基于雌激素受体探讨梓醇对抗糖剥夺心肌细胞损伤的作用
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摘要
目的基于雌激素受体,探讨梓醇介导自噬对糖剥夺心肌细胞氧化损伤的作用和机制。方法采用糖剥夺6 h复制大鼠心肌细胞(H9c2)损伤模型,观察中药地黄中有效成分梓醇对其保护作用及机制。将细胞分为5组:对照组、模型组、梓醇(0. 28、2. 8、28μmol·L~(-1))组。检测梓醇对细胞中活性氧(ROS)、丙二醛(MDA)、超氧化物歧化酶(SOD)的影响;电镜观察梓醇对细胞自噬的影响。利用雌激素受体(ER)阻断剂,探究梓醇对抗氧化损伤机制是否由ER介导。进一步采用慢病毒转染使ERα低表达后,观察梓醇作用前后氧化损伤、自噬水平的变化。结果与空白对照组相比,模型组ROS、MDA水平升高,SOD水平降低,自噬相关蛋白表达无明显差异;与模型组比较,梓醇能够升高SOD水平降低氧化损伤,升高自噬水平。采用慢病毒转染ERα后,与空载病毒梓醇组相比,梓醇抑制氧化损伤、促进自噬的作用被逆转。结论梓醇能通过上调ERα表达激活自噬,发挥抗氧化损伤、保护心肌的作用。
Aim To investigate the effect of catalpolmediated autophagy on oxidative damage of cardiomyocytes induced by glucose deprivation based on estrogen receptor( ER) and the related mechanism. Methods The cardiomyocytes( H9 c2) injury model in rat was induced by glucose deprivation for 6 h. The protective effect of catalpol on H9 c2 injury and its mechanism were observed. The cells were divided into five groups: control group,model group,catalpol group( 0. 28,2. 8,28 μmol·L~(-1)). The effects of catalpol on reactive oxygen species( ROS),malondialdehyde( MDA) and superoxide dismutase( SOD) in cells were detected,and the effect of catalpol on autophagy was assessed by electron microscopy. ER blockers was used to detect whether the protective effect of catalpol on oxidative damage was related to ER. Lentivirus transfection was used to lower the expression of ER alpha,and the oxidative damage and autophagy of H9 c2 treated with catapol or not were observed. Results Compared with control group,the levels of ROS and MDA increased and SOD decreased in model group,and there was no significant difference in the expression of autophagy related proteins. Compared with model group,catalpol could reduce oxidative damage and increase autophagy level. After transfection of ER alpha with lentivirus,catalpol inhibited the level of oxidative damage and promoted the effect of autophagy compared with the empty virus catalpol group. Conclusions Catalpol can activate autophagy by up-regulating the expression of ER alpha and inhibiting the oxidative damage of cardiomyocytes.
Aim To investigate the effect of catalpolmediated autophagy on oxidative damage of cardiomyocytes induced by glucose deprivation based on estrogen receptor( ER) and the related mechanism. Methods The cardiomyocytes( H9 c2) injury model in rat was induced by glucose deprivation for 6 h. The protective effect of catalpol on H9 c2 injury and its mechanism were observed. The cells were divided into five groups: control group,model group,catalpol group( 0. 28,2. 8,28 μmol·L~(-1)). The effects of catalpol on reactive oxygen species( ROS),malondialdehyde( MDA) and superoxide dismutase( SOD) in cells were detected,and the effect of catalpol on autophagy was assessed by electron microscopy. ER blockers was used to detect whether the protective effect of catalpol on oxidative damage was related to ER. Lentivirus transfection was used to lower the expression of ER alpha,and the oxidative damage and autophagy of H9 c2 treated with catapol or not were observed. Results Compared with control group,the levels of ROS and MDA increased and SOD decreased in model group,and there was no significant difference in the expression of autophagy related proteins. Compared with model group,catalpol could reduce oxidative damage and increase autophagy level. After transfection of ER alpha with lentivirus,catalpol inhibited the level of oxidative damage and promoted the effect of autophagy compared with the empty virus catalpol group. Conclusions Catalpol can activate autophagy by up-regulating the expression of ER alpha and inhibiting the oxidative damage of cardiomyocytes.
引文
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[13]Ma Y,Gai Y,Yan J,et al.Puerarin attenuates anoxia/reoxygenation injury through enhancing Bcl-2 associated athanogene 3 expression,a modulator of apoptosis and autophagy[J].Med Sci Monit,2016,22:977-83.
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[2]Zhang Y,Su W,Zhang Q,et al.Glycine protects H9C2 cardiomyocytes from high glucose-and hypoxia/reoxygenation-induced injury via inhibiting PKCβ2 activation and improving mitochondrial quality[J].J Diabetes Res,2018,2018:1-8.
[3]Yang Y,Karakhanova S,Hartwig W,et al.Mitochondria and mitochondrial ROS in cancer:novel targets for anticancer therapy[J].J Cell Physiol,2016,231(12):2570-81.
[4]Chen X,Tan M,Xie Z,et al.Inhibiting ROS-STAT3-dependent autophagy enhanced capsaicin-induced apoptosis in human hepatocellular carcinoma cells[J].Free Radic Res,2016,50(7):744-55.
[5]彭辉.地黄多糖对心肌缺血/再灌注损伤大鼠保护作用研究[J].辽宁中医药大学学报,2016,18(6):39-42.[5]Peng H.Protection of Rehmannia glutinosa polysaccharides on rats with myocardial ischemia-reperfusion injury[J].J Liaoning Univ Tradit Chin Med,2016,18(6):39-42.
[6]Wang Y,Liao D,Qin M,Li X.Simultaneous determination of catalpol,aucubin,and geniposidic acid in different developmental stages of Rehmannia glutinosa Leaves by high performance liquid chromatography[J].J Anal Methods Chem,2016,2016(2):1-6.
[7]Lu SF,Huang Y,Wang N,et al.Cardioprotective effect of electroacupuncture pretreatment on myocardial ischemia/reperfusion injury via antiapoptotic signaling[J].Evid Based Complement Alternat Med,2016,2016(25):1-9.
[8]Dutta D,Xu J,Kim JS,et al.Upregulated autophagy protects cardiomyocytes from oxidative stress-induced toxicity[J].Autophagy,2013,9(3):328-44.
[9]Emani SM,Mccully JD.Mitochondrial transplantation:applications for pediatric patients with congenital heart disease[J].Transl Pediatr,2018,7(2):169.
[10]左玮,梅丹.高血糖通过抑制线粒体自噬加重大鼠脑缺血/再灌注损伤[J].中国药理学通报,2016,32(6):846-53.[10]Zuo W,Mei D.Hyperglycemia aggravates cerebral ischemia/reperfusion injury in rats by inhibiting mitochondrial autophagy[J].Chin Pharmacol Bull,2016,32(6):846-53.
[11]Pons DG,Nadal-Serrano M,Torrens-Mas M,et al.The phytoestrogen genistein affects breast cancer cells treatment depending on the ERα/ERβratio[J].J Cell Biochem,2016,117(1):218-29.
[12]Lin C,Lu Y,Yan X,et al.Catalpol protects glucose-deprived rat embryonic cardiac cells by inducing mitophagy and modulating estrogen receptor[J].Biomed Pharmacother,2017,89:973.
[13]Ma Y,Gai Y,Yan J,et al.Puerarin attenuates anoxia/reoxygenation injury through enhancing Bcl-2 associated athanogene 3 expression,a modulator of apoptosis and autophagy[J].Med Sci Monit,2016,22:977-83.
[14]Duan W J,Li Y F,Liu F L,et al.A SIRT3/AMPK/autophagy network orchestrates the protective effects of trans-resveratrol in stressed peritoneal macrophages and RAW264.7 macrophages[J].Free Radic Biol Med,2016,95:230-42.