Involvement of apoptosis antigen Fas in clonal deletion of human thymocytes

Apoptosis appears to play a major role in the differentiationand selection of T and B lymphocytes, but the mechanisms ofclonal deletion of T cells in thymus are not well understood.We have prepared an anti-human Fas IgM mAb with associated apoptosis-inducingactivity in Fas antigen-positive target cells including humanT cells. We analyzed the expression of apoptosis antigen Fason human thymocytes by cytofluorometry showing low, but significantamounts of Fas antigen on double-negative and double-positiveundifferentiated thymocytes. On the contrary, most of the differentiatedthymocytes (single-positive or CD3-brightest) expressed undetectablelevels of Fas antigen. About 1-2% of thymocytes expressed highamounts of Fas antigen, and these cells, which were CD3-bright,were CD4-bright and CD8-low at the stage of late double-positivelineage. Immunohlstologlcal analysis shows these Fas-brightcells on the edge of the medulla. Stimulation through the TCRcomplex was shown to induce the expression of Fas antigen onthymocytes at the late double-positive stage and prolonged stimulationthrough the TCR complex rendered the Fas-bright thymocytes sensitiveto apoptosis-induclng activity of anti-Fas. To show the involvementof the Fas system in the negative selectlon/clonal deletionof thymocytes, we organ-cultured human thymus in the presenceof the superantigen, staphylococcus enterotoxin B (SEB), andnew antagonistic anti-Fas mAb, which can inhibit the apoptosis-induclngactivity of the original anti-Fas mAb. The SEB-reactJve TCRcomplex on thymocytes was at first down-regulated by SEB, thenthe SEB-reactlve clone was deleted by apoptosis, which was inhibitedby an antagonistic anti-Fas mAb. Thus, Fas antigen is shownto be involved in the negative selection/clonal deletion ofsuperantlgen-reactive thymocytes.