摘要
Abstract
[Objective]To study the effects of Moringa oleifera leaf extract on oxidative stress injury,and expression of apoptosis and autophagy proteins in H9c2 cardiomyocytes exposed to hypoxia.[Method]H9c2 cardiomyocytes were divided into normoxia control group,hypoxia control group,and Moringa oleifera leaf extract 20,50,100,and 200 μg/mL groups,6 wells in each group.The normoxia control group was cultured in 37 ℃,5%CO2 incubator,and the hypoxic control group and Moringa oleifera leaf extract groups were cultured in 94.5%N2,0.5%O2,5%CO2 hypoxia workstation for 24 h and 48 h,respectively.CCK-8 method was used to detect cell viability and determine the best experimental conditions.H9c2 cardiomyocytes were divided into normoxia control group,hypoxia control group and Moringa oleifera leaf extract 100 μg/mL group.After cultured for 24 hours,the cells were collected and the activities of GSH-Px and SOD were measured by kits,the level of ROS was measured by flow cytometry method,the mitochondrial membrane potential by JC-1 probe,the level of autophagy by MDC assay,the protein expression by Western Blot and the protein quantity by BCA method.[Result]According to the results of cell viability test,the optimal experimental conditions were determined as follows:the exposure time to hypoxia was 24 hours,and the concentration of Moringa oleifera leaf extract was 100 μg/mL.Compared with the normoxia control group,the activities of SOD and GSH-Px and mitochondrial membrane potential were significantly reduced(P<0.05),the levels of ROS and autophagy were significantly increased(P<0.05),and the expression of apoptosis-related protein cleaved-caspase3 and the protein ratio of Bax to Bcl-2 were significantly increased(P<0.05).The expression level of autophagy-related protein Beclin-1 was increased(P<0.05),while the expres-sion level of p62 was decreased(P<0.05)in H9c2 cells of hypoxic control group.Compared with the hypoxic control group,the activity of SOD and the mitochondrial membrane potential were significantly increased,ROS and autophagy levels were decreased,the expression levels of antioxidant proteins Nrf2 and HO-1 were up-regulated(P<0.05).The expression level of apoptosis-related protein cleaved-caspase3 and the protein ratio of Bax to Bcl-2 was decreased(P<0.05).The expression level of autophagy-related protein Beclin-1 decreased(P<0.05),while that of p62 increased(P<0.05)in Moringa oleifera leaf extract group.[Conclusion]The extract of Moringa oleifera leaves can reduce oxidative stress,and could decrease the level of apoptosis and autophagy by regulating the expression of related proteins in H9c2 cardiomyocytes exposed to hypoxia.关键词
辣木叶提取物/H9c2心肌细胞/氧化应激/凋亡/自噬Key words
Moringa oleifera leaf extract/H9c2 cardiomyocytes/oxidative stress/apoptosis/autophagy
