Kinase-dependent activation of voltage-gated Ca2+ channels by ET-1 in pulmonary arterial myocytes during chronic hypoxia.
Kinase-dependent activation of voltage-gated Ca2+ channels by ET-1 in pulmonary arterial myocytes during chronic hypoxia.
复制标题
DOI：
10.1152/ajplung.00396.2011
复制
发表时间：
2012-05
期刊：
影响因子：
--
通讯作者：
Trevor Luke;J. Maylor;Clark Undem;J. Sylvester;L. Shimoda
中科院分区：
文献类型：
--
作者：
Trevor Luke;J. Maylor;Clark Undem;J. Sylvester;L. Shimoda
暴露于慢性缺氧（CH）会导致肺动脉高压。血管收缩剂内皮素-1（ET-1）被认为在缺氧性肺动脉高压的发展中起作用。在慢性缺氧大鼠肺动脉平滑肌细胞（PASMCs）中，ET-1的信号转导被改变，即使ET-1通过抑制K（+）通道引起的去极化作用丧失，ET-1诱导的细胞内钙离子浓度（Δ[Ca（2+）]（i））的变化也通过激活电压依赖性Ca（2+）通道（VDCC）而发生。这种反应背后的机制尚不清楚。我们推测CH后ET-1对VDCCs的激活可能是由蛋白激酶C（PKC）和/或Rho激酶介导的，这两种激酶都能磷酸化并激活VDCCs。为了验证这一假设，我们用钙敏感染料fura 2-AM和荧光显微镜技术检测了PKC和Rho激酶抑制剂对CH（10%O（2），3 wk）暴露大鼠PASMC中ET-1诱导的Δ[Ca（2+）]（i）的影响。我们发现PKC抑制剂staurosporine和GF 109203 X以及Rho激酶抑制剂Y-27632和HA 1077使ET-1诱导的Δ[Ca（2+）]（i）降低> 70%。用染料木黄酮或酪氨酸磷酸化抑制剂A23抑制酪氨酸激酶（TK），或联合抑制PKC、TK和Rho激酶，降低Δ[Ca（2+）]（i）的程度与单独抑制PKC或Rho激酶相似。用佛波醇12-肉豆蔻酸酯13-乙酸酯和GTP-γ-S验证了PKC或Rho激酶激活我们细胞中VDCCs的能力。这些结果表明，CH后，ET-1诱导的PASMCs的Δ[Ca（2+）]（i）通过主要由PKC、TK和Rho激酶介导的VDCCs的Ca（2+）内流发生。
Exposure to chronic hypoxia (CH) causes pulmonary hypertension. The vasoconstrictor endothelin-1 (ET-1) is thought to play a role in the development of hypoxic pulmonary hypertension. In pulmonary arterial smooth muscle cells (PASMCs) from chronically hypoxic rats, ET-1 signaling is altered, with the ET-1-induced change in intracellular calcium concentration (Δ[Ca(2+)](i)) occurring through activation of voltage-dependent Ca(2+) channels (VDCC) even though ET-1-induced depolarization via inhibition of K(+) channels is lost. The mechanism underlying this response is unclear. We hypothesized that activation of VDCCs by ET-1 following CH might be mediated by protein kinase C (PKC) and/or Rho kinase, both of which have been shown to phosphorylate and activate VDCCs. To test this hypothesis, we examined the effects of PKC and Rho kinase inhibitors on the ET-1-induced Δ[Ca(2+)](i) in PASMCs from rats exposed to CH (10% O(2), 3 wk) using the Ca(2+)-sensitive dye fura 2-AM and fluorescent microscopy techniques. We found that staurosporine and GF109203X, inhibitors of PKC, and Y-27632 and HA 1077, Rho kinase inhibitors, reduced the ET-1-induced Δ[Ca(2+)](i) by >70%. Inhibition of tyrosine kinases (TKs) with genistein or tyrphostin A23, or combined inhibition of PKC, TKs, and Rho kinase, reduced the Δ[Ca(2+)](i) to a similar extent as inhibition of either PKC or Rho kinase alone. The ability of PKC or Rho kinase to activate VDCCs in our cells was verified using phorbol 12-myristate 13-acetate and GTP-γ-S. These results suggest that following CH, the ET-1-induced Δ[Ca(2+)](i) in PASMCs occurs via Ca(2+) influx through VDCCs mediated primarily by PKC, TKs, and Rho kinase.