Cellular Mechanisms of Lymphatic Muscle Contractility

淋巴肌收缩力的细胞机制

基本信息

批准号：
7806457
负责人：
Michael John Davis
金额：
$ 33.34万
依托单位：
UNIVERSITY OF MISSOURI-COLUMBIA
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-05-01 至 2013-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7806457
关键词：
Abbreviations Address Adenoviruses Affect Agonist Behavior Blood Vessels Blood capillaries Caliber Cardiac Chronic Contractile Proteins Contracts Coupled Down-Regulation Drainage procedure Edema Equilibrium Exhibits Extracellular Fluid Figs - dietary Financial compensation Fluid Balance Frequencies Functional disorder Heart Hybrids Immunoblotting Immunofluorescence Microscopy Impairment In Vitro Infection Isometric Exercise Isotonic Exercise Laboratories Lead Length Light Liquid substance Lymphangiogenesis Lymphatic Lymphatic Capillaries Lymphatic System Lymphatic vessel Lymphedema MYLK gene Mastectomy Measures Mediating Mesentery Methods Modeling Molecular Profiling Molecular Target Monitor Muscle Muscle Cells Muscle Contraction Muscle function Myocardium Myosin Heavy Chains Myosin Light Chain Kinase Output Pacemakers Pain Patients Phenotype Physiological Physiology Play Preparation Protein Isoforms Protein Overexpression Proteins Protocols documentation Pump Rattus Reconstructive Surgical Procedures Regulation Relaxation Resistance Risk Role Running Small Interfering RNA Smooth Muscle Smooth Muscle Myosins Speed Striated Muscles Substance P System Testing Therapeutic Therapeutic Agents Therapeutic Intervention Thick Filament Thin Filament Time Tissues Transfection Transgenic Mice Translating Tropomyosin Troponin C Vascular Smooth Muscle Venous Western Blotting Work base citrate carrier genetic regulatory protein improved in vivo knock-down lymph flow lymphatic pump mouse model non-muscle myosin heavy chain-B overexpression pressure protein expression vector

项目摘要

DESCRIPTION (provided by applicant): Over ten million people in the US currently suffer from some form of lymphedema, including a high percentage of patients recovering from mastectomy or reconstructive surgery. In conjunction with methods to promote lymphangiogenesis in chronically edematous tissue, strategies to enhance lymphatic pump function and drainage of the affected regions are necessary. Lymphatics display a dramatically different contractile phenotype than blood vessels, in which contractions are characterized by both phasic and tonic components; blood vessels exhibit predominantly tonic behavior. Rat mesenteric lymphatics serve as a prototypical collecting lymphatic vessel model, allowing both in vivo and in vitro studies. Surprisingly, those vessels express contractile protein isoforms typically found only in striated muscle: troponin C (cTn-C), 1-striated tropomyosin (1-TMstr), and the fast, 2B isoform of myosin heavy chain (SM-B). Their functional roles in lymphatics are unknown. This unique expression profile and our recent findings that lymphatic muscle has a much higher shortening velocity than arterial or venous smooth muscle, suggest that lymphatic muscle functions as a hybrid between vascular smooth muscle and cardiac muscle. We propose to test the hypothesis that expression of SM-B MHC, cTn-C and 1- TMstr enable collecting lymphatic vessels to undergo the rapid, phasic contractions and relaxations required for normal lymphatic pump function. We will use a combination of isobaric, isometric and isotonic lymphatic preparations unique to our laboratory that enable us to comprehensively assess both phasic and tonic components of the lymphatic pump. These methods will be combined with short-term vessel culture and adenoviral transfection methods that allow protein overexpression or siRNA-mediated protein knockdown to change the expression of SM-B, Tn-C and 1-TM, alone and in combination, over a period of up to 14 days. In conjunction with functional tests to assess phasic and tonic components of contractility in vitro, message/protein expression of the targets will be monitored by RT/PCR, Western blotting and immunofluorescence microscopy. We predict that the expression of these three proteins imparts the uniquely high rate of lymphatic muscle contraction/relaxation required for intrinsic pacemaker activity to be translated into efficient lymphatic pumping. Completion of this work will advance our understanding of lymphatic contraction and lead to therapeutic strategies whereby lymphatic pump function can be enhanced in the absence of collateral effects on blood vessels. Project Narrative: Lymphatic capillaries run in parallel to blood vessels and capture excess fluid filtered out of blood capillaries. Lymphatic vessels move fluid uphill against a pressure gradient and therefore require robust, heart-like, pumping activity of the muscle cells in their walls. Dysfunction of the lymphatic pump system is associated with edema, pain, lack of mobility, and increased risk of infection conditions that affect more than 10 million people in the USA. These studies will investigate which proteins allow these vessels to contract so that specific therapeutic agents can be developed to correct lymphatic pump dysfunction and drainage of edematous tissues.

描述（由申请人提供）：美国目前有超过一千万人患有某种形式的淋巴水肿，包括从乳房切除术或重建手术中恢复过的患者中有很高的比例。结合促进慢性水肿组织中淋巴管生成的方法，必须采用增强淋巴泵功能和受影响区域的排水的策略。淋巴管的收缩表型与血管的收缩表型截然不同，在该血管中，收缩的特征是阶段性和补品成分。血管主要表现出强调行为。大鼠肠系膜淋巴管是一种典型的收集淋巴管模型，可以在体内和体外研究。令人惊讶的是，这些血管表达通常仅在横纹肌肉中发现的收缩蛋白同工型：肌钙蛋白C（CTN-C），1型肌球蛋白（1-TMSTR）和肌球蛋白重链（SM-B）的快速2B同工型。它们在淋巴管中的功能作用尚不清楚。这种独特的表达曲线以及我们最近的发现，即淋巴肌比动脉或静脉平滑肌的缩短速度要高得多，这表明淋巴肌肉起着血管平滑肌和心脏肌肉之间的杂种。我们建议测试以下假设：SM-B MHC，CTN-C和1-TMSTR的表达使收集淋巴管的表达能够经历正常淋巴泵功能所需的快速，体性收缩和弛豫。我们将结合我们实验室独有的等速剂，等距和等渗淋巴制剂，使我们能够全面评估淋巴泵的阶段和滋补成分。这些方法将与短期血管培养和腺病毒转染方法结合使用，这些方法允许蛋白过表达或siRNA介导的蛋白质敲低，以更改SM-B，TN-C和1-TM的表达，单独和组合，在长达14天的时间内。与功能测试一起评估体外收缩性的阶段和补品成分，目标的信息/蛋白质表达将通过RT/PCR，蛋白质印迹和免疫荧光显微镜监测。我们预测，这三种蛋白质的表达赋予了固有的起搏器活性所需的淋巴肌肉收缩/松弛率，以便将其转化为有效的淋巴泵送。这项工作的完成将提高我们对淋巴收缩的理解，并导致治疗策略，在这些策略中，在没有对血管附带影响的情况下，可以增强淋巴泵功能。项目叙述：淋巴毛细血管与血管并行运行，并捕获从毛细血管中过滤的多余液体。淋巴管在压力梯度上移动液体上坡，因此需要坚固的，类似心脏的泵送活性的肌肉细胞在其壁中。淋巴泵系统功能障碍与水肿，疼痛，流动性不足以及影响美国超过1000万人的感染状况的增加有关。这些研究将研究哪种蛋白质允许这些血管收缩，以便可以开发特定的治疗剂来纠正淋巴泵功能障碍和水肿组织的排水。