Monocyte chemoattractant Proteins and Vascular Injury

项目概要/摘要 2017年，约有75万美国人被诊断患有终末期肾病（ESKD），每年增长3%。87%会 接受血液透析（HD）肾脏替代治疗，首选通过动静脉瘘建立血管通路 （AVF）.由于静脉狭窄（VS）和新生内膜增生（VNH），AVF的年通畅率约为62%， 血流量减少和次优HD，采用经皮腔内血管成形术（PTA）治疗，费用> 30亿美元/年。 狭窄动静脉内瘘PTA术后损伤血管壁单核巨噬细胞聚集 (AVF)MCP-1表达增加导致VS/VNH。Bindarit是MCP-1的口服选择性抑制剂， 2，和-3，我们将其封装在聚乳酸-羟基乙酸（PLGA）纳米颗粒中，嵌入热敏 Pluronic F127水凝胶（BN NP）用于外膜周输送至流出静脉，以在本提案中进行试验。扫描 用电子显微镜和动态光散射对BN纳米粒子和对照纳米粒子（NP）进行了表征 C）。采用液相色谱-串联质谱法（LC-MS/MS）研究药物释放动力学。 在AVF狭窄的小鼠模型中，PTA后立即将BN NP或NP C施用到外膜周围 流出静脉在3天和21天后处死动物，进行基因表达、组织形态计量学和免疫组化。 免疫组织化学分析。每周进行多普勒超声检查。在大小上没有区别， BN NP的储能模量与对照相比。药代动力学分析表明，药物从 与对照组相比，BN NP。BN NP处理的血管中MCP-1、MCP-2和MCP-3基因和蛋白表达降低 水平，降低的CCR 2，增加的FABP 4/IL 8，巨噬细胞/单核细胞丰度，促炎细胞因子， CD 4（+）细胞减少，内皮炎症和静脉纤维化减少，导致血管重塑 和改善的开放性，降低VS/VNH。PTA后21天，BN NP的峰值流速增加 组在PTA后，将BN NP外膜周围给药至流出静脉导致VS/VNH降低。 中心假设PTA后将Bindarit NP外膜周围输送至AVF流出静脉可降低Mcp-1、-2、-3， FABP 4/IL 8增加的CCR 2表达导致免疫、巨噬细胞浸润减少， 平滑肌细胞、纤维化和VS/VNH。我们提出三个具体目标： 目的1：确定Bindarit NPs如何减少MCP-1、MCP-2和MCP-3，导致单核细胞-巨噬细胞减少 分化、迁移、增殖、内皮细胞对白细胞的化学吸引以及炎症 细胞因子表达 目的2：评估Bindarit NP对CCR 2和FABP 4/IL 8在狭窄AVF PTA后降低VS/VNH的作用。 目的3：确定Bindarit NP在CKD猪PTA后降低VS/VNH的安全性和有效性。

PROJECT SUMMARY/ABSTRACT In 2017, ~750K Americans were diagnosed with end-stage kidney disease (ESKD), which rises 3% yearly. 87% will have renal replacement therapy with hemodialysis (HD), with preferred vascular access through arteriovenous fistula (AVF). AVFs have ~62% year patency due to venous stenosis (VS) and neointimal hyperplasia (VNH) causing reduced blood flow and suboptimal HD which is treated with percutaneous transluminal angioplasty (PTA) at >$3B/yr. Monocyte and macrophage recruitment occurs to the injured vessel wall after PTA of stenotic arteriovenous fistulas (AVF) through increased expression of MCP-1 leading to VS/VNH. Bindarit is an oral selective inhibitor of MCP-1, - 2, and -3 and we encapsulated it in polylactic-co-glycolic acid (PLGA) nanoparticles embedded in a thermosensitive Pluronic F127 hydrogel (BN NP) for periadventitial delivery to the outflow vein to test in this proposal. Scanning electron microscope and dynamic light scattering were used to characterize the BN NP and control nanoparticles (NP C). Liquid chromatography with tandem mass spectrometry (LC-MS/MS) was used to study drug release kinetics. Immediately after PTA, in a murine model of AVF stenosis, BN NP or NP C was administrated to the periadventitia of outflow veins. Animals were sacrificed 3 and 21 days later for gene expression, histomorphometric, and immunohistochemical analyses. Doppler ultrasound was performed weekly. There was no difference in the size and storage modulus of BN NP compared controls. Pharmacokinetic analysis demonstrated increased drug release from BN NP when compared to controls. BN NP treated vessels had reduced MCP-1, MCP-2 and MCP-3 gene and protein levels, reduced CCR2, increased FABP4/IL8, macrophage/monocyte abundance, proinflammatory cytokines, reduced CD4 (+) cells, reduced endothelial inflammation, and venous fibrosis resulting in positive vascular remodeling and improved patency with reduced VS/VNH. There was increased peak velocity 21 days after PTA in the BN NP group. Periadventitial administration of BN NP to the outflow vein after PTA results in decreased VS/VNH. Central Hypothesis. Periadventitial delivery of Bindarit NPs to AVF outflow vein after PTA decreases Mcp-1, -2, -3, CCR2 expression with increased FABP4/IL8 leading to less immune, macrophage cell infiltration with reduction in smooth muscle cells, fibrosis, and VS/VNH. We propose three specific aims: Aim 1: Determine how Bindarit NPs reduce MCP-1, -2, and -3 leading to decreased monocyte to macrophage differentiation, migration, proliferation, leukocyte chemoattraction by endothelial cells and inflammatory cytokine expression. Aim 2: Assess the role(s) of Bindarit NPs on CCR2 and FABP4/IL8 on reducing VS/VNH after PTA of stenotic AVFs. Aim 3: Ascertain the safety and efficacy of Bindarit NPs on reducing VS/VNH after PTA in pigs with CKD.