KATP channels and lymphatic dysfunction in Cantu Syndrome

坎图综合征中的 KATP 通道和淋巴功能障碍

基本信息

批准号：
10229489
负责人：
Michael John Davis
金额：
$ 37.38万
依托单位：
UNIVERSITY OF MISSOURI-COLUMBIA
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-01 至 2023-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10229489
关键词：
Active Biological Transport Acute Affect Attenuated CRSP3 gene Caliber Cannulations Cantu syndrome Cells Chronic Chronic Disease Data Defect Disease Down-Regulation EFRAC Electrodes Endothelium Exhibits Fatty acid glycerol esters Fluid Balance Functional disorder Genetic Human Hydrostatic Pressure Hyperactivity Ileitis Impairment Inflammatory Inflammatory Bowel Diseases Intercellular Fluid Intervention Ion Channel Lead Limb structure Link Liquid substance Lymph Lymphatic Lymphatic Endothelial Cells Lymphatic Endothelium Lymphatic function Lymphedema Measures Mediating Membrane Potentials Methods Molecular Mus Mutant Strains Mice Mutation Near-infrared optical imaging Nitric Oxide Operative Surgical Procedures Pacemakers Patients Peripheral Pharmacology Pinacidil Process Production Prostaglandins Protein Isoforms Pump Role Secondary to Smooth Muscle Smooth Muscle Myocytes Source Testing Tissues base cancer therapy cell type functional restoration gain of function gain of function mutation in vivo indexing inhibitor/antagonist loss of function lymphatic dysfunction lymphatic pump lymphatic vessel mutant novel novel therapeutic intervention novel therapeutics patch clamp pressure primary lymphedema voltage

项目摘要

A striking feature of Cantú Syndrome(CS) is that over 50% of patients present with some form of lymphedema. Although the underlying disease process in CS is known to involve gain-of-function (GOF) mutations in the KATP channel, the mechanism by which lymphedema develops in CS patients is unknown and has not been studied previously. Because spontaneous contractions of collecting lymphatic vessels underlie ? of normal lymph transport, we propose that lymphedema in CS, and potentially other forms of lymphedema, result from reduced excitability of lymphatic smooth muscle. Our central hypothesis is that CS-related GOF mutations in Kir6 and/or SUR subunits result in hyperactivation of KATP current, hyperpolarization of lymphatic smooth muscle and inhibition of the intrinsic electrical pacemaker that drives spontaneous lymphatic contractions. We have pioneered methods to test our hypothesis using popliteal lymphatic vessels of the mouse. Diameter and indices of pumping efficiency are measured in single, cannulated vessels under defined pressure / flow conditions ex vivo. We record membrane potential in either lymphatic smooth muscle or endothelium using sharp electrodes, and dissociate and patch clamp either cell type in order to selectively assess KATP current. In complementary in vivo studies using conventional or near-infrared fluorescence imaging, we assess the function of multi-valve chains of popliteal vessels so that potentially beneficial effects of classic and novel KATP channel antagonists can be tested. Our ability to study mouse lymphatic vessels allows us to take advantage of the global and tissue- specific KATP channel gain- or loss-of-function mutants generated by our collaborators. We have collected a substantial amount of preliminary data to show that Kir6.1 and SUR2 isoforms of KATP are functionally expressed in mouse lymphatic vessels and that spontaneous lymphatic contractions in Kir6.1 or SUR2 GOF mice are severely attenuated but can be rescued by inhibition of KATP channels. These preliminary data are the first demonstration of an underlying contractile defect leading to primary lymphedema. The central hypothesis will be tested with 3 aims. 1) Delineate how KATP channels normally regulate lymphatic pumping. We will assess the roles of LSM and/or LEC KATP channels in mediating inhibition of pumping by nitric oxide and prostanoids. 2) Determine how CS GOF KATP mutations impair lymphatic pumping by expressing the mutant channels in the LEC / LSM layers and measuring excitability and contractile function. 3) Assess whether lymphatic dysfunction can be rescued in mice with CS GOF KATP mutations. The identified source of lymphatic dysfunction in CS (a possible contractility deficit) provides a novel opportunity to test if pharmacologic intervention can be effective in reversing CS-related lymph transport dysfunction. Accomplishment of these aims will elucidate the role of KATP in lymphatic vessels, uncover the mechanism of lymphatic dysfunction in CS, and test a novel therapeutic method to treat lymphedema in CS and other patients with reduced lymphatic smooth muscle excitability.

Cantú综合征（CS）的一个惊人特征是，超过50％的患者患有某种形式的淋巴水肿。尽管已知CS中的潜在疾病过程涉及KATP中的功能奖励（GOF）突变渠道，CS患者淋巴水肿的机制尚不清楚，尚未是以前研究了。因为收集淋巴血管的赞助收缩是？正常淋巴运输，我们提出，CS中的淋巴水肿以及潜在的其他形式的淋巴水肿是由淋巴平滑肌的兴奋减少。我们的中心假设是与CS相关的GOF突变 KIR6和/或SUR亚基导致KATP电流过度激活，淋巴平滑肌的超极化并抑制驱动赞助淋巴收缩的内在电气起搏器。我们有使用小鼠的popliteal淋巴视频来检验我们的假设的开创性方法。直径和指数在定义的压力 /流条件下，在单个插管容器中测量抽水效率的效率体内。我们使用锋利的电子记录淋巴平滑肌或内皮的膜电位，并分离和斑块夹任何一个单元格类型，以选择性评估KATP电流。在完善中使用常规或近红外荧光成像的体内研究，我们评估了多阀的功能 Popliteal血管的链条，因此经典和新型KATP通道拮抗剂的潜在有益作用可以进行测试。我们研究小鼠淋巴视频的能力使我们能够利用全球和组织我们的合作者产生的特定KATP渠道获得或功能丧失突变体。我们收集了一个大量的初步数据表明，KATP的Kir6.1和Sur2同工型在功能上表达在小鼠淋巴视频和Kir6.1或Sur2 GoF小鼠中的赞助淋巴收缩中严重减弱，但可以通过抑制KATP通道做出反应。这些初步数据是第一个导致原发性淋巴水肿的潜在收缩缺陷的证明。中央假设将通过3个目标进行检验。 1）描述KATP通道通常如何调节淋巴抽气。我们将评估LSM和/或LEC KATP渠道在中介抑制中的作用一氧化氮和前列腺素泵送。 2）确定CS GOF KATP突变如何损害淋巴管通过在LEC / LSM层中表达突变通道并测量令人兴奋和收缩的泵送功能。 3）评估是否可以在CS GOF KATP突变的小鼠中检索淋巴功能障碍。 CS中鉴定出的淋巴功能障碍的来源（可能的收缩力防御）提供了一种新的机会为了测试药理学干预是否可以有效地反向CS相关的淋巴转运功能障碍。这些目标的实现将阐明KATP在淋巴管中的作用，发现的机制 CS中的淋巴功能障碍，并测试一种治疗CS和其他患者淋巴水肿的新型治疗方法令人兴奋的淋巴平滑肌降低。