急性肺栓塞后富半胱氨酸蛋白1的表达变化及其机制研究  

The expression changes of cysteine-rich protein 1 and its mechanism after acute pulmonary embolism

作　　者：李圣青[1] 李树钧[1] 刘阿茹[1] 赵峰[1] 简文[1] 遆新宇[1] 欧阳海峰[1] 

机构地区：[1]第四军医大学西京医院呼吸科,西安市710032

出　　处：《中华老年多器官疾病杂志》2008年第3期215-219,共5页Chinese Journal of Multiple Organ Diseases in the Elderly

摘　　要：目的研究大鼠急性肺栓塞模型肺组织中富半胱氨酸蛋白1(CRP1)的表达变化以及内皮素-1(ET-1)和血小板源性生长因子(PDGF)对肺动脉平滑肌细胞(PASMCs)中CRP1表达的影响。方法建立大鼠急性肺栓塞模型,分别在急性肺栓塞后1,8,24,48h开胸取出肺组织。常规提取肺组织的总RNA和总蛋白,以正常组为对照,采取半定量RT-PCR和Westernblot方法研究CRP1在mRNA水平和蛋白水平的表达变化;采用免疫组织化学的方法检测大鼠肺组织中CRP1在肺栓塞前后表达的变化及其组织分布情况。分离培养PASMCs,在培养液中分别加入1μmol/LET-1和10μg/LPDGF,分别采用半定量RT-PCR、Westernblot和免疫荧光染色的方法研究不同时间点CRP1在PASMCs中的表达。采用Westernblot方法研究SRF-CRP1-GATA6转录蛋白复合体中SRF和GATA6蛋白在不同时间点的表达。结果在大鼠急性肺栓塞后的不同时间点,CRP1的mRNA水平和蛋白水平均逐渐升高,免疫组化研究表明CRP1主要分布在PASMCs,在急性肺栓塞后表达显著升高。PASMCs在ET-1和PDGF的作用下CRP1的表达随着时间的延长均显著升高。Westernblot检测发现PASMCs内SRF和GATA6蛋白在不同时间点的表达均明显升高。结论大鼠急性肺栓塞后肺组织内PASMCs的CRP1表达明显升高。离体实验表明ET-1和PDGF显著促进了PASMCs内CRP1的表达。CRP1可能通过SRF-CRP1-GATA6转录蛋白复合体参与PASMCs特异性基因表达的调控。Objective To study the expression changes of cysteine-rich protein 1 （CRP1） in the lung tissues of acute rat puhnonary embolism （PE） model and the effects of endothelin-1 （ET-1） and platelet-derived growth factor （PDGF） on the expression of CRP1 in pulmonary artery smooth muscle cells （PASMCs）. Methods Acute rat PE model was established. The samples of lung tissues were collected at different time points： 1h, 8h, 24h and 48h alter PE. The normal rats were used as controls. The mRNA level changes of CRP1 were identified by semi-quantitative RT-PCR, and the protein level changes of it were identified by Western blot method. The immunohistochemical method was employed to study the distribution and expression changes of CRP1 in the lung tissues after acute PE. PASMCs were isolated and cultured with 1μmol L ET-1 and 10μg/L PDGF. Semi-quantitative RT-PCR, Western blot and immunofluorescent staining were employed to study the expression changes in PASMCs at different time points. The expression changes of SRF and GATA6 inPASMCs, the components of the transcriptosomeSRF-CRPl- GATA6, were studied by the methods of Western blot. Results At different time points after PE, the mRNA levels and protein levels of CRP1 increased in the lung tissues of acute rat PE models. The immunohistochemical study indicated that CRP1 was mainly expressed in PASMCs of the fumica media of the pulmonary arteries, and the expression of it was up-regulated after acute PE. The expression of CRP1 in PASMCs stimulated by ET-1 or PDGF increased gradually at different time points and so did the expression of SRF and GATA6. Conclusion The expression of CRP1 in PASMCs was up-regulated after acute PE. ET-1 and PDGF could significantly increase the expression of CRP1 in PASMCs in vitro. The protein CRP1 might activate expression of PASMCs-specific gene through the SRF-CRP1-GATA6 transcriptosome.

关 键 词：肺栓塞 肺动脉 平滑肌细胞 富半胱氨酸蛋白1 血小板源性生长因子 内皮素-1 

分 类 号：R543.2[医药卫生—心血管疾病] R563.5[医药卫生—内科学]