巨噬细胞移动抑制因子靶向FAK信号通路调控气道平滑肌细胞粘附、迁移和增殖的实验研究

It is well known that the adhesion,migration and proliferation of Airway smooth muscle cells (ASMCs) play the pivotal role of in airway remodeling of asthma.. Macrophage migration inhibitory fact(MIF) is a kind of cytokines with enzyme activity. It has been confirmed that MIF can regulate adhesion,migration and proliferation of the vascular smooth muscle and tumor cells through MAPK, Akt and FAK pathways.Several studies have shown that the expression of MIF in peripheral blood and lung tissue are higher in asthmatic patients. Our initial study found that rMIF can Significantly activated the phosphorylation of FAK/Akt in ASMC, small molecule inhibitors of MIF can reduce the airway wall thickening of asthmatic mice.Accordingly, we proposed the hypothesis: MIF can target FAK signal ,regulate the Akt,MAPK and Rho pathways, influence the adhesion,migration and proliferation of ASMC, participate the occurrence, development in airway remodeling of asthma.In order to confirm the hypothesis,we establish the models of asthma mice and human ASMC in vitro,Then using recombinant adenovirus containing wild-type MIF cDNA (Ad-MIF), Ad shRNA-MIF and external MIF factor intervene in the models, clarify the molecular mechanism how the MIF regulate the adhesion, migration and proliferation of ASMC through receptors, intracellular signal molecules and effects.We expect to further explore the mechanism of airway remodeling in asthma, hope to open a new way to control the asthma.

气道平滑肌细胞(ASMC)的粘附、迁移和增生在哮喘气道重塑过程中起重要作用。巨噬细胞移动抑制因子(MIF)是一种有酶活性的细胞因子，可通过 MAPK、Akt、FAK等途径调控血管平滑肌细胞、肿瘤细胞等的粘附、迁移和增殖。既往研究显示哮喘患者外周血和肺组织中MIF增高；我们前期研究发现，重组MIF能显著激活ASMC内FAK/Akt磷酸化，MIF小分子抑制剂阻断MIF效应能减轻哮喘小鼠气道壁增厚。据此,我们提出:MIF 可通过靶向FAK分子，调控Akt、MAPK及Rho等信号途径, 影响ASMC的粘附、迁移和增殖,参与哮喘气道重塑的发生、发展。为证实上述假说，本项目拟建立哮喘小鼠及体外人ASMC模型，通过 MIF 基因过表达及干扰、外源性MIF因子干预，从受体、细胞内信号分子及效应三个层面阐明MIF调控ASMC粘附、迁移和增殖的分子机制，为防治哮喘气道重塑开辟提供新的理论基础和研究方向。

气道平滑肌细胞(ASMC)的粘附、迁移和增生在哮喘气道重塑过程中起重要作用。巨噬细胞移动抑制因子(MIF)是一种有酶活性的细胞因子，可通过 MAPK、Akt、FAK 等途径调控血管平滑肌细胞、肿瘤细胞等的粘附、迁移和增殖。本项目通过建立体内外ASMC 模型，通过 MIF 基因过表达及干扰、外源性 MIF 因子干预，从受体、细胞内信号分子及效应三个层面阐明 MIF调控 ASMC 粘附、迁移和增殖的分子机制，期望为防治哮喘气道重塑开辟提供新的理论基础和研究方向。. 本研究采用分子生物学技术首先构建人巨噬细胞移动抑制因子基因腺病毒表达载体，检测 AD-MIF 腺病毒转染人气道平滑肌细胞后蛋白表达，成功构建 MIF腺病毒，病毒浓度达 2E+9PFU/ml。. 在我们的研究中，我们发现MIF过表达，能促进ASMC增殖，增加G2期的ASMC细胞，下调P21，上调cyclin D1、cyclin D3、 Cdk6的表达，且能使ASMC的凋亡明显减少，且过表达MIF能通过上调基质金属蛋白酶2（MMP)-2促进细胞迁移。最后，我们的研究还证实，过表达MIF能通过增加磷酸化的ERK和FAK水平促进细胞增殖和迁移。总之，我们的研究显示，过表达MIF能通过上调ERK和FAK活性促进ASMC的的增殖和迁移，从而提示我们抑制MIF水平可能为哮喘病人的气道重塑治疗提供有效的策略。