基于Rho/ROCK通路介导的细胞凋亡探讨补肝散对衰老大鼠心肌损伤的影响  

Effects of Bugan Powder on Myocardial Injury of Aging Rats Based on Cardiomyocyte Apoptosis Mediated by Rho/ROCK Pathway

作　　者：白德高 张月 霍冉 王杰鹏[1,2] 方芳 方朝义[1,2] BAI Degao;ZHANG Yue;HUO Ran;WANG Jiepeng;FANG Fang;FANG Chaoyi(Hebei University of Chinese Medicine,Shijiazhuang 050200 Hebei,China;Hebei Key Laboratory of Integrated Chinese and Western Medicine for Lung Disease Research,Shijiazhuang 050091 Hebei,China)

机构地区：[1]河北中医学院,河北石家庄050200 [2]河北省中西医结合肺病研究重点实验室,河北石家庄050091

出　　处：《中药新药与临床药理》2022年第8期1032-1039,共8页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：河北省中医药类科研计划项目(2020136);河北中医学院省属高校基本科研业务费专项项目(YJZ2019006);河北中医学院研究生创新资助项目(XCXZZSS2021010);2022年度河北省高等学校科学技术研究项目(ZD2022025)。

摘　　要：目的观察衰老大鼠心肌损伤特征并探讨补肝散的干预机制。方法将32只SPF级SD雄性大鼠随机分为空白组、模型组、补肝散组和维生素E组,除空白组腹腔注射等量生理盐水外,其余各组均腹腔注射D-半乳糖(D-galactose,D-gal)制备衰老大鼠模型。模型复制当日起,补肝散组与维生素E组分别灌胃浓度为14.06 g·kg^(-1)的补肝散和27 mg·kg^(-1)的维生素E,空白组和模型组给予等量生理盐水。以上均每日1次,连续8周,并于模型复制末日测定抓力。取血清及心脏组织,测定心脏脏器指数;检测血清丙二醛(MDA)含量和超氧化物歧化酶(SOD)活性变化;检测血清谷草转氨酶(AST)、肌酸激酶(CK)、乳酸脱氢酶(LDH)含量;苏木素-伊红(HE)染色观察大鼠心肌组织病理形态学变化;Masson染色观察大鼠心肌组织胶原沉积和纤维化情况;TUNEL染色观察心肌细胞凋亡情况;免疫组织化学染色(ABC法)观察Ras同源基因A(RhoA)、Rho相关卷曲螺旋形成蛋白激酶1(ROCK1)、Bcl-2相关X蛋白(Bax)和B细胞淋巴瘤2(Bcl-2)表达特征;免疫印迹法(Western Blot)检测心肌组织RhoA、ROCK1、Bax和Bcl-2的蛋白表达。结果与空白组比较,模型组大鼠心肌细胞形态改变,呈现嗜酸性变性,胞质染色不均,部分心肌细胞坏死,心肌组织胶原沉积和纤维化明显;抓力及心脏脏器指数降低,MDA含量升高,SOD活性降低,心肌细胞凋亡指数及AST、CK、LDH含量升高(均P<0.05);RhoA、ROCK1、Bax蛋白表达增加,Bcl-2蛋白表达减少(均P<0.05)。与模型组比较,补肝散组大鼠心肌细胞轻度嗜酸性变性,偶有坏死、肥大情况,心肌组织胶原沉积较少,偶有纤维化产生,损伤情况较轻;大鼠抓力及心脏脏器指数升高,MDA含量降低,SOD活性升高,心肌细胞凋亡指数及AST、CK、LDH含量降低(均P<0.05);RhoA、ROCK1、Bax蛋白表达减少,Bcl-2蛋白表达增加(均P<0.05)。结论补肝散可通过调节Rho/ROCK介导的凋亡途径减轻衰老大鼠�Objective To observe the characteristics of myocardial injury in aging rats and explore the interventional mechanism of Bugan Powder.Methods Thirty-two specific pathogen-free(SPF)grade Sprague-Dawley(SD)male rats were randomly divided into blank group,model group,Bugan Powder group and vitamin E group.The aging rat model was established by intraperitoneal injection of D-galactose(D-gal).The blank group was injected intraperitoneally with equal amount of normal saline at the same time.The Bugan powder group and the vitamin E group were respectively given 14.06 g·kg^(-1) of Bugan powder and 27 mg·kg^(-1) of vitamin E after modeling,and the blank group and the model group were given the same amount of normal saline.All the groups were administered once a day for 8 consecutive weeks,and the grasping force was measured at the end of the model repliction.Serum and cardiac tissue were collected to measure cardiac index,activity of superoxide dismutase(SOD),the contents of malondialdehyde(MDA),serum aspartate aminotransferase(AST),creatine kinase(CK)and lactate dehydrogenase(LDH).Hematoxylin-eosin(HE)staining was used to observe the pathological changes of myocardial tissue of rats.Masson staining was used to observe the collagen deposition and fibrosis in myocardial tissue of rats.Cardiomyocyte apoptosis was detected by TUNEL staining.The expression characteristics of Ras homolog gene A(RhoA),Rhoassociated coiled-coil forming protein kinase 1(ROCK1),Bcl-2-associated X protein(Bax)and B-cell lymphoma-2(Bcl-2)were observed by immunohistochemical staining(ABC method).Western Blot assay was used to detect the protein expression of RhoA,ROCK1,Bax and Bcl-2 in myocardial tissue.Results Compared with the blank group,the model group obviously showed morphological changes in the cardiomyocytes of rats,eosinophilic degeneration,uneven cytoplasmic staining,focal necrosis of the cardiomyocytes,collagen deposition and fibrosis in myocardial tissue.The grasping force and cardiac index decreased,the content of MDA increased,SOD acti

关 键 词：衰老 RHO/ROCK信号通路 细胞凋亡 补肝散 大鼠 心肌损伤 

分 类 号：R285.5[医药卫生—中药学]