Generation of natural killer cells from long-term cultures of mouse bone marrow

The features of a mouse long-term bone marrow culture (LTBMC) system that produces natural killer (NK) cell activity are described. Over a 4-week period in the NK-LTBMC, cellularity dropped from approximately 2.5 x 10(7) to 8 x 10(5) cells/25-cm2 flask. About 3 x 10(5) of these cells were loosely adherent. The cultures at this time contained about one-third the spleen colony forming units, one-tenth the granulocyte macrophage colony forming units and about one-third the transplantable NK progenitor activity of fresh bone marrow (BM), and no detectable NK cell activity. In the 4-week NK-LTBMC, IL-2-responsive precursor cells appeared to be selectively maintained and gave an 8-fold higher activity after culture with human recombinant IL-2 (rIL-2) than did fresh BM. The addition of 50-5,000 IU/ml of rIL-2 resulted, after a minimal 3-day lag, in progressively increased cellularity for as long as 13 days. The percentage and staining intensity of NK-1.1+ cells increased with time after addition of rIL-2. CD3 epsilon + cells were occasionally seen and B220+ cells were present in low numbers at day 7 and slowly increased through day 13. The stroma was necessary for IL-2-dependent development of NK activity.