Role of TILRR in modulating inflammatory responses relevant to vaginal HIV-1 acquisition

Abstract

TILRR (Toll-like Interleukin 1 Receptor Regulator) is a transcript variant of FREM1 (Fras-related extracellular matrix 1) and the expression of TILRR is observed in a broad range of human cells including cervicovaginal epithelial cells. TILRR is reported as a co-receptor of IL-1R1 (Interleukin-1 receptor type 1) and potentiates NF-κB activation and inflammatory responses. Because TILRR expresses in cervicovaginal epithelial cells and modulates inflammatory response, it may play a role in enhancing the risk of vaginal HIV infection. To investigate the potential role of TILRR on vaginal HIV transmission, I used two cervicovaginal epithelial cell lines, HeLa and VK2/E6E7, and investigated the influence of TILRR on the expression of genes in the NF-κB inflammatory pathway, production of inflammatory cytokines/chemokines, and the migration of immune cells. I also investigated whether TILRR exists in human blood and whether the blood TILRR level is correlated with blood inflammatory mediators and susceptibility to HIV infection. I overexpressed TILRR in cell lines and examined the mRNA expression of 84 genes associated with the NF-κB signaling pathway by RT2 profiler PCR array. The production of cytokines/chemokines in cell culture supernatants was examined by an in-house developed multiplex bead array system. The influence of TILRR overexpression on the migration of immune cells was evaluated by a Transwell assay and a novel microfluidic device. Additionally, the existence of soluble TILRR protein in human blood plasma was examined by an in-house developed multiplex bead array method, affinity purification, and Western blot analysis. My study data showed that overexpression of TILRR in cervicovaginal epithelial cell lines significantly upregulated the mRNA expression of many inflammation-responsive genes in the NF-κB signal transduction pathway, increased the production of several pro-inflammatory mediators in cervicovaginal epithelial cell culture supernatants, and influenced the migration of immune cells, the CD4+ HIV target cells. TILRR also exists as a soluble protein in human blood, and high plasma TILRR level is correlated with high plasma inflammatory cytokines/chemokines and is associated with FREM1 SNP rs1552896 genotypes and HIV seroconversion. Because systemic and mucosal inflammation increases susceptibility to HIV infection, TILRR could be a novel target to reduce inflammation and HIV infection.