Differential Contribution of Fas- and Perforin-Mediated Mechanisms to the Cell-Mediated Cytotoxic Activity of Naive and In Vivo-Primed Intestinal Intraepithelial Lymphocytes

Abstract

Intestinal intraepithelial lymphocytes (IELs) are known to exert strong constitutive cytotoxic activity. In the present study we compared the Ag-specific cytotoxic activity and the effector mechanisms involved in non-Ag-primed, naive and in in vivo-primed IELs and splenic CD8 T cells. Ex vivo isolated naive CD8αα TCRαβ IELs, CD8αβ IELs, and splenocytes from lymphocytic choriomeningitis virus (LCMV)-specific TCR transgenic mice exert Ag-specific cytotoxic activity in a long-term, but not in a short-term, cytotoxicity assay. This cytotoxic activity is mainly Fas-Fas ligand mediated and is significantly reduced in the presence of 20 μg/ml Fas-Fcγ1 fusion protein. Both CD8αβ IELs and CD8αβ splenocytes isolated from LCMV-infected C57BL/6 mice exert potent perforin-dependent cell-mediated cytotoxicity. CD8αα TCRαβ IELs from LCMV-infected animals, however, show only minimal Ag-specific cytotoxicity. The potent cytotoxic activity of in vivo activated CD8αβ IELs is not affected by the addition of Fas-Fcγ1. Nevertheless CD8αβ IELs from LCMV-infected perforin-deficient mice exert Ag-specific cytotoxicity in a short-term cytotoxicity assay, and this cytotoxicity is almost completely blocked by the addition of Fas-Fcγ1. These results demonstrate that naive CD8αβ IELs exert Ag-specific, Fas-Fas ligand-mediated, constitutive cytotoxic activity in a long-term cytotoxicity assay, whereas primed CD8αβ IELs primarily use the perforin-dependent exocytosis pathway to exert their potent cytotoxic activity. Furthermore, these results clearly illustrate the requirement for Ag-specific determination of IEL-mediated cytotoxicity, because the elevated, but variable, frequencies of memory-type T cells in this compartment may lead to ambiguous results when polyclonal activation or redirected assays are used.