Liu Xiaoqiu, Thangavel Muthusamy, Sun Shu Qiang, Kaminsky Joseph, Mahautmr Penden, Stitham Jeremiah, Hwa John, Ostrom Rennolds S
Department of Pharmacology, University of Tennessee Health Science Center, 874 Union Ave., Crowe 115, Memphis, TN 38163, USA.
Naunyn Schmiedebergs Arch Pharmacol. 2008 Jun;377(4-6):359-69. doi: 10.1007/s00210-007-0196-0. Epub 2007 Oct 13.
Cardiac fibroblasts produce and degrade extracellular matrix and are critical in regulating cardiac remodeling and hypertrophy. Fibroblasts are activated by factors such as transforming growth factor beta and inhibited by agents that elevate 3',5'-cyclic adenosine monophosphate (cAMP) levels. cAMP signal generation and response is known to be compartmentalized in many cell types in part through the colocalization of receptors and specific adenylyl cyclase isoforms in lipid rafts and caveolae. The present study sought to define the localization of key G protein-coupled receptors with adenylyl cyclase type 6 (AC6) in lipid rafts of rat cardiac fibroblasts and to determine if this colocalization was functionally relevant. We found that cardiac fibroblasts produce cAMP in response to agonists for beta-adrenergic (isoproterenol), prostaglandin EP2 (butaprost), adenosine (adenosine-5'-N-ethylcarboxamide, NECA), and prostacyclin (beraprost) receptors. Overexpression of AC6 increased cAMP production stimulated by isoproterenol and beraprost but not by butaprost or NECA. A key function of fibroblasts is the production of collagen. Isoproterenol- and beraprostmediated inhibition of collagen synthesis was also enhanced by AC6 overexpression, while inhibition by butaprost and NECA were unaltered. Lipid raft fractions from cardiac fibroblasts contain the preponderance of beta-adrenergic receptors and AC6 but exclude EP2 receptors. While we could not determine the localization of native prostacyclin receptors, we were able to determine that epitope-tagged prostanoid IP receptors (IPR) expressed in COS7 cells did localize, in part, in lipid raft fractions. These findings indicate that IP receptors are expressed in lipid rafts and can activate raft-localized AC isoforms. AC6 is completely compartmentized in lipid raft domains where it is activated solely by coresident G protein-coupled receptors to regulate cardiac fibroblast function.
心脏成纤维细胞产生并降解细胞外基质，在调节心脏重塑和肥大过程中起关键作用。成纤维细胞可被诸如转化生长因子β等因子激活，并被能提高3',5'-环磷酸腺苷（cAMP）水平的物质抑制。已知在许多细胞类型中，cAMP信号的产生和反应是分区进行的，部分原因是受体和特定腺苷酸环化酶同工型在脂筏和小窝中共定位。本研究旨在确定大鼠心脏成纤维细胞脂筏中关键G蛋白偶联受体与6型腺苷酸环化酶（AC6）的定位，并确定这种共定位是否具有功能相关性。我们发现，心脏成纤维细胞在受到β-肾上腺素能（异丙肾上腺素）、前列腺素EP2（布他前列素）、腺苷（5'-N-乙基甲酰胺腺苷，NECA）和前列环素（贝拉前列素）受体激动剂刺激时会产生cAMP。AC6的过表达增加了异丙肾上腺素和贝拉前列素刺激产生的cAMP，但未增加布他前列素或NECA刺激产生的cAMP。成纤维细胞的一个关键功能是产生胶原蛋白。AC6的过表达也增强了异丙肾上腺素和贝拉前列素介导的胶原蛋白合成抑制作用，而布他前列素和NECA的抑制作用未改变。心脏成纤维细胞的脂筏组分含有大量β-肾上腺素能受体和AC6，但不含有EP2受体。虽然我们无法确定天然前列环素受体的定位，但我们能够确定在COS7细胞中表达的表位标记的类前列腺素IP受体（IPR）部分定位于脂筏组分中。这些发现表明，IP受体在脂筏中表达，并可激活定位于脂筏的AC同工型。AC6完全定位于脂筏结构域，在那里它仅被共定位的G蛋白偶联受体激活，以调节心脏成纤维细胞的功能。
