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It has been shown that hematopoietic progenitors can be expanded ex vivo in the presence of various cytokine combinations. Glucocorticoids (GC) are involved in the self-renewal of erythroid progenitors in chicken. To see whether GC have a similar effect on hematopoiesis in humans, CD34(+) peripheral blood stem cells were cultured in serum free medium in the presence of a GC, triamcinolone acetonide. However, our results demonstrate an inhibition of both erythroid and granulocyte-macrophage (GM) proliferation and a modification of erythroid colony morphology. Furthermore, RU38486 (Mifepristone), a potent GC antagonist, was unable to reverse the inhibitory effect of triamcinolone acetonide. We also identified and characterized another steroid subfamily, the mineralocorticoid (MC) subfamily, in human PB CD34(+) cells. The MC, aldosterone, significantly enhanced GM colony formation and diminished the erythroid colony number. Neither of effects were inhibited by ZK91587, an antagonist specific to the MC receptor (MCR). In contrast, ZK91587 reversed the stimulatory effect of deoxycorticosterone on GM colony formation. Cytoplasmic staining for MCR was observed in CD34(+) cells incubated with a polyclonal antiserum raised against human MCR. To our knowledge, this is the first demonstration of the presence of MCR in human PB CD34(+) cells.

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Interleukin-12 (IL-12) or natural killer cell stimulatory factor (NKSF), has multiple effects on T lymphocytes and natural killer cells. In this study, the effect of IL-12 on human hematopoiesis was studied by analyzing the growth of CD34+ peripheral blood stem cells (PBSC), in steady state. In the presence of Epo, IL-12 alone or in combination with IL-3 or SCF had no effect on the formation of colonies from CD34+ cells. In culture with Epo, G-CSF, and IL-3, the effect of Flt3-ligand (FL) on CD34+ PBSC was investigated in the presence or absence of IL-12. No additional effect of IL-12 was observed when combined with FL. We evaluated 5-FU-treated human CD34+ PBSC proliferation in cultures with Epo, G-CSF, and IL-3, in the presence or absence of IL-12. No cytokine combination enhanced colony formation from 5-FU-treated CD34+ cells. However, in cultures of 5-FU-treated human CD34+ cells, the most efficient combination was IL-3 + Epo + G-CSF + Accessory cells (CD34-). Furthermore, IL-12 enhanced this colony formation significantly. To investigate whether immature CD34+ cells were responsible for FL or SCF, 5-FU-treated human CD34+ cells were cultured with or without IL-12. Whereas no synergistic effect was observed in combination with IL-12, SCF alone significantly enhanced colony formation. However, the colony number was found to be smaller than with the potent combination of accessory cells in the presence of IL-12. These results indicate that accessory cells, lost in CD34+ cell purification, could be partly responsible for an IL-12 effect on immature human PBSC proliferation.