Interactions between cyclic AMP and calcium signalling pathways in airway smooth muscle cells.

由于空间组织的信号和它们之间的相互作用广泛存在，钙离子和cAMP被传递到细胞内。例如，在支气管呼吸道，通过激活肌球蛋白轻链激酶，钙离子是收缩的主要控制因素，而引起cAMP形成的受体则引起松弛。这两条通路都是用于治疗哮喘和慢性阻塞性肺疾病(COPD)的药物的靶点，β2受体激动剂和M3受体拮抗剂在临床上得到了广泛的应用。在联合治疗中，这些药物至少是相加的，也许是协同作用的，但相互作用的性质尚不确定。在人支气管气道平滑肌细胞(BASMC)中，cAMP减弱了Ca~(2+)信号和收缩机构对Ca~(2+)的敏感性；而Ca~(2+)则减弱了cAMP信号。这些生理和临床上重要的相互作用背后的机制尚未解决。到目前为止，M3和Beta2受体之间的相互作用依赖于细胞系中的异源表达，但它们需要在保留适当信号蛋白队列的天然人类组织中进行探索。诺华公司正在开发COPD的联合疗法，但目前还没有评估其疗效的人类临床前模型，也没有足够的知识来确定受体下游可能的药物靶点。培养的人支气管气道平滑肌细胞(BASMC)在传代10代后基本保持其天然表型，但需要转基因才能维持正常的M3受体水平。这些单元目前为BASMC中的信号分析提供了最佳模型。我们的目的是确定β2和M3受体激动剂刺激的人BASMC中cAMP和钙离子的相互作用部位。

Ca2+ and cAMP are delivered to cells as spatially organised signals and reciprocal interactions between them are widespread. In bronchial airways, for example, Ca2+ is the primary control of contraction, via activation of myosin light chain kinase, while receptors that evoke cAMP formation cause relaxation. Both pathways are targets of drugs used to treat asthma and chronic obstructive pulmonary disease (COPD), with agonists of beta2-receptors and antagonists of M3 muscarinic receptors in widespread clinical use. In combination therapy, these drugs act at least additively and perhaps synergistically,but the nature of the interactions is undefined. In human bronchial airway smooth muscle cells (BASMC), cAMP attenuates Ca2+ signals and the Ca2+-sensitivity of the contractile apparatus; and Ca2+ attenuates cAMP signalling. The mechanisms underlying these physiologically and clinically important interactions are unresolved. Hitherto interactions between M3 and beta2 receptors have relied on heterologous expression in cell lines, but they need to be explored in native human tissues that retain an appropriate cohort of signalling proteins. Novartis is developing combination therapies for COPD, but there is presently no human pre-clinical model for assessment of their efficacy, and limited knowledge from which to identify possible drug targets downstream of receptors. Human bronchial airway smooth muscle cells (BASMC) in culture largely retain their native phenotype for up to 10 passages, but transfection is required to maintain normal levels of M3 receptors. These cells currently provide the best model for signalling analyses in BASMC. Our aims are to identify sites of interaction between cAMP and Ca2+ in human BASMC stimulated with agonists of beta2 and M3 receptors.