Estimation of effector cytotoxic lymphocytes against male H-Y antigens induced by two-step stimulations as CD8⁺NK1.1⁻TCR^int and CD8⁺NK1.1⁺TCR^int cells

Abstract

C57BL/6 (B6) female mice were injected with 10⁷ splenocytes of B6 male mice to immunize male H-Y antigens. This stimulation induced a slight increase in the proportions of CD3^intIL-2Rβ⁺ cells, CD3^intNK1.1⁺ cells (i.e., NKT cells), and CD4⁺T cells in the liver and spleen. However, cytotoxic activity against male H-Y antigens was not detected in hepatic and splenic lymphocytes. When these in vivo primed hepatic or splenic lymphocytes were cultured in vitro with irradiated splenocytes of male mice (i.e., mixed lymphocyte reaction, MLR), potent cytotoxicity against male H-Y antigens was induced. Unprimed (in vivo) hepatic and splenic lymphocytes did not acquire such activity even by in vitro stimulation. If the primed mice were in vivo stimulated again with male lymphocytes, such cytotoxicity was not obtained. In other words, in vivo priming and in vitro culture stimulation were both required for the induction of cytotoxicity. Phenotypic characterization and cell-depletion study using normal B6 mice and NKT-deficient mice revealed that effector cells were both CD8⁺NK1.1⁻TCR^int and CD8⁺NK1.1⁺TCR^int cells. These results suggest that recognition of H-Y antigens and effector function against H-Y antigens may be events of innate immunity similar to the case of other self-related antigens.