骨桥蛋白趋化和趋触作用对FAK和ILK磷酸化修饰的影响

The Roles of Osteopontin in Chemotaxis and Haptotaxis Involve the Focal Adhesion Kinase and Integrin-linked Kinase

为探讨可溶性(趋化作用)或锚定(趋触作用)形式的骨桥蛋白(OPN)与整合素相互作用对下游信号分子黏着斑激酶(FAK)和整合素偶联激酶(ILK)磷酸化修饰的影响,分别用包被于培养瓶上锚定型或加在培养液中的可溶性OPN刺激血管平滑肌细胞(VSMC)后,观察FAK和ILK的磷酸化及FAK与ILK相互作用的变化。结果显示,包被于培养瓶上的OPN通过趋触作用促进VSMC黏附和伸展,接种45min时,黏附细胞数达对照组的2.4倍(P<0.05);OPN的趋触及趋化作用均可诱导FAK磷酸化、ILK去磷酸化并抑制FAK与ILK结合;转染可表达整合素β3亚单位胞内区的表达质粒pEGFP-C3-β3CD能阻断OPN与整合素相互作用所引发的FAK磷酸化及ILK去磷酸化。研究结果表明,OPN的趋触和趋化作用对整合素下游信号分子FAK和ILK的影响是一致的,且这些作用是由整合素β3亚单位胞内区所介导的。

To study the effect of the soluble or anchoring osteopontin （OPN） on focal adhesion kinase （FAK） and integrin-linked kinase （ILK） in vascular smooth muscle cells （VSMCs）, VSMCs were treated with OPN coated on culture flasks or soluble OPN, then the level of FAK and ILK phosphorylation, as well as their interactions were detected. The results showed that OPN coated on culture flasks promoted VSMC adhesion and spreading through haptotaxis. After seeded for 45 min, the number of attached cells on OPN-coated flasks was increased by 1.4 folds compared with control, OPN added in culture media or coated on the culture flasks induced FAK phosphorylation and ILK dephosphorylation, and inhibited FAK interaction with ILK through chemotaxis or haptotaxis. Transfection of the eukaryotic expression vector, pEGFP-C3-β3CD, harboring the cytoplasmic domain of human integrin β3 subunit cDNA inhibited these effects. These results suggest that both soluble and anchoring OPN regulate similarly the integrin downstream molecules FAK and ILK through the cytoplasmic domain of integrin β3 subunit.