Daily Moderate Ethanol Ingestion Attenuates Postischemic Microvascular Dysfunctio

描述(由申请人提供)：经常摄入低到中等水平的乙醇(Etoh)可以保护器官和微血管免受缺血和再灌注(I/R)的有害影响。最近，我们发现，先天摄入乙醇通过一种新的机制引起抗炎表型的发展(缺血后炎症介质的形成减少，黏附分子的表达、氧化剂的产生以及毛细血管后小静脉中白细胞的卷曲、黏附和迁移)，在存在心血管疾病危险因素的情况下仍然有效。令人惊讶的是，我们的工作发现，在乙醇暴露期间，促炎性降钙素基因相关肽和肿瘤坏死因子-�介导的中性粒细胞激活在启动这种适应性转化中发挥了重要作用，当组织暴露于无水乙醇24小时后，这种适应性转化在毛细血管后小静脉中变得明显。在目前的方案中，我们试图建立在这些基本观察的基础上，以评估总体假设，即每天适度的乙醇诱导感觉神经元释放依赖于TRPV1的降钙素基因相关肽，进而激活CD_4+T淋巴细胞表达肿瘤坏死因子-�。肿瘤坏死因子依赖的中性粒细胞蛋白水解酶介导的间质信号生成参与内皮整合素�v�3诱导HO-1表达/活性增加，从而限制缺血后微血管功能障碍。为了解决这一假设，我们建议确定：(1)乙醇诱导的、降钙素基因相关肽依赖的T淋巴细胞激活，随后产生肿瘤坏死因子，激活组织驻留的中性粒细胞，蛋白水解性地产生信号，触发抗炎表型的发展，以响应先天乙醇；(2)中性粒细胞蛋白酶启动，�v�3整合素依赖的HO-1表达和活性增加，作为I/R期间抗炎表型的下游调节因子。活体显微镜方法将用于量化缺血后白细胞/内皮细胞的相互作用。此外，还将研究乙醇升高血浆和组织中CGRP和肿瘤坏死因子水平，诱导T细胞和中性粒细胞活化，增加HO-1的表达和活性，以防止I/R诱导的内皮黏附分子和炎性介质表达的作用。意义：这项工作将确定降钙素基因相关肽激活的T细胞、肿瘤坏死因子激活的中性粒细胞的蛋白分解活性产生的信号和�v�3依赖的HO-1表达/活性之间的新联系，从而通过预先摄入乙醇获得对I/R的耐受。完成这些研究将为在相关患者群体中发展转化疗法提供机制基础。

DESCRIPTION (provided by applicant): Regular consumption of ethanol (EtOH) at low to moderate levels protects organs and microvasculature from the deleterious effects of ischemia and reperfusion (I/R). Recently, we discovered that antecedent ethanol ingestion provokes the development of an anti-inflammatory phenotype (reduced postischemic formation of inflammatory mediators and markedly attenuated adhesion molecule expression, oxidant production, and leukocyte rolling, adhesion, and emigration in postcapillary venules) via a novel mechanism that remains effective in the presence of co-existing risk factors for cardiovascular disease. Surprisingly, our work uncovered important roles for proinflammatory calcitonin gene-related peptide (CGRP) and tumor necrosis factor-� (TNF) mediated neutrophil activation during the period of ethanol exposure in initiating this adaptive transformation that becomes apparent in postcapillary venules when tissues are exposed to I/R 24 hrs after EtOH. In the current proposal, we seek to build on these fundamental observations to evaluate the overall hypothesis that daily moderate EtOH induces TRPV1-dependent CGRP release from sensory neurons, which in turn activates CD4+ T lymphocytes to express tumor necrosis factor-� (TNF). TNF-dependent, neutrophil proteasemediated generation of signals in the interstitium engage endothelial integrin �v�3 to induce increased HO-1 expression/activity to limit postischemic microvascular dysfunction. To address this postulate, we propose to determine the roles of: (1) EtOH-induced, CGRP-dependent activation of T lymphocytes, which subsequently produce TNF to activate tissue resident neutrophils to proteolytically generate signals that trigger the development of an anti-inflammatory phenotype in response to antecedent ethanol; and (2) neutrophil protease-initiated, �v�3 integrin-dependent increased HO-1 expression and activity as downstream mediators of the anti-inflammatory phenotype seen during I/R. Intravital microscopic approaches will be used to quantify postischemic leukocyte/endothelial cell interactions. The effects of EtOH to elevate plasma and tissue CGRP and TNF levels, induce T cell and neutrophil activation, and increase HO-1 expression and activity to prevent I/R-induced endothelial adhesion molecule and inflammatory mediator expression will also be investigated. Significance: This work will identify new links between CGRP-activated T cells, signals generated by the proteolytic activity of TNF-activated neutrophils, and �v�3-dependent HO-1 expression/activity in the acquisition of tolerance to I/R by antecedent ethanol ingestion. Completing these studies will provide the mechanistic basis for development of translational therapeutics in relevant patient populations.