Effects of SCF on Erythroid Myeloid and Megakaryocytic Lineages In Vitro

In erythroid and myeloid liquid and semisolid cultures (i.e., culture conditions include erythroid and myeloid lineage-associated cytokines) of committed progenitors of varying purity from bone marrow, cord blood, fetal liver, and peripheral blood, SCF can dramatically enhance the expansion and differentiation of CFUs (4-7). The culture conditions often include one or more of the other relatively early-acting cytokines as well. The progeny obtained typically reflect the later acting lineage-associated cytokines that are present. In semisolid cultures, the effects of SCF are often seen as dramatic increases in the number and size of CFU-C, probably resulting from direct synergistic action of SCF on c-kit+ lineage-committed progenitors. The synergy by SCF has been demonstrated repeatedly with erythropoietin (EPO) for BFU-E; with granulocyte (G)-CSF and GM-CSF for CFU-G and CFU-GM; with M-CSF for CFU-M; with thrombopoietin/megakaryocyte growth and development factor (TPO/MGDF) for CFU-megakaryocyte (CFU-Meg); and with cytokine combinations for CFU-Mix (4-7,51,57,71,75-78). That the action of SCF is largely direct has been demonstrated by studies with purified cell populations, sequential additions of cytokines (57,75,77,79), limiting dilution (76), single-cell plating (69,72,77,80), and autoradiography using 125I-SCF (50,57,65).

The expression of c-kit and action of SCF during human erythroid lineage maturation in culture have been dissected in some detail (65,76,79,81). Blocking Ab against c-kit inhibits BFU-E formation but does not inhibit CFU-E formation in bone marrow cultures containing EPO and SCF. c-Kit expression, and the ability of SCF to enhance the generation of BFU-E, starts to decline after approximately d 6-9 of in vitro culture, i.e., at about the time of BFU-E to CFU-E transitioning. Expression of transferrin receptor is downregulated in parallel with downregulation of c-kit expression, whereas glycophorin expression is upregulated in parallel. Observations that SCF can stimulate production of fetal hemoglobin by human erythroid cells in culture suggest effects occurring after the onset on hemoglobinization (4,5,7,80,81).

The etiologies of the (relatively rare) bone marrow failure syndromes such as Diamond-Blackfan anemia (DBA), Fanconi's anemia, aplastic anemia, and myelodysplasia syndromes (MDSs) are quite heterogeneous and poorly understood, but SCF/c-kit defects have not been implicated. DBA (a congenital pure red blood cell aplasia) was initially considered a candidate for SCF and/or c-kit defects because of the erythroid specificity, but Southern blotting analyses and sequencing of cDNA from patients have revealed no alterations in these genes (82,83). The hematopoietic progenitors that are present, usually at much reduced levels, in the bone marrow of patients with these disorders generally appear to express c-kit normally and can be expanded and directed toward erythroid and myeloid differentiation at least somewhat in vitro in response to SCF along with other cytokines such as EPO, IL-3, GM-CSF, and G-CSF (82,84-87).

The same is true for bone marrow from patients with human immunodeficiency virus (HlV)-associated myelosuppression (88).

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