Cueille Carine, Birot Olivier, Bigard Xavier, Hagner Stefanie, Garel Jean-Michel
INSERM U-606, Hôpital Lariboisière, 75475 Paris-Cedex 10, France.
Biochem Biophys Res Commun. 2005 Jan 7;326(1):23-9. doi: 10.1016/j.bbrc.2004.10.205.
Adrenomedullin and CGRP are two potent vasodilator peptides, and their receptors are formed by heterodimerization of the CRLR and a RAMP molecule. Hypoxia is associated with many diseases of the cardiovascular system. It was recently shown that the human CRLR gene promoter contains an HIF-1alpha regulatory element, and that CRLR mRNA was increased by hypoxia in human endothelial cells. In the present work, we have assessed the effect of hypoxia on CRLR expression both in vivo and in vitro using two different experimental models. We have also investigated the effect of hypoxia on RAMP expression. (1) We analyzed the effects of a chronic hypobaric hypoxia on rat ventricle expression of RAMPs and CRLR. (2) Acute hypoxia was studied in human vascular smooth cells from coronary artery (CASMC) exposed for 6h to 2% O(2). RT-PCR was used to analyze the mRNA expression, and protein levels were determined by Western blotting. A sharp increase in HIF-1alpha protein levels was induced by hypoxia in CASMC, and 3.5-fold rise of the CRLR protein occurred after 1h of hypoxia in face of unchanged mRNA levels. The CRLR mRNA levels were only elevated later. A clear decrease of the CRLR protein level occurred after 3 and 6h of hypoxia. Thus, acute hypoxia in CASMC induced a rapid change of the CRLR protein amount independently of changes in the CRLR mRNA. This finding suggested a major post-transcriptional effect of hypoxia on CRLR expression in CASMC. RAMP2 and adrenomedullin mRNAs were increased after 4h, but no change was observed for RAMP1. Chronic hypoxia in rats enhanced both mRNA and protein levels of the three RAMPs and CRLR in right and left ventricles. Together, our in vivo and in vitro data suggested that hypoxia up-regulates both adrenomedullin and its receptor (CRLR/RAMP2) to enhance the signaling at the target cell.
肾上腺髓质素和降钙素基因相关肽是两种强效血管舒张肽，它们的受体由降钙素受体样受体（CRLR）和受体活性修饰蛋白（RAMP）分子异源二聚化形成。缺氧与许多心血管系统疾病相关。最近研究表明，人类CRLR基因启动子含有缺氧诱导因子-1α（HIF-1α）调控元件，并且在人内皮细胞中缺氧可使CRLR mRNA增加。在本研究中，我们使用两种不同的实验模型评估了缺氧对体内和体外CRLR表达的影响。我们还研究了缺氧对RAMP表达的影响。（1）我们分析了慢性低压缺氧对大鼠心室中RAMP和CRLR表达的影响。（2）对来自冠状动脉的人血管平滑肌细胞（CASMC）进行急性缺氧研究，将其暴露于2%氧气环境6小时。采用逆转录聚合酶链反应（RT-PCR）分析mRNA表达，通过蛋白质印迹法测定蛋白质水平。缺氧诱导CASMC中HIF-1α蛋白水平急剧增加，缺氧处理1小时后CRLR蛋白水平升高3.5倍，而mRNA水平未改变。CRLR mRNA水平仅在随后升高。缺氧处理3小时和6小时后CRLR蛋白水平明显下降。因此，CASMC中的急性缺氧诱导了CRLR蛋白量的快速变化，而与CRLR mRNA的变化无关。这一发现提示缺氧对CASMC中CRLR表达具有主要的转录后效应。RAMP2和肾上腺髓质素mRNA在4小时后增加，但RAMP1未观察到变化。大鼠慢性缺氧增强了左右心室中三种RAMP和CRLR的mRNA和蛋白水平。总之，我们的体内和体外数据表明，缺氧上调肾上腺髓质素及其受体（CRLR/RAMP2），以增强靶细胞处的信号传导。
