title: Interleukin-2- and mitogen-activated NK-like killer cells from highly purified human peripheral blood T cell (CD3+ N901-) cultures.
abstract: In this study, highly purified (HP) CD3-positive N901-negative T lymphocytes could be induced to become natural killer (NK)-like in culture in the presence of recombinant interleukin-2 (rIL-2) and phytohemagglutinin (PHA). Thus, purified CD3+ N901- T cells from fresh human peripheral blood were obtained by negative selection using an indirect panning technique. To ensure that T lymphocyte fractions were completely devoid of any detectable NK cells, two additional purification procedures were employed: incubation of post-pan T cells with the NK-cytotoxic lysomotropic agent L-leucinemethylester, and complement-mediated lysis using the NK cell specific NKH1a monoclonal antibody. Purity of CD3+ N901- cells could be confirmed by surface marker analysis, whereby two NK-associated antigens, N901 and H-25, were undetectable, while 94 +/- 1% of cells expressed the CD3 (Leu-4) antigen. On functional analysis, fresh HP CD3+ N901- cells exhibited no cytotoxic activity against the standard NK target K562. When HP NK-depleted T lymphocytes were cultured for 7 days in the presence of rIL-2 (100 U/ml), neither surface antigen expression nor cytotoxic activity against K562 changed significantly. However, significant cytotoxicity against K562 [18 +/- 5% specific lysis at 25:1 effector:target (E/T) ratio] could be induced when HP CD3+ N901- cells were grown for 7 days in the presence of rIL-2 and PHA (0.5% v/v). Concomitantly, antigens N901 and H-25 were found to be coexpressed on a minor proportion (22 +/- 16 and 22 +/- 6%, respectively) of CD3+ (88 +/- 2% on day 7) cells. Four-week long-term culture of HP NK-depleted T cells in the presence of rIL-2 and PHA yielded a continuous increase in cytotoxicity against K562 cells (0 up to 46% specific lysis at 25:1 E/T ratio). Of particular interest was the emergence of cytotoxicity against the NK-resistant Daudi cell target (15 +/- 8% specific lysis at 25:1 E/T ratio on day 21). Expression of antigens N901 and H-25 as well as CD3 remained essentially unchanged in long-term culture. In sorting experiments, the H-25+ cell fraction was significantly enriched for cytotoxicity against K562, when compared to both H-25- and unseparated cell fractions. In summary, our results suggest that a proportion of HP CD3+ N901- T lymphocytes may give rise to cells that exhibit NK-like functional and phenotypic properties.
mesh_terms: D000945:Antigens, Differentiation, T-Lymphocyte; D000954:Antigens, Surface / Q000032:analysis; D002454:Cell Differentiation; D002469:Cell Separation; D002478:Cells, Cultured; D003602:Cytotoxicity, Immunologic; D006801:Humans; D007111:Immunity, Cellular; D007376:Interleukin-2 / Q000502:physiology*; D007694:Killer Cells, Natural / Q000276:immunology*; D010835:Phytohemagglutinins / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000494:pharmacology; D013601:T-Lymphocytes / Q000145:classification* / Q000276:immunology
chemical_list: D000945:Antigens, Differentiation, T-Lymphocyte; D000954:Antigens, Surface; D007376:Interleukin-2; D010835:Phytohemagglutinins; D011994:Recombinant Proteins