Asthma is a chronic respiratory disease characterized by reversible airway obstruction and hyper- responsiveness of the airways to bronchoconstrictor stimuli. Veterans represent a large cohort of asthma sufferers, many of whom do not achieve relief of symptoms with available therapies. Reversible airway obstruction in asthma results primarily from hypercontraction of airway smooth muscle cells (ASMCs) in the bronchioles of the lung. The mechanisms involved in ASMC hypercontraction are poorly characterized but likely involve exaggerated bronchoconstrictor responses to Gq/11-coupled receptor agonists, which include acetylcholine, histamine, endothelin-1, and leukotriene D4. Increased Ca2+ influx via voltage-sensitive Ca2+ channels (VSCCs) contributes to the exaggerated bronchoconstrictor responses. The voltage change required to activate VSCCs can be provided by suppressing potassium (K+) channel activity. We recently discovered that KCNQ (Kv7 family) K+ channels are expressed and active in freshly isolated bronchiolar myocytes and that their activity is suppressed by multiple bronchoconstrictor agonists. We further demonstrated that Kv7 current suppression is sufficient to induce airway constriction and that pharmacological Kv7 channel activators significantly attenuate airway constriction in precision-cut human lung slices. Based on these discoveries, we propose to examine the hypothesis that bronchoconstrictor agonists suppress the activity of Kv7 family K+ channels in ASMCs to induce membrane depolarization and ASMC hypercontraction. Furthermore, we will explore two additional hypotheses that have important clinical/translational relevance for our understanding and treatment of asthma. We hypothesize that inflammatory mediators alter Kv7 channel expression, which plays a role in the etiology of airway hyperresponsiveness in asthma. Furthermore, because Kv7 channels are established drug targets, we hypothesize that clinically used Kv7 channel activators can be used as a monotherapy or they may be combined with other therapies to prevent or reduce excessive airway constriction in asthma. To address our hypotheses, three Specific Aims are proposed.
Aim 1) Elucidate the mechanisms by which Gq/11-coupled receptor agonists regulate Kv7 currents in ASMCs. The role of protein kinase C will be determined by using a variety of biochemical, pharmacological, and molecular biological approaches in combination with patch clamp electrophysiology and measurements of airway smooth muscle function in human and guinea pig ASMCs and lung tissues.
Aim 2) Test two novel hypotheses linking airway inflammation to suppression of Kv7 channel expression in ASMCs and increased sensitivity to bronchoconstrictor stimuli. Hypothesis 1: inflammatory mediators, such as IL-13 and IL-1?, upregulate repressor element 1 silencing transcription factor (REST), which then inhibits transcription of KCNQ genes in ASMCs. Hypothesis 2: micro RNA 146a (miR146a), which is also increased in response to inflammatory mediators, induces post- transcriptional repression of Kv7.5 expression in ASMCs. REST and miR146a expression will be measured/altered in ASMCs from normal and asthmatic human and guinea pig lungs to determine their relationship to altered expression/function of Kv7 channels and the link to airway hyperresponsiveness.
Aim 3) Evaluate the therapeutic benefits of Kv7 channel activators, alone or combined with ?2-adrenergic receptor agonists, on airway function in vivo in normal and antigen-sensitized guinea pigs. Our proposed studies are significant because they will elucidate previously unknown mechanisms for ASMC hypercontraction and identify a new therapeutic strategy to provide better relief of airway hyperresponsiveness in asthma patients. This outcome would directly benefit thousands of veterans and civilians who suffer from asthma or other airway diseases.
Asthma is a common disabling condition among veterans. Our proposed studies address both the causes of asthma (a novel central role for Kv7 channels in ASMC hypercontraction) and its treatment (identification of improved therapeutic strategies based on drugs already in clinical use that can be applied in novel ways to relieve asthma symptoms). In summary, the proposed research will provide important benefits that directly serve the stated mission of the VA Office of Research and Development to promote 'fundamental research on disorders of significance to the veteran population' and support research on the causes, progression, diagnosis, and treatment of diseases and disorders affecting veterans.