Role of immunocompetent cells in the regulation of human megakaryocytopoiesis in vitro.

T cells and monocytes/macrophages (Mo) have been shown to play important roles in modulating the growth and differentiation of human erythroid and myeloid progenitors and have been implicated in the mechanisms of gamma interferon (gamma-IFN) mediated suppression of normal human marrow erythroid progenitors in vitro. In order to assess the importance of T cells and Mo in the growth of human megakaryocytic progenitors (CFU-Mk) in vitro and to investigate gamma-IFN effect on human megakaryocytopoiesis, normal human marrow (BM) was cultured in plasma clot in the presence and absence T cells, Mo and gamma-IFN under conditions that support the formation of CFU-Mk derived colonies. The removal of T cells from BM (BM-T) caused a significant decrease (71.3 +/- 3.2 colonies observed vs 231.2 +/- 38.5 colonies predicted) in both the number and size of CFU-Mk derived colonies, and no such changes were seen with Mo depletion (BM-Mo); co-culture of autologous T cells with BM depleted of both Mo and T cells (BM-Mo-T) caused a significant increase in CFU-Mk derived colonies and restored colony size. The addition of gamma-IFN (less than 50-10,000 IU/ml) to BM caused a dose dependent inhibition of CFU-Mk (0-90%) as evidenced by decreased colony numbers and reduced colony size. The addition of gamma-IFN (50-10,000 IU/ml) to BM-T caused reduced inhibition of CFU-Mk (0-60%); co-culture of T cells (but not Mo) pre-incubated with gamma-IFN (10,000 IU/ml; 1 hour, 37 C followed by washing X 3) resulted in supression of CFU-Mk (80% inhibition with the addition of 1:4 T cells:marrow cells). The results demonstrate that T cells have the ability to modulate the growth of human CFU-Mk in vitro and may, under appropriate conditions, either promote (normal T cells) or inhibit (gamma-IFN activated T Cells) human megakaryocytopoiesis.

Suppression of normal human erythropoiesis by human recombinant DNA-produced alpha-2-interferon in vitro.

Interferons (IFN) have been shown to suppress the proliferation of human erythroid progenitors (BFU-E, CFU-E) in vitro. We have previously demonstrated that the inhibition of erythroid colony formation by gamma-IFN in vitro is mediated, in part, through the activation of monocytes and T-lymphocytes. In order to examine the mechanism(s) underlying the inhibitory action of one type of recombinant alpha-IFN (alpha-2-IFN) on erythropoiesis, the effect of different doses (80-10,000 U) of alpha-2-IFN on erythroid colony formation by normal human bone marrow cells in the presence or absence of monocytes and/or T cells was studied. The addition of alpha-2-IFN to whole marrow caused the suppression of BFU-E (10%-68%) and CFU-E (5%-75%) in a dose-dependent fashion. This inhibition occurred with the direct addition of alpha-2-IFN to culture plates but not with brief preincubation of marrow cells with alpha-2-IFN followed by washing of the cells. By contrast, brief exposure of marrow cells to gamma-IFN resulted in significant suppression of erythroid colony formation. The inhibitory action of alpha-2-IFN was not influenced by erythropoietin. Removal of monocytes and/or T cells prior to the addition of alpha-2-IFN failed to significantly reduce the suppressive effects of this molecule (BFU-E: 21%-66%; CFU-E: 20%-83%). Coculture of purified monocytes or T-lymphocytes preexposed to alpha-2-IFN with autologous bone marrow cells did not cause suppression of erythropoiesis; monocytes or T cells similarly treated with gamma-IFN, however, inhibited autologous BFU-E and CFU-E in vitro. These results demonstrate that, unlike gamma-IFN, the inhibitory effect of alpha-2-IFN on erythroid colony formation in vitro is not mediated to any significant degree through monocytes and T-lymphocytes. The suppressive effect of alpha-2-IFN occurs either directly at the erythroid progenitor(s) level and/or through accessory cell(s) other than monocytes and T cells.

Suppression of normal human erythropoiesis by gamma interferon in vitro. Role of monocytes and T lymphocytes.

Interferons (IFN) have been shown to suppress the proliferation of human erythroid progenitors (erythroid burst-forming units [BFU-E] and colony-forming units [CFU-E]) in vitro. To examine the mechanism(s) underlying this inhibitory activity, the effect of different doses (50-10,000 U) of a highly purified preparation of recombinant DNA produced human gamma-IFN on erythroid colony formation by normal human bone marrow BFU-E and CFU-E in the presence and absence of monocytes and/or T lymphocytes was studied. The addition of gamma-IFN to whole marrow caused suppression of BFU-E (6-65%) and CFU-E (31-79%) in a dose-dependent fashion. This inhibition occurred both with the direct addition of gamma-IFN to the culture plates as well as by the preincubation of marrow cells with gamma-IFN followed by the washing of the cells; at the highest concentration of gamma-IFN (10,000 U), near-maximal inhibition of colony formation occurred with as little as 15 min of preexposure (BFU-E, 50%; CFU-E, 81%). Removal of monocytes and/or T lymphocytes before the addition of gamma-IFN significantly reduced the inhibitory effects of this lymphokine (BFU-E, -1 to 38%; CFU-E, -8 to 67%). Co-culture of purified autologous monocytes or T cells preexposed to gamma-IFN with monocyte and T cell-depleted marrow cells resulted in highly significant inhibition of erythroid colony formation even when these treated cells comprised less than 1% of the total nucleated cell populations in culture. The inhibitory action of gamma-IFN was not prevented or reversed by erythropoietin. These results demonstrate that the inhibitory effects of gamma-IFN on erythropoiesis are mediated to a significant degree through accessory cell populations, and suggest that gamma-IFN may represent a useful tool in the study of the role of immunocompetent cells in the regulation of erythropoiesis in vitro.