Molecular regulation of vein graft adapation

描述(申请人提供)：静脉移植物成功地长期适应动脉环境而不过度增厚内膜的机制仍不完全清楚。最近，一些EPhin-Eph途径的成员被描述为发育特定的血管识别的关键决定因素，其中EPhin-B2配体是动脉的胚胎决定因素，Eph-B4受体是静脉的胚胎决定因素。我们最近在人类和大鼠身上都发现，在静脉移植适应过程中，Eph-B4的表达减少和静脉认同性的丧失与内膜增厚有关，而血管内皮生长因子-A是内膜增厚的上游抑制因子。此外，我们还发现，在人和大鼠静脉移植适应过程中，血管保护蛋白Nogo-B的数量都有所增加。然而，目前尚不能确定成人血管特异性基因是否存在可塑性，EPhin-Eph通路是否在静脉移植物适应动脉环境中发挥机制作用，或者Nogo-B是否与Eph-B4信号通路相关联。我们提供了令人兴奋的新数据：1)在静脉移植适应的小鼠模型中，Eph-B4基因的表达减少和免疫反应蛋白与内膜增厚相关；2)我们可以在体内操纵Eph-B4信号来直接检测Eph-B4在小鼠静脉移植适应中的作用；3)Nogo-B抑制静脉移植适应过程中的内膜增厚；以及4)针对血管内皮生长因子-A的siRNA处理的移植静脉中，Nogo-B蛋白表达增加，从而将Nogo-B途径与VEGF-A-Eph-B4途径联系起来。我们假设静脉规范基因的表达减少而动脉规范基因的表达不同时会导致过度的内膜增厚，最终导致静脉移植失败。我们将通过以下具体目标来验证我们的假设：目的1.确定Eph-B4信号是否在静脉移植适应过程中介导内膜增厚。目的II.确定血管内皮生长因子-A抑制血管内膜增厚是否通过调节血管特性和Eph-B4的表达来实现。目的III.确定Nogo-B在静脉移植适应过程中的血管保护作用是否通过调节Eph-B4功能来实现。这些研究是创新的，因为之前还没有在小鼠身上进行静脉移植物身份的分析，而且之前还没有在成人中证明Eph-B4途径的可塑性。在这些研究完成后，我们希望确定Eph-B4在静脉移植适应中的作用，并确定通过改变血管特性来限制静脉移植适应期间内膜增厚的策略是否将是一种新的人类临床试验方法。公共卫生相关性：大量经手术放置的冠状动脉和外周静脉移植物由于过度的内膜增厚而继续失败，从而导致患者的发病率和死亡率。长期心肌血运重建和保肢效果的改善仍取决于对移植静脉适应动脉环境时内膜过度增厚的机制的了解。这一建议采用了一种新的方法，即操纵血管身份，来控制内膜增厚。

DESCRIPTION (provided by applicant): The mechanisms of successful long-term vein graft adaptation to the arterial environment without excessive intimal thickening are still not completely understood. Several members of the Ephrin-Eph pathway have recently been described as developmentally specified critical determinants of vessel identity, with Ephrin- B2 ligand an embryonic determinant of arteries and Eph-B4 receptor an embryonic determinant of veins. We have recently shown in both humans and rats that diminished Eph-B4 expression and loss of venous identity is associated with intimal thickening during vein graft adaptation, and that VEGF-A is an upstream inhibitor of intimal thickening. In addition, we showed increased amounts of the vasculoprotective protein Nogo-B during both human and rat vein graft adaptation. However, it is currently not established whether plasticity exists in vessel specification genes in adults, whether the Ephrin-Eph pathway plays a mechanistic role during vein graft adaptation to the arterial environment, or whether Nogo-B is linked to Eph-B4 signaling pathways. We present our exciting new data that: 1) diminished Eph-B4 gene expression and immunoreactive protein is associated with intimal thickening in the mouse model of vein graft adaptation; 2) we can manipulate Eph-B4 signaling in vivo to directly test the role of Eph-B4 in vein graft adaptation in mice; 3) Nogo-B inhibits intimal thickening during vein graft adaptation; and 4) Nogo-B protein is increased in vein grafts treated with siRNA directed against VEGF-A, linking the Nogo-B pathway with the VEGF-A-Eph-B4 pathway. We hypothesize that decreased expression of venous specification genes without concomitant expression of arterial specification genes leads to excessive intimal thickening, and ultimately vein graft failure. We will test our hypothesis with the following specific aims: Aim I. To determine whether Eph-B4 signaling mediates intimal thickening during vein graft adaptation. Aim II. To determine whether VEGF-A inhibition of intimal thickening is mediated by regulation of vessel identity and Eph-B4 expression. Aim III. To determine whether the vasculoprotective effects of Nogo-B during vein graft adaptation are mediated by regulation of Eph-B4 function. The studies are innovative in that analyses of vein graft identity have not been previously performed in mice, and plasticity of the Eph-B4 pathway has not been previously demonstrated in adults. At the completion of these studies, we expect to identify a role for Eph-B4 in vein graft adaptation and to determine whether strategies to limit intimal thickening during vein graft adaptation by altering vessel identity will be a novel approach for a human clinical trial. PUBLIC HEALTH RELEVANCE: Significant numbers of surgically placed coronary and peripheral vein grafts continue to fail due to excessive intimal thickening with resultant patient morbidity and mortality. Improving results of long-term myocardial revascularization and limb salvage remain critically dependent on understanding the mechanisms of excessive intimal thickening during vein graft adaptation to the arterial environment. This proposal takes a novel approach, manipulation of vessel identity, to control intimal thickening.