Misra Sanjay, Kilari Sreenivasulu, Yang Binxia, Sharma Amit, Wu Chih-Cheng, Vazquez-Padron Roberto I, Broadwater John
Vascular and Interventional Radiology Translational Laboratory, Department of Radiology, Mayo Clinic, Rochester, Minnesota.
Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, Minnesota.
J Am Soc Nephrol. 2021 Jul;32(7):1630-1648. doi: 10.1681/ASN.2020101458. Epub 2021 Apr 23.
Fractalkine receptor 1 (CX3CR1) mediates macrophage infiltration and accumulation, causing venous neointimal hyperplasia (VNH)/venous stenosis (VS) in arteriovenous fistula (AVF). The effect of blocking CX3CR1 using an anti-human variable VHH molecule (hCX3CR1 VHH, BI 655088) on VNH/VS was determined using a humanized mouse in which the human () gene was knocked in (KI).
Whole-transcriptomic RNA sequencing with bioinformatics analysis was used on human stenotic AVF samples, C57BL/6J, KI mice with AVF and CKD, and in experiments to identify the pathways involved in preventing VNH/VS formation after hCX3CR1 VHH administration.
Accumulation of CX3CR1 and CD68 was significantly increased in stenotic human AVFs. In C57BL/6J mice with AVF, there was increased , and gene expression, and increased immunostaining of CX3CR1 and CD68. In hCX3CR1-KI mice treated with hCX3CR1 VHH molecule (KI-A), compared with vehicle controls (KI-V), there was increased lumen vessel area and patency, and decreased neointima in the AVF outflow veins. RNA-seq analysis identified TNF- and NF-B as potential targets of CX3CR1 inhibition. In KI-A-treated vessels compared with KI-V, there was decreased gene expression of , , and ; with reduction of , NF-B, and ; decreased M1, Ly6C, smooth muscle cells, fibroblast-activated protein, fibronectin, and proliferation; and increased TUNEL and M2 staining. In cell culture, monocytes stimulated with PMA and treated with hCX3CR1 VHH had decreased , , proliferation, and migration.
CX3CR1 blockade reduces VNH/VS formation by decreasing proinflammatory cues.
趋化因子受体1（CX3CR1）介导巨噬细胞浸润和聚集，导致动静脉内瘘（AVF）出现静脉内膜增生（VNH）/静脉狭窄（VS）。使用抗人可变VHH分子（hCX3CR1 VHH，BI 655088）阻断CX3CR1对VNH/VS的影响，通过敲入（KI）人（）基因的人源化小鼠来确定。
对人狭窄AVF样本、C57BL/6J、患有AVF和慢性肾脏病的KI小鼠进行全转录组RNA测序及生物信息学分析，并在实验中确定hCX3CR1 VHH给药后预防VNH/VS形成所涉及的途径。
狭窄的人AVF中CX3CR1和CD68的积累显著增加。在患有AVF的C57BL/6J小鼠中， 、 和基因表达增加，CX3CR1和CD68的免疫染色增加。在用hCX3CR1 VHH分子处理的hCX3CR1-KI小鼠（KI-A）中，与载体对照（KI-V）相比，AVF流出静脉的管腔血管面积增加、通畅性增加，内膜减少。RNA测序分析确定肿瘤坏死因子（TNF）和核因子κB（NF-κB）是CX3CR1抑制的潜在靶点。与KI-V相比，在KI-A处理的血管中， 、 和 的基因表达降低； 、NF-κB和 减少；M1、Ly6C、平滑肌细胞、成纤维细胞活化蛋白、纤连蛋白和增殖减少；TUNEL和M2染色增加。在细胞培养中，用佛波酯（PMA）刺激并用hCX3CR1 VHH处理的单核细胞， 、 、增殖和迁移减少。
CX3CR1阻断通过减少促炎信号来减少VNH/VS形成。
