目的:探讨心脏上皮/内皮间充质转化(EMT/EndMT)与病毒性心肌炎心肌纤维化的关系.方法:40只BALB/c 小鼠随机分为2组,分别为对照组(n=16)和心肌炎组(n=24).2组小鼠每周1次分别腹腔注射无病毒培养液或柯萨奇病毒B3(CVB3);于第7 d后随机处死2组小鼠各8只,30 d后处死其余存活小鼠.以苦味酸天狼星红染色检测小鼠心脏胶原容积分数(CVF),ELISA法检测血清胶原前肽的含量变化,以实时RT-PCR法和Western blotting方法检测小鼠心脏中EMT/EndMT蛋白组包括启动因子转化生长因子β1(TGF-β1)和Wnt1、转录因子Twist1、上皮细胞标志物上皮细胞钙黏素(E-cadherin)、内皮细胞表面标志血管内皮钙黏素(VE-cadherin)、间充质蛋白如成纤维细胞特异蛋白(FSP-1)和α-平滑肌肌动蛋白(α-SMA)等的基因和蛋白表达情况.结果:急性病毒性心肌炎时,出现EMT/EndMT现象,其特征为VE-cadherin和E-cadherin丢失,FSP-1和α-SMA表达上调,启动因子TGF-β1和Wnt1表达增高,转录因子Twist1表达增加,胶原合成增多;慢性期未发现上皮/内皮细胞表型丢失,间充质标志蛋白仍表达上调,胶原合成增多,TGF-β1、Twist1和Wnt1表达依然增加.结论:EMT/EndMT参与了急性病毒性心肌炎心肌纤维化的形成,慢性期未发现EMT/EndMT参与心肌纤维化.%AIM: To investigate the relationship between epithelial/endothelial to mesenchymal transition ( EMT/EndMT ) and myocardial fibrosis in viral myocarditis.METHODS: Forty BALB/c mice were randomized to control group ( n = 16, intraperitoneally injected with virus - free vehicle weekly ) and viral myocarditis group [n = 24, intraperitoneally injected with coxsackievirus B3 ( CVB3 ) weekly].Eight mice in each group were sacrificed on day 7 and the remaining survived mice were sacrificed on day 30 after inoculation.The cardiac collagen volume fraction ( CVF ) was calculated by picrosirius red - stained sectioning, and the serum concentrations of propeptides of procollagen ( indicating collagen synthesis ) were detected by enzyme - linked immunosorbent assay.The mRNA expression and protein levels of EMT/EndMT proteomes including initiating factors such as transforming growth factor β1 ( TGF - β1 ) and Wnt1, transcripting factor Twist1, epithelial/endothelial markers ( E - cadherin and VE - cadherin ), mesenchymal markers such as fibroblast specific protein 1 ( FSP - 1 ) and α - smooth muscle actin (α - SMA ) in the hearts were detected by real - time RT - PCR and Western blotting, respectively.RESULTS: The changes of EMT/EndMT, characterized by loss of VE - cadherin and E -cadherin, gain of FSP - 1 and α - SMA, over - expression of initiating factors ( TGF - β1 and Wnt1 ) and transcripting factor Twist1, and increased collagen synthesis, were observed in acute viral myocarditis.Loss of epithelial/endothelial markers was not found in chronic myocarditis while over - expression of mesenchymal markers,TGF - β1 , Twist1 and Wnt1, and increased collagen synthesis remained.CONCLUSION: EMT/EndMT participates in myocardial fibrosis in acute stage but not in chronic stage of viral myocarditis.
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