ARNT: A novel regulator of cardiac vascular endothelial barrier function in heart failure

摘要 心脏病发作后，恢复心脏的血流量(即再灌注)对于限制心脏的量是至关重要的。 指因缺血损伤而受损的心肌组织；然而，再灌流本身会导致其自身类型的 心肌损伤(缺血-再灌注[IR]损伤)，没有治疗方法，可能导致 慢性心力衰竭(HF)。因此，对心脏疾病诱导的分子机制的了解 迫切需要再灌注来确定预防IR损伤的新策略。心源性水肿，这是 主要由心脏血管通透性增加引起，是IR损伤的主要因素之一， 我的初步研究结果表明，血管通透性受到以下表达的限制 芳香烃受体核转运体(ARNT)，又称缺氧诱导因子1β，在 内皮细胞(ECs)。我最近培育了一系列携带可诱导的EC特异性ARNT的小鼠 基因敲除突变(ecARNT-/-)，对这些小鼠的实验结果表明， ECARNT在再灌注期间的表达会导致心脏出血，而当ECARNT-/-突变是 在成年动物中诱导，心脏水肿发展并逐渐发展为心衰。因此，ecARNT似乎 是内皮屏障功能的重要调节因子，但ecARNT在心脏病中的作用目前还不是 正在接受调查。本提案中描述的实验通过研究 血管紧张素转换酶调节血管内皮细胞屏障功能的机制及其对损伤进展的影响 氢氟酸。此外，我们的初步研究表明，ARNT还调节基质的表达 金属蛋白酶3(MMP3)，它裂解形成相邻内皮细胞连接的蛋白质；因此，我们将 对培养的内皮细胞进行体外实验，以确定抑制MMP3是否可以逆转受损的内皮细胞 与ecARNT缺失相关的内皮屏障功能，如果是这样的话，阐明支持 这一观察结果。我们还将在ecARNT-/-小鼠身上使用MMP3抑制剂，并产生一系列 EcARNT/MMP3双基因敲除小鼠以确定抑制MMP3是否可以恢复血管完整性和 限制IR诱导的心衰的进展。完成后，我们预计我们的研究结果将有 确定新的治疗目标和策略，通过减少心脏水肿来改善患者的预后 在心脏病发作恢复的早期阶段，预防(或延缓)心脏的进展 疾病。

Abstract After a heart attack, restoring blood flow to the heart (i.e., reperfusion) is critically necessary to limit the amount of myocardial tissue that is damaged by ischemic injury; however, reperfusion itself leads to its own type of myocardial damage (ischemia-reperfusion [IR] injury) for which there is no treatment and that can lead to chronic heart failure (HF). Thus, knowledge of the molecular mechanisms that are induced by cardiac reperfusion is urgently needed to identify novel strategies for preventing IR injury. Cardiac edema, which is mainly caused by an increase in cardiac vascular permeability, is one of the primary contributors to IR injury, and the results from my preliminary studies indicate that vascular permeability is limited by the expression of Aryl Hydrocarbon Receptor Nuclear Translocator (ARNT), also known as hypoxia-inducible factor 1β, in endothelial cells (ECs). I have recently generated a line of mice carrying an inducible, EC-specific ARNT knockout mutation (ecARNT–/–), and the results from experiments with these mice indicate that the loss of ecARNT expression during reperfusion leads to cardiac hemorrhage, and that when the ecARNT–/– mutation is induced in adult animals, cardiac edema develops and gradually progresses to HF. Thus, ecARNT appears to be a crucial regulator of endothelial barrier function, but the role of ecARNT in heart disease is not currently being investigated. The experiments described in this proposal address this unmet need by studying the mechanisms of ARNT-regulated endothelial barrier function after IR injury and their impact on the progression of HF. Furthermore, our preliminary studies suggest that ARNT also regulates the expression of matrix metalloproteinase 3 (MMP3), which cleaves proteins that form junctions between adjacent ECs; thus, we will conduct in-vitro experiments with cultured ECs to determine whether MMP3 inhibition can reverse the impaired endothelial barrier function associated with ecARNT deletion and if so, elucidate the mechanisms that support this observation. We will also use the MMP3 inhibitors in ecARNT–/– mice and generate a line of ecARNT/MMP3 double-knockout mice to determine whether MMP3 inhibition can restore vascular integrity and limit the progression of IR-induced HF. Upon completion, we expect the results from our studies to have identified new therapeutic targets and strategies for improving patient outcomes by reducing cardiac edema during the early stages of recovery from a heart attack and preventing (or delaying) the progression of heart disease.