Murasawa S, Mori Y, Nozawa Y, Gotoh N, Shibuya M, Masaki H, Maruyama K, Tsutsumi Y, Moriguchi Y, Shibazaki Y, Tanaka Y, Iwasaka T, Inada M, Matsubara H
Department of Medicine II, Kansai Medical University, Moriguchi, Osaka, Japan.
Circ Res. 1998 Jun 29;82(12):1338-48. doi: 10.1161/01.res.82.12.1338.
The signaling cascade elicited by angiotensin II (Ang II) resembles that characteristic of growth factor stimulation, and recent evidence suggests that G protein-coupled receptors transactivate growth factor receptors to transmit mitogenic effects. In the present study, we report the involvement of epidermal growth factor receptor (EGF-R) in Ang II-induced extracellular signal-regulated kinase (ERK) activation, c-fos gene expression, and DNA synthesis in cardiac fibroblasts. Ang II induced a rapid tyrosine phosphorylation of EGF-R in association with phosphorylation of Shc protein and ERK activation. Specific inhibition of EGF-R function by either a dominant-negative EGF-R mutant or selective tyrphostin AG1478 completely abolished Ang II-induced ERK activation. Induction of c-fos gene expression and DNA synthesis were also abolished by the inhibition of EGF-R function. Calmodulin or tyrosine kinase inhibitors, but not protein kinase C (PKC) inhibitors or downregulation of PKC, completely abolished transactivation of EGF-R by Ang II or the Ca2+ ionophore A23187. Epidermal growth factor (EGF) activity in concentrated supernatant from Ang II-treated cells was not detected, and saturation of culture media with anti-EGF antibody did not affect the Ang II-induced transactivation of EGF-R. Conditioned media in which cells were incubated with Ang II could not induce phosphorylation of EGF-R on recipient cells. Platelet-derived growth factor-beta receptor was not phosphorylated on Ang II stimulation, and Ang II-induced c-jun gene expression was not affected by tyrphostin AG1478. Our results demonstrated that in cardiac fibroblasts Ang II-induced ERK activation and its mitogenic signals are dominantly mediated by EGF-R transactivated in a Ca2+/calmodulin-dependent manner and suggested that the effects of Ang II on cardiac fibroblasts should be interpreted in association with the signaling pathways regulating cellular proliferation and/or differentiation by growth factors.
血管紧张素II（Ang II）引发的信号级联反应类似于生长因子刺激的特征，最近的证据表明，G蛋白偶联受体可使生长因子受体反式激活，从而传递促有丝分裂效应。在本研究中，我们报告了表皮生长因子受体（EGF-R）参与Ang II诱导的心脏成纤维细胞细胞外信号调节激酶（ERK）激活、c-fos基因表达和DNA合成。Ang II诱导EGF-R快速酪氨酸磷酸化，同时伴有Shc蛋白磷酸化和ERK激活。通过显性负性EGF-R突变体或选择性酪氨酸激酶抑制剂AG1478特异性抑制EGF-R功能，可完全消除Ang II诱导的ERK激活。抑制EGF-R功能也可消除c-fos基因表达和DNA合成的诱导。钙调蛋白或酪氨酸激酶抑制剂可完全消除Ang II或Ca2+离子载体A23187对EGF-R的反式激活，而蛋白激酶C（PKC）抑制剂或PKC下调则不能。未检测到Ang II处理细胞浓缩上清液中的表皮生长因子（EGF）活性，用抗EGF抗体饱和培养基不影响Ang II诱导的EGF-R反式激活。用Ang II孵育细胞的条件培养基不能诱导受体细胞上EGF-R的磷酸化。血小板衍生生长因子-β受体在Ang II刺激下未发生磷酸化，Ang II诱导的c-jun基因表达不受酪氨酸激酶抑制剂AG1478的影响。我们的结果表明，在心脏成纤维细胞中，Ang II诱导的ERK激活及其促有丝分裂信号主要由以Ca2+/钙调蛋白依赖性方式反式激活的EGF-R介导，提示Ang II对心脏成纤维细胞的作用应与生长因子调节细胞增殖和/或分化的信号通路相关联来解释。
