Isolation of T-cell clones cytotoxic agcirst nemotopoietic progenitor cells from a patienl with aplastic aremia

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Although immune-mediated suppression of hematopoiesis has been considered to be the most important mechanism for the development of idiopathic aplastic anemia, direct evidence for such immunologic attack has not been obtained. To identify T lymphocytes that may contribute to the development of bone marrow failure, we established T-cell clones from the bone marrow of an untrasnfused patient with aplastic anemia, who owned HLA-DRB1^<**>1501 and DRB1^<**>0405. Among CD4^+ T-cell clones isolated, a Vbeta21 T-cell clone, NT4.2, not only secreted interferon-gamma in response to irradiated autologous CD34^+ cells or EBV-transformed lymphoblastoid cell line (LCL) but also lysed the autologous LCL and allogneic LCL owning HLA-DRB1^<**>0405. Single strand conformation polymorphism analysis of amplified Vbeta21 cDNA products revealed that NT4.2 was the most dominant clone among Vbeta21^+ T cells in the bone marrow of the patient. The cytotoxicity against LCL cells by NT4.2 was blocked by the addition of anti HLA-DR monoclonal antibodies (mAb) or anti CD3 mAb into the culture. NT4.2 cells also lysed autologous CD34^+ cells as well as allogeneic CD34^+ cells of a normal individual owining HLA-DRB1^<**>0405 when the CD34^+ cells were cultured in the presence of various colony-stimulaing factors for one week prior to the ^<51>Cr release assay. The NT4.2 cells strongly inhibited colony formation by autologous hematopoietic progenitor cells or by the allogeneic progenitor cells with HLA-DRB1^<**>0405 when they were incubated with the cultured CD34^+ cells prior to plating with the methylcellulose medium. These findings suggest that this T-cell clone may contribute to the suppression of hematopoiesis in aplastic anemia by recognizing a self-peptide on hematopoietic progenitor cells bound to HLA-DR4.