Graft versus host disease, an alloimmune attack on host tissues mounted by donor T cells, is the most important toxicity of allogeneic bone marrow transplantation. The mechanism by which allogeneic T cells are initially stimulated is unknown. In a murine allogeneic bone marrow transplantation model it was found that, despite the presence of numerous donor antigen-presenting cells, only host-derived antigen-presenting cells initiated graft versus host disease. Thus, strategies for preventing graft versus host disease could be developed that are based on inactivating host antigen-presenting cells. Such strategies could expand the safety and application of allogeneic bone marrow transplantation in treatment of common genetic and neoplastic diseases.

Mice lacking interleukin-2 (IL-2) developed a severe hematopoietic disorder characterized by the abnormal development of myeloid cells and neutropenia. Analysis of the bone marrow of IL-2–deficient (IL-2−/−) mice showed that the number of mature polymorphonuclear cells was decreased by 65% to 75%, and granulocyte/macrophage precursor cells were reduced by 50%. Bone marrow cells from IL-2−/− mice were unable to sustain myelopoiesis in lethally irradiated mice and in long-term bone marrow cultures (LTBMC). The addition of exogenous IL-2 to LTBMC of IL-2−/− cells partially restored hematopoietic progenitor activity. In the bone marrow of wild-type mice, immature (Mac-1lo) myeloid cells, including myeloblasts and promyelocytes, constitutively expressed the β-chain of the IL-2R, and the number of Mac-1loIL-2Rβ+ cells was increased by twofold to threefold in IL-2−/− mice. During culture in the presence of IL-2 and the absence of stromal cells, Mac-1loIL-2Rβ+ immature myeloid cells proliferated and gave rise to mature granulocytes and macrophages. Collectively, these observations indicate that defective myelopoiesis in IL-2−/− mice is at least in part a consequence of their direct dependency on IL-2, and by regulating the growth of immature myeloid cells, IL-2 plays an important role in the homeostatic regulation of myelocytic cell generation.

In order to study a previously described genetic difference manifested in stem cell kinetics of specific mouse strains, effects of this putative gene, stk, were measured on growth and expansion of stem and progenitor cell populations ex vivo. Bone marrow cells from each of two inbred mouse strains, C57BL/6J and DBA/2J, were placed into separate bioreactor cultures perfused continuously with growth medium containing erythropoietin (Epo), interleukin-3 (IL-3), granulocyte-macrphage colony stimulating factor (GM-CSF), and Kit ligand as well as 5% CO2. Expansion of cell numbers reached 20-fold for DBA/2J and 10-fold for C57BL/6J marrow within about 1 week of culture. Significant production was also seen of colony-forming unit (CFU)-GM (up nine-fold from input levels) just prior to the cell production peak, and, importantly, moderate expansion of day 12 colony-forming unit-spleen (CFU-S; two- to threefold) occurred as well, although CFU-S production peaked at a relatively short 4 days. CFU-S and CFU-GM levels declined rapidly in culture, either because of unfavorable growth conditions or terminal differentiation. Attempts to remove toxic metabolites by increasing the media perfusion rate resulted in a boost in cell expansion capability by DBA/2J marrow. In bioreactors in which stromal cells were established before marrow inoculation, there was greater expansion of CFU-S (especially by DBA/2J) and CFU-GM, although total cell yield appeared to be unaffected, perhaps because the maximum cell density had already been reached. The relative high potential for CFU-S expansion measured in DBA/2J marrow over that of C57BL/6J will be useful in following genetic contributions to bone marrow production capacity.

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