摘要
目的:探讨补阳还五汤对糖尿病心肌病(DCM)小鼠心肌组织保护作用及炎症和凋亡因子的影响.方法:30只7～8周龄db/db小鼠饲养至10周使其发展为DCM并随机分为模型组、补阳还五汤组、舒洛地特组,每组10只.7～8周龄db/m组小鼠10只作为对照组.给予相应药物治疗12周后,检测小鼠血糖(FBG)、肌酸激酶同工酶(CK-MB)、白细胞介素-6(IL-6)、单核细胞趋化因子-1(MCP-1)、肿瘤坏死因子-α(TNF-α)、甘油三脂(TG)、高密度脂蛋白胆固醇(HDL-C)、低密度脂蛋白胆固醇(LDL-C)水平;qRT-PCR检测Toll样受体4(TLR4)、核转录因子-KB p65(NF-κB p65)mRNA表达;Western Blot检测小鼠心脏组织TLR4、NF-κB p65、Cleaved caspase-3和Cleaved PARP蛋白表达;免疫组化检测心脏组织TLR4、NF-κB p65、Ⅰ型胶原蛋白、Ⅲ型胶原蛋白表达;HE、Masson和天狼星红染色观察心脏病理形态学变化.结果:与对照组比较,模型组小鼠心肌纤维数量明显减少,心肌组织排列紊乱,胶原纤维明显增多;血清FBG、CK-MB、IL-6、MCP-1、TNF-α、TG、LDL-C含量显著升高(P＜0.01,P＜0.05),HDL-C水平显著降低(P＜0.01),TNF-α、IL-6、MCP-1显著升高(P＜0.01);心脏组织TLR4、NF-κBp65、Cleaved caspase-3、Cleaved PARP、Ⅰ 型胶原蛋白、Ⅲ型胶原蛋白表达显著增强(P＜0.01,P＜0.05).与模型组比较,补阳还五汤干预后,DCM小鼠心脏病理损伤程度明显改善;血清CK-MB、IL-6、MCP-1、TNF-α、TG、LDL-C水平显著下降(P＜0.01,P＜0.05),HDL-C水平显著上调(P＜0.05),TNF-α、IL-6、MCP-1 显著降低(P＜0.01,P＜0.05);心肌组织TLR4、NF-κB p65、Cleaved caspase-3、Cleaved PARP、Ⅰ型胶原蛋白、Ⅲ型胶原蛋白表达显著降低(P＜0.01,P＜0.05).结论:补阳还五汤可通过抑制TLR4/NF-κB通路及促凋亡因子表达,对DCM小鼠发挥抗炎和抗凋亡作用,减轻心脏病理损伤,从而发挥心脏保护作用.
Abstract
Objective:To explore the protective effect of Buyang Huanwu Decoction on myocardial tissue and the influence of inflammatory and apoptotic factors in mice with diabetes cardiomyopathy(DCM).Methods:A total of 30 db/db mice(7-8 weeks old)were raised for 10 weeks to develop DCM and randomly divided into model group,Buyang Huanwu Decoction group,and Shuluodite group,with 10 mice in each group.Another 10 db/m mice(7-8 weeks old)were included in the control group.Corresponding medication treatment were provided for 12 weeks,the levels of blood glucose(FBG),creatine kinase isoenzyme(CK-MB),interleukin-6(IL-6),monocyte chemokine 1(MCP-1),tumor necrosis factor-α(TNF-α),triglycerides(TG),high-density lipoprotein cholesterol(HDL-C),and low-density lipoprotein cholesterol(LDL-C)were detected.The mRNA expression of Toll like receptor(TLR4)and nuclear transcription factor-κB p65(NF-κB p65)were detected by qRT-PCR.Western Blot was used to detect the expression of TLR4,NF-κB p65,Cleaved caspase-3 and Cleaved PARP in mouse cardiac tissue.Immunohistochemical was used to detect the expression of TLR4,NF-κB p65,Ⅰ and Ⅲ collagen in cardiac tissue.HE,Masson,and Sirius red staining were used to observe the pathological and morphological changes of the heart.Results:Compared with the control group,the model group mice showed a significant decrease in the number of myocardial fibers,disordered arrangement of myocardial tissue and a significant increase in collagen fibers,the contents of FBG,CK-MB,IL-6,MCP-1,TNF-α,TG,LDL-C were significantly increased(P＜0.01,P＜0.05),while the HDL-C levels was decreased(P＜0.01),TNF-α,IL-6,and MCP-1 were significantly elevated(P＜0.01),at the same time,the expression of TLR4,NF-κB p65,Cleaved caspase-3,Cleaved PARP,and Ⅰ and Ⅲ collagen proteins was significantly enhanced in cardiac tissue(P＜0.01,P＜0.05).Compared with the model group,after intervention with Buyang Huanwu Decoction,the degree of cardiac pathological damage was significantly improved in DCM mice,the levels of CK-MB,IL-6,MCP-1,TNF-α,TG,and LDL-C significantly decreased(P＜0.01,P＜0.05),while the levels of HDL-C were significantly upregulated(P＜0.01),TNF-α,IL-6,and MCP-1 were decreased(P＜0.01,P＜0.05),the expression of TLR4,NF-κB p65,Cleaved caspase-3,Cleaved PARP,Ⅰ and Ⅲ collagen was significantly reduced(P＜0.01,P＜0.05).Conclusion:Buyang Huanwu Decoction can exert anti-inflammatory and anti-apoptotic effects on DCM mice by inhibiting the TLR4/NF-κB pathway and pro-apoptotic factor expression,reducing cardiac pathological damage,and thus exerting cardioprotective effects.
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