目的 探讨黄芩素对自身免疫性心肌炎(experimental autoimmune myocarditis，EAM)模型小鼠心肌组织纤维化的影响及其机制。方法 将20只BalB/C小鼠随机分为空白组、EAM组、黄芩素组、地塞米松组，每组5只。在饲养至第0、7天时，除空白组小鼠外余均皮下注射200μL完全弗氏佐剂与α-MyHC肽混合乳剂，饲养至第8天时开始对EAM组、黄芩素组及地塞米松组小鼠分别给予羧甲基纤维素钠、黄芩素、地塞米松灌胃，给药至第21天时行超声心动图检测各组小鼠左心室射血分数(LVEF)、缩短分数(LVFS)、舒张末期内径(LVIDd)及收缩末期内径(LVIDs)。第21天时将各组小鼠脱颈处死后取心脏组织进行HE染色和Masson染色;采用流式细胞术检测各组小鼠脾细胞中Th17/Treg细胞比例;采用实时荧光定量聚合酶链反应(RT-qPCR)和免疫印迹法检测各组小鼠心脏组织中的胶原Ⅰ(Collagen Ⅰ)、基质金属蛋白酶-1(MMP-1)、白细胞介素17A(IL-17A)、Treg转录因子(FOXP3)及白细胞介素10(IL-10)的mRNA及蛋白表达水平。结果 超声心动图显示，黄芩素组小鼠LVEF、LVFS 显著高于 EAM 组(F=51。55、35。38，q=8。08、5。97，P＜0。05)，LVIDd、IVIDs 显著低于 EAM 组(F=4。64、17。72，q=2。88、4。77，P＜0。05)。HE及Masson染色结果显示，黄芩素组及地塞米松组小鼠心脏组织炎症细胞的浸润和胶原纤维的沉积较EAM组有所改善。流式细胞术检测结果显示，黄芩素组及地塞米松组小鼠脾细胞中Th17/Treg细胞比例显著低于EAM组(F=6。80，q=3。60、3。06，P＜0。05)。RT-qPCR结果显示，黄芩素组小鼠心脏组织中 Collagen Ⅰ、ILL-17A mRNA 水平显著低于 EAM 组(F=112。40、77。49，q=14。57、8。27，P＜0。05)，MMP-1、FOXP3、IL-10 mRNA 水平显著高于 EAM 组(F=19。05～144。60，q=5。37～11。21，P＜0。05)。免疫印迹法检测结果显示，黄芩素组小鼠心脏组织中Collagen Ⅰ、IL-17A蛋白表达显著低于EAM组(F=13。70、13。97，q=4。72、5。64，P＜0。05)，MMP-1、FOXP3、IL-10 蛋白表达显著高于 EAM 组(F=6。77～17。21，q=3。32～6。14，P＜0。05)。结论 黄芩素通过调控脾脏中Th17/Treg平衡减轻EAM小鼠心肌损伤，进而减轻心肌组织纤维化进程。
Effect of baicalein on fibrosis of myocardial tissue in EAM model mice
Objective To explore the effect of baicalein on myocardial fibrosis in experimental autoimmune myocarditis(EAM)mice and its mechanism.Methods Twenty BalB/C mice were divided into blank group,EAM group,baicalein group,and dexamethasone group,with 5 mice in each group.On days 0 and 7,all the mice except those in the blank group were subcuta-neously injected with 200 μL of complete Freund's adjuvant and α-MyHC peptide emulsion.Starting from day 8,the mice in the EAM group,baicalein group,and dexamethasone group were intragastrically administered sodium carboxymethyl cellulose,baica-lein,and dexamethasone,respectively.Echocardiography was performed on day 21 of administration to determine left ventricular ejection fraction(LVEF),shortening fraction(LVFS),left ventricular end-diastolic internal diameter(LVIDd),and left ventricu-lar end-systolic internal diameter(LVIDs)in mice across all groups.On day 21,the mice from all groups were euthanized by cervi-cal dislocation and their heart tissues were collected for HE staining and Masson staining.Flow cytometry was used to detect the proportion of Th17/Treg cells among the splenocytes.Real-time fluorescence quantitative PCR(RT-qPCR)and Western blot were used to analyze the mRNA and protein expression of Collagen Ⅰ,matrix metalloproteinase-1(MMP-1),interleukin 17A(IL-17A),Treg transcription factor(FOXP3),and 1L-10 in the cardiac tissue of mice.Results Echocardiographic results showed that the LVEF and LVFS of mice in baicalein group were significantly higher than those in the EAM group(F=51.55,35.38,q=8.08,5.97,P＜0.05),while LVIDd and LVIDs were significantly lower than the EAM group(F=4.64,17.72,q=2.88,4.77,P＜0.05).HE and Masson staining showed that infiltration of inflammatory cells and deposition of collagen fibers in the cardiac tissue of mice were significantly mitigated in the baicalein and dexamethasone groups as compared with the EAM group.Flow cytometry showed that the proportion of Th17/Treg cells was significantly higher in the EAM group than in the baicalein and dexamethasone groups(F=54.75,q=3.60,3.06,P＜0.05).RT-qPCR showed that the mRNA levels of Collagen 1 and IL-17A in the heart tissue of mice were significantly lower in the baicalein group than in the EAM group(F=112.40,77.49,q=14.57,8.27,P＜0.05),while the mRNA levels of MMP-1,FOXP3,and IL-10 were significantly higher in the baicalein group than in the EAM group(F=19.05-144.60,q=5.37-11.21,P＜0.05).Western blot showed that the expression of Collagen Ⅰ and IL-17A was significantly lower in the heart tissue of mice in the baicalein group than that in the EAM group(F=13.70,13.97,q=4.72,5.64,P＜0.05),while the expression of MMP-1,FOXP3,and IL-10 was significantly higher in the baicalein group than in the EAM group(F=6.77-17.21,q=3.32-6.14,P＜0.05).Conclusion Baicalein alleviates myo-cardial damage in EAM mice by regulating the balance between Th17/Treg in the spleen,thereby reducing the progression of myo-cardial tissue fibrosis.
国家科技期刊平台
NSTL
万方数据
维普
