摘要
目的探索三七总皂苷(panax notoginseng saponins,PNS)预处理心肌细胞减少缺氧/复氧(hypoxiareoxygenation,H/R)损伤的作用机制。方法设置对照组(常规培养H9c2细胞)、模型组(H/R损伤的H9c2细胞)、PNS组(加入PNS处理的H/R损伤的H9c2细胞)、二甲氧基雌二醇(2-methoxyestradiol,2-ME2)组(加入2-ME2处理的H/R损伤的H9c2细胞)和PNS+2-ME2组心肌细胞(同时加入PNS和2-ME2处理的H/R损伤的H9c2细胞),采用酶联免疫吸附法检测心肌细胞损伤,蛋白质印迹法检测线粒体自噬信号通路表达,免疫荧光法检测心肌细胞自噬活性。结果模型组高于对照组的指标有肌酸激酶(creatine kinase,CK)(0.37±0.05 vs.2.41±0.66)、乳酸脱氢酶(lactate dehydrogenase,LDH)(2.20±0.69 vs.4.05±1.08)、天门冬氨酸氨基转移酶(aspartate amino transferase,AST)(38.17±9.02 vs.107.83±20.89)、细胞自噬标志物LC3(0.17±0.05 vs.0.63±0.66)、低氧诱导因子-1α(hypoxic inducible factor-1α,HIF-1α)(0.22±0.09 vs.0.66±0.08)和B淋巴细胞瘤-2/腺病毒E1B 19kDa相互作用蛋白3(B lymphocytoma-2/adenovirus E1B 19kDa interacting protein 3,BNIP3)(0.15±0.02 vs.0.59±0.13),差异均有统计学意义(P<0.05)。PNS组低于模型组的指标有CK(2.41±0.66 vs.1.55±0.39)、LDH(4.05±1.08 vs.3.19±0.88)、AST(107.83±20.89 vs.66.52±11.43),差异均有统计学意义(P<0.05);PNS组高于模型组的指标有LC3(0.63±0.66 vs.0.85±0.23)、HIF-1α(0.66±0.08 vs.0.91±0.22)和BNIP3(0.59±0.13 vs.0.82±0.17),差异均有统计学意义(P<0.05)。2-ME2+PNS组低于模型组的指标有CK(2.41±0.66 vs.2.08±0.54)、LDH(4.05±1.08 vs.3.51±0.92)、AST(107.83±20.89 vs.83.49±19.53);2-ME2+PNS组高于模型组的指标有LC3(0.63±0.66 vs.0.70±0.54)、HIF-1α(0.66±0.08 vs.0.72±0.10)和BNIP3(0.59±0.13 vs.0.64±0.12),差异均有统计学意义(P<0.05)。2-ME2组高于模型组的指标有CK(2.41±0.66 vs.3.62±0.98)、LDH(4.05±1.08 vs.5.67±1.39)、AST(107.83±20.89 vs.192.13±28.97),差异均有统计学意义(P<0.05);2-ME2组低于模型组的指标有HIF-1α(0.66±0.08 vs.0.27±0.03)和BNIP3(0.59±0.13 vs.0.25±0.07),差异均有统计学意义(P<0.05)。与对照组相比,模型组心肌细胞LC3荧光斑点数明显增加(P<0.05);与模型组相比,PNS组和2-ME2+PNS组心肌细胞LC3荧光斑点数明显增加,2-ME2组LC3荧光斑点数明显减少(P<0.05)。结论PNS能够通过激活HIF-1α/BNIP3线粒体自噬信号通路,减少心肌细胞H/R损伤。
Objective To explore the action mechanisms of panax notoginseng saponins(PNS)pretreatment on reducing hypoxia/reoxygenation(H/R)injury in cardiomyocytes.Methods The control group(routinely cultured H9c2 cells),model group(H9c2 cells with H/R injury),PNS group(H9c2 cells with H/R injury treated with PNS),2-methoxyestradiol(2-ME2)group(H9c2 cells with H/R injury treated with 2-ME2)and PNS+2-ME2 group(H9c2 cells with H/R injury treated with PNS and 2-ME2).The cardiomyocytes damage was detected by ELISA.The expressions of mitochondrial autophagy signaling pathways were detected by Western Blot.The activity of cardiomyocytes autophagy was detected by immunofluorescence method.Results Creatine kinase(CK)(0.37±0.05 vs.2.41±0.66),lactate dehydrogenase(LDH)(2.20±0.69 vs.4.05±1.08),aspartate amino transferase(AST)(38.17±9.02 vs.107.83±20.89),LC3(0.17±0.05 vs.0.63±0.66),hypoxic inducible factor-1 alpha(HIF-1α)(0.22±0.09 vs.0.66±0.08)and b lymphocytoma-2/adenovirus E1B 19kDa interacting protein 3(BNIP3)(0.15±0.02 vs.0.59±0.13),all with statistically significant differences(P<0.05).The PNS group was lower than the model group in CK(2.41±0.66 vs.1.55±0.39),LDH(4.05±1.08 vs.3.19±0.88)and AST(107.83±20.89 vs.66.52±11.43),all with statistically significant differences(P<0.05).The PNS group was higher than the model group in LC3(0.63±0.66 vs.0.85±0.23),HIF-1α(0.66±0.08 vs.0.91±0.22)and BNIP3(0.59±0.13 vs.0.82±0.17),and the differences were statistically significant(P<0.05).2-ME2+PNS group was lower than the model group in CK(2.41±0.66 vs.2.08±0.54),LDH(4.05±1.08 vs.3.51±0.92),AST(107.83±20.89 vs.83.49±19.53);the indexes that were higher in the 2-ME2+PNS group than in the model group were LC3(0.63±0.66 vs.0.70±0.54),HIF-1α(0.66±0.08 vs.0.72±0.10)and BNIP3(0.59±0.13 vs.0.64±0.12),all statistically significant(P<0.05).2-ME2 group was higher than the model group in terms of CK(2.41±0.66 vs.3.62±0.98),LDH(4.05±1.08 vs.5.67±1.39),AST(107.83±20.89 vs.192.13±28.97),all with statistically significant differences(P<0.05).The indicators that were lower in the 2-ME2 group than the model group were HIF-1α(0.66±0.08 vs.0.27±0.03)and BNIP3(0.59±0.13 vs.0.25±0.07),all with statistically significant differences(P<0.05).Compared with the control group,the number of LC3 fluorescent spots in cardiomyocytes was significantly increased in the model group(P<0.05);compared with the model group,the number of LC3 fluorescent spots in cardiomyocytes was significantly increased in the PNS and 2-ME2+PNS groups and significantly decreased in the 2-ME2 group(P<0.05).Conclusion PNS can reduce H/R injury in cardiomyocytes by activating HIF-1α/BNIP3 mitochondrial autophagy signaling pathways.
作者
王丽红
韩勇
王超海
詹舟虹
卢梦恺
金佳炜
刘新文
WANG Lihong;HAN Yong;WANG Chaohai;ZHAN Zhouhong;LU Mengkai;JIN Jiawei;LIU Xinwen(Department of Pharmacy,Affiliated Hospital of Shaoxing University of Arts and Sciences(Shaoxing Municipal Hospital),Zhejiang,Shaoxing 312000,China)
出处
《中国现代医生》
2022年第35期15-19,共5页
China Modern Doctor
基金
浙江省医药卫生科技计划项目(2019RC297)。
