心康冲剂对慢性心力衰竭大鼠心肌细胞焦亡的影响  

Effect of Xinkang Granule(心康冲剂)on Cardiomyocyte Pyroptosis in Rats with Chronic Heart Failure

作　　者：郭冰 贺璇 余意 刘蓉芳 王敏[1] 李园 李亮[3] 毛以林[1] GUO Bing;HE Xuan;YU Yi;LIU Rongfang;WANG Min;LI Yuan;LI Liang;MAO Yilin(The Second Affiliated Hospital of Hunan University of Chinese Medicine,Changsha 410005,Hunan,China;Wuyi Hospital of Traditional Chinese Medicine,Jiangmen 529099,Guangdong,China;Provincial Key Laboratory of TCM Diagnostics,Hunan University of Chinese Medicine,Changsha 410208,Hunan,China)

机构地区：[1]湖南中医药大学第二附属医院,湖南长沙410005 [2]江门市五邑中医院,广东江门529099 [3]湖南中医药大学中医诊断学湖南省重点实验室,湖南长沙410208

出　　处：《中华中医药学刊》2024年第4期108-112,I0022-I0024,共8页Chinese Archives of Traditional Chinese Medicine

基　　金：国家自然科学基金青年科学基金项目(82004283);湖南省自然科学基金面上项目(2020JJ4473);湖南省中医药管理局科研计划重点项目(C2022004);湖南中医药大学“一方”研究生创新项目(2021YF05)。

摘　　要：目的探讨心康冲剂对慢性心力衰竭大鼠心肌细胞焦亡的调控作用。方法46只SD大鼠,其中随机取6只大鼠为空白组,余40只经腹腔注射阿霉素药物建立慢性心力衰竭模型,模型成功后将32只大鼠随机分为模型组12只、心康冲剂组10只、缬沙坦组10只。缬沙坦组以等效剂量0.03 g/(kg·d)灌胃、心康冲剂组给予0.5 g/(kg·d)等效剂量灌胃。空白组与模型组每日等量生理盐水灌胃,连续4周。实验结束后,心脏超声检测各组大鼠心功能,HE、Masson染色观察心脏组织形态,透射电镜下观察心肌组织超微结构,免疫组化法检测焦亡底物消皮素D-N端片段(GasderminD-N,GSDMD-N)表达,酶联免疫吸附法(Enzyme linked immunosorbent assay,ELISA)检测大鼠血清白介素1β(Interleukin-1β,IL-1β)、白介素18(Interleukin-18,IL-18)表达,实时荧光定量PCR法检测各组大鼠心肌组织核苷酸结合寡聚化结构域样受体3(NOD-like receptor thermal protein domain associated protein 3,NLRP3)通路各mRNA,Western blot法检测NLRP3通路各蛋白表达量。结果与空白组相比,模型组大鼠左室射血分数(Left ventricular ejection fractions,LVEF)、左心室短轴缩短率(Left ventricular fractional shortening,LVFS)均明显降低(P<0.01),染色结果显示心肌细胞死亡增多、纤维化严重,且GSDMD-N焦亡蛋白表达在胞膜和胞浆,透射电镜结果显示模型组中心肌细胞膜有破裂,有孔隙形成,线粒体肿胀严重,血液中炎症因子IL-18、IL-1β明显升高(P<0.01),心肌组织中NLRP3、半胱氨酸天冬氨酸特异性蛋白酶-1(Cysteine-containing aspartate-specific proteases-1,Caspase-1)、GSDMD mRNA及GSDMD-N蛋白表达均明显升高(P<0.01)。与模型组相比,心康冲剂、缬沙坦干预后心肌细胞焦亡减轻,LVEF、LVFS都不同程度地增加(P<0.01);染色及电镜结果显示心肌病理学损伤减轻;炎症因子IL-18、IL-1β及细胞焦亡通路NLRP3、Caspase-1、GSDMD/GSDMD-N的mRNA及蛋白都有所降低(P<0.01)Objective To investigate the regulatory effect of Xinkang Granule(心康冲剂)on cardiomyocyte pyroptosis in rats with chronic heart failure(CHF).Methods Six of 46 SPF male and famale SD rats were randomly selected as the blank group,and the other 40 rats were injected with adriamycin intraperitoneally to establish CHF model.After the model was successful,32 rats were randomly divided into model group(n=12),Xinkang Granule group(n=10)and valsartan group(n=10).The valsartan group was given 0.03 g/(kg·d)equivalent dose by gavage,and the Xinkang Granule group was given 0.5 g/(kg·d)equivalent dose by gavage.The blank group and model group were given the same amount of normal saline daily for 4 weeks.At the end of the experiment,the cardiac function of each group was detected by cardiac ultrasound.HE and Masson staining were used to observe the morphology of heart tissue.The ultrastructure of myocardial tissue was observed under transmission electron microscope(TEM).The expression site of pyroptosis substrate gasdermin D-N(GSDMD-N)was detected by immunohistochemistry,and enzyme-linked immunosorbent assay was used to detect the expressions of interleukin-1β(IL-1β)and interleukin-18(IL-18)in serum of rats.Real-time PCR was used to detect mRNA of NOD-like receptor thermal protein domain associated protein 3(NLRP3)pathway in each group.Western blot was used to detect the protein expression levels of NLRP3 pathway.Results Compared with those of the blank group,left ventricular ejection fractions(LVEF)and left ventricular fractional shortening(LVFS)were significantly decreased(P<0.01)in the model group.The results of staining showed the increased number of myocardial cell death and the promotion of fibrosis.Moreover,GSDMD-N was expressed in the cell membrane and cytosol.In the model group,the results of TEM showed that the myocardial membrane was broken,the pore formation was observed,and the mitochondrial swelling was severe.Inflammatory factors IL-18 and IL-1βin blood were significantly increased(P<0.01).The express

关 键 词：心康冲剂 慢性心力衰竭 NLRP3 心肌细胞焦亡 

分 类 号：R289.5[医药卫生—方剂学]