Background: The aim of this study was to investigate whether the secretory granules of human mast cells store stem cell factor (SCF). We also addressed the question whether mast cell chymase, a chymotrypsin–like protease, also present in the secretory granules of human mast cells cleaves SCF at the peptide bound between Phe 158 and Met159. Methods: The skin samples were obtained from patients with mastocytosis, undergoing skin biopsy for diagnostic purposes. Mast cells were isolated and purified from human lung parenchyma (human lung mast cells, HLMC) by countercurrent elutriation followed by discontinuous Percoll density gradient. SCF contents of human mast cells were assessed for immunoreactive SCF by ELISA. Western blot analysis of SCF and its cleavage products were performed with the MoAb anti–SCF 7H6. SCF and its proteolytic fragment were characterized by electrospray mass spectrometry (ES/MS). Results: SCF is present in the secretory granules of human skin and lung mast cells. Immunoreactive SCF (iSCF) was detected in the cell lysates of HLMC, but not in basophils. iSCF was rapidly (3 min) released after challenge with anti–IgE, and iSCF in supernatants rapidly declined after 30 min. ES/MS analysis of rhSCF1–166 treated with recombinant human chymase showed a polypeptide of 17,977.1±0.6 Da and a minor component of 697.4±0.1 Da generated by specific cleavage at Phe159. SCF1–166 and SCF1–159 similarly activated HLMC and potentiated anti–IgE–induced activation of these cells. The cleavage product SCF160–166 had no effect on mast cells. Western blot analysis of supernatants of anti–IgE activated HLMC incubated for various intervals with rhSCF1–166 showed that rhSCF1–166 was converted to a faster–migrating form with a molecular weight compatible with SCF1–159 and to several SCF species. Conclusion: SCF is stored in human mast cell secretory granules and is immunologically released by mast cells. SCF1–166 is rapidly cleaved by chymase and other proteases to several SCF species.

Keywords:
Basophils, Chymase, Histamine, Mast cells, Stem cell factor
1.
Qiu FH, Ray P, Brown K, Barker PE, Jhanwar S, Ruddle FH, Besmer P: Primary structure of c–kit: Relationship with the CSF–1/PDGF receptor kinase family–oncogenic activation of c–kit involves deletion of extracellular domain and C terminus. EMBO J 1988;7:1003–1011.
2.
Zsebo KM, Wypych J, McNiece IK, Lu HS, Smith KA, Karkare SB, Sachdev RK, Yuschenkoff VN, Birkett NC, Williams LR, Satyagal VN, Tung W, Bosselman RA, Mendiaz EA, Langley KE: Identification, purification, and biological characterization of hematopoietic stem cell factor from buffalo rat liver–conditioned medium. Cell 1990;63:195– 201.
3.
Geissler EN, Liao M, Brook JD, Martin FH, Zsebo KM, Housman DE, Galli SJ: Stem cell factor (SCF), a novel hematopoietic growth factor and ligand for c–kit tyrosine kinase receptor, maps on human chromosome 12 between 12q14.3 and 12qter. Somat Cell Mol Genet 1991;17:207–214.
4.
Flanagan JG, Chan DC, Leder P: Transmembrane form of the kit ligand growth factor is determined by alternative splicing and is missing in the Sld mutant. Cell 1991;64:1025– 1035.
5.
Zsebo KM, Williams DA, Geissler EN, Broudy VC, Martin FH, Atkis HL, Hsu R–Y, Birkett NC, Okino KH, Murdock DC, Jacobsen FW, Langley KE, Smith KA, Takeishi T, Cattanach BM, Galli SJ, Suggs SV: Stem cell factor (SCF) is encoded at the Sl locus of the mouse and is the ligand for the c–kit tyrosine kinase receptor. Cell 1990;63:213–224.
6.
Toksoz D, Zsebo KM, Smith KA, Hu S, Brankow D, Suggs SV, Martin FH, Williams DA: Support of human hematopoiesis in long–term bone marrow cultures by murine stromal cells selectively expressing the membrane–bound and secreted forms of the human homolog of the steel gene product, stem cell factor. Proc Natl Acad Sci USA 1992;89:7350– 7354.
7.
Longley BJ Jr, Morganroth GS, Tyrrell L, Ding TG, Anderson DM, Williams DE, Halaban R: Altered metabolism of mast–cell growth factor (c–kit ligand) in cutaneous mastocytosis. N Eng J Med 1993;328:1302–1307.
8.
Hamann K, Haas N, Grabbe J, Czarnetzki BM: Expression of stem cell factor in cutaneous mastocytosis. Br J Dermatol 1995;133:203– 208.
9.
Beck D, Gross N, Beretta Brognara C, Perruisseau G: Expression of stem cell factor and its receptor by human neuroblastoma cells and tumors. Blood 1995;86:3132–3136.
10.
Turner Am, Zsebo KM, Martin F, Jacobsen FW, Bennett LG, Broudy VC: Nonhematopoietic tumor cell lines express stem cell factor and display c–kit receptors. Blood 1992;80: 374–381.
11.
Funasaka Y, Boulton T, Cobb M, Yarden Y, Fan B, Lyman SD, Williams DE, Anderson DM, Zakut R, Mishima Y, Halaban R: c–kit–kinase induces a cascade of protein tyrosine phosphorylation in normal human melanocytes in response to mast cell growth factor and stimulates mitogen–activated protein kinase but is down–regulated in melanomas. Mol Biol Cell 1992;3:197–209.
12.
Columbo M, Horowitz EM, Botana LM, MacGlashan DW Jr, Bochner BS, Gillis S, Zsebo KM, Galli SJ, Lichtenstein LM: The human recombinant c–kit receptor ligand, rhSCF, induces mediator release from human cutaneous mast cells and enhances IgE–dependent mediator release from both skin mast cells and peripheral blood basophils. J Immunol 1992;149: 599–608.
13.
de Paulis A, Ciccarelli A, Cirillo R, de Crescenzo G, Marone G: Modulation of human lung mast cell function by the c–kit receptor ligand. Int Arch Allergy Immunol 1992; 99:326–329.
14.
Valent P, Spanblochl E, Sperr WR, Sillaber C, Zsebo KM, Agis H, Strobl H, Geissler K, Bettlheim P, Lechner K: Induction of differentiation of human mast cells from bone marrow and peripheral blood mononuclear cells by recombinant human stem cell factor/kit–ligand in long term culture. Blood 1992;80:2237–2245.
15.
Saito H, Ebisawa M, Tachimoto H, Shichijo M, Fukagawa K, Matsumoto K, Iikura Y, Awaji T, Tsujimoto G, Yanagida M, Uzumaki H, Takahashi G, Tsuji K, Nakahata T: Selective growth of human mast cells induced by steel factor, IL–6, and prostaglandin E2 from cord blood mononuclear cells. J Immunol 1996; 157:343–350.
16.
Marone G, Casolaro V, Patella V, Florio G, Triggiani M: Molecular and cellular biology of mast cells and basophils. Int Arch Allergy Immunol 1997;114:207–217.
17.
Patella V, Marinò I, Arbustini E, Lamparter–Schummert B, Verga L, Adt M, Marone G: Stem cell factor in mast cells and increased mast cell density in idiopathic and ischemic cardiomyopathy. Circulation 1998;97:971– 978.
18.
Zhang S, Anderson DF, Bradding P, Coward WR, Baddeley SM, MacLeod JDA, McGill JI, Church MK, Holgate ST, Roche WR: Human mast cells express stem cell factor. J Pathol 1998;186:59–66.
19.
Goldstein SM, Leong J, Schwartz LB, Cooke D: Protease composition of exocytosed human skin mast cell protease–proteoglycan complexes. Tryptase resides in a complex distinct from chymase and carboxypeptidase. J Immunol 1992;148:2475–2482.
1999
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