McLees A, Graham A, Malarkey K, Gould G W, Plevin R
Department of Physiology and Pharmacology, University of Strathclyde, Glasgow, U.K.
Biochem J. 1995 May 1;307 ( Pt 3)(Pt 3):743-8. doi: 10.1042/bj3070743.
In the endothelial cell line EAhy 926, 1-oleoyl-lysophosphatidic acid (LPA) stimulated the tyrosine phosphorylation of the pp42 isoform of mitogen-activated protein (MAP) kinase. Maximum phosphorylation was observed within 5 min of LPA addition, but the response was sustained for up to 120 min. Re-addition of LPA after 60 min stimulated a further sustained increase in the tyrosine phosphorylation of MAP kinase. In cells pretreated with phorbol 12-myristate 13-acetate (PMA; 24 h) or preincubated with the protein kinase C inhibitor Ro-318220, LPA-induced tyrosine phosphorylation of pp42 MAP kinase was substantially reduced at 2 min but potentiated at 60 min. Ro-318220 in combination with either PMA or pertussis toxin pretreatment abolished the LPA response at all time points, suggesting an involvement of protein kinase C in the pertussis toxin-sensitive part of the pathway. Agents which raised intracellular cyclic AMP levels did not affect the initial phase of LPA-stimulated MAP kinase activation, but abolished the late phase. However, this effect was prevented by Ro-318220, implicating a greater role for protein kinase C than protein kinase A in the regulation of sustained MAP kinase responses. LPA stimulated an increase in the tyrosine phosphorylation of focal adhesion kinase pp125 (pp125FAK) in EAhy 926 cells which was both protein kinase C- and pertussis toxin-independent. These results are discussed in terms of the pathways regulating both MAP kinase and pp125FAK in response to LPA in the EAhy 926 endothelial cells line.
在内皮细胞系EAhy 926中，1-油酰基-溶血磷脂酸（LPA）刺激了丝裂原活化蛋白（MAP）激酶pp42亚型的酪氨酸磷酸化。在添加LPA后5分钟内观察到最大磷酸化，但该反应持续长达120分钟。60分钟后重新添加LPA刺激了MAP激酶酪氨酸磷酸化的进一步持续增加。在用佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA；24小时）预处理的细胞中或用蛋白激酶C抑制剂Ro-318220预孵育的细胞中，LPA诱导的pp42 MAP激酶酪氨酸磷酸化在2分钟时显著降低，但在60分钟时增强。Ro-318220与PMA或百日咳毒素预处理联合使用在所有时间点均消除了LPA反应，表明蛋白激酶C参与了该途径中对百日咳毒素敏感的部分。提高细胞内环磷酸腺苷水平的试剂不影响LPA刺激的MAP激酶活化的初始阶段，但消除了后期阶段。然而，Ro-318220可阻止这种作用，这表明在持续的MAP激酶反应调节中，蛋白激酶C比蛋白激酶A发挥更大的作用。LPA刺激了EAhy 926细胞中粘着斑激酶pp125（pp125FAK）的酪氨酸磷酸化增加，这与蛋白激酶C和百日咳毒素无关。根据EAhy 926内皮细胞系中响应LPA调节MAP激酶和pp125FAK的途径对这些结果进行了讨论。
