Calcium Channels in Neonatal Pulmonary Hypertension

描述（由申请人提供）：本试点项目的目标是确定电压门控的l型Ca2+ (CaL)通道在新生儿肺动脉高压（PH）发病机制中的作用。利用一种完善的新生儿仔猪缺氧诱导PH模型，体内血流动力学显示，暴露于慢性缺氧（CH） 21天的新生儿仔猪静息肺张力增加。在同一只CH仔猪的离体灌注肺中进行的研究表明，硝苯地平敏感的钙通道导致血管张力异常，并且在小肺动脉的贴片夹紧血管平滑肌细胞中观察到钙电流密度升高。随后的研究显示，CH仔猪肺血管中钙通道的成孔阿尔法1c亚基显著上调，这与促进钙通道运输的辅助p2a亚基表达增加相对应。有趣的是，在培养的小肺动脉中，血栓素A2和内皮素-1（两种与新生儿ph有关的血管收缩物质）也诱导了相同模式的CaL通道异常。在21天CH仔猪中，预防性治疗降低血栓素A2的可用性可阻止肺血管中钙通道的上调，并减轻PH的发展。基于这些关键发现，血管收缩物质对钙通道的上调有助于新生儿PH，我们将追求特定的目标，旨在：(a)确定新生仔猪小肺动脉CaL通道的分子组成；(b)确定血栓素A2和内皮素-1上调CH仔猪小肺动脉CaL通道的机制基础；(c)确定血栓素A2和内皮素-1是否在体内上调肺血管中的钙通道，从而促进PH的发展。这些研究将首次提供有关新生儿PH期间促进肺血管中钙通道异常表达的刺激和机制的详细信息，并有助于确定减轻婴幼儿PH的治疗靶点。

DESCRIPTION (provided by applicant): The goal of this pilot project is to define the contribution of voltage-gated, L-type Ca2+ (CaL) channels to the pathogenesis of pulmonary hypertension (PH) in the neonate. Using a well established model of hypoxia-induced PH in neonatal piglets, in-vivo hemodynamics revealed an increased resting pulmonary tone in neonatal piglets exposed to chronic hypoxia (CH) for 21 days. Studies in isolated, perfused lungs from the same CH piglets demonstrated that nifedipine-sensitive CaL, channels contributed to the anomalous vascular tone, and an elevated density of CaL current was observed in patch-clamped vascular smooth muscle cells from small pulmonary arteries. Subsequent studies revealed a striking upregulation of the pore-forming alpha1C subunit of the CaL channel in the pulmonary vasculature of CH piglets, which corresponded to an increased expression of the ancillary p2a subunit that promotes CaL channel trafficking. Interestingly, this same pattern of CaL channel abnormalities was induced in cultured small pulmonary arteries by thromboxane A2 and endothelin-1, two vasoconstrictor substances implicated in neonatal PH. Finally, prophylactic therapy to lower thromboxane A2 availability in 21-day CH piglets prevented the upregulation of CaL channels in the pulmonary vasculature and mitigated the development of PH. Based on these key findings suggesting that the upregulation of CaL channels by vasoconstrictor substances contributes to neonatal PH, we will pursue specific aims designed to: (a) identify the molecular composition of CaL channels in small pulmonary arteries of neonatal piglets, (b) define the mechanistic basis by which thromboxane A2 and endothelin-1 upregulate CaL channels in small pulmonary arteries of CH piglets, and (c) determine if thromboxane A2 and endothelin-1 upregulate CaL channels in the pulmonary vasculature in vivo to contribute to the development of PH. These studies will provide the first detailed information on the stimuli and mechanisms that promote abnormal CaL, channel expression in the pulmonary vasculature during neonatal PH, and will assist in identifying therapeutic targets to mitigate PH in infants and young children.