Velarde V, Ullian M E, Morinelli T A, Mayfield R K, Jaffa A A
Department of Medicine, Medical University of South Carolina and Ralph H. Johnson Department of Veterans Affairs Medical Center, Charleston, South Carolina 29425, USA.
Am J Physiol. 1999 Aug;277(2):C253-61. doi: 10.1152/ajpcell.1999.277.2.C253.
Vascular smooth muscle cell (VSMC) proliferation is a prominent feature of the atherosclerotic process occurring after endothelial injury. A vascular wall kallikrein-kinin system has been described. The contribution of this system to vascular disease is undefined. In the present study we characterized the signal transduction pathway leading to mitogen-activated protein kinase (MAPK) activation in response to bradykinin (BK) in VSMC. Addition of 10(-10)-10(-7) M BK to VSMC resulted in a rapid and concentration-dependent increase in tyrosine phosphorylation of several 144- to 40-kDa proteins. This effect of BK was abolished by the B(2)-kinin receptor antagonist HOE-140, but not by the B(1)-kinin receptor antagonist des-Arg(9)-Leu(8)-BK. Immunoprecipitation with anti-phosphotyrosine antibodies followed by immunoblot revealed that 10(-9) M BK induced tyrosine phosphorylation of focal adhesion kinase (p125(FAK)). BK (10(-8) M) promoted the association of p60(src) with the adapter protein growth factor receptor binding protein-2 and also induced a significant increase in MAPK activity. Pertussis and cholera toxins did not inhibit BK-induced MAPK tyrosine phosphorylation. Protein kinase C downregulation by phorbol 12-myristate 13-acetate and/or inhibitors to protein kinase C, p60(src) kinase, and MAPK kinase inhibited BK-induced MAPK tyrosine phosphorylation. These findings provide evidence that activation of the B(2)-kinin receptor in VSMC leads to generation of multiple second messengers that converge to activate MAPK. The activation of this crucial kinase by BK provides a strong rationale to investigate the mitogenic actions of BK on VSMC proliferation in disease states of vascular injury.
血管平滑肌细胞（VSMC）增殖是内皮损伤后发生的动脉粥样硬化过程的一个显著特征。已有关于血管壁激肽释放酶 - 激肽系统的描述。该系统对血管疾病的作用尚不清楚。在本研究中，我们对血管平滑肌细胞中缓激肽（BK）刺激下导致丝裂原活化蛋白激酶（MAPK）激活的信号转导途径进行了表征。向血管平滑肌细胞中添加10^(-10) - 10^(-7) M的BK会导致几种144至40 kDa蛋白质的酪氨酸磷酸化迅速且呈浓度依赖性增加。BK的这种作用被B(2) - 激肽受体拮抗剂HOE - 140消除，但未被B(1) - 激肽受体拮抗剂去 - Arg(9) - Leu(8) - BK消除。用抗磷酸酪氨酸抗体进行免疫沉淀后再进行免疫印迹显示，10^(-9) M的BK诱导粘着斑激酶（p125(FAK)）的酪氨酸磷酸化。10^(-8) M的BK促进了p60(src)与衔接蛋白生长因子受体结合蛋白 - 2的结合，并且还诱导MAPK活性显著增加。百日咳毒素和霍乱毒素不抑制BK诱导的MAPK酪氨酸磷酸化。佛波醇12 - 肉豆蔻酸酯13 - 乙酸酯下调蛋白激酶C和/或蛋白激酶C、p60(src)激酶及MAPK激酶的抑制剂可抑制BK诱导的MAPK酪氨酸磷酸化。这些发现提供了证据，表明血管平滑肌细胞中B(2) - 激肽受体的激活导致多种第二信使的产生，这些第二信使汇聚以激活MAPK。BK对这种关键激酶的激活为研究BK在血管损伤疾病状态下对血管平滑肌细胞增殖的促有丝分裂作用提供了有力的理论依据。
