Molecular Mechanisms of Airway Smooth Muscle Relaxation

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Although inhaled beta-agonist Is a first line therapy for treatment and prophylaxis of acute bronchospasm that occurs with asthma, its mechanisms of action are poorly understood. In this ROO application we propose of series of hypothesis-driven experiments, as well as the development of new approaches, to clarify mechanisms mediating airway smooth muscle (ASM) relaxation. The proposed studies will utilize both murine and human tissue and cells to explore mechanisms by which two relevant agents, beta-agonist and prostaglandin E2 (PGE2), mediate inhibition of ASM tension generation. Specific Aims include: (Aim 1) Identify the specific targets through which beta-agonist and PGE2 attenuate ASM calcium flux and ainway contraction; and (Aim 2) Identify the PKA substrates in ASM treated with ISO and PGE2, and those genes whose expression is altered by chronic agonist treatment. The proposed studies will provide greater mechanistic insight into the Phase I outcomes by identifying the specific intracellular targets of beta-agonist signaling that antagonize calcium flux and contraction, and how modulation of these targets is influenced with chronic agonist treatment. Additional aims include: (Aim 3) Establish the application of lentiviral infection for heterologous expression in murine and human airways in culture; and (Aim 4) Generate preliminary analyses of genotype effects on primary outcomes. The latter 2 Aims will develop new, powerful tools for enable further mechanistic insight into ASM contractile regulation, and identify genotype effects enabling subsequent studies in Functional Genomics. Upon completion of these Phase II studies, the PI should have both an empirical basis and powerful methodologies to pursue highly relevant research in the field of ASM physiology and biology. LAY SUMMARY: Studies in this proposal will help us understand how a common asthma therapy, beta-agonist, works at a cellular and molecular level thus improving our ability to develop similar, more effective therapies.