El-Haroun H, Clarke D L, Deacon K, Bradbury D, Clayton A, Sutcliffe A, Knox Alan J
Division of Respiratory Medicine, University of Nottingham, Clinical Science Bldg., City Hospital, Nottingham, NG5 1PB, United Kingdom.
Am J Physiol Lung Cell Mol Physiol. 2008 Mar;294(3):L553-62. doi: 10.1152/ajplung.00044.2006. Epub 2007 Dec 21.
We have previously shown that interleukin (IL)-1beta, transforming growth factor (TGF)-beta1, or bradykinin (BK) impair cAMP generation in response to prostacyclin analogs in human pulmonary artery smooth muscle (PASM), suggesting that inflammation can impair the effects of prostacyclin analogs on PASM in pulmonary hypertension. Here we explored the biochemical mechanisms involved. We found that IL-1beta, BK, and TGF-beta1 reduced adenylyl cyclase isoform 1, 2, and 4 mRNA, increased Galphai protein levels, and reduced prostacyclin receptor (IP receptor) mRNA expression. In contrast, Galphas protein levels were unchanged. Protein kinase A (PKA) (H-89, KT-2750, PKIm) and p38 mitogen-activated protein (MAP) kinase (SB-202190) inhibitors attenuated these effects, but protein kinase C (bisindolylmaleide) or phosphoinositol 3-kinase (LY-294002) inhibitors did not. Fluorescent kemptide assay and Western blotting confirmed that PKA and p38 MAP kinase were activated by IL-1beta, BK, and TGF-beta1. These studies suggest that IL-1beta, BK, and TGF-beta1 impair IP receptor-mediated cAMP accumulation by multiple effects on different components of the signaling pathway and that these effects are PKA and p38 MAP kinase dependent.
我们之前已经表明，白细胞介素（IL）-1β、转化生长因子（TGF）-β1或缓激肽（BK）会削弱人肺动脉平滑肌（PASM）中前列环素类似物诱导的环磷酸腺苷（cAMP）生成，这表明炎症会损害前列环素类似物对肺动脉高压中PASM的作用。在此，我们探究了其中涉及的生化机制。我们发现，IL-1β、BK和TGF-β1会降低腺苷酸环化酶同工型1、2和4的mRNA水平，增加Gαi蛋白水平，并降低前列环素受体（IP受体）的mRNA表达。相比之下，Gαs蛋白水平未发生变化。蛋白激酶A（PKA）（H-89、KT-2750、PKIm）和p38丝裂原活化蛋白（MAP）激酶（SB-202190）抑制剂可减弱这些作用，但蛋白激酶C（双吲哚马来酰胺）或磷酸肌醇3激酶（LY-294002）抑制剂则无此作用。荧光kemptide检测和蛋白质印迹法证实，PKA和p38 MAP激酶被IL-1β、BK和TGF-β1激活。这些研究表明，IL-1β、BK和TGF-β1通过对信号通路不同组分的多种作用，损害IP受体介导的cAMP积累，且这些作用依赖于PKA和p38 MAP激酶。
