Role of Fibroblasts in Cardiac Remodeling and Regulation of Cardiomyocytes

成纤维细胞在心脏重塑和心肌细胞调节中的作用

• 批准号: 8834587
• 负责人: Celinda Michelle Kofron
• 金额: $ 5.77万
• 依托单位: RHODE ISLAND HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-01 至 2016-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8834587
• 关键词: Action Potentials; Adult; Affect; Angiotensin II; Area; Arrhythmia; Biochemical; Biomimetics; Calcium; Cardiac; Cardiac Myocytes; Cardiovascular Diseases; Cell Communication; Cell Count; Cells; Cessation of life; Chemicals; Coronary Arteriosclerosis; Coupling; Defect; Deposition; Development; Dyes; Endothelin-1; Event; Experimental Models; Extracellular Matrix; Fibroblasts; Fibrosis; Gap Junctions; Gene Transfer; Generations; Healed; Heart; Heart failure; Hereditary Disease; Histology; Homeostasis; Hypertension; Hypertrophy; Immunohistochemistry; Individual; Infarction; Injury; Laboratories; Lead; Location; Maps; Mechanics; Mediating; Mediator of activation protein; Modeling; Morphology; Muscle; Myocardial; Myocardial Infarction; Myocardial tissue; Myocardium; Myofibroblast; Neonatal; Optics; Outcome; Performance; Physiological; Population; Prevention; Proliferating; Property; Rattus; Regulation; Research; Research Design; Role; Shapes; Signal Pathway; Signal Transduction; Stress; Testing; Therapeutic; Tissues; Ventricular; Work; cardiovascular risk factor; healing; hemodynamics; improved; in vitro Model; in vivo; inhibitor/antagonist; interstitial; mortality; mutant; overexpression; paracrine; pressure; prevent; protein expression; public health relevance; response; response to injury; scaffold; subcutaneous; sudden cardiac death; three dimensional structure; three-dimensional modeling

 DESCRIPTION (provided by applicant): Cardiac fibroblasts (CFs), along with cardiac myocytes (CMs), maintain the three-dimensional structure and electrical, chemical, and biochemical homeostasis of the myocardium. In cardiovascular disease, CF become activated, proliferate, and produce more extracellular matrix (ECM), leading to the development of fibrosis. Fibrosis can be divided in reactive (or interstitial) fibrosis, a common response to excessive myocardial loads that causes excess ECM deposition between muscle bundles, and reparative (or compact) fibrosis, a healing response to maintain structural integrity after CM death in response to injury or infarct. CFs (20% of the volume and 40-60% of the total cardiac cell population) are interwoven in densely packed CMs (75% of tissue volume but less than 50% in cells number) and can influence CMs via paracrine effects, direct cell-cell interactions, and indirect cell-ECM interactions. While regulation of CMs by CFs is known to occur in the myocardium in vivo, the mechanisms and functional significance of CF-to-CM cross talk under physiological and pathophysiological conditions are not well understood. The objective of my proposal is to determine the functional consequences and mechanisms of CF-to-CM regulation using a scaffold-free 3D model developed in my sponsor's lab that intermingles CFs and CMs as in the healthy myocardium. I will generate microtissues with interstitial fibrosis by co-seeding neonatal rat ventricular CMs with CFs that are selectively activated (Aim 1). To that end, I will utilize adenoviral gene transfer to overexpress a constitutively active mutant Gαq, a key mediator of CF activation, or use CFs from rats infused with Ang II for 2 weeks in vivo. In Aim 2, I will develop 3D microtissues with activated CFs in compact configuration that separate CM mimicking compact fibrosis using individual microtissues as building blocks. In both Aims, I will test my central hypothesis is that the number of CFs, the concentration of CFs in a given area (or type of fibrosis: compact or interstitial), and the activation state of CFs influence the morphology and electrical activity in cardiac microtissues. Morphological changes of the microtissues will be characterized with histology and immunohistochemistry. Action potentials and calcium transients will be investigated as key parameters of CM function using optical mapping. Potential mechanisms by which the functional and morphological changes are driven will be investigated using immunohistochemistry, protein expression analysis, dye transfer, and gap junction inhibitors. The proposed study is expected to advance understanding of the impact of CFs on CMs the integrated functional response of cardiac microtissues and may elucidate new avenues for therapeutic strategies to treat and/or prevent structural and electrical remodeling in the diseased heart.

 描述（由申请人提供）：心脏成纤维细胞（CF）与心肌细胞（CM）一起维持心肌的三维结构以及电、化学和生化稳态。在心血管疾病中，CF被激活、增殖和增殖。产生更多的细胞外基质（ECM），导致纤维化的发展可分为反应性（或间质性）纤维化，这是过度反应的常见反应。心肌负荷导致肌束之间过量的 ECM 沉积，以及修复性（或致密性）纤维化，这是因损伤或梗塞而导致 CM 死亡后维持结构完整性的愈合反应（体积的 20% 和纤维化的 40-60%）。心肌细胞总数）交织在密集的 CM 中（占组织体积的 75%，但细胞数量不到 50%），并且可以通过旁分泌效应、直接细胞间作用影响 CM虽然已知 CF 对 CM 的调节发生在体内心肌中，但 CF 与 CM 相互作用在生理和病理生理条件下的机制和功能意义尚不清楚。我的建议是使用我的赞助者实验室开发的无支架 3D 模型来确定 CF 到 CM 调节的功能后果和机制，该模型将 CF 和 CM 混合在一起，就像在健康心肌 I 中一样。将通过共同播种产生间质纤维化的微组织 具有选择性激活的 CF 的新生大鼠心室 CM（目标 1）为此，我将利用腺病毒基因转移过表达组成型活性突变体 Gαq（CF 激活的关键介质），或使用来自注入 Ang II 的大鼠的 CF。在目标 2 中，我将在体内开发 3D 微组织，其具有紧凑结构的激活 CF，可使用个体模拟紧凑纤维化来分离 CM。在这两个目标中，我将测试我的中心假设，即 CF 的数量、给定区域中 CF 的浓度（或纤维化类型：致密型或间质型）以及 CF 的激活状态会影响形态。心脏微组织的形态变化将通过组织学和免疫组织化学来表征，动作电位和钙瞬变将作为 CM 功能的关键参数进行研究。将使用免疫组织化学、蛋白质表达分析、染料转移和间隙连接抑制剂来研究驱动功能和形态变化的潜在机制。这项研究有望促进对 CF 对 CM 的影响的理解。心脏微组织的功能反应，并可能阐明治疗和/或预防患病心脏结构和电重塑的治疗策略的新途径。