薯蓣皂苷元对心肌缺血再灌注损伤大鼠细胞凋亡的影响及相关机制的分析

Effects of Diosgenin on Apoptosis and NLRP3/caspase-1 Pathway in Rats With Myocardial Ischemiareperfusion Injury

目的:探究薯蓣皂苷元对心肌缺血再灌注损伤(MIRI)大鼠细胞凋亡的影响,并初步分析其对NOD样受体蛋白3(NLRP3)/半胱氨酸天冬氨酸特异性蛋白酶1(Caspase-1)信号通路的作用。方法:采用左冠状动脉前降支结扎法建立MIRI大鼠模型,将造模成功的72只大鼠,按照随机数字表法分成MIRI组、薯蓣皂苷元低剂量组(50 mg/kg)、薯蓣皂苷元中剂量组(75 mg/kg)和薯蓣皂苷元高剂量组(100 mg/kg),每组18只;另取18只大鼠为假手术组。造模完成后,给予相应药物灌胃,每日1次。末次给药后,记录每组大鼠在不同时间段的心脏血液动力学参数;三苯基四氮唑(TTC)法检测大鼠心肌梗死面积;苏木素-伊红染色检测各组心肌组织病理学变化;TUNEL法检测心肌细胞凋亡情况;ELISA法检测大鼠血清中白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)的含量;免疫蛋白印迹法检测大鼠心肌组织中NLRP3、Caspase-1、凋亡相关斑点样蛋白(ASC)、IL-1β蛋白表达水平。结果:与假手术组相比,MIRI组大鼠心肌梗死面积更大,心肌细胞凋亡率、左心室舒张末期压力(LVEDP)、血清IL-6、IL-1β、TNF-α含量及心肌组织中NLRP3、Caspase-1、ASC、IL-1β蛋白表达水平均显著升高(P均<0.05),左心室收缩压(LVSP)、左心室射血分数(LVEF)、短轴缩短率(FS)及最大收缩/舒张期压力变化速率(±dp/dtmax)均显著降低(P均<0.05);与MIRI组相比,薯蓣皂苷元不同剂量干预组大鼠心肌梗死面积较小,细胞凋亡率、LVEDP、血清IL-6、IL-1β、TNF-α含量及心肌组织中NLRP3、Caspase-1、ASC、IL-1β蛋白表达水平均显著降低(P均<0.05),LVSP、LVEF、FS及±dp/dtmax均显著升高(P均<0.05)。结论:薯蓣皂苷元可改善MIRI大鼠的心功能,减少炎症反应和细胞凋亡,可能与抑制NLRP3/caspase-1信号通路有关。

Objectives:To investigate the effect of diosgenin on apoptosis in rats with myocardial ischemia-reperfusion injury(MIRI),and analyze its effect on the signaling pathway of NOD-Like receptor protein 3(NLRP3)/cysteinyl aspartate specific proteinase-1(Caspase-1).Methods:The rat model of MIRI was established by ligation of left anterior descending coronary artery.72 rats with successful modeling were randomly divided into MIRI group,low-dose diosgenin group(50 mg/kg),medium-dose diosgenin group(75 mg/kg)and high-dose diosgenin group(100 mg/kg),with 18 rats in each group.Another 18 rats were taken as sham operation group.After modeling,the rats were given corresponding diosgenin by gavage once daily for 4 weeks.After the last administration,echocardiography was performed and cardiac hemodynamic parameters of rats in each group were recorded The infarct size was measured by triphenyltetrazolium(TTC)method.The pathological changes of myocardium were detected by HE staining.The apoptosis of cardiomyocytes was detected by TUNEL method.The contents of interleukin-6(IL-6),interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α)in serum were detected by ELISA.The protein expression levels of NLRP3,caspase-1,apoptosis associated speck-like protein containing a CARD(ASC)and IL-1βwere detected by Western blot.Results:The infarct size,myocardial apoptosis rate,left ventricular end-diastolic pressure(LVEDP),contents of serum IL-6,IL-1βand TNF-α,and the protein expression levels of NLRP3,caspase-1,ASC and IL-1βin myocardial tissue were significantly higher(all P<0.05),while left ventricular systolic pressure(LVSP),left ventricular ejection fraction(LVEF),fractional shortening(FS)and maximum systolic/diastolic pressure change rate(±dp/dtmax)were significantly lower in the MIRI group than in the sham operation group(all P<0.05).The infarct size,myocardial apoptosis rate,LVEDP,contents of serum IL-6,IL-1βand TNF-α,and the protein expression levels of NLRP3,caspase-1,ASC and IL-1βin myocardial tissue were significantly lower(all P<0.05),while LVSP,LVEF,FS and±dp/dtmax were significantly higher in diosgenin-treated groups than in MIRI group(all P<0.05).Conclusions:Diosgenin can improve the cardiac function of MIRI rats,reduce inflammatory response and cell apoptosis,which may be related to the inhibition of NLRP3/Caspase-1 signaling pathway.