大鼠急性肺栓塞后TGF-beta对SM_(22)-alpha表达的调节

Expression of SM22-alpha regulated by TGF-beta after acute PE

目的:研究大鼠急性肺栓塞模型肺组织中SM22-alpha的表达变化以及TGF-β信号转导系统对SM22-alpha表达的调节作用。方法:建立大鼠急性肺栓塞模型,分别在急性肺栓塞后1、8、24和48h开胸取出肺组织。常规提取肺组织总RNA和总蛋白,以正常组为对照,采取半定量RT-PCR的方法研究SM22-alpha,TGF-β1和TGF-βR在mRNA水平表达的变化;采用Western-blot方法进一步验证SM22-alpha,TGF-β1,TGF-βR,Smad7以及磷酸化的pSmad2和pSmad3在蛋白水平的表达变化;采用免疫组织化学方法检测大鼠肺组织中SM22-alpha,TGF-β1和TGF-βR在肺栓塞前后表达的变化及其组织分布情况。结果:在大鼠急性肺栓塞后的不同时间点,SM22-alpha的mRNA水平和蛋白水平均逐渐降低,在24和48h下降最为明显。参与SM22-alpha表达调节的TGF-β1和TGF-βR在mRNA水平和蛋白水平的表达也逐渐减低;免疫组化研究表明SM22-alpha主要分布在肺动脉中膜的平滑肌细胞(SMCs),在急性肺栓塞后表达显著降低。TGF-β1和TGF-βR主要分布在肺动脉壁、支气管壁和肺泡壁,急性肺栓塞后TGF-β1和TGF-βR在上述组织内的表达均明显降低。Western-blot检测发现急性肺栓塞后TGF-β信号转导分子中磷酸化的pSmad2和pSmad3在蛋白水平的表达逐渐减低,而对于TGF-β信号转导系统起抑制作用的Smad7分子的表达却明显上升。结论:大鼠急性肺栓塞后肺组织内TGF-β信号转导系统的抑制导致了SM22-alpha的表达下降,从而影响了肺动脉系统内SMCs的收缩功能。

Objective： To study the expression changes of SM22-alpha in the lung tissues of a rat acute pulmonary embolism （PE） model and its regulation by TGF-beta signal transducting system. Methods： We constructed a rat acute PE model. The samples of lung tissues were collected at different time points lh, 8h, 24h and 48h, The normal rats were used as control. The mRNA level changes of SM22- alpha, TGF-β and TGF-βR Ⅱ were identified by semi-quantitative RT-PCR, and the protein level changes of SM22-alpha, TGF-β1, TGF-βR Ⅱ , Smad7 and phosphorylated pSmad2 and pSmad3 were identified by western blot methods. The immunohistochemical method was employed to study the distribution and expression changes of SM22-alpha, TGF-β2 and TGF-βR 1 in the lung tissues after aerate PE. Results： ;At different time points, the mRNA levels and protein levels of SM22-alpha decreased in the lung tissues of rat acute PE models, obviously at 24h and 48h. The mRNA levels and protein levels of TGF-β1 and TGF-βR Ⅱ , which play an important role in the regulation of the expression of SM22-alpha, were also downregulated after acute PE. The immunohistoehemical study indicated that SM22-alpha was mainly expressed in the smooth muscle cells（SMCs） of the media of the pulmonary arteries. The expression of it was downregulated after acute PE. The protein of TGF-β1 and TGF-βR Ⅱ were mainly distributed in the pulmonary arteries, bronchus and alveolar walls, and were all down-regulated after acute PE. The protein pSmad2 and pSmad3, which are the downstream signaling intermediates of TGF-beta signal transducting system, were all down-regulated after acute PE, but the expression of Smad7, which has the ability of blocking TGF-β receptor signaling system, was up-regulated obviously after acute PE. Conclusion： The blocking of TGF-beta signal transdueting system resulted in the down-regulation of SM22-alpha, which would affect the contraction ability of SMCs in the pulmonary artery system.