Role of Toll-Like Receptor Signaling in Cardiac Ischemia

描述（由申请人提供）：心血管疾病是美国的头号杀手。toll样受体（TLR）介导的先天免疫和炎症反应在心肌缺血/再灌注（I/R）损伤的病理生理机制中起核心作用。众所周知，激活tlr4介导的NF-kB信号通路在心肌I/R损伤中起有害作用，而激活PI3K/Akt信号通路可保护心肌I/R损伤。我们已经报道了TLR4缺乏或TLR2调节通过PI3K/Akt依赖机制保护心肌免受I/R损伤。我们发现TLR4缺失会增加心肌中microRNA-146a （miR-146a）和miR-486的表达。我们还发现，TLR2调节增加了心肌、循环外泌体和骨髓间充质干细胞分泌的外泌体中miR-146a和miR-486的数量。我们已经证明，心肌中miR-146a表达的增加可以减少心肌梗死面积，改善心肌I/R后的心功能。MiR-146a靶向IRAK1和TRAF6，而IRAK1和TRAF6是tlr介导NF-kB激活的重要中间体，从而减弱I/ r诱导的NF-kB激活。此外，我们发现miR-486抑制PTEN表达，导致PI3K/Akt信号通路激活。我们的研究结果表明，TLR4缺陷或TLR2调节通过上调抑制NF-kB激活和激活PI3K/Akt信号的microRNA表达来诱导心脏保护。综合考虑，我们的数据和文献清楚地表明，tlr4介导的NF-kB激活与心肌I/R损伤的疾病进展密切相关，而PI3K/Akt信号通路的激活与心肌损伤的衰减密切相关。目前，我们尚未完全了解NF-kB和PI3K/Akt在I/R损伤中的差异调控机制。该应用程序旨在破译I/R诱导心肌损伤中差异调节这些信号通路的细胞和分子机制。然后，我们将以实用的方式应用这些机械知识来开发新的心脏病发作治疗方法。具体来说，我们将定义microRNA在心肌I/R损伤中NF-kB和PI3K/Akt的差异调节中的作用。我们还将尝试将这些知识应用于开发新的基于miR的心脏病发作治疗方法。我们假设i) mirna在心肌i /R损伤中以不同的方式激活PI3K/Akt信号并抑制NF-kB的激活；ii) mirna携带在外泌体中，被心肌细胞有效摄取，通过不同的激活保护性和有害的细胞内信号来保护心脏。特异性目的1将阐明NF-kB和PI3K/Akt调控心肌细胞中miR-146a和miR-486表达的机制。特异性Aim 2将明确miR-146a和miR-486对心肌损伤和功能恢复的影响。特异性目标3将探讨合成的作用

DESCRIPTION (provided by applicant): Cardiovascular disease is the number one killer in the United States. Toll-like receptor (TLR)-mediated innate immune and inflammatory responses play a central role in the pathophysiologic mechanisms of myocardial ischemia/reperfusion (I/R) injury. It is well known that activation of TLR4-mediated NF-kB signaling plays a deleterious role in myocardial I/R injury, while activation of the PI3K/Akt signaling pathway protects against myocardial I/R injury. We have reported that either TLR4 deficiency or TLR2 modulation protects the myocardium from I/R injury via a PI3K/Akt dependent mechanism. We have discovered that TLR4 deficiency increases the expression of microRNA-146a (miR-146a) and miR-486 in the myocardium. We have also found that TLR2 modulation increases the amount of miR-146a and miR-486 in the myocardium, in circulating exosomes, and in exosomes secreted by bone marrow mesenchymal stem cells. We have demonstrated that increased expression of miR-146a in the myocardium decreases myocardial infarct size and improves cardiac function following myocardial I/R. MiR-146a targets IRAK1 and TRAF6 which are the important intermediates in TLR-mediated NF-kB activation, thus, attenuating I/R-induced NF-kB activation. In addition, we have found that miR-486 inhibits PTEN expression, leading to activation of PI3K/Akt signaling. Our findings suggest that either TLR4 deficiency or TLR2 modulation induce cardioprotection via up-regulation of microRNA expression that inhibits NF-kB activation and activates PI3K/Akt signaling. When considered together, our data and the literature clearly show that TLR4-mediated NF-kB activation is closely linked to disease progression in myocardial I/R injury, while activation of PI3K/Akt signaling pathways is closely associated with attenuation of myocardial injury. At present, we do not fully understand the mechanisms of differential regulation of NF-kB and PI3K/Akt in I/R injury. This application is directed at deciphering the cellular and molecular mechanisms that differentially regulate these signaling pathways in I/R induced myocardial injury. We will then apply that mechanistic knowledge in a practical fashion to develop new and novel therapies for heart attack. Specifically, we will define the role of microRNA in differential regulation of NF-kB and PI3K/Akt in myocardial I/R injury. We will also attempt to apply this knowledge for the development of new and novel miR based therapies for heart attack. We hypothesize that i) miRNAs differentially activate PI3K/Akt signaling and inhibit NF-kB activation in myocardial I/R injury and ii) miRNAs are carried in exosomes which are efficiently uptaken by cardiac myocytes, resulting in cardioprotection via differential activation f protective versus deleterious intracellular signaling. Specific Aim 1 will elucidate the mechanisms by which modulation of NF-kB and PI3K/Akt regulates the expression of miR-146a and miR-486 in cardiac myocytes. Specific Aim 2 will define the effect of miR-146a and miR-486 on myocardial injury and functional recovery. Specific Aim 3 will investigate the role of synthetic exosomes carrying miR-146a and miR-486 as therapeutics in myocardial I/R injury.