑤HG+sirtinol (SIRT1 inhibitor) group,⑥HG+apocynin (NOX4 inhibitor) group, and HUVECs were further cultured for 24 hours. The cell proliferation was examined by CCK-8 kit, ROS level was detected by lfow cytometry method, protein expressions of SIRT1 and NOX4 were measured by Western blot analysis. Results: ① Compared with Normal group, HG group had decreased HUVECs proliferation, Atv improved the HG inhibited proliferation in a does dependent manner. ② HG group had the higher level of ROS, increased NOX4 protein expression and decreased SIRT1 protein expression. ③ In HG condition, Atv up-regulated SIRT1 expression and down-regulated ROS and NOX4 expressions in a does dependent manner.④In HG condition, sirtinol decreased SIRT1 expression, increased NOX4 expression, and apocynin decreased NOX4 expression, while it had no inlfuence on SIRT1 expression. Conclusion: Atorvastatin could resist HG-induced oxidative stress injury in HUVECs, which might be related to up-regulated SIRT1 expression, and SIRTI plays the role in NADPH oxidase at upstream.