黄芪多糖调节HMGB1-RAGE信号通路对自身免疫性心肌炎大鼠心肌损伤的影响

The Effect of Astragalus Polysaccharides on Myocardial Injury in Rats with Experimental Autoimmune Myocarditis by Regulating HMGB1-RAGE Signaling Pathway

目的:探讨黄芪多糖(AP)调节高迁移率族蛋白Box-1-晚期糖基化终产物受体(HMGB1-RAGE)信号通路对自身免疫性心肌炎(EAM)大鼠心肌损伤的影响。方法:取50只SPF级SD大鼠,适应性喂养1周,随机分为对照组、EAM组、AP组(30 g/kg)、rHMGB1组(8μg/kgr)、AP+rHMGB1组[Ap(30 g/kg)+rHMGB1(8μg/kg)],除对照组(弗氏完全佐剂)外,其余各组通过左、右后足垫皮下注射抗原佐剂乳化液方法建立EAM大鼠模型并按照上述剂量进行干预;3周后,超声仪检测心率(HR)、左心室射血分数(LVEF)和舒张末期内径(LVEDs);酶联免疫吸附(ELISA)检测各组大鼠血清中炎症因子水平;马松(Masson′s)及苏木精-伊红(HE)染色检测心肌组织纤维化及病理学变化;免疫印迹(Western blotting)及实时荧光定量PCR(qRT-PCR)检测HMGB1、RAGE蛋白及mRNA表达。结果:与对照组比较,EAM组心肌结构紊乱及纤维化严重,病理评分、肿瘤坏死因子-α(TNF-α)水平、白细胞介素-6(IL-6)水平、HR、LVEDs、HMGB1、RAGE蛋白及mRNA表达显著增加,LVEF显著下降(P<0.05);与EAM组比较,AP组病理损伤及纤维化得到改善,病理评分、TNF-α水平、IL-6水平、HR、LVEDs、HMGB1、RAGE蛋白及mRNA表达显著降低,LVEF显著增加(P<0.05),但rHMGB1组病理损伤及纤维化进一步加重,病理评分、TNF-α水平、IL-6水平、HR、LVEDs、HMGB1、RAGE蛋白及mRNA表达显著增加,LVEF显著下降(P<0.05);与AP组比较,AP+rHMGB1组病理损伤及纤维化稍加重,病理评分、TNF-α水平、IL-6水平、HR、LVEDs、HMGB1、RAGE蛋白及mRNA表达显著增加,LVEF显著下降(P<0.05);与rHMGB1组比较,AP+rHMGB1组病理损伤及纤维化得到改善,病理评分、TNF-α、IL-6水平、HR、LVEDs、HMGB1、RAGE蛋白及mRNA表达显著降低,LVEF显著增加(P<0.05)。结论:AP可能通过抑制HMGB1-RAGE信号通路改善EAM大鼠心肌损伤。

Objective:To investigate the effect of astragalus polysaccharides(AP)on myocardial injury in rats with experimental autoimmune myocarditis(EAM)by regulating high mobility group Box-1-receptor for advanced glycation end products(HMGB1-RAGE)signaling pathway.Methods:Totally 50 rats were fed adaptively for one week and randomly separated into control group,EAM group,AP group(30 g/kg),rHMGB1 group(8μg/kg),and AP+rHMGB1 group[AP(30 g/kg)+rHMGB1(8μg/kg)].Except for the control group(Freund's complete adjuvant),other groups established EAM rat models by subcutaneous injection of antigen adjuvant emulsion into the left and right hind foot pads,and intervened according to the above dosage.After three weeks,ultrasound was applied to detect heart rate(HR),left ventricular ejection fraction(LVEF)and end-diastolic diameter(LVEDs).Enzyme-linked immunosorbent(ELISA)was applied to detect the levels of inflammatory factors in the serum of rats in each group.Masson's and Hematoxylin-eosin(HE)staining were applied to detect myocardial tissue fibrosis and pathological changes.Western blotting and real-time fluorescence quantitative PCR(qRT-PCR)were applied to detect the protein and mRNA expression of HMGB1 and RAGEA.Results:Compared with control group,the EAM group showed severe myocardial structural disorder and fibrosis.And the pathological score,levels of TNF-αand IL-6,HR,LVEDs,and protein and mRNA expression of HMGB1,RAGE obviously increased in EAM group,while LVEF obviously decreased in EAM group(P<0.05).Compared with EAM group,the pathological damage and fibrosis were improved in AP group.And the pathological score,levels of TNF-αand IL-6,HR,LVEDs,and protein and mRNA expression of HMGB1,RAGE obviously decreased in AP group,while LVEF obviously increased in AP group(P<0.05).However,the pathological damage and fibrosis further worsened in rHMGB1 group.And the pathological score,levels of TNF-αand IL-6,HR,LVEDs,and protein and mRNA expression of HMGB1,RAGE obviously increased in rHMGB1 group,while LVEF obviously decreased in rHMGB1 group(P<0.05).Compared with AP group,the pathological damage and fibrosis were slightly aggravated in AP+rHMGB1 group.And the pathological score,levels of TNF-αand IL-6,HR,LVEDs,and protein and mRNA expression of HMGB1,RAGE obviously increased in AP+rHMGB1 group,while LVEF obviously decreased in AP+rHMGB1 group(P<0.05).Compared with rHMGB1 group,the pathological damage and fibrosis were improved in AP+rHMGB1 group.And the pathological score,levels of TNF-αand IL-6,HR,LVEDs,and protein and mRNA expression of HMGB1,RAGE obviously decreased in AP+rHMGB1 group,while LVEF obviously increased in AP+rHMGB1 group(P<0.05).Conclusion:AP may improve myocardial injury in EAM rats by inhibiting HMGB1-RAGE signaling pathway.