Reversible expression of mouse mast cell protease 2 mRNA and protein in cultured mast cells exposed to IL-10.
BALB/cJ mouse mast cells derived by culturing bone marrow progenitor cells in WEHI-3 cell-conditioned medium (BMMCW) do not contain mouse mast cell protease 2 (mMCP-2) mRNA, but these cells can be induced to express this transcript after exposure to rIL-10. To study the translation and granule accumulation of mMCP-2 in rIL-10-treated BMMC (BMMCW+IL-10), a rabbit antibody was developed to a synthetic peptide that corresponds to the novel amino acid sequence in mMCP-2 at residues 56 to 71. After affinity purification, this antibody, anti-mMCP-2(56-71) IgG, reacted in SDS-PAGE/immunoblots against a 28-kDa protein in BMMCW+IL-10 that had the N-terminal amino acid sequence of mMCP-2. As assessed immunohistochemically, mMCP-2 protein accumulated in the secretory granules of Kirsten sarcoma virus-immortalized mouse mast cells, BMMCW+IL-10, and the mucosal mast cells present in the jejunum of Trichinella spiralis-infected BALB/cJ mice. Time course analyses of the induction of mMCP-2 mRNA and protein in BMMCW+IL-10 revealed that these cells contain a high steady-state level of mMCP-2 mRNA 24 h after their exposure to rIL-10. Although a small amount of immunodetectable mMCP-2 protein is present in the cells treated for 24 h, large amounts of this protease are not obtained until 7 days of treatment of the cells with rIL-10. Time course analyses of the loss of mMCP-2 mRNA and protein in BMMCW+IL-10 revealed that the steady-state level of mMCP-2 mRNA decreased dramatically 24 h after rIL-10 was removed from the culture medium, but that the level of mMCP-2 protein did not decline measurably until day 5 of culture. The fact that the steady-state levels of mMCP-2 mRNA and protein in BMMC can both be reversibly altered by culturing these mast cells in the presence and absence of rIL-10 suggests that the phenotype of mast cells is not fixed. Rather, it is in a dynamic state regulated by the cytokine network to which mast cells are exposed in their different microenvironments.
Ghildyal
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Department of Medicine, Harvard Medical School, Boston, MA 02115.
Friend
D S
DS
Nicodemus
C F
CF
Austen
K F
KF
Stevens
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RL
D000595:Amino Acid Sequence; D000818:Animals; D002478:Cells, Cultured; D053818:Chymases; D003594:Cytoplasmic Granules / Q000201:enzymology; D015870:Gene Expression; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D016753:Interleukin-10 / Q000494:pharmacology*; D008407:Mast Cells / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010455:Peptides / Q000276:immunology; D012333:RNA, Messenger / Q000235:genetics; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000235:genetics* / Q000378:metabolism*; D017160:Trichinella spiralis; D014235:Trichinellosis / Q000276:immunology
D010455:Peptides; D012333:RNA, Messenger; D016753:Interleukin-10; D012697:Serine Endopeptidases; C083294:chymase 2; C057818:cytotoxic T lymphocyte-specific serine protease; D053818:Chymases; C509229:Cma2 protein, mouse