Anti human CX3CR1 VHH molecule attenuates venous neointimal hyperplasia of arteriovenous fistula in mouse model

Sanjay Misra, Sreenivasulu Kilari, Binxia Yang, Amit Sharma, Chih Cheng Wu, Roberto I. Vazquez-Padron, John Broadwater

Research output: Contribution to journalArticlepeer-review


Background 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 CX3CR1 (hCX3CR1) gene was knocked in (KI). Methods Whole-transcriptomic RNA sequencing with bioinformatics analysis was used on human stenotic AVF samples, C57BL/6J, hCX3CR1 KI mice with AVF and CKD, and in in vitro experiments to identify the pathways involved in preventing VNH/VS formation after hCX3CR1 VHH administration. Results Accumulation of CX3CR1 and CD68 was significantly increased in stenotic human AVFs. In C57BL/6J mice with AVF, there was increased Cx3cr1, Cx3cl1, Cd68, and Tnf-a 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-a and NF-kB as potential targets of CX3CR1 inhibition. In KI-A-treated vessels compared with KI-V, there was decreased gene expression of Tnf-a, Mcp-1, and Il-1b; with reduction of Cx3cl1, NF-kB, and Cd68; 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 TNF-a, CD68, proliferation, and migration. Conclusions CX3CR1 blockade reduces VNH/VS formation by decreasing proinflammatory cues.

Original languageEnglish (US)
Pages (from-to)1630-1648
Number of pages19
JournalJournal of the American Society of Nephrology
Issue number7
StatePublished - Jul 2021

ASJC Scopus subject areas

  • Nephrology


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