Effects of TanshinoneⅡA on hypoxia/reperfusion human kidney-2 cells and their mechanism

Abstract:

Objective：To determine the effect and mechanism of TanshinoneⅡA（TSNⅡA） on oxidative stress and apoptosis of hypox－ia/reperfusion（H/R） HK-2 cells. Methods：HK-2 cells were cultured in vitro for establishment of H/R-induced models. HK-2 cells were divided into the control group，H/R group，H/R+20 μg/mL TSN ⅡA group，H/R+10 μg/mL TSN ⅡA group，and H/R+5 μg/mL TSN ⅡA group. The morphological changes of all groups were detected by light microscope. The cell proliferation of H/R HK-2 cells was measured by CCK-8，and the cell apoptosis by flow cytometry. The levels of ROS were observed by fluorescence microscope，and the expressions of bcl-2，bax，and Cleaved Caspase-3 by Western blot assay. Results：The cell proliferation of H/R group decreased ob－viously compared with that of the control group[（43.6±10.1）% vs. 100%]. Low and medium concentrations of TSN IIA could improve H/R cell proliferation，among which，10 μg/ml TSN ⅡA group presented significant proliferation protection effects compared with H/R group[（79.1±11.1）% vs. （43.6±10.1）%]（P<0.05）. In contrast，high concentration of TSN IIA（20 μg/mL） might reduce the cell proliferation as compared with 10 μg/mL TSN ⅡA group[（51.0±10.4）% vs. （79.1±11.1）%]（P<0.05）. Compared with H/R group，the levels of ROS detected by fluorescence microscope were decreased. The flow cytometry showed the apoptosis of all TSNIIA-treat－ed groups were prohibited，especially for the 20 μg/mL TSNⅡA group（P<0.05）. The Western blot results indicated that TSNⅡA up-regulated the expression of bcl-2 and meanwhile down-regulat－ed the expression of Bax（P<0.05）. Conclusion：Tanshinone I－IA may protect the H/R injury by inhibiting oxidative stress and attenuating cell apoptosis.