The Role of Macrophage-Derived MMP-9 in LV Remodeling

描述(由申请人提供)： 这是一份A2重新提交的更新申请，用于研究心肌梗死(MI)后的左心室重构。即使采用目前的治疗策略，心肌梗死仍然是心力衰竭的主要原因。识别心肌梗死后刺激左心室(LV)不利重构的事件可能为预防、减缓或逆转心力衰竭的进展提供治疗靶点。心肌梗死后，细胞外基质(ECM)的周转是LV重塑的驱动因素，炎症反应与ECM周转之间存在明确的相关性。对基质金属蛋白酶-9(MMP9)功能的初步分析表明，这种特殊的MMPs主要通过改变巨噬细胞的反应来影响重构，因为MMP9基因缺失的小鼠表现出巨噬细胞在MI后进入左室的功能受损。已有研究表明，基质金属蛋白酶-9可以裂解细胞外基质产生生物活性多肽，并激活转化生长因子b(TGFb)，从而可能将基质金属蛋白酶-9定位于巨噬细胞下游和涉及心脏成纤维细胞的关键事件的上游。因此，该项目的长期目标是了解巨噬细胞和巨噬细胞来源的基质金属蛋白酶-9在左心室对心肌梗死的反应中的作用。这项建议将集中于阐明巨噬细胞和基质金属蛋白酶-9驱动的机制，以关键地检验这一假说，即巨噬细胞通过对细胞外基质和转化生长因子-b的作用调节心肌梗死后的左室反应。使用一种独特的细胞特异性转基因小鼠模型，该模型仅在巨噬细胞中过度表达人基质金属蛋白酶-9，并进行特定的基质金属蛋白酶-9和肿瘤生长因子b干预，我们将确定基质金属蛋白酶-9介导的事件，最大程度地影响左室重构。为了验证我们的中心假说，我们将1)确定巨噬细胞的水平和激活状态是否调节成纤维细胞的激活和左室重构；2)确定基质金属蛋白酶-9和转化生长因子b是否调节巨噬细胞表型、成纤维细胞的激活和左室重构；以及3)确定由基质金属蛋白酶-9产生的生物活性ECM多肽是否通过巨噬细胞和成纤维细胞的激活来调节心肌梗死后的左室重构。我们将使用综合生理学、细胞生物学、生物化学、质谱学和组织学方法的多学科方法来揭示机制，并量化巨噬细胞激活状态和基质金属蛋白酶-9水平对左室重构过程的影响。这一建议是创新的，因为大多数研究使用基质金属蛋白酶-9作为输出测量，只确定基质金属蛋白酶-9水平是否随着刺激的反应而变化，而不是确定酶如何调节细胞外基质重塑。这些研究的结果将阐明巨噬细胞来源的基质金属蛋白酶-9对心肌梗死后重塑的影响。我们的多方面方法将进一步促进对心肌梗死后左室重构事件的机械性理解，这可能为翻译研究提供靶点。

DESCRIPTION (provided by applicant): This is an A2 resubmission of a renewal application to study left ventricular remodeling following myocardial infarction (MI). MI, even with current therapeutic strategies, remains a leading cause of heart failure. The identification of events that stimulate adverse remodeling of the left ventricle (LV) post-MI may provide therapeutic targets to prevent, slow, or reverse the progression to heart failure. Post-MI, extracellular matrix (ECM) turnover is a driving event in LV remodeling, and there is a well- established association between the inflammatory response and ECM turnover. An initial analysis of matrix metalloproteinase-9 (MMP-9) functions suggests that this particular MMP predominantly influences remodeling by altering the macrophage response, as MMP-9 null mice show impaired macrophage influx into the LV post- MI. MMP-9 has been shown to cleave ECM to generate bioactive peptides and to activate transforming growth factor b (TGFb), which potentially places MMP-9 downstream of the macrophage and upstream of key events that involve the cardiac fibroblast. The long-term goals of this project, accordingly, are to understand the roles of macrophages and macrophage-derived MMP-9 in the LV response to MI. This proposal will focus on elucidating macrophage and MMP-9 driven mechanisms to critically test the hypothesis that macrophages modulate the LV response to MI through MMP-9 effects on ECM substrates and transforming growth factor-b. Using a unique cell specific transgenic mouse model that overexpresses human MMP-9 only in macrophages and specific MMP-9 and TGFb interventions, we will determine the MMP-9 mediated events that most influence LV remodeling. To test our central hypothesis, we will 1) determine whether macrophage levels and activation status regulate fibroblast activation and LV remodeling; 2) determine whether MMP-9 and TGFb regulate macrophage phenotype, fibroblast activation, and LV remodeling; and 3) determine whether bioactive ECM peptides generated by MMP-9 regulate LV remodeling post-MI through macrophage and fibroblast activation. We will use a multi-discipline approach that integrates physiology, cell biology, biochemistry, mass spectrometry, and histological approaches to unveil mechanisms and quantify the LV remodeling process as a function of macrophage activation status and MMP-9 levels. This proposal is innovative because most studies use MMP-9 as an output measurement and only determine whether MMP-9 levels change in response to a stimulus, not how the enzyme regulates ECM remodeling. The results of these studies will clarify the consequences of macrophage-derived MMP-9 on post- MI remodeling. Our multi-faceted approach will further advance the mechanistic understanding of the events that initiate post-MI LV remodeling, which may provide targets for translational research.