Intracameral Injection of Antigen Attracts Circulating Monocytic Cells Associated With Anterior Chamber-Associated Immune Deviation to Iris, Blood and Thymus

Abstract

Purpose:: Production and/or activation of splenic regulatory T cells by the injection of foreign antigen into the Anterior Chamber is, in part, dependent on F4/80⁺ peripheral blood mononuclear cells (AC-PBMC), and on thymic NKT cells (Thyreg) that migrate to the spleen. Here we investigate (i) the phenotype of circulating AC-PBMC that induce Thyreg after intrathymic (it) or iv injection and (ii) the migration of PBMC to the iris after intracameral injection of antigen.

Methods:: PBMC enriched by centrifugation over ficoll were labeled with CFSE, and injected iv into mice that then received an intracameral injection of antigen or PBS. Twenty-four hr after GFP-transgenic mice received intracameral antigen, GFP-AC-PBMC were recovered and injected iv into naïve mice. The homing of GFP-AC-PBMC to the thymus was determined by flow cytometry. Irides were recovered 24hr after mice had received iv CFSE-labeled PBMC plus an intracameral injection of TNP-BSA,ovalbumin (OVA) or PBS. A suspension of these iris cells stained with anti-F4/80 was analyzed by FACS. The functional phenotype of the Thyreg-inducing-AC-PBMC was determined by an intrathymic (it) or iv injection of immunomagnetic bead- isolated AC-PBMC. Thymocytes recovered 24hr following these injections were assayed for Thyreg by iv transfer into TNP-BSA-immunized mice. DTH to TNP was assayed one week later.

Results:: 24hr after mice that received CFSE-labeled PBMC received an intracameral injection of antigen, CFSE-labeled cells appeared in the iris. Mice had a marked increase in F4/80⁺, Qa-1⁺ cells in the thymus, or blood 24hr after the intracameral injection of antigen. The activation of Thyreg was associated with the migration of F4/80⁺, CD11c⁺, B220^- cells and F4/80⁺,CD11c^-,B220^- cells to the thymus.

Conclusions:: Circulating PBMC able to induce the suppression of DTH appear to cycle through the anterior chamber after the intracameral injection of antigen, then recirculate and home to the thymus and spleen.

Keywords: ACAID • immunomodulation/immunoregulation • anterior chamber