摘要
目的观察柴胡三参胶囊对心肌缺血再灌注损伤(MIRI)大鼠心肌细胞焦亡的影响,从ROS/TXNIP/NLRP3通路探讨其作用机制。方法将大鼠随机分为假手术组、模型组、曲美他嗪组和柴胡三参胶囊低、中、高剂量组,每组10只。柴胡三参胶囊低、中、高剂量组分别按0.15、0.3、0.6 g/kg灌胃,曲美他嗪组按5.4 mg/kg灌胃,假手术组和模型组予等体积生理盐水灌胃,28 d后造模。除假手术组外,其余各组采用左冠状动脉前降支结扎法建立MIRI大鼠模型。HE染色观察大鼠心肌组织病理变化,透射电镜观察心肌组织超微结构,ELISA检测血清肌酸激酶同工酶(CK-MB)、乳酸脱氢酶(LDH)、白细胞介素(IL)-18、IL-1β含量,DCFH-DA荧光探针检测心肌组织活性氧(ROS)含量,免疫荧光染色检测心肌细胞焦亡,Western blot和RT-PCR分别检测心肌组织硫氧还蛋白互作蛋白(TXNIP)、核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)、半胱氨酸蛋白酶1(Caspase-1)、消皮素D(GSDMD)蛋白和mRNA表达。结果与假手术组比较,模型组大鼠心肌纤维结构不完整,细胞出现脂肪样变、间质水肿及出血,伴有少量炎性细胞浸润,胞质疏松,胞膜破裂严重、有大量孔隙形成;血清CK-MB、LDH、IL-18、IL-1β含量显著增加;心肌组织ROS含量显著增加,细胞焦亡率显著升高,TXNIP、NLRP3、Caspase-1、GSDMD蛋白和mRNA表达显著升高,差异均有统计学意义(P<0.01)。与模型组比较,各给药组大鼠心肌纤维结构均有不同程度改善,心肌细胞脂肪样变减少,间质水肿及出血程度减轻,无明显炎性细胞浸润,胞质较致密,胞膜相对完整、孔隙减少;柴胡三参胶囊中、高剂量组和曲美他嗪组大鼠血清CK-MB、LDH、IL-18、IL-1β含量显著减少,心肌组织ROS含量显著减少,细胞焦亡率显著降低,TXNIP、NLRP3、Caspase-1、GSDMD蛋白和mRNA表达显著降低,差异均有统计学意义(P<0.05,P<0.01)。结论柴胡三参胶囊可有效减轻MIRI大鼠心肌细胞损伤,其机制可能与下调ROS/TXNIP/NLRP3信号通路,抑制心肌细胞焦亡有关。
Objective To observe the effects of Chaihu Sanshen Capsules on cardiomyocyte pyroptosis in rats with myocardial ischemia-reperfusion injury(MIRI);To explore its mechanism based on ROS/TXNIP/NLRP3 pathway.Methods The rats were randomly divided into sham-operation group,model group,trimetazidine group,and Chaihu Sanshen Capsules low-,medium-and high-dosage groups,with 10 rats in each group.The Chaihu Sanshen Capsules low-,medium-and high-dosage groups were given 0.15,0.3 and 0.6 g/kg by gavage respectively,and the trimetazidine group was given 5.4 mg/kg by gavage;the sham-operation group and the model group were given the same volume of normal saline by gavage.Except for the sham-operation group,the model was established by ligation of the anterior descending branch of the left coronary artery after gavage for 28 days.HE staining was used to observe the pathological changes of myocardial tissue;the ultrastructure of myocardium was observed by transmission electron microscope;the contents of serum CK-MB,LDH,IL-18 and IL-1βwere detected by ELISA;the content of ROS in myocardial tissue was detected by DCFH-DA fluorescent probe;the pyroptosis of cardiomyocytes was detected by immunofluorescence staining;the expressions of TXNIP,NLRP3,Caspase-1,GSDMD protein and mRNA in myocardium were detected by Western blot and RT-PCR.Results Compared with the sham-operation group,the structure of myocardial fibers were not complete,with cellular steatosis,interstitial edema and bleeding,accompanied by a small amount of inflammatory cell infiltration,the cytoplasm of cardiomyocytes was loose,the cell membrane was broken seriously,and a large number of pores were formed;the contents of serum CK-MB,LDH,IL-18,IL-1βincreased significantly;the content of ROS in myocardial tissue increased significantly,the pyroptosis rate of cardiomyocytes increased significantly,the expressions of TXNIP,NLRP3,Caspase-1 and GSDMD protein and mRNA in myocardial tissue increased significantly,with statistical significance(P<0.01).Compared with the model group,the myocardial fibers were improved in varying degrees in the administration groups,the steatosis of cardiomyocytes was reduced,the degree of interstitial edema and bleeding was alleviated,and there was no obvious inflammatory cell infiltration,the cytoplasm of cardiomyocytes was dense,the cell membrane was relatively complete,and the pore of cell membrane was less than that of the model group;the contents of serum CK-MB,LDH,IL-18 and IL-1βin Chaihu Sanshen Capsules medium-and high-dosage groups,and trimetazidine group decreased significantly,the content of ROS in myocardial tissue was significantly reduced,the pyroptosis rate of cardiomyocytes decreased significantly,the expressions of TXNIP,NLRP3,Caspase-1 and GSDMD protein and mRNA decreased significantly,with statistical significance(P<0.05,P<0.01).Conclusion Chaihu Sanshen Capsules can effectively reduce cardiomyocyte injury in MIRI rats,and its mechanism may be related to downregulating ROS/TXNIP/NLRP3 signaling pathway and inhibiting cardiomyocyte pyroptosis.
作者
张杼惠
曹蛟
刘蓉芳
陈程
杨耀闾
何涛
谭彩
谭琦
刘建和
ZHANG Zhuhui;CAO Jiao;LIU Rongfang;CHEN Cheng;YANG Yaolü;HE Tao;TAN Cai;TAN Qi;LIU Jianhe(The First Hospital of Hunan University of Chinese Medicine,Changsha 410007,China;Sub-center of National Clinical Research Center for Cardiovascular Diseases of Traditional Chinese Medicine,Changsha 410007,China;Wuyi Hospital of Traditional Chinese Medicine,Jiangmen 529000,China;Huangpu Hospital of Guangdong Second Hospital Traditional Chinese Medicine,Guangzhou 510530,China;Guangdong Province Engineering Technology Research Institute of Traditional Chinese Medicine,Guangzhou 510530,China)
出处
《中国中医药信息杂志》
CAS
CSCD
2023年第5期114-120,共7页
Chinese Journal of Information on Traditional Chinese Medicine
基金
国家自然科学基金(82004283、81774130)
湖南省中医药科研计划重点项目(2021011)
湖南省卫健委科研计划项目(202116001377)
湖南省教育厅科学研究项目(21A0234、19C1434)
长沙市自然科学基金重点项目(kq2202450)。
作者简介
通讯作者:刘建和,E-mail:ljhtcm1@163.com。
