Mast cells expressing chymase but not tryptase can be derived by culturing human progenitors in conditioned medium obtained from a human mastocytosis cell strain with c-kit ligand.
Human mast cells (MC) were derived from umbilical cord blood and bone marrow progenitors cultured in the presence of a conditioned medium from a human mastocytosis cell strain and recombinant human kit ligand. MCs were studied using a sequential double immunoenzymatic analysis to determine the heterogeneity of expression of tryptase and chymase, two MC-specific proteases. The conditioned medium and kit ligand promoted the development of distinct MC subtypes from bone marrow and umbilical cord blood progenitors. kit ligand induced MC from umbilical cord blood predominantly of two immunophenotypes, MCT positive for tryptase but negative for chymase, and MCTC positive for tryptase and chymase. In contrast, the conditioned medium induced MC subtype MCTC and a third subtype, MCC, positive for chymase but negative for tryptase from both bone marrow and umbilical cord blood progenitors. In situ hybridization analyses confirmed the existence of a chymase-positive, tryptase-negative human MC in the culture. In umbilical cord blood cultures supplemented with both conditioned medium and kit ligand, the number of MCC cells was up-regulated from 2.7 to 34.0% of the total cells in the culture. In contrast, the number of MCT cells declined from 54.3 to 21.5% on day 49 of culture. In bone marrow cultures supplemented with conditioned medium alone, the MCC subtype represented 100% of the MCs on day 10 of culture. This study clearly demonstrates that a third type of MC, MCC, expressing chymase without concomitant expression of tryptase can be induced in vitro from normal human progenitors. In addition, it shows that tryptase and chymase, two MC-specific proteases, can be differentially expressed in in vitro derived human MCs by changing the cytokine combination of the culture.
Li
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L
University of New South Wales, School of Medicine and Department of Immunology, Australia.
Meng
X W
XW
Krilis
S A
SA
D001483:Base Sequence; D001854:Bone Marrow Cells; D002478:Cells, Cultured; D053818:Chymases; D003470:Culture Media; D004790:Enzyme Induction; D005312:Fetal Blood; D006801:Humans; D007124:Immunoenzyme Techniques; D017403:In Situ Hybridization; D008407:Mast Cells / Q000201:enzymology*; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D012697:Serine Endopeptidases / Q000378:metabolism*; D019089:Stem Cell Factor / Q000494:pharmacology*; D053802:Tryptases
D003470:Culture Media; D009838:Oligodeoxyribonucleotides; D019089:Stem Cell Factor; D012697:Serine Endopeptidases; C083294:chymase 2; D053818:Chymases; D053802:Tryptases