Dendritic epidermal gamma/delta T cells (DETC) activated in vivo proliferate in vitro in response to Mycobacterium leprae antigens.

Abstract

gamma/delta T-cell receptor (TCR)+ dendritic epidermal T cells (DETC) are part of a primitive defense system in the skin; they are capable of responding only to a limited number of antigens. The aim of the present study was to test whether DETC can proliferate in vitro in response to antigens of Mycobacterium leprae. DETC were obtained from CBA mouse ear skin by trypsinization and Histopaque gradient centrifugation. The resulting epidermal cell suspension contained up to 20% DETC, as analyzed by the fluorescence activated cell sorter (FACS) after staining with anti-Thy-1 or anti-gamma/delta TCR monoclonal antibodies (mAbs). The freshly isolated cells, or DETC cultured up to 4 weeks with interleukin-2 (IL-2), were exposed in vitro for up to 6 days to varying doses of the following M. leprae antigens: (1) integral (live) M. leprae bacilli; (2) Dharmendra antigen; and (3) PGL-1 (phenolic glycolipid of M. leprae). The DETC response was assessed by tritiated thymidine (3H-TdR) incorporation. The freshly isolated DETC, or DETC cultured up to 4 weeks with IL-2, did not respond significantly to any of the M. leprae antigens, although at the same time they were able to respond vigorously to concanavalin A (Con A), as positive control. If, however, DETC were isolated from skin, painted 7 days before with croton oil (10 microL/cm2 to cause irritant dermatitis, they were able to respond to all M. leprae antigens by a 3-4-fold incrase in the 3H-TdR uptake. The most effective stimulator was a 1 : 1 mixture of Dharmendra and PGL-1 (0. 01 microg/mL), which was as effective as 10-fold higher doses of either antigen alone. Cell counts confirmed that increased DNA synthesis was associated with cell proliferation. Experiments employing alpha/beta-TCR CBA murine spleen cells and epidermal cell suspension treated with anti-gamma/delta or antialpha/beta mAbs + C' proved that only the gamma/delta DETC were the responder cells to M. leprae antigens. The results suggest that activation of DETC in vivo may make them responsive to M. leprae antigens. A significant increase in the number of class II major histocompatibility complex (MHC) positive, nondendritic cells was observed in the croton oil-treated epidermis. We hypothesize that croson oil-induced upregulation of class II MHC expression, which endows epidermal cells with antigen-presenting capabilities, might be an important factor in vivo in delivering an immunogenic signal to resident DETC in the skin.