Novel Molecular Target to Prevent Maturation Failure of Arteriovenous Fistula

摘要 自体动静脉瘘(AVF)是血液透析的首选血管通路。然而，较高的 由于流出静脉血流不足而导致的成熟失败，使得内瘘不适用于血液透析。 新生内膜增生和外向重构失败是AVF成熟失败的主要原因。 是由于炎症、增殖、迁移和血管平滑肌细胞的表型变化。 (VSMCs)，以及由于基质金属蛋白酶(MMPs)增加而导致的细胞外重构。我们发现 髓系细胞-1(TREM-1)、TLR4及相关蛋白表达增加 在未成熟的房室瘘中。根据我们的新发现，中心假设是血液动力学损伤 在AVF产生过程中诱导炎症上调TREM-1和TLR4以增强新生内膜 增殖和血管重塑，拮抗TREM-1和TLR4将促进AVF的成熟。 这一假设将通过以下目的进行检验：目标1：我们的推论假设预测 应用TREM-1和TLR4拮抗剂可防止猪动静脉瘘的成熟失败。我们会 检测一种有效的抑制性TREM-1多肽在猪动静脉瘘模型中的作用。因为TREM-1可以 与TLR4协同介导AVF成熟失败的病理，强大的TLR4拮抗剂的作用将 也进行检查，以防止AVF的成熟失败。结果参数将包括新生内膜 AVF流入段和流出段的增生，AVF的血管造影，彩色多普勒超声， 光学相干断层扫描，与组织学、免疫组织化学染色分析炎症反应，表达各种 巨噬细胞和中性粒细胞的介质和渗透、VSMC的凋亡和血管重塑。目标2： 我们的推论预测，TREM-1和TLR4拮抗作用可以抑制炎症，从而 通过减少内膜增生和血管的发展来预防动静脉瘘的成熟失败 重塑主要由炎性细胞、组织蛋白酶L、IL-8和基质金属蛋白酶-12引起。这些研究将是 取AIM 1猪股动脉和股静脉的血液和分离的VSMCs。 机制研究将检验在IL-8存在的情况下抑制TREM-1和TLR4对中性粒细胞的影响， 单核细胞分化的巨噬细胞和血管平滑肌细胞及组织蛋白酶L介导的弹性蛋白和胶原降解 弹性蛋白衍生肽对单核细胞向巨噬细胞和VSMC分化的影响 扩散和迁徙。其他机制研究将包括TLR4和TREM-1之间的联系 促进基质重塑，促进中性粒细胞和巨噬细胞释放炎性细胞因子 TALK诱导VSMC表型转换和巨噬细胞极化。 这项研究的发现将证实TREM-1是否是治疗干预和扩展的新靶点 开发更好的分子来对抗TREM-1和设计I期临床试验的知识。

ABSTRACT Autologous arteriovenous fistula (AVF) is the preferred vascular access in hemodialysis. However, high rate of maturation failure due to inadequate blood flow in the outflow vein renders the fistula not useful for hemodialysis. Neointimal hyperplasia and failure of outward remodeling are the major causes of AVF maturation failure which is due to inflammation, proliferation, migration, and phenotypic changes of vascular smooth muscle cells (VSMCs), and extracellular remodeling due to increased matrix metalloproteinases (MMPs). We discovered increased expression of triggering receptor expressed on myeloid cells-1 (TREM-1), TLR4 and related proteins in the immature AV fistula. Based on our novel findings, the central hypothesis is that hemodynamic injury during AVF creation induces inflammation to upregulate TREM-1 and TLR4 to enhance neointimal hyperplasia and vascular remodeling, and antagonizing TREM-1 and TLR4 will enhance AVF maturation. This hypothesis will be tested with the following Aims: Aim 1: Our corollary hypothesis predicts that the administration of TREM-1 and TLR4 antagonists will prevent maturation failure of AVF in swine. We will examine the effect of a potent inhibitory TREM-1 peptide in the AVF model in pigs. Since TREM-1 could synergize with TLR4 to mediate the pathology of AVF maturation failure, effect of a potent TLR4 antagonist will also be examined to prevent maturation failure of AVF. The outcome parameters will include neointimal hyperplasia in the inflow and outflow segments in the AVF, angiography of the AVF, color Doppler ultrasound, optical coherence tomography, and histology, immunostaining to analyze inflammation, expression of various mediators and infiltration of macrophages and neutrophils, VSMC apoptosis, and vascular remodeling. Aim 2: Our corollary hypothesis predicts that the TREM-1 and TLR4 antagonism inhibits inflammation and thus prevents maturation failure of AVF by reducing the development of intimal hyperplasia and vascular remodeling primarily due to inflammatory cells, cathepsin L, IL-8 and MMP-12. These studies will be performed in the blood and isolated VSMCs of femoral artery and femoral vein of the pigs from Aim 1. Mechanistic studies will examine the effect of TREM-1 and TLR4 inhibition in the presence of IL-8 on neutrophils, monocyte-differentiated macrophages and VSMCs, and cathepsin L-mediated elastin and collagen degradation in VSMCs, and the effect of elastin-derived peptides on monocyte differentiation into macrophages and VSMC proliferation and migration. Additional mechanistic studies will include the link between TLR4 and TREM-1 in promoting matrix remodeling, release of inflammatory cytokines from neutrophils and macrophages in the cross- talk inducing phenotype switch in VSMCs and macrophage polarization. The findings from this study will confirm if TREM-1 is a novel target for therapeutic intervention and extend the knowledge to develop better molecules to antagonize TREM-1 and design phase I clinical trials.