The Kallikrein/Kinin-Mast Cell Interaction in Volume Overload

容量超负荷时激肽释放酶/激肽-肥大细胞的相互作用

• 批准号: 8597411
• 负责人: Chih-Chang Kevin Wei
• 金额: --
• 依托单位: BIRMINGHAM VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-10-01 至 2014-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8597411
• 关键词: Acute; Adult; Affect; Angiotensin II; Angiotensin-Converting Enzyme Inhibitors; Aortic Valve Insufficiency; Aprotinin; BDKRB2 gene; Biochemical; Blood Pressure; Bradykinin; Cardiac; Cardiac Myocytes; Cardiovascular Diseases; Cell Communication; Cell Degranulation; Cell Survival; Cell surface; Cells; Chronic; Chymase; Complex; Congenital Abnormality; Data; Dilatation - action; Down-Regulation; Event; Evolution; Extracellular Matrix; Extracellular Matrix Proteins; Failure; Fibroblasts; Fibronectins; Fibrosis; Fistula; Focal Adhesions; Gene Expression; Heart; Heart Valve Diseases; Heart failure; Homeostasis; In Vitro; Individual; Infection; Inflammatory; Infusion procedures; Intercellular Fluid; Kallikrein-Kinin System; Kidney Diseases; Kininogenase; Kinins; Lead; Left; Left Ventricular Function; Left Ventricular Remodeling; Matrix Metalloproteinases; Mediating; Microdialysis; Mitral Valve Insufficiency; Molecular; Motion; Muscle Cells; Neonatal; Patients; Peptide Hydrolases; Production; Rattus; Receptor Activation; Rheumatic Fever; Role; Serine Protease; Simulate; Source; Staging; Stretching; Symptoms; Therapeutic; Time; Tissues; United States; Ventricular; Veterans; Wild Type Mouse; aging population; cell growth; improved; in vivo; insight; interstitial; mast cell; mortality; novel; patient population; pressure; prevent; public health relevance; receptor; response

DESCRIPTION (provided by applicant): The onset of volume overload (VO) sets in motion a sequence of biochemical and inflammatory events in the cardiac interstitial fluid (ISF) space that interact with cell surface molecules to dictate extracellular matrix (ECM) turnover with subsequent maladaptive left ventricular (LV) remodeling. In response to pressure overload there is an increase ECM production while in the pure VO there is a decrease in ECM. The molecular mechanisms that control ECM response in VO are not understood. Currently, therapy with angiotensin-converting enzyme (ACE) inhibitors, which decreases angiotensin II and increases bradykinin (BK), is ineffective in a pathophysiologic LV VO (1). Therefore, the purpose of the current proposal is to understand the mechanisms of ECM loss in order to prevent progressive LV remodeling and failure in VO. We have shown that the acute VO of aortocaval fistula (ACF) in the rat has increased ISF BK (2). BK through activation of the kallikrein/kinin system and BK2 receptor prevents fibrosis and decreases blood pressure in cardiovascular and renal disease (3). In acute VO, BK2 receptor antagonism prevents ECM loss, mast cell (MC) accumulation and LV dilatation (2, 4). However, there was an increase in mean arterial pressure, which precludes long term therapy with BK2 antagonism in VO. In addition to its antifibrotic effect, BK is chemotactic for inflammatory cells, in particular mast cells (MCs), which are the source of the serine protease chymase. We have shown MCs have BK2 receptors and direct BK interstitial infusion causes MC degranulation and chymase release into the ISF space in wild type mice in vivo. Besides its Ang II-forming capacity, chymase activates kallikrein which can further increase ISF BK (5). Kallikrein and chymase also activate matrix metallo- proteinases (MMPs) (6) and directly degrade fibronectin (7), an important component of the focal adhesion complex that controls cell growth and survival. ISF BK and kallikrein gene expression, as well as increased MCs and chymase activity, are increased at both early and late stages of VO, suggesting cardiac BK formation in response to VO. Taken together, our data suggest a vicious cycle of local production of cardiac kallikrein and BK, which in turn, stimulates release of MC-chymase and activates kallikrein, thus perpetuating the increase in cardiac BK in VO. Thus, we hypothesize that the pure stretch of VO induces cardiac kallikrein/kinin production that produces both a decrease in ECM synthesis and increase MC degranulation. Kallikrein inhibition and downregulation will decrease ISF BK formation and MC chymase release and further prevent MMP activation, resulting in improved LV remodeling and function in rat with the VO of ACF. Aim 1 Determine whether kallikrein activation mediates ECM loss in the heart through MC activation. Using in vivo microdialysis, we have shown that interstitial infusion of BK in the normal rat is associated with increase in MCs and ECM loss. We will determine whether kallikrein infusion induces BK2 receptor activation and subsequently stimulates MC-chymase release and mediates ECM loss. To define the role of mast cell chymase, we will treat mast cell deficient rats with ISF BK infusion or kallikrein infusion in the normal rat heart and in the rat heart subjected to VO with ACF. Aim 2 Determine whether cardiomyocytes or fibroblasts are the source of increased kallikrein expression and BK release in response to stretch in vitro and VO in vivo. To isolate the effect of stretch alone, neonatal LV myocytes and adult LV fibroblasts will be stretched to simulate VO to determine their differential ability to produce kallikrein and BK. To dissect out the direct vs. systemic effects of kallikrein inhibition (aprotinin) in VO, we will knockdown kallikrein in the acute ACF in vivo and study effects on ISF BK and chymase activity, MMP/TIMPs, ECM and non ECM proteins. Aim 3 Determine whether inhibition of kallikrein will improve LV remodeling and function in the VO rat heart. We will study the effects of kallikrein inhibition on in vivo LV function, ECM homeostasis, ISF BK and chymase activity, and mast cell degranulation in response to VO in the early and chronic time points. PUBLIC HEALTH RELEVANCE: Project Narrative In the United States, heart failure affects more than 4 million people, who display 6-year mortality rates of greater than 65% once failure develops. The number of affected individuals is expected to rise significantly in our aging population over the next 30 years. Heart failure has many causes, congenital defects and infections induced rheumatic fever, are among the most common symptoms lead to heart valve disease. Our studies will provide novel mechanistic insights into the differential adaptive responses of the circulating versus tissue specific kallikrein/kinin generating mechanism in the evolution of cardiovascular disease and the impact of targeted bradykinin therapy on such changes. Our current proposal will uncover new targets that lead to improved therapeutic strategies for the management of patients with the volume overload of aortic regurgitation and mitral regurgitation which constitutes a large portion of our Veteran patient population.

描述(由申请人提供)： 容量超负荷(VO)的发生在心脏间质液体(ISF)间隙启动了一系列的生化和炎症事件，这些事件与细胞表面分子相互作用，决定了细胞外基质(ECM)的周转，从而导致随后的左心室(LV)重构。在压力过载时，ECM产量增加，而在纯VO中，ECM产量减少。控制VO中ECM反应的分子机制尚不清楚。目前，血管紧张素转换酶(ACE)抑制剂可降低血管紧张素II，增加缓激肽(BK)，对病理性LV VO无效(1)。因此，本研究的目的是为了了解ECM丢失的机制，以防止VO的进行性左室重构和失败。我们发现，大鼠主动脉腔内瘘(ACF)的急性VO增加了ISF BK(2)。BK通过激活激肽释放酶/激动素系统和BK2受体，在心血管和肾脏疾病中预防纤维化和降低血压(3)。在急性VO中，BK2受体拮抗剂可防止ECM丢失、肥大细胞(MC)聚集和左室扩张(2，4)。然而，平均动脉压升高，这排除了长期使用BK2拮抗剂治疗VO的可能性。除了抗纤维化作用外，BK还对炎症细胞，特别是肥大细胞(MC)具有趋化作用，而肥大细胞(MC)是丝氨酸蛋白酶糜酶的来源。我们已经证明MCs具有BK2受体，在体内，直接BK间质输注可以引起MC脱颗粒和糜酶释放到ISF间隙。除了Ang II的形成能力外，糜酶还能激活激肽释放酶，从而进一步增加ISF BK(5)。激肽释放酶和糜酶还激活基质金属蛋白酶(MMPs)(6)并直接降解纤维连接蛋白(7)，纤维连接蛋白(7)是控制细胞生长和存活的焦点黏附复合体的重要组成部分。在VO的早期和晚期，ISF BK和激肽释放酶基因的表达，以及MCs和糜酶活性的增加，提示VO导致心脏BK的形成。综上所述，我们的数据提示了心脏激肽释放酶和BK的局部产生的恶性循环，这反过来刺激MC-糜酶的释放并激活激肽释放酶，从而使VO中心脏BK的增加永久化。因此，我们假设，单纯的VO牵张可诱导心脏激肽释放酶/激动素的产生，从而导致ECM合成减少和MC脱颗粒增加。激肽释放酶抑制和下调将减少ISF BK的形成和MC糜酶的释放，进一步阻止MMPs的激活，从而改善ACF VO大鼠的左室重构和功能。目的1确定激肽释放酶激活是否通过MC激活介导心肌细胞外基质丢失。利用体内微透析，我们已经证明，在正常大鼠体内，间质内注入BK与MCs增加和ECM丢失有关。我们将确定激肽释放酶是否诱导BK2受体激活，继而刺激MC-糜酶释放，并介导细胞外基质丢失。为了明确肥大细胞糜酶的作用，我们将在正常大鼠心脏和VO合并ACF的大鼠心脏中用ISF BK或激肽释放酶输注治疗肥大细胞缺陷大鼠。目的2确定体外牵张和体内VO引起激肽释放酶表达和BK释放增加的来源是心肌细胞还是成纤维细胞。为了分离单独拉伸的影响，我们将拉伸新生LV心肌细胞和成年LV成纤维细胞来模拟VO，以确定它们产生激肽释放酶和BK的不同能力。为了探讨激肽释放酶抑制(抑肽酶)在VO中的直接和全身作用，我们将在体内敲除急性ACF中的激肽释放酶，并研究其对ISF、BK和Chymase活性、MMP/TIMPs、ECM和非ECM蛋白的影响。目的3确定抑制激肽释放酶是否能改善VO大鼠心脏的重塑和功能。我们将在早期和慢性时间点研究激肽释放酶抑制对体内左心功能、细胞外基质稳态、ISF-BK和糜酶活性以及肥大细胞脱颗粒的影响。 公共卫生相关性： 在美国，心力衰竭影响了400多万人，一旦发生心力衰竭，他们的6年死亡率超过65%。预计未来30年，在我们老龄化的人口中，受影响的个人数量将大幅上升。心力衰竭的原因很多，先天性缺陷和感染导致的风湿热是导致心脏瓣膜疾病的最常见症状之一。我们的研究将为心血管疾病进化中循环和组织特异性激肽释放酶/激动素产生机制的差异适应反应以及靶向缓激肽治疗对这些变化的影响提供新的机制见解。我们目前的提案将发现新的目标，从而改进治疗策略，以管理主动脉瓣反流和二尖瓣反流患者的容量超负荷，这在我们的资深患者群体中占很大比例。