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: 8198360
• 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/8198360
• 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（ISF）空间中启动了一系列生化和炎症事件，这些事件与细胞表面分子相互作用，决定细胞外基质（ECM）的更新，随后发生适应不良的左心室（LV）重塑。响应于压力过载，ECM产生增加，而在纯VO中ECM减少。VO中控制ECM反应的分子机制尚不清楚。目前，使用血管紧张素转换酶（ACE）抑制剂治疗，降低血管紧张素II和增加缓激肽（BK），对病理生理性LV VO无效（1）。因此，本提案的目的是了解ECM损失的机制，以防止进行性LV重构和VO失败。 我们已经证明，大鼠下腔静脉瘘（ACF）的急性VO增加了ISF BK（2）。BK通过激活激肽释放酶/激肽系统和BK 2受体预防纤维化并降低心血管和肾脏疾病的血压（3）。在急性VO中，BK 2受体拮抗作用可防止ECM损失、肥大细胞（MC）蓄积和LV扩张（2，4）。然而，平均动脉压升高，这排除了VO中BK 2拮抗剂的长期治疗。 除了其抗纤维化作用外，BK还对炎性细胞，特别是肥大细胞（MC）具有趋化性，肥大细胞是丝氨酸蛋白酶糜酶的来源。我们已经表明MC具有BK 2受体，并且直接BK间质输注导致MC脱粒和糜酶释放到体内野生型小鼠的ISF空间中。除了其血管紧张素II形成能力外，糜蛋白酶还激活激肽释放酶，这可以进一步增加ISF BK（5）。激肽释放酶和糜蛋白酶还激活基质金属蛋白酶（MMP）（6）并直接降解纤连蛋白（7），纤连蛋白是控制细胞生长和存活的粘着斑复合物的重要组分。ISF BK和激肽释放酶基因表达，以及增加的MC和糜酶活性，在早期和晚期阶段的VO增加，表明心脏BK形成响应VO。 两者合计，我们的数据表明，一个恶性循环的本地生产的心脏激肽释放酶和BK，这反过来又刺激释放MC-糜蛋白酶和激活激肽释放酶，从而永久增加心脏BK在VO。因此，我们假设，VO的纯拉伸诱导心脏激肽释放酶/激肽生产，产生减少ECM合成和增加MC脱粒。激肽释放酶的抑制和下调将减少ISF BK的形成和MC糜酶的释放，并进一步阻止MMP的激活，从而改善ACF VO大鼠的LV重构和功能。目的1确定激肽释放酶激活是否通过MC激活介导心脏ECM丢失。使用体内微透析，我们已经表明，BK在正常大鼠的间质输注与MC和ECM损失的增加。我们将确定激肽释放酶输注是否诱导BK 2受体活化，随后刺激MC-糜酶释放并介导ECM损失。为了明确肥大细胞糜酶的作用，我们将在正常大鼠心脏和用ACF进行VO的大鼠心脏中用ISF BK输注或激肽释放酶输注治疗肥大细胞缺陷大鼠。目的2确定心肌细胞或成纤维细胞是否是体外拉伸和体内VO引起的激肽释放酶表达和BK释放增加的来源。为了分离单独拉伸的效果，新生儿LV肌细胞和成人LV成纤维细胞将被拉伸以模拟VO，以确定其产生激肽释放酶和BK的差异能力。为了剖析VO中激肽释放酶抑制（抑肽酶）的直接与全身效应，我们将在体内敲除急性ACF中的激肽释放酶，并研究对ISF BK和糜酶活性、MMP/TIMP、ECM和非ECM蛋白的影响。目的3确定抑制激肽释放酶是否会改善VO大鼠心脏的左室重构和功能。我们将研究激肽释放酶抑制剂对体内LV功能、ECM稳态、ISF BK和糜酶活性以及肥大细胞脱颗粒在早期和慢性时间点对VO的反应的影响。 公共卫生相关性： 在美国，心力衰竭影响超过400万人，一旦发生心力衰竭，6年死亡率超过65%。预计在未来30年内，我国老龄化人口中受影响的人数将大幅增加。心力衰竭的病因很多，先天性缺陷和感染诱发的风湿热，都是导致心脏瓣膜病的最常见症状。我们的研究将提供新的机制的差异适应性反应的循环与组织特异性激肽释放酶/激肽产生机制的心血管疾病的演变和靶向缓激肽治疗对这种变化的影响的见解。我们目前的提案将发现新的靶点，从而改善治疗策略，用于管理主动脉瓣返流和二尖瓣返流容量超负荷患者，这些患者占我们退伍军人患者人群的很大一部分。