Chemokines in healing myocardial infarction

摘要： 转化生长因子-超家族成员在梗死心脏重塑中发挥核心作用，调节 参与心肌炎症、修复和纤维化的所有细胞类型的表型和功能。转化生长因子- S通过激活涉及受体激活的SMADs(R-SMADs)的分子级联发挥作用，或 通过独立于SMAD的机制。然而，有效的修复需要对转化生长因子-的严格调控。 行动，以防止导致不良适应心脏重塑的不良后果。 过度激活或延长的转化生长因子-信号可促进心肌细胞、成纤维细胞和巨噬细胞 激活，刺激持续性纤维化、肥大和吞噬反应。内生性 负调控转化生长因子-在组织修复、重塑和修复中的作用机制 纤维化仍不清楚。 目前的提案探索了负责抑制和 心肌梗死和重构心肌中转化生长因子-超家族信号的终止。我们的 未发表的初步数据表明，两种不同的机制在负调控中起着重要作用 转化生长因子-超家族信号。首先，抑制性Smads的细胞特异性诱导(I-Smads)， Smad7和Smad6对心肌梗死后转化生长因子-和骨形态发生蛋白的负性调节作用 脑梗塞。第二，转化生长因子-信号可能通过细胞特异性在受体水平进行调节。 BMP和激活素结合的跨膜假性受体Bambi的诱导 抑制物)，它缺乏细胞内的激酶域，并且至少部分地抑制转化生长因子-信号转导， 通过与i-SMADS的交互。负调控的细胞特异性机制的作用 转化生长因子-的研究将在4个具体目标中进行： 特异性目的1：探讨Smad7在心肌细胞、成纤维细胞调节中的作用 和脑梗塞后巨噬细胞表型。我们的初步研究表明Smad7是 在交界区心肌细胞、梗死区肌成纤维细胞和巨噬细胞中显著上调， Smad7基因敲除或过表达调节转化生长因子-驱动的细胞反应。因此， 我们将使用心肌细胞、成纤维细胞/肌成纤维细胞和髓系细胞特异的Smad7基因敲除小鼠， 最近由我们的实验室开发，以探索Smad7对梗塞心脏的细胞效应。 具体目标2：剖析导致这些效应的分子机制。 Smad7在体内和体外的表达。Smad7操作可能涉及抑制依赖于Smad或不依赖于Smad的 SmAD途径，可能涉及与T受体、R-SMADs或转化生长因子-非依赖性信号的相互作用。我们的 心脏成纤维细胞的初步研究表明，Smad7在没有Smad7的情况下抑制Smad2/3的激活 影响T-受体的磷酸化。Smad7依赖于细胞周期调控的分子机制 心肌细胞、成纤维细胞和巨噬细胞的表型将在体外和体内进行研究，使用 功能损失法和功能增益法。 特异目的3：探讨Smad6在脑梗塞修复和重塑中的作用 心。我们的初步研究表明，Smad6在心肌细胞、成纤维细胞和巨噬细胞中诱导 并证明成纤维细胞Smad6的作用不同于 Smad7介导的效应。将产生有条件的Smad6基因敲除小鼠来解剖细胞- Smad6在心肌梗死和重构中的作用及其机制 负责Smad6介导的效应将在体内和体外进行探索。 特异性目标4：研究BAMBI在调节血管内皮细胞转化生长因子-反应中的作用 心肌梗死和心脏重塑。我们的初步研究表明，Bambi在脑梗塞中表达上调 肌成纤维细胞和巨噬细胞。因此，我们将使用成纤维细胞和巨噬细胞特异性的Bambi 基因敲除和体外实验，以研究Bambi在调节细胞特异性转化生长因子-α中的作用。 在梗塞心脏中的作用。此外，我们还将探讨Bambi与I- 斯马德。 拟议的研究将提供第一次系统的机制调查。 在组织损伤模型中负调控转化生长因子-的作用。

ABSTRACT: TGF- superfamily members play a central role in remodeling of the infarcted heart, modulating phenotype and function of all cell types involved in myocardial inflammation, repair, and fibrosis. TGF- s function by activating molecular cascades involving Receptor-activated Smads (R-Smads), or through Smad-independent mechanisms. However, effective repair requires tight regulation of TGF- actions, in order to prevent adverse consequences that lead to maladaptive cardiac remodeling. Overactive, or prolonged TGF- signaling can promote cardiomyocyte, fibroblast and macrophage activation, stimulating persistent fibrotic, hypertrophic and phagocytic responses. The endogenous mechanisms responsible for negative regulation of TGF- responses in tissue repair, remodeling and fibrosis remain unknown. The current proposal explores the molecular mechanisms responsible for suppression and termination of TGF- superfamily signaling in the infarcted and remodeling myocardium. Our unpublished preliminary data suggest an important role for 2 distinct mechanisms in negative regulation of TGF- superfamily signaling. First, cell-specific induction of the inhibitory Smads (I-Smads), Smad7 and Smad6 may negatively regulate TGF- and BMP responses following myocardial infarction. Second, TGF- signaling may be regulated at the receptor level through cell-specific induction of the transmembrane pseudoreceptor BAMBI (BMP and activin membrane-bound inhibitor), which lacks an intracellular kinase domain, and inhibits TGF- signaling, at least in part, through interactions with I-Smads. The role of the cell-specific mechanisms for negative regulation of TGF- will be explored in 4 specific aims: Specific aim 1: to investigate the role of Smad7 in regulation of cardiomyocyte, fibroblast and macrophage phenotype following infarction. Our preliminary studies show that Smad7 is markedly upregulated in border zone cardiomyocytes, and in infarct myofibroblasts and macrophages, and that Smad7 knockdown or overexpression modulate TGF--driven cellular responses. Accordingly, we will use cardiomyocyte-, fibroblast/myofibroblast- and myeloid cell-specific Smad7 knockout mice, recently generated by our laboratory to explore the cellular effects of Smad7 on the infarcted heart. Specific aim 2: to dissect the molecular mechanisms responsible for the effects of Smad7 in vivo and in vitro. Smad7 actions may involve suppression of Smad-dependent or non- Smad pathways and may involve interactions with TRs, R-Smads or TGF--independent signals. Our preliminary studies in cardiac fibroblasts suggest that Smad7 restrains Smad2/3 activation without affecting phosphorylation of TRs. The molecular mechanisms for Smad7-dependent regulation of cardiomyocyte, fibroblast and macrophage phenotype will be studied in vitro and in vivo, using both loss and gain-of-function approaches. Specific aim 3: to investigate the role of Smad6 in repair and remodeling of the infarcted heart. Our preliminary studies show Smad6 induction in cardiomyocytes, fibroblasts, and macrophages infiltrating the healing infarct, and demonstrate that fibroblast Smad6 exerts actions distinct from Smad7-mediated effects. Conditional Smad6 knockout mice will be generated to dissect the cell- specific actions of Smad6 in the infarcted and remodeling myocardium, and the mechanisms responsible for Smad6-mediated effects will be explored in vivo and in vitro. Specific aim 4: to study the role of BAMBI in regulation of TGF- responses in the infarcted and remodeling heart. Our preliminary studies show that BAMBI is upregulated in infarct myofibroblasts and macrophages. Accordingly, we will use fibroblast- and macrophage-specific BAMBI knockouts and in vitro experiments, in order to study the role of BAMBI in modulating cell-specific TGF-  actions in the infarcted heart. Moreover, we will explore interactions between BAMBI and the I- Smads. The proposed studies will provide the first systematic investigation of the mechanisms responsible for negative regulation of TGF- in a model of tissue injury.