还原型谷胱甘肽对脓毒症大鼠心肌组织中Toll样受体4表达的影响

Effects of Glutathione on the Expression of Toll-Like Receptor 4 in Myocardial Tissues in Rats with Sepsis

目的:探讨还原型谷胱甘肽(glutathione,GSH)对脓毒症大鼠急性心肌损伤的保护作用及其机制。方法:采用盲肠结扎穿孔术(cecal ligation and puncture,CLP)建立SD大鼠脓毒症致急性心肌损伤模型。按照随机数字表将大鼠随机分为正常组(n=6)、假手术组(n=18)、脓毒症组(n=18)、GSH干预组(n=18)。GSH干预组于CLP术后经尾静脉注射GSH60 mg/kg,共0.1 mL;假手术组和脓毒症组则给予等量0.9%氯化钠溶液。除正常组外,每组大鼠再按术后6 h、12 h、24 h分为3个亚组(每组各6只),各亚组组大鼠在CLP术后相应时间点采集血清及心肌组织标本。所有大鼠均于血标本采集完毕后处死。HE染色观察心肌组织病理改变;采用自动生化分析仪检测血清肌酸激酶同工酶(creatine kinase MB isoenzyme,CK-MB)的水平,采用实时荧光定量逆转录–聚合酶链反应法检测心肌组织Toll样受体4(Toll-like receptor 4,TLR4)的mRNA表达。结果:与假手术组及正常组比较,脓毒症组大鼠术后6 h血清CK-MB水平和心肌组织TLR4 mRNA表达开始升高,12 h达到高峰,24 h仍明显升高,差异有统计学意义(P<0.05),且术后24 h心肌组织HE染色显示肌纤维结构排列疏松紊乱,间质充血水肿,见大量炎性反应细胞浸润等损伤性改变;与脓毒症组比较,GSH干预组大鼠术后6 h血清CK-MB水平和心肌组织TLR4 mRNA表达已有所下降,术后12 h、24 h明显下降,差异均有统计学意义(P<0.05),且术后24 h时心肌组织上述病理学损伤性改变也明显减轻。结论:心肌组织TLR4 mRNA的高表达在脓毒症致急性心肌损伤中起着重要的作用,而在脓毒症早期应用GSH干预可抑制心肌组织TLR4 mRNA的表达,对脓毒症致急性心肌损伤有保护作用。

Objeetive..To investigate the protective role of glutathione（GSH） in myocardial tissues against sepsis-induced myocardial injury in rats. Methods：Sepsis-induced myocardial injury was induced by cecal ligation and puncture（CLP） in SD rats. Rats were randomly divided into normal group（n = 6） ,sham group（n = 18） ,CLP group（n = 18） and GSH ＋ CLP group（n = 18）, using a random number table. Rats in the latter three groups were sacrificed at 6 h, 12 h and 24 h after the operation（n = 6）. Rats in GSH ＋ CLP group were intravenously administered with GSH （60 mg/kg） after the operation and the rats in the other two groups were treated with same dose of 0.9% sodium chloride solution. The samples of blood and myocardial tissues were harvested at each time point. Toll-like receptor 4（TLR4） mRNA in myocardial tissues was examined by real-time fluorescence reverse transcription-polymerase chain reaction（RT-PCR）, and histological alterations were examined by HE staining. The serum creatine kinase MB isoenzyme（CK-MB） activities was determined by auto analyzer. Results： Compared with normal group and sham group, in CLP group serum CK-MB level and TLR4 mRNA expression in myocardial tissues increased at 6 h after operation, up to top at 12 h and kept high until 24 h（P〈0.05）, and the myocardial injury was confirmed by histological examination. Compared with CLP group, the increases of CK-MB levels and the TLR4 mRNA expression were attenuated at 6 h, 12 h and 24 h after operation in GSH ＋ CLP group（P〈0.05）. Meanwhile, GSH could reversed the microscopic damages, demonstrating its protective effects against sepsis-induced myocardial injury. Conclusions： The increased TLR4 mRNA expression has an important role in the sepsis-induced myocardial injury. GSH ameliorate myocardial injury by suppressing TLR4 mRNA expression in myocardial tissues. Thus, supplementing antiseptic myocardial injury treatment with GSH may be beneficial in the clinical setting.