微创医学

【摘要】目的探讨3-甲基腺嘌呤(3-MA)对风湿性心脏病（RHD）瓣膜纤维化相关指标的影响。方法取30只Lewis雌性大鼠，利用电脑随机数将大鼠分为正常组、模型组和实验组，每组10只。模型组和实验组通过后足垫、腹部皮下注射抗原Ⅰ、抗原Ⅱ造模，大鼠多处关节出现红肿，B超检查显示二尖瓣有不同程度反流，表明RHD模型建立成功。建模成功后，第8~9周，实验组大鼠于腹部皮下注射3-MA 2.5 mg/kg，1次/周，模型组大鼠腹部皮下注射等量生理盐水；第10~24周，模型组、实验组大鼠腹部皮下注射0.5 mL抗原Ⅱ，1次/周，正常组大鼠在相同时间以同样方式注射等量生理盐水。采用免疫印迹（Western blot）法检测3组大鼠二尖瓣组织中PI3K、Akt、α-平滑肌肌动蛋白（α-SMA）、LC3Ⅱ及Ⅰ型胶原蛋白的相对表达量；采用ELISA法检测肿瘤坏死因子-α（TNF-α）、白介素-6（IL-6）、白介素-8（IL-8）水平。结果正常组、实验组、模型组大鼠的二尖瓣组织中PI3K、Akt、LC3Ⅱ的相对表达量依次升高，组间差异均有统计学意义(均P＜0.05)；模型组α-SMA、Ⅰ型胶原蛋白的相对表达量均高于正常组、实验组，差异均有统计学意义（均P＜0.05）；正常组与实验组的α-SMA、Ⅰ型胶原蛋白相对表达量比较，差异均无统计学意义（均P＞0.05）。正常组、实验组、模型组大鼠的二尖瓣组织中TNF-α、IL-6、IL-8水平依次升高，组间差异均有统计学意义(均P＜0.05)。结论3-MA能够有效抑制RHD大鼠二尖瓣组织自噬，减轻心脏瓣膜纤维化；调控PI3K/Akt信号通路可能是3-MA起抑制纤维化的作用机制之一；自噬作用的调控可能与TNF-α、IL-6及IL-8水平具有一定的相关性。

【Abstract】 ObjectiveTo investigate the effect of 3-methyladenine (3-MA) on related indexes of valve fibrosis in rheumatic heart disease (RHD). MethodsThirty female Lewis rats were divided into normal group, model group and experimental group by computer random number method, with 10 rats in each group. The model group and experimental group were modeled by subcutaneous injection of antigen Ⅰ and antigen Ⅱ into the hind foot pad and abdomen. Multiple joints of the rats showed redness and swelling, and two-dimensional ultrasonography showed different degrees of mitral valve regurgitation, indicating that the model was successfully established. After successful modeling, rats in the experimental group were injected subcutaneously with 3-MA 2.5 mg/kg once a week in the abdomen at the 8th to 9th week, while rats in the model group were injected subcutaneously with the same amount of normal saline in the abdomen. From 10th to 24th week, rats in the model group and experimental group were injected with 0.5 mL antigen Ⅱ subcutaneously in the abdomen once a week, while rats in the normal group were injected with the same amount of normal saline at the same time and in the same way. The relative expressions of PI3K, Akt, alpha-smooth muscle actin (α-SMA,) LC3 Ⅱ and type Ⅰ collagen in the mitral valve tissues of the three groups were detected by Western blot. The levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-8 (IL-8) were detected by ELISA. ResultsThe relative expressions of PI3K, Akt and LC3Ⅱ in mitral valve tissue of normal group, experimental group and model group increased in turn, and the differences among the groups were statistically significant (all P＜0.05). Compared with the normal group and the experimental group, the relative expressions of α-SMA, type Ⅰ collagen were higher in the model group, with statistically significant differences (all P＜0.05). There were no statistically significant differences in the relative expressions of α-SMA and type Ⅰ collagen between the normal group and the experimental group (all P＞0.05). The levels of TNF-α, IL-6 and IL-8 in mitral valve tissue of normal group, experimental group and model group increased in turn, and the differences among the groups were statistically significant (all P＜0.05). Conclusion3-MA can effectively inhibit autophagy of mitral valve tissue in RHD rats and reduce heart valve fibrosis. The regulation of PI3K/Akt signaling pathway may be one of the mechanisms of 3-MA inhibiting fibrosis. The regulation of autophagy may be related to the levels of TNF-α, IL-6 and IL-8.