miRNAs在肺纤维化中的表达及调控机制研究

发布时间：2018-04-03 15:06

  本文选题：特发性肺纤维化　切入点：EMT　出处：《滨州医学院》2012年硕士论文

【摘要】：目的观察miRNAs (let-7d、miR-140、miR-192)在肺纤维化大鼠中的表达变化,研究其对TGF-β1/Smad3信号通路的调控作用及其对下游靶基因表达的影响,探讨miRNA在肺纤维化发病中的作用机制。 方法SD大鼠60只随机分为对照组及模型组,BLM气管注入法建立肺纤维化动物模型。分别于7d、14d、28d处死大鼠10只留取肺组织,HE染色和masson染色动态观察肺组织病理学变化,电镜观察肺组织超微结构的变化,羟脯氨酸碱水解法测定胶原的合成；免疫组化法测定α-SMA、HMGA2、Thy-1、胶原蛋白I的表达；Real-time RT-PCR检测miRNAs (let-7d、miR-140、miR-192)和TGF-β1/Smad3通路相关基因的表达量；Western Blotting检测HMGA2、E-钙粘蛋白的表达。 结果HE染色显示BLM组肺组织按时间顺序呈现由炎症至纤维化的动态改变；masson染色和羟脯氨酸含量检测显示肺组织胶原纤维的含量随肺组织纤维化的程度逐渐增加；电镜下可见Ⅱ型细胞数目较前减少,纤维母细胞增生,胶原纤维明显增多；免疫组化结果示BLM组肺组织HMGA2、α-SMA、胶原蛋白I的表达明显增加而thy-1蛋白的表达逐步下降；Real-time RT-PCR结果显示,Let-7d、miR-140的表达随纤维化程度的加重逐步下降,而miR-192的的表达量逐渐上升,三种miRNA的表达在肺纤维初期即变化明显；TGF-β1、ZEB1、ZEB2, α-SMA的表达的升高,E-Cad基因mRNA的逐步下降。Western Blot结果示HMGA2的表达上升明显、E-钙粘蛋白逐步下降。 结论miRNA (let-7d、miR-140、miR-192)在肺纤维中通过smad3信号通路对EMT过程进行调控。
[Abstract]:Objective to observe the expression of miRNAs let-7dnmiR-140 miR-1922 in rats with pulmonary fibrosis, to study its regulatory effect on TGF- 尾 1/Smad3 signaling pathway and its effect on downstream target gene expression, and to explore the mechanism of miRNA in the pathogenesis of pulmonary fibrosis.Methods 60 SD rats were randomly divided into control group and model group.Ten rats were killed on day 14 and 28, respectively. The pathological changes of lung tissue were observed by HE staining and masson staining. The ultrastructure of lung tissue was observed by electron microscope, and collagen synthesis was measured by hydrolysate of hydroxyproline.The expression of 伪 -SMA-HMGA _ 2 Thy-1, real-time RT-PCR of collagen I, miRNAs let-7dnmiR-140 miR-1922) and TGF- 尾 1/Smad3 pathway related genes were detected by immunohistochemistry. The expression of HMGA _ 2 and E-cadherin was detected by Western Blotting.Results HE staining showed that lung tissue in BLM group showed dynamic changes from inflammation to fibrosis in chronological order. Mass on staining and hydroxyproline content detection showed that the content of collagen fibers in lung tissue increased with the degree of pulmonary fibrosis.Under electron microscope, the number of type 鈪，

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