骨桥蛋白与整合素相互作用调节血管平滑肌细胞黏附和迁移的分子机制
发布时间：2018-12-09 18:23
【摘要】： 血管平滑肌细胞(vascular smooth muscle cells,VSMCs)的黏附、增殖与迁移是高血压、动脉粥样硬化和血管成形术后再狭窄等血管重塑性疾病的细胞病理学基础。多种生长因子和细胞外基质(extracellular matrix,ECM),如骨桥蛋白(osteopontin,OPN)、纤连蛋白(fibronectin,FN)等均参与VSMC生物学行为的调节。整合素作为ECM蛋白的受体不仅介导细胞与ECM之间的相互作用,而且还通过双向信号转导功能,控制细胞黏附、迁移、分化、增殖和凋亡。因而,研究整合素与其ECM配体相互作用所触发的信号转导途径,以及对细胞生物学行为的影响有助于阐明血管重塑性疾病的分子机制。 为了阐明OPN与整合素相互作用调节VSMC生物学行为的分子机制,本研究构建整合素β3亚单位胞内区肽段真核表达载体(pEGFP-C3-β3CD),并人工合成含有β3胞内区不同保守序列(NXXY)的寡肽(肽-747和肽-759),通过导入VSMC,观察β3亚单位胞内区及其不同区段在OPN诱导VSMC黏附和迁移功能中所起的作用;并观察OPN与整合素相互作用对信号分子Src、黏着斑激酶(focal adhesion kinase,FAK)、整合素耦联激酶(integrin-linked kinase,ILK)的影响,如FAK、ILK的磷酸化水平以及两者相互作用的变化。在此基础上,用Src特异性抑制剂PP2、FAK磷酸化特异性抑制剂黏着斑相关非激酶(focal adhesion related non-kinase,FRNK)、ILK反义RNA或siRNA(ILK siRNAⅠ和Ⅱ),分别阻断Src、FAK磷酸化或ILK表达,进一步探讨三者在OPN诱导VSMC黏附和迁移中所起的作用以及三个信号分子的上、下游关系。 1整合素β3亚单位胞内区在OPN诱导VSMC黏附和迁移中的作用 ECM蛋白作为整合素的配体,通过与细胞膜上的整合素受体结合而将与整合素β亚单位胞内区相互作用的蛋白募集到胞膜内侧,共同形成黏着斑(focal adhesion)复合物,并启动多条信号转导途径,从而导致细胞生物学行为改变。本部分实验观察整合素β3亚单位胞内区及其保守序列
[Abstract]:Adhesion, proliferation and migration of vascular smooth muscle cells (vascular smooth muscle cells,VSMCs) are the cytopathological basis of vascular remodeling diseases such as hypertension, atherosclerosis and restenosis after angioplasty. A variety of growth factors and extracellular matrix (extracellular matrix,ECM), such as osteopontin (osteopontin,OPN) and fibronectin (fibronectin,FN), are involved in the regulation of the biological behavior of VSMC. Integrin, as the receptor of ECM protein, not only mediates the interaction between cells and ECM, but also controls cell adhesion, migration, differentiation, proliferation and apoptosis through two-way signal transduction. Therefore, the study of the signal transduction pathway triggered by the interaction of integrin with its ECM ligands and the effects on cell biological behavior can help to elucidate the molecular mechanism of vascular remodeling diseases. In order to elucidate the molecular mechanism of the interaction between OPN and integrin in regulating the biological behavior of VSMC, an eukaryotic expression vector (pEGFP-C3- 尾 3CD) of integrin 尾 3 subunit was constructed. Oligopeptides (peptide-747 and peptide-759) containing different conserved (NXXY) sequences of 尾 _ 3 intracellular region were synthesized. The role of 尾 _ 3 subunit intracellular region and its different regions in the adhesion and migration of VSMC induced by OPN was observed by introducing VSMC,. The effects of the interaction of OPN and integrin on the signal molecule Src, focal adhesion kinase (focal adhesion kinase,FAK) and integrin coupled kinase (integrin-linked kinase,ILK), such as the phosphorylation level of FAK,ILK and the interaction between them, were observed. On this basis, the phosphorylation or ILK expression of Src,FAK was blocked by (focal adhesion related non-kinase,FRNK), ILK antisense RNA or siRNA (ILK siRNA 鈪，
本文编号：2369833
[Abstract]:Adhesion, proliferation and migration of vascular smooth muscle cells (vascular smooth muscle cells,VSMCs) are the cytopathological basis of vascular remodeling diseases such as hypertension, atherosclerosis and restenosis after angioplasty. A variety of growth factors and extracellular matrix (extracellular matrix,ECM), such as osteopontin (osteopontin,OPN) and fibronectin (fibronectin,FN), are involved in the regulation of the biological behavior of VSMC. Integrin, as the receptor of ECM protein, not only mediates the interaction between cells and ECM, but also controls cell adhesion, migration, differentiation, proliferation and apoptosis through two-way signal transduction. Therefore, the study of the signal transduction pathway triggered by the interaction of integrin with its ECM ligands and the effects on cell biological behavior can help to elucidate the molecular mechanism of vascular remodeling diseases. In order to elucidate the molecular mechanism of the interaction between OPN and integrin in regulating the biological behavior of VSMC, an eukaryotic expression vector (pEGFP-C3- 尾 3CD) of integrin 尾 3 subunit was constructed. Oligopeptides (peptide-747 and peptide-759) containing different conserved (NXXY) sequences of 尾 _ 3 intracellular region were synthesized. The role of 尾 _ 3 subunit intracellular region and its different regions in the adhesion and migration of VSMC induced by OPN was observed by introducing VSMC,. The effects of the interaction of OPN and integrin on the signal molecule Src, focal adhesion kinase (focal adhesion kinase,FAK) and integrin coupled kinase (integrin-linked kinase,ILK), such as the phosphorylation level of FAK,ILK and the interaction between them, were observed. On this basis, the phosphorylation or ILK expression of Src,FAK was blocked by (focal adhesion related non-kinase,FRNK), ILK antisense RNA or siRNA (ILK siRNA 鈪，
本文编号：2369833
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