Determining the Physiological Mechanism of CFTR Inhibition by Sphingomyelinase

项目总结 在本申请中提出的研究试图解决抑制囊状细胞的生理基础 纤维跨膜传导调节因子(CFTR)是由鞘磷脂酶(SMase)介导的。CFTR是 在常见遗传病囊性纤维化(CF)中有缺陷的蛋白质。目前的主要死因是 Cf是由频繁的细菌感染和持续的肺部炎症引起的肺功能不全。 有趣的是，感染和炎症都促进了sMase的分泌，我们的实验室已经证明了这一点。 降低CFTRs的功能。为了了解sMase是否在CF疾病严重程度恶化中起作用，我们 将首先利用质谱学和免疫组织化学的方法对CF肺中sMase的数量进行定量。我们 然后将通过电生理学来研究SMASE抑制CFTR的机制 Cftr的行为和消除sMase对cftr活性影响的药理学方法。最后，我们 将确定sMase介导的CFTR抑制对唯一临床批准的 CFTR增效剂药物(VX-770)。根据我们以前的发现，我们假设sMase在 Cf肺通过一种新的机制抑制CFTR，并阻止VX-770挽救CFTR的活性。一起， 这些数据将有助于确定CF患者是否需要联合用药来抵消SMASE的影响 CFTR。

PROJECT SUMMARY The research proposed in this application seeks to address the physiological basis of inhibition of the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) by the enzyme sphingomyelinase (SMase). CFTR is the protein defective in the common genetic disease Cystic Fibrosis (CF). The current major cause of death for CF is pulmonary insufficiency caused by frequent bacterial infections and persistent inflammation of the lungs. Interestingly, both infection and inflammation promote secretion of the enzyme SMase, which our lab has shown to decrease the function of CFTR. To understand if SMase plays a role in worsening disease severity in CF, we will first quantify the amount of SMase in CF lungs by using mass spectrometry and immunohistochemistry. We will then determine the mechanism by which SMase inhibits CFTR by using electrophysiology to study the behavior of CFTR and pharmacological methods to negate the effects of SMase on CFTR activity. Lastly, we will determine the effect SMase-mediated inhibition of CFTR has on the efficacy of the only clinically-approved CFTR potentiator drug (VX-770). Based on our previous findings, we hypothesize that SMase is upregulated in the CF lung, inhibits CFTR via a novel mechanism, and prevents VX-770 from rescuing CFTR activity. Together, these data will help determine if CF patients need co-administration of a drug to nullify the effects of SMase on CFTR.