Lymphokine-activated killer cells in rats. IV. Developmental relationships among large agranular lymphocytes, large granular lymphocytes, and lymphokine-activated killer cells

The developmental relationships among large agranular lymphocytes (LAL) large granular lymphocytes (LGL) and the activation of these cells into lymphokine-activated killer (LAK) cells by rIL-2 was investigated. Highly enriched populations of LAL were isolated from Fischer 344 spleen cells by a combination of nylon-wool filtration (to remove B cells and macrophages), treatment with a pan T cell antibody plus complement (to remove T cells) and incubation in L-leucine methyl ester (to remove LGL). The resultant cells were highly enriched in morphologically identifiable LAL which expressed asialo GM1 and partially expressed the OX8 surface marker. The enriched LAL did not contain detectable NK cytotoxic activity, did not express pan T cell (OX19), Ia, Ig, or laminin surface markers and contained less than 0.2% LGL. Incubation of LAL in a low dose of rIL-2 (100 U/ml) induced the generation of LGL having NK activity within 24 h of culture. Longer culture periods (48 h) resulted in a continued increase in the percentage of LGL and higher levels of NK activity. However, with this low dose of rIL-2, little or no LAK activity (i.e., reactivity against NK-resistant target cells) was generated. With a high dose of rIL-2 (500 U/ml), LAL responded by first generating LGL with NK activity (within 24 h), with subsequent generation of LAK activity by 48 h. Evidence that the development of granular lymphocytes from LAL was responsible first for NK activity and then LAK activity was demonstrated by depletion of the generated granular NK or LAK effector cells by second treatments with L-leucine methyl ester. Concomitant with the induction of LGL with NK or LAK activity, rIL-2 also caused LGL to proliferate and expand four- to five-fold in 48 h. This occurred in the presence of high or low dose rIL-2. These results indicate that LAL are the precursors of LGL/NK cells, that LAL, LGL/NK cells and LAK cells appear to represent sequential developmental or activation stages and that LAL may comprise major source of LAK progenitors in lymphoid populations having few LGL or mature active NK cells.