摘要
目的研究肉苁蓉提取物松果菊苷(Echinacoside,ECH)对糖尿病心肌病db/db小鼠模型心肌细胞是否具有保护作用及可能的作用机制。方法将29只SPF级6周龄雄性db/db小鼠和db/m小鼠适应性饲养1周后,按照随机数字表法取9只db/m小鼠作为正常对照组(db/m组,n=9),将db/db小鼠分为糖尿病心肌病组(db/db组,n=10)与松果菊苷干预组(db/db+ECH组,n=10)。其中db/db组和db/m组每日按照0.05 ml/10 g标准进行生理盐水灌胃2周。db/db+ECH组小鼠按松果菊苷300 mg/(kg·d)灌胃2周。每周监测三组的小鼠体重,观察小鼠摄食、饮水及活动情况。于第10周用2%戊巴比妥钠腹腔注射麻醉小鼠并处死,取尾静脉血测定空腹血糖(FBG)。采用HE染色观察心肌组织病理改变,采用Tunel染色法测定心肌细胞凋亡率,采用免疫印迹(Western blot)法检测凋亡相关蛋白p53、p38MAPK、caspase-3和caspase-8的表达水平,采用real-time PCR测定心肌细胞p53、p38MAPK mRNA和caspase mRNA、Bcl-2、Bax mRNA的表达水平。结果与db/m组相比,db/db组小鼠的体重、血糖和血脂水平显著升高(P<0.05或P<0.01)。HE染色切片显示,与db/m组比较,db/db组小鼠的心肌细胞可见明显肥大、坏死,心肌细胞排列结构紊乱;而db/db+ECH组小鼠的心肌细胞形态有所缓解,细胞间隙减少,排列较规则。Tunel染色观察心肌细胞凋亡指数显示,相较于db/m组,db/db组小鼠的心肌细胞凋亡程度显著增高(P<0.01),同时伴有p53蛋白、p38MAPK蛋白、caspase-3和caspase-8蛋白表达量显著升高(P<0.01),且p53、p38MAPK m RNA和caspase mRNA、Bax mRNA表达量明显上升,Bcl-2 mRNA水平下调即Bcl-2/Bax水平降低(P<0.01)。与db/db组比较,db/db+ECH组上述指标均有明显降低,且Tunel染色观察小鼠心肌细胞凋亡指数显著下降(P<0.01)。结论松果菊苷能够明显改善db/db小鼠心肌细胞凋亡,其机制可能与通过p53/p38MAPK/caspase信号转导途径有关。
Objective To study the protective effect of Echinacoside(ECH)from Cistanche deserticola extract on myocardial cells of diabetic cardiomyopathy db/db mice and its possible mechanism.Methods A total of 29 SPF grade 6-week-old male db/db mice and db/m mice were fed adaptively for one week,and 9 cses of db/m mice were selected as the normal control group(the db/m group,n=9)according to random number table method.The db/db mice were divided into the diabetic cardiomyopathy group(the db/db group,n=10)and the Echinacoside intervention group(the db/db+ECH group,n=10).The db/db group and the db/m group were given saline for 2 weeks daily according to 0.05 ml/10 g standard.The mice in the db/db+ECH group were intragastrically administered with Echinacoside 300 mg/(kg·d)for 2 weeks.The weight of mice in three groups was monitored weekly,and the feeding,drinking and activity of mice were observed.At the 10th week,mice were anesthetized by intraperitoneal injection of 2% Pentobarbital Sodium and executed.The fasting blood glucose(FBG)was measured by tail vein blood.HE staining was used to observe the pathological changes of myocardial tissue,Tunel staining was used to determine the apoptotic rate of myocardial cells,Western blot was used to detect the expression levels of apoptotic-related proteins p53,p38MAPK,caspase-3 and caspase-8,and real-time PCR was used to detect the expression levels of p53,p38MAPK mRNA and caspase mRNA,Bcl-2 and Bax mRNA in myocardial cells.Results Compared with the db/m group,the body weight,blood glucose and blood lipid levels of mice in the db/db group increased significantly(P<0.05 or P<0.01).HE staining section showed that compared with db/m group,cardiac myocytes in the db/db group were obviously hypertrophic and necrotic,and the arrangement of cardiac myocytes was disordered.In the db/db+ECH group,the morphology of myocardial cells was alleviated,the intercellular space was reduced and the arrangement was regular.Tunel staining showed that the apoptotic index of cardiomyocytes in the db/db group was significantly higher than that in the db/m group(P<0.01).At the same time,the expression of p53 protein,p38MAPK protein,caspase-3 and caspase-8 protein increased significantly(P<0.01).The expression of p53,p38MAPK mRNA,caspase mRNA and Bax mRNA increased significantly,while the level of Bcl-2 decreased,that was the level of Bcl-2/Bax decreased(P<0.01).Compared with the db/db group,the above indexes in the db/db+ECH group significantly decreased,and the apoptotic index of cardiomyocyte in mice was significantly decreased by Tunel staining(P<0.01).Conclusion ECH can significantly improve the apoptosis of myocardial cells in db/db mice,and its mechanism may be related to the p53/p38MAPK/caspase signal transduction pathway.
作者
张祥
杨洛
徐雄峰
唐凤娟
郝亚荣
ZHANG Xiang;YANG Luo;XU Xiong-feng;TANG Feng-juan;HAO Ya-rong(Department of Geriatrics,Renmin Hospital of Wuhan University,Hubei Province,Wuhan 430060,China)
出处
《中国当代医药》
2019年第13期13-18,共6页
China Modern Medicine
基金
湖北省自然科学基金面上项目(2016CFB673)