Kelsen S G, Higgins N C, Zhou S, Mardini I A, Benovic J L
Department of Medicine, Temple University School of Medicine, Philadelphia, Pennsylvania, USA.
Am J Respir Crit Care Med. 1995 Dec;152(6 Pt 1):1774-83. doi: 10.1164/ajrccm.152.6.8520736.
Beta-adrenergic agonist-mediated activation of the beta receptor coupled-adenylyl cyclase (beta AR-AC) system expressed by human airway epithelial cells alters airway function. However, little is known about the magnitude of expression, subtype, and function of the beta receptor-adenylyl cyclase (beta AR-AC) system in human airway epithelial cells from healthy, nonsmoking subjects. Therefore, we characterized beta AR number and subtype and the cAMP response to isoproterenol (iso) in acutely dissociated human tracheocytes harvested from 22 healthy, nonsmoking adults during fibroptic bronchoscopy. Moreover, because the regulation of beta AR-AC system function in response to beta-agonists or inflammatory mediators released into the airway in asthma is poorly understood, we examined the cAMP response to iso after 30 min exposure of cells to iso or the protein kinase C activator, sn-1,2-dioctanoyl glycerol (diC8). The beta AR-AC system was highly expressed and functional in human airway epithelial cells. Group mean beta AR density (i.e., Bmax), equilibrium dissociation constant (Kd), and the percentage of beta 2AR subtypes assessed by radioligand binding were approximately 8,900 receptors/cell, 45 pM, and approximately 80%, respectively. Mean maximum cAMP production was approximately 42 pmol/10(5) cells, and the mean EC50 of the response to iso was 131 nM. However, Bmax and cAMP responses to iso varied considerably across subjects. For example, Bmax varied ninefold, and the EC50 of the cAMP response varied 39-fold interindividually. The EC50 was inversely related to beta AR density (r = -0.81, p < 0.05), suggesting that sensitivity of the cAMP response to iso was in part dependent on beta AR density. In all experiments, cAMP responses to iso stimulation were markedly desensitized in dose-dependent fashion by 30 min pretreatment with iso or diC8. For example, pretreatment with iso 10 microM or diC8 100 microM reduced maximum cAMP production to 22 and 63% of control values, respectively. These data indicate that: (1) the beta AR-AC system is highly expressed on acutely dissociated airway epithelial cells from normal adult, but beta AR expression and its functional coupling to adenylyl cyclase vary considerably interindividually; and (2) the beta AR-AC system of normal human airway epithelial cells is rapidly desensitized by exposure to beta-adrenergic agonists or activators of PKC.
β-肾上腺素能激动剂介导的人呼吸道上皮细胞表达的β受体偶联腺苷酸环化酶（βAR-AC）系统的激活会改变气道功能。然而，对于健康、不吸烟受试者的人呼吸道上皮细胞中β受体-腺苷酸环化酶（βAR-AC）系统的表达水平、亚型和功能了解甚少。因此，我们对在纤维支气管镜检查期间从22名健康、不吸烟成年人采集的急性解离的人气管细胞中βAR的数量和亚型以及对异丙肾上腺素（iso）的cAMP反应进行了表征。此外，由于对哮喘中βAR-AC系统功能对释放到气道中的β-激动剂或炎症介质的反应调节了解不足，我们在细胞暴露于iso或蛋白激酶C激活剂sn-1,2-二辛酰甘油（diC8）30分钟后，检测了对iso的cAMP反应。βAR-AC系统在人呼吸道上皮细胞中高度表达且具有功能。通过放射性配体结合评估的组平均βAR密度（即Bmax）、平衡解离常数（Kd）和β2AR亚型的百分比分别约为8900个受体/细胞、45 pM和约80%。平均最大cAMP产生量约为42 pmol/10⁵个细胞，对iso反应的平均EC50为131 nM。然而，Bmax和对iso的cAMP反应在不同受试者之间差异很大。例如，Bmax相差9倍，cAMP反应的EC50个体间相差39倍。EC50与βAR密度呈负相关（r = -0.81，p < 0.05），表明cAMP对iso反应的敏感性部分取决于βAR密度。在所有实验中，用iso或diC8预处理30分钟以剂量依赖性方式使对iso刺激的cAMP反应明显脱敏。例如，用10 μM的iso或100 μM的diC8预处理分别将最大cAMP产生量降低至对照值的22%和63%。这些数据表明：（1）βAR-AC系统在正常成年人急性解离的气道上皮细胞上高度表达，但βAR表达及其与腺苷酸环化酶的功能偶联在个体间差异很大；（2）正常人呼吸道上皮细胞的βAR-AC系统通过暴露于β-肾上腺素能激动剂或PKC激活剂而迅速脱敏。
