Pp125FAK is required for stretch dependent morphological response of endothelial cells
Pp125FAK is required for stretch dependent morphological response of endothelial cells
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DOI：
10.1038/sj.onc.1201950
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发表时间：
1998-07-30
期刊：
影响因子：
8
通讯作者：
Sokabe, M
中科院分区：
文献类型：
--
作者：
Naruse, K;Yamada, T;Sokabe, M
本研究探讨了牵张诱导人脐静脉内皮细胞（HUVECs）形态学变化的关键信号通路。在弹性硅膜上培养的细胞经1Hz，20%长度的单轴周期性拉伸后，细胞形态逐渐由多边形变为长轴垂直于拉伸轴的细长纺锤形。无论是用钆（Gd 3+）（拉伸激活通道的有效阻断剂）处理细胞，还是去除细胞外Ca 2+，都阻断了蛋白质的酪氨酸磷酸化，这表明拉伸激活（SA）离子通道调节拉伸特异性酪氨酸磷酸化。主要磷酸化蛋白质的分子量约为120 - 135 kDa和70 kDa，免疫沉淀实验表明，桩蛋白，黏着斑激酶，pp125（FAK））和pp130（CAS）分别出现在70 kDa和120 - 135 kDa的条带中。提示酪氨酸磷酸化是形态学改变所必需的。此外，观察到pp125（FAK）的激酶活化响应于周期性拉伸。此外，反义硫代磷酸寡脱氧核苷酸（S-ODN）抑制pp125（FAK）表达在HUVECs中，导致桩蛋白酪氨酸磷酸化的抑制和牵张依赖的形态学变化。这些结果表明，激活酪氨酸激酶（S）的细胞内Ca 2+和pp125（FAK）的增加发挥了关键作用，在独特的形态学变化，特别是在内皮细胞进行单轴循环拉伸。
In this study, critical signaling pathway required for the stretch induced morphological changes of human umbilical endothelial cells (HUVECs) was investigated. Uniaxial cyclic stretch (1 Hz, 20% in length) of the cells cultured on an elastic silicon membrane induced a gradual morphological change in the cells from a polygonal shape to an elongated spindle-like shape whose long axis was aligned perpendicular to the stretch axis, We found that protein tyrosine phosphorylation of cellular proteins increased and peaked at 20 min in response to cyclic stretch. Either treatment of cells with gadolinium (Gd3+), a potent blocker for stretch-activated channels, or removal of extracellular Ca2+ blocked the tyrosine phosphorylation of the proteins, suggesting that stretch-activated (SA) ion channels regulated stretch specific tyrosine phosphorylation, The major phosphorylated proteins had molecular masses of approximately 120-135 kDa, and 70 kDa, Immunoprecipitation experiments revealed that paxillin, focal adhesion kinase (pp125(FAK)) and pp130(CAS) were included in the 70 kDa and 120-135 kDa bands, respectively, The morphological change was inhibited by herbimycin A and genistein, inhibitors of tyrosine kinases, suggesting that tyrosine phosphorylation was required for the morphological change, In addition, the kinase activation of pp125(FAK) was observed in response to cyclic stretch, Moreover, suppression of pp125(FAK) expression by the antisense phosphorothioate oligodeoxynucleotides (S-ODN) in HUVECs resulted in inhibition of tyrosine phosphorylation of paxillin and the stretch-dependent morphological changes. These results suggest that an activation of tyrosine kinase(s) by an increase in intracellular Ca2+ and pp125(FAK) play a critical role in the unique morphological change specifically observed in endothelial cells subjected to uni-axial cyclic stretch.