Rho kinase is an effector underlying Ca2+-desensitizing hypoxic relaxation in porcine coronary artery.
Rho kinase is an effector underlying Ca2+-desensitizing hypoxic relaxation in porcine coronary artery.
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Rho 激酶是猪冠状动脉 Ca2 脱敏缺氧舒张的潜在效应器。
DOI：
10.1152/ajpheart.01158.2006
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发表时间：
2007
期刊：
影响因子：
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通讯作者：
Paul,RichardJ
中科院分区：
文献类型：
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作者：
Wardle,RobertL;Gu,Min;Ishida,Yukisato;Paul,RichardJ
急性缺氧使大多数全身动脉扩张，导致组织灌注增加。我们之前已经证明，在高刺激条件下，猪冠状动脉被缺氧放松，而细胞内[Ca2+]没有变化。这种Ca2+脱敏性缺氧松弛（CDHR）在通透性猪冠状动脉平滑肌（PCASM）中得到了验证，尽管固定[Ca2+]，但缺氧降低了力和肌球蛋白调节轻链磷酸化（p-MRLC）。肌球蛋白磷酸酶靶向亚基1 （p-MYPT1）的Rho激酶依赖性磷酸化与MRLC磷酸酶活性降低和p-MRLC和力的Ca2+敏感性增加有关。我们最近报道了p-MYPT1去磷酸化是CDHR的关键效应因子。在目前的研究中，我们验证了Rho激酶而不是p-MYPT1磷酸酶是参与CDHR的调节酶的假设。我们用α-毒素使去内皮化的PCASM通透化。在Rho激酶抑制剂Y-27632存在的情况下，CDHR在肌球蛋白轻链激酶（MLCK）引起的收缩中减弱。相比之下，缺氧时，由于Rho激酶磷酸化MYPT1和MRLC或MRLC单独（在MLCK抑制剂ML7存在的情况下）而使收缩放松。通过原位分析，我们发现Rho激酶活性（通过MYPT1和MRLC的硫代磷酸化来测量）几乎被缺氧所消除。Rho激酶催化活性片段的体外活性不受缺氧的影响。我们的证据强烈暗示缺氧直接抑制Rho激酶依赖性的MYPT1磷酸化。这是p-MYPT1和p-MRLC减少的基础，从而导致Ca2+脱敏性缺氧松弛。
Acute hypoxia dilates most systemic arteries leading to increased tissue perfusion. We have previously shown that at high-stimulus conditions, porcine coronary artery was relaxed by hypoxia without a change in intracellular [Ca2+] . This Ca2+-desensitizing hypoxic relaxation (CDHR) was validated in permeabilized porcine coronary artery smooth muscle (PCASM) in which hypoxia decreased force and myosin regulatory light chain phosphorylation (p-MRLC) despite fixed [Ca2+] . Rho kinase-dependent phosphorylation of myosin phosphatase-targeting subunit 1 (p-MYPT1) is associated with decreased MRLC phosphatase activity and increased Ca2+sensitivity of both p-MRLC and force. We recently reported that p-MYPT1 dephosphorylation was a key effector in CDHR . In the current study, we tested the hypothesis that Rho kinase and not p-MYPT1 phosphatase is the regulated enzyme involved in CDHR. We used α-toxin to permeabilize deendothelialized PCASM. CDHR was attenuated in contractions attributable to myosin light chain kinase (MLCK, in the presence of the Rho kinase inhibitor Y-27632). In contrast, hypoxia relaxed contractions attributable to Rho kinase phosphorylation of MYPT1 and MRLC or MRLC alone (in the presence of the MLCK inhibitor ML7). Using an in situ assay, we showed that Rho kinase activity, measured as thiophosphorylation of MYPT1 and MRLC, was nearly abolished by hypoxia. The in vitro activity of the catalytically active fragment of Rho kinase was not affected by hypoxia. Our evidence strongly implicates that hypoxia directly inhibits Rho kinase-dependent phosphorylation of MYPT1. This underlies the decreases in both p-MYPT1 and p-MRLC and thereby leads to the Ca2+-desensitizing hypoxic relaxation.