基于AMPK/SIRT1/PGC-1α信号通路研究香青兰总黄酮对大鼠心肌缺血再灌注损伤的保护机制
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篇名: | 基于AMPK/SIRT1/PGC-1α信号通路研究香青兰总黄酮对大鼠心肌缺血再灌注损伤的保护机制 |
TITLE: | Study on Protective Mechanism of Dracocephalum moldavica Total Flavonoids against Myocardial Ischemia- reperfusion Injury in Rats Based on AMPK/SIRT 1/PGC-1α Signaling Pathway |
摘要: | 目的:研究香青兰总黄酮(TFDM)对腺苷酸活化蛋白激酶(AMPK)/沉默信息调节因子1(SIRT1)/过氧化物酶体增殖物激活受体γ辅激活因子1α(PGC-1α)信号通路的影响,探究其保护大鼠心肌缺血再灌注损伤(MIRI)的作用机制。方法:将50只健康雄性SD大鼠随机分为假手术组、模型组、TFDM组[60mg/(kg·d),以提取物计]、CompoundC+TFDM组[灌胃60mg/(kg·d)TFDM+再灌注前15min尾静脉注射250μg/kgCompoundC(AMPK抑制剂)]、EX-527+TFDM组[灌胃60mg/(kg·d)TFDM+再灌注前20min腹腔注射5mg/kgEX-527(SIRT1抑制剂)],每组10只。每天灌胃给药1次,连续7d。末次灌胃给药后,假手术组大鼠行假手术,其余4组大鼠均采用结扎冠状动脉左前降支缺血30min、再灌注2h构建MIRI模型。再灌注结束后,采用苏木精-伊红染色法观察大鼠心肌组织病理学变化;采用反相高效液相色谱法测定其心肌组织中腺苷三磷酸(ATP)、腺苷二磷酸(ADP)、腺苷一磷酸(AMP)及烟酰胺腺嘌呤二核苷酸(NAD+)的含量;采用实时荧光定量-聚合酶链式反应法检测大鼠心肌组织中AMPK、SIRT1和PGC-1αmRNA表达水平;采用Westernblotting法检测大鼠心肌组织中AMPK蛋白的磷酸化水平和SIRT1、PGC-1α蛋白表达水平。结果:与假手术组比较,模型组大鼠心肌纤维排列紊乱、横向条纹消失,细胞肿胀破裂、坏死,细胞核变形移位;心肌组织中ATP、NAD+含量和AMPK、SIRT1、PGC-1αmRNA表达水平以及SIRT1、PGC-1α蛋白表达水平均显著降低(P<0.05或P<0.01),ADP、AMP含量及AMPK蛋白的磷酸化水平均显著升高(P<0.01)。与模型组比较,TFDM组大鼠心肌病理学形态明显改善;心肌组织中ATP、NAD+含量和AMPK、SIRT1、PGC-1αmRNA表达水平以及AMPK蛋白的磷酸化水平和SIRT1、PGC-1α蛋白表达水平均显著升高(P<0.05或P<0.01),ADP、AMP含量均显著降低(P<0.01)。与TFDM组比较,CompoundC+TFDM组和EX-527+TFDM组大鼠上述指标的改善作用均被逆转(P<0.05或P<0.01)。结论:TFDM可能是通过激活AMPK/SIRT1/PGC-1α信号通路,调节能量代谢,从而发挥其对心肌的保护作用。 |
ABSTRACT: | OBJECTIVE:To s tudy the effects of Dracocephalum moldavica total flavonoids (TFDM)on AMPK/SIRT 1/PGC-1α signaling pathway ,and to explore the mechanism of its protective effect on myocardial ischemia reperfusion injury (MIRI)rats. METHODS:Totally 50 healthy male SD rats were randomly divided into sham operation group ,model group ,TFDM group [ 60 mg/(kg·d),by extract] ,Compound C+TFDM group [ig administration of 60 mg/(kg·d)TFDM+intravenous injection of 250 μg/kg Compound C(AMPK inhibitor )via tail vein 15 min before reperfusion] ,EX-527+TFDM group [ig administration of 60 mg/ (kg·d)TFDM+ip injection of 5 mg/kg EX- 527(SIRT1 inhibitor)20 min before reperfusion] ,with 10 rats in each group. They were given relevant medicine intragastrically ,once a day ,for consecutive 7 days. After last ig administration ,sham operation group underwent sham operation ,other 4 groups were established MIRI model by ligating left anterior descending coronary artery , ischemia for 30 min and reperfusion for 2 h. After reperfusion ,the myocardial histopathological changes were observed by HE staining;RP-HPLC method was used to determine the contents of ATP ,ADP,AMP and NAD + in cardiac tissue. mRNA expressions of AMPK ,SIRT1 and PGC- 1α were detected by quantitative real-time PCR assay. Western blotting assay was the expressions of SIRT 1 and PGC- 1α protein in myocardium. RESULTS: Compared with sham operation group , model group showed myocardial fib ers arranged disorder and horizontal stripes disappearance ,cell swelling burst and necrosis ,and nuclei deformation displacement ;the contents of ATP and NAD+,mRNA expression of AMPK ,SIRT1 and PGC- 1α,protein expression of SIRT 1 and PGC- 1α in cardiac tissue were decreased significantly (P<0.05 or P<0.01);the contents of ADP and AMP ,the phosphorylation level of AMPK protein were increased significantly (P<0.01). Compared with model group ,myocardial pathological morphology were improved significantly in TFDM group ;the contents of ATP and NAD + in cardiac tissue ,mRNA expression of AMPK ,SIRT1 and PGC- 1α,the phosphorylation level of AMPK protein ,the protein expression of SIRT 1 and PGC- 1α were increased significantly(P<0.05 or P< 0.01),while the contents of ADP and AMP were decreased significantly (P<0.01). Compared with TFDM group ,improvement effects of Compound C + TFDM group and EX- 527 + TFDM group on above indexes were reversed (P<0.05 or P<0.01). CONCLUSIONS:TFDM may play a protective role on myocardium by activating AMPK/SIRT 1/PGC-1α signaling pathway and regulating energy metabolism. |
期刊: | 2021年第32卷第03期 |
作者: | 赵云丽,袁勇,马晓莉,黄川生,文志萍,郭新红,王新春 |
AUTHORS: | ZHAO Yunli,YUAN Yong,MA Xiaoli,HUANG Chuansheng,WEN Zhiping,GUO Xinhong,WANG Xinchun |
关键字: | 香青兰总黄酮;能量代谢;腺苷酸活化蛋白激酶 ;沉默信息调节因子 1;过氧化物酶体增殖物激活受体 γ辅激活因子 1α; |
KEYWORDS: | Dracocephalum moldavica total flavonoids ;Energy metabolism ;AMPK;SIRT1;PGC-1α;Mechanism |
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