基于AMPK/TGF-β1信号通路的万金文武汤改善糖尿病心肌病大鼠心肌纤维化研究

Study of Wanjinwenwu decoction on the improvement of myocardial fibrosis in rats with diabetic cardiomyopathy based on AMPK/TGF-β1 signaling pathway

[目的]探讨万金文武汤(WWD)对改善糖尿病心肌病(DCM)大鼠心肌纤维化的作用机制.[方法]采用高脂高糖饲料喂养加单次腹腔注射给予链脲佐菌素方法建立大鼠2型糖尿病心肌病模型.将大鼠随机分为5组,即正常组(Ctrl)、模型组(DCM)、二甲双胍组(MET)、万金文武汤低剂量组(WWDL)及高剂量组(WWDH),除Ctrl组和DCM组外,其余各组分别灌胃给予200 mg·kg^(-1)MET、300 mg·kg^(-1 )WWD、600 mg·kg^(-1)WWD.每周检测各组大鼠的血糖变化;采用ELISA法检测血清中肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)、活性氧(ROS)、超氧化物歧化酶(SOD)、丙二醛(MDA)及血脂含量变化;采用免疫蛋白印迹法检测心肌组织中腺苷酸活化蛋白激酶(AMPK)、转化生长因子β1(TGF-β1)、胶原蛋白(Collagen)Ⅰ及CollagenⅢ等的表达水平.利用Masson染色法评价大鼠心肌纤维化程度.[结果]与DCM组比较,WWDH、WWDL组及MET组血糖水平、心体比值及血清中TNF-α、IL-6、ROS、MDA水平均得到显著改善,SOD活性显著增强(P<0.05),p-AMPK蛋白表达水平显著升高,TGF-β1、CollagenⅠ、CollagenⅢ等蛋白表达水平明显降低(P<0.05).Masson染色结果显示,WWD可逆转原本增多的蓝色胶原纤维,降低心肌纤维化程度.[结论]WWD可显著改善DCM大鼠心肌纤维化,其作用机制认为可能与调控AMPK/TGF-β1信号通路、拮抗心肌结构受损及重塑心肌功能有关系.

OBJECTIVE To explore the mechanism of Wanjinwenwu decoction(WWD)on myocardial fibrosis in rats with diabetic cardiomyopathy(DCM).METHODS The type 2 DCM model was established by feeding rats with high-fat and high-sugar diet and single intraperitoneal injection of streptozotocin(STZ).Rats were randomly divided into 5 groups,control group(Ctrl),model group(DCM),metformin group(MET),Wanjinwenwu decoction low-dose group(WWDL)and high-dose group(WWDH).Except for Ctrl and DCM groups,the other groups were given 200 mg·kg^(-1) MET,300 mg·kg^(-1) WWD and 600 mg·kg^(-1) WWD by gavage,respectively.The changes of blood glucose were detected weekly.The serum levels of tumor necrosis factor-α(TNF-α),interleukin-6(IL-6),reactive oxygen species(ROS),superoxide dismutase(SOD),malonaldehyde(MDA)and blood lipid were detected by ELISA.Western blot was used to detect the expressions of AMPactivated protein kinase(AMPK),transforming growth factor-β1(TGF-β1),collagenⅠand collagenⅢin myocardial tissue.Masson staining was used to evaluate the degree of myocardial fibrosis.RESULTS Compared with the DCM group,the level of blood glucose,heart-to-body ratio,the levels of TNF-α,IL-6,ROS and MDA in the serum were significantly improved,and the activity of SOD was significantly enhanced(P<0.05),and p-AMPK protein expression level was significantly increased,and the protein expression levels of TGF-β1,CollagenⅠand CollagenⅢwere significantly decreased in the WWDH,WWDL and MET groups(P<0.05).Masson staining showed that WWD could reverse the increase of blue collagen fibers and reduce the degree of myocardial fibrosis.CONCLUSION WWD can significantly improve myocardial fibrosis in DCM rats,and its mechanism may be related to regulating AMPK/TGF-β1 signaling pathway,antagonizing myocardial structural damage and remodeling myocardial function.