Human c-kit ligand (stem cell factor) induces platelet Fc receptor expression in megakaryoblastic cells.
Platelets and megakaryocytes express Fc receptors for IgG which are encoded by the Fc gamma RIIA gene. In an effort to establish a cellular model for induction of Fc gamma RIIA expression during megakaryocyte development by hematopoietic growth factors, steady-state Fc gamma RIIA mRNA levels were monitored in c-kit receptor-positive megakaryocytic cells (M07e, HEL, and Dami) in response to c-kit ligand (KL; also known as stem cell factor, mast cell growth factor, or Steel factor). Northern blot analysis showed that exposure of cells to KL led to significant increases in Fc gamma RIIA levels in M07e (15 x at 24 hours), with smaller increases in HEL (1.9 x at 2 hours) and Dami (1.6 x at 24 hours) cells. K562 cells, which lack c-kit receptor, showed no effect of KL on modulating Fc gamma RIIA mRNA levels. The effects of KL were specific for Fc gamma RIIA, as there were no effects on platelet factor 4 (PF4), gamma-globin, or GATA-1 mRNA levels. Effects of KL, alone and in combination with granulocyte-macrophage colony-stimulating factor (GM-CSF) and gamma-interferon (IFN-gamma), on surface Fc gamma RIIA expression were assessed by flow cytometry using anti-Fc gamma RII monoclonal antibody IV.3. In M07e cells, KL alone and in combination led to significant increases in the percentage of cells positive for surface Fc gamma RIIA and the mean cell fluorescence intensity. Transient transfection studies of an Fc gamma RIIA promoter-luciferase reporter gene in the presence or absence of KL showed increased reporter gene expression in KL-treated cells, with the largest increase (3.7-fold) in the M07e cells. In HEL and Dami cells, other cytokines active in megakaryocytopoiesis when used alone (interleukin-3 [IL-3], IL-6, IL-11, GM-CSF) had negligible activity in increasing reporter gene activity. These results suggest that increased levels of Fc gamma RIIA mRNA after KL treatment of M07e cells are a result, in part, of increased Fc gamma RIIA gene transcription. Our results indicate that M07e cells represent a cellular model for KL-induced Fc gamma RIIA expression in early megakaryocyte development.
Kiss
C
C
Division of Hematology, Children's Hospital of Philadelphia, PA 19104, USA.
Surrey
S
S
Schreiber
A D
AD
Schwartz
E
E
McKenzie
S E
SE
D000818:Animals; D000911:Antibodies, Monoclonal; D001792:Blood Platelets / Q000276:immunology*; D002460:Cell Line; D016207:Cytokines / Q000494:pharmacology*; D005434:Flow Cytometry; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006133:Growth Substances / Q000494:pharmacology*; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology; D017370:Interleukin-11 / Q000494:pharmacology; D007700:Kinetics; D008156:Luciferases / Q000096:biosynthesis; D008533:Megakaryocytes / Q000187:drug effects / Q000276:immunology*; D011401:Promoter Regions, Genetic; D017452:Receptors, IgG / Q000096:biosynthesis*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000494:pharmacology; D015183:Restriction Mapping; D019089:Stem Cell Factor / Q000494:pharmacology*
D000911:Antibodies, Monoclonal; D016207:Cytokines; D006133:Growth Substances; D017370:Interleukin-11; D017452:Receptors, IgG; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D019089:Stem Cell Factor; D007371:Interferon-gamma; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D008156:Luciferases