The effects of recombinant rat stem cell factor (SCF/c-kit ligand) on murine megakaryocytopoiesis were studied using partially purified bone marrow cells derived from normal and 5-fluorouracil (5-FU)-treated mice in a serum-free culture system. SCF alone did not support the formation of megakaryocyte (M) and granulocyte-macrophage-megakaryocyte (GMM) colonies. However, the addition of SCF to cultures containing interleukin-3 (IL-3) resulted in a significant increase in the number of M and GMM colonies formed by bone marrow cells from normal mice, whereas IL-6 augmented only M colony growth. The stimulatory effect of SCF was approximately three to four times as high as that of IL-6 on the primitive progenitors capable of megakaryocytic-lineage expression derived from 5-FU-treated mice. In addition, SCF, but not IL-6, significantly increased the number of constituent cells in the individual M colonies supported by IL-3. On the other hand, SCF did not exert any effect on the size and DNA content of megakaryocytes in IL-3- dependent M and GMM colonies, whereas IL-6 enhanced the maturation of megakaryocytes. These results suggest that SCF stimulates the proliferative process in megakaryocytic progenitors and that the main activity of IL-6 is the promotion of megakaryocyte maturation.

We examined the effects of interferon-gamma (IFN-gamma) on the growth of murine hematopoietic progenitors supported by interleukin-3 (IL-3) or stem cell factor (SCF) in a serum-free culture system. IFN-gamma inhibited IL-3-dependent granulocyte-macrophage colony growth by normal bone marrow cells, but increased the number of pure and mixed megakaryocyte colonies by post-5-fluorouracil bone marrow cells. The addition of IFN-gamma to the culture containing SCF resulted in a synergistic action on the development of primitive hematopoietic progenitors as well as on the development of mature populations. Primitive progenitors responding to SCF + IFN-gamma were suggested to be supported by SCF in the early stage of development and require IFN- gamma for subsequent growth. Replating experiments of blast cell colonies and comparison of total colony growth among SCF + IFN-gamma, SCF + IL-3, and SCF + IFN-gamma + IL-3 suggest that multipotential progenitors supported by SCF + IFN-gamma are a part of those reactive to SCF + IL-3. These findings suggest that IFN-gamma has bifunctional activity on murine hematopoiesis.

Severe bacterial infections sometimes occur and spread rapidly in SGD. Detection of neutrophil granules by flow cytometry is useful for a rapid diagnosis and a screening of SGD.

The effects of recombinant human erythropoietin (rEp) on murine hematopoietic progenitors were studied using a serum-free culture. A high concentration of rEp stimulated the formation of mixed erythroid- megakaryocyte colonies (EM colonies) and blast cell colonies, as well as erythroid colonies, erythroid bursts, and megakaryocyte colonies from normal mouse bone marrow cells. Direct effects of rEp on EM colony, megakaryocyte colony, and erythroid burst formation were confirmed by depletion of accessory cells such as T cells, B cells, and macrophages from crude bone marrow cells, and inhibition of the colonies by the addition of rabbit anti-rEp antibody to the culture in a dose-dependent fashion. Replating experiments were performed to confirm the differentiating ability of blast cell colonies grown in the presence of rEp. Most of the blast cell colonies yielded not only secondary erythroid colonies but also megakaryocyte colonies in the presence of 2 IU/mL rEp. Some of the blast cell colonies produced secondary EM colonies in the presence of 16 IU/ml rEp of 2 IU/mL rEp plus interleukin-3, although no granulocyte-macrophage colonies were found in the secondary culture. These results suggest that Ep acts not only as a late-acting factor that is specific for erythroid progenitors, but also as a bipotential EM-stimulating factor for murine hematopoietic cells.