Endothelin-mediated vascular growth requires p42/p44 mitogen-activated protein kinase and p70 S6 kinase cascades via transactivation of epidermal growth factor receptor
Endothelin-mediated vascular growth requires p42/p44 mitogen-activated protein kinase and p70 S6 kinase cascades via transactivation of epidermal growth factor receptor
复制标题
DOI：
10.1210/en.140.10.4659
复制
发表时间：
1999-10-01
期刊：
影响因子：
4.8
通讯作者：
Hirata, Y
中科院分区：
文献类型：
--
作者：
Iwasaki, H;Eguchi, S;Hirata, Y
内皮素-1（ET-1）是一种有效的内皮源性缩血管肽，对血管平滑肌细胞具有促生长作用，在血管重构中起致病作用。为了深入了解ET-1诱导血管生长的细胞和分子机制，我们研究了受体酪氨酸激酶如表皮生长因子受体（EGFR）和血小板衍生生长因子受体的反式激活是否是激活p42/p44促分裂原活化蛋白（MAP）激酶和p70 S6激酶（p70（S6 K））所必需的，ET-1对培养的大鼠血管平滑肌细胞的促生长作用。抗酪氨酸磷酸化抗体免疫印迹显示ET-1可迅速（2 min内）瞬时诱导多种蛋白的酪氨酸磷酸化，其中180-kDa蛋白为EGFR。ET-1迅速增加EGFR和Shc与谷胱甘肽-S-转移酶-Grb 2融合蛋白的结合。ET-1诱导的MAP激酶激活被EGFR激酶抑制剂（AG 1478）降低，但血小板衍生生长因子受体激酶抑制剂（AG 1296）不降低。AG 1478剂量依赖性地降低ET-1刺激的MAP激酶活性以及[H-3]亮氨酸和[H-3]胸苷摄取。ET-1诱导的EGFR酪氨酸磷酸化，以及MAP激酶激活，抑制ETA受体拮抗剂和细胞内Ca 2+拮抗剂，但不抑制ETB受体拮抗剂，百日咳毒素，或蛋白激酶C抑制剂。此外，H-Ras的显性失活突变体和MAP激酶激酶（MEK-1）抑制剂（PD 98059）完全阻断ET-1诱导的MAP激酶激活以及[H-3]亮氨酸和[H-3]胸苷摄取。AG 1478和PD 98059均抑制ET-1诱导的p70（S6 K）磷酸化和活化。此外，雷帕霉素，哺乳动物雷帕霉素靶的选择性抑制剂，完全阻断ET-1刺激的[H-3]亮氨酸和[H-3]胸苷摄取。这些结果表明，ETA受体介导的血管生长的ET-1需要MAP激酶和p70（S6 K）级联介导的部分通过Ca 2+依赖的EGFR反式激活。
Endothelin-1 (ET-1), a potent endothelium-derived vasoconstrictor peptide, exerts a growth-promoting effect on vascular smooth muscle cells, implicating its pathogenic role in vascular remodeling. To gain insight into the cellular and molecular mechanism whereby ET-1 induces vascular growth, we studied whether transactivation of receptor tyrosine kinases, such as epidermal growth factor receptor (EGFR) and platelet-derived growth factor receptor, are required for activation of p42/p44 mitogen-activated protein (MAP) kinase and p70 S6 kinase (p70(S6K)), and subsequent growth-promotion by ET-1 in cultured rat vascular smooth muscle cells. Immunoblotting with antiphosphotyrosine antibody revealed that ET-1 rapidly (within 2 min) and transiently induced tyrosine phosphorylation of several proteins, among which 180-kDa protein was shown to be EGFR. ET-1 rapidly increased association of EGFR and Shc with glutathione-S-trans ferase-Grb2 fusion protein. The ET-1-induced activation of MAP kinase was reduced by an EGFR kinase inhibitor (AG1478) but not by a platelet-derived growth factor receptor kinase inhibitor (AG1296). AG1478 dose-dependently decreased ET-1-stimulated MAP kinase activity as well as [H-3]leucine and [H-3]thymidine uptake. The ET-1-induced tyrosine phosphorylation of EGFR, as well as MAP kinase activation, was inhibited by an ETA receptor antagonist and intracellular Ca2+ antagonists but not by an ETB receptor antagonist, pertussis toxin, or protein kinase C inhibitors. In addition, dominant negative mutant of H-Ras and a MAP kinase kinase (MEK-1) inhibitor (PD98059) completely blocked ET-1-induced MAP kinase activation as well as [H-3]leucine and [H-3]thymidine uptake. Both AG1478 and PD98059 inhibited ET-1-induced phosphorylation and activation of p70(S6K). Furthermore, rapamycin, a selective inhibitor of mammalian target of rapamycin, completely blocked ET-1-stimulated [H-3]leucine and [H-3]thymidine uptake. These results suggest that ETA receptor-mediated vascular growth by ET-1 requires both MAP kinase and p70(S6K) cascades mediated partly via Ca2+-dependent EGFR transactivation.