Characterization of cultured mast cells derived from Ws/Ws mast cell-deficient rats with a small deletion at tyrosine kinase domain of c-kit.
The Ws mutant allele of rats represents a 12-base deletion at the tyrosine kinase domain of the c-kit gene. Although homozygous Ws/Ws rats were deficient in both connective tissue-type mast cells (CTMC) and mucosal-type mast cells (MMC), mast cells did develop when bone marrow cells of Ws/Ws rats were cultured in the presence of concanavalin A-stimulated spleen cell conditioned medium (ConA-SCM). Although the proliferative response of rat cultured mast cells (RCMC) derived from Ws/Ws rats to ConA-SCM was comparable to that of RCMC derived from control normal (+/+) rats, the proliferative response of Ws/Ws RCMC to rat recombinant stem cell factor (rrSCF; a ligand for the c-kit receptor tyrosine kinase) was much lower than that of +/+ RCMC. However, a slight c-kit kinase activity was detectable in Ws/Ws RCMC, and the proliferation of Ws/Ws RCMC was accelerated when rrSCF was added to ConA-SCM. Because CTMC contain rat mast cell protease-I (RMCP-I) and MMC contain RMCP-II, the phenotype of +/+ and Ws/Ws RCMC in various culture conditions was evaluated by immunohistochemistry of RMCPs. Both +/+ and Ws/Ws RCMC showed the MMC-like phenotype (RMCP-I-/II+) when they were cultured with ConA-SCM alone. Most +/+ RCMC and about half of Ws/Ws RCMC acquired a novel protease (RMCP-I+/II+) phenotype when they were cultured with rrSCF alone. However, because the number of Ws/Ws RCMC dropped to one-tenth in the medium containing rrSCF alone, the absolute number of Ws/Ws RCMC with the RMCP-I+/II+ phenotype did not increase significantly. The effect of rrSCF in inducing the novel phenotype was suppressed when ConA-SCM was added to rrSCF. In contrast, +/+ and Ws/Ws RCMC cocultured with +/+ fibroblasts showed the RMCP-I+/II+ phenotype even in the presence of ConA-SCM. Moreover, a fibroblast cell line derived from SI/SI mouse embryos that did not produce SCF did not support the survival of both +/+ and Ws/Ws RCMC but did induce the RMCP-I+/II+ phenotype in about half of +/+ and Ws/Ws RCMC when their survival was supported by the addition of ConA-SCM. The normal signal transduction through the c-kit receptor did not appear to be prerequisite for the acquisition of the RMCP-I+/II+ phenotype.
Tei
H
H
Department of Pathology, Osaka University Medical School, Suita, Japan.
Kasugai
T
T
Tsujimura
T
T
Adachi
S
S
Furitsu
T
T
Tohya
K
K
Kimura
M
M
Zsebo
K M
KM
Newlands
G F
GF
Miller
H R
HR
D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001854:Bone Marrow Cells; D015815:Cell Adhesion Molecules / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D003208:Concanavalin A; D017077:Culture Media, Conditioned; D010450:Endopeptidases / Q000096:biosynthesis / Q000378:metabolism; D005347:Fibroblasts / Q000502:physiology; D005455:Fluorescent Antibody Technique; D016298:Hematopoietic Cell Growth Factors / Q000494:pharmacology; D007700:Kinetics; D008213:Lymphocyte Activation; D008214:Lymphocytes / Q000276:immunology / Q000502:physiology; D008407:Mast Cells / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D009414:Nerve Growth Factors / Q000276:immunology / Q000502:physiology; D010641:Phenotype; D011505:Protein-Tyrosine Kinases / Q000096:biosynthesis / Q000235:genetics*; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000235:genetics*; D019009:Proto-Oncogene Proteins c-kit; D011519:Proto-Oncogenes; D051381:Rats; D011922:Rats, Mutant Strains; D020794:Receptor Protein-Tyrosine Kinases / Q000096:biosynthesis / Q000235:genetics*; D016184:Receptors, Colony-Stimulating Factor / Q000096:biosynthesis / Q000235:genetics*; D017384:Sequence Deletion; D013154:Spleen / Q000276:immunology / Q000502:physiology; D019089:Stem Cell Factor; D013997:Time Factors
D000911:Antibodies, Monoclonal; D015815:Cell Adhesion Molecules; D017077:Culture Media, Conditioned; D016298:Hematopoietic Cell Growth Factors; D009414:Nerve Growth Factors; D011518:Proto-Oncogene Proteins; D016184:Receptors, Colony-Stimulating Factor; D019089:Stem Cell Factor; D003208:Concanavalin A; D011505:Protein-Tyrosine Kinases; D019009:Proto-Oncogene Proteins c-kit; D020794:Receptor Protein-Tyrosine Kinases; D010450:Endopeptidases