Galectin-3 modulates monocyte infiltration in the injured cornea by inducing sialyl Lewis^X expression

Abstract

Purpose : Galectin-3 has been associated with many proinflammatory functions by triggering the migration of monocytes; nevertheless, the mechanism underlying this process has not been fully established. Monocyte recruitment is initiated by multiple adhesive interactions between vascular selectins and the sialyl Lewis ^X (sLe^X) glycan on circulating monocytes. Here, we investigated whether galectin-3 regulates the expression of sLe^X in activated monocytes and, therefore, their recruitment following injury.

Methods : Human monocytes were purified from the blood of healthy donors. Mouse monocytes were purified from the bone marrow of galectin-3 knockout (Gal3^−/−) and wild-type (WT) mice. Galectin-3 expression and binding in human monocytes were inhibited using galectin-3 siRNA (siGal3) and the thiodigalactoside analog TD139, respectively. Competitive migration assays were carried out in immunocompromised WT chimera by intravenous injection with fluorescently labeled WT and Gal3^-/- monocytes. The glycogene expression profile of mouse monocytes was evaluated using real-time PCR array. Parallel plate flow chamber assays were performed on vascular endothelial cells using galectin-3 deficient monocytes glycoengineered to express human FUT7.

Results : Inhibition of galectin-3 expression or binding in activated human and murine monocytes led to impaired biosynthesis of sLe^X. Conversely, the addition of human recombinant galectin-3 led to increased levels of sLe^X. Injured corneas of Gal3^-/- mice displayed lower numbers of CD45⁺ CD11b⁺ sLe^X cells compared to WT mice. Deficiency in sLe^X expression was concomitant with impaired recruitment of Gal3^-/- monocytes into wounded corneas of WT chimeric mice. Using a PCR array, we found that galectin-3 regulates the biosynthesis of sLe^X on activated monocytes by inducing the expression of FUT7. Enforced fucosylation of human galectin-3 deficient monocytes resulted in augmented rolling capability on activated endothelial cells under flow conditions in vitro.

Conclusions : Overall, these data demonstrate a novel mechanism by which galectin-3 positively regulates the recruitment of monocytes into inflamed tissues by inducing the expression of FUT7.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.