Metalloelastase expression in a mouse macrophage cell line--regulation by 4beta-phorbol 12-myristate 13-acetate, lipopolysaccharide and dexamethasone.
The regulation of the expression of mouse macrophage elastase (MME) was investigated using the murine tumor cell line P388D1. The effects of three factors were studied: a phorbol ester (4beta-phorbol 12-myristate 13-acetate, PMA), an endotoxin (lipopolysaccharide, LPS) and a corticosteroid (dexamethasone). Both in situ hybridization and northern blot analysis showed that P388D1 cells constitutively express the MME gene. Quantification of the MME mRNA by northern blot analysis showed that only PMA and dexamethasone significantly regulate MME gene expression in a time-dependent and dose-dependent manner. After PMA treatment, the MME mRNA level was maximal between 4 h and 9 h (medium-term response), and the mean amplitude of the response to a concentration of 100 nM was 2.5-fold (P<0.01). LPS did not induce any significant change in MME mRNA level even when 1% serum was added to the cultures. Following dexamethasone treatment, the MME mRNA level was minimal between 21 h and 33 h (long-term response), and the mean amplitude of the response to a concentration of 100 nM was 0.49-fold (P < 0.05). Using actinomycin D, it appeared that the inhibition of RNA synthesis reduces the ulterior stimulating effect of PMA from 184% to 121%, and that MME mRNA has a half-life longer than 8 h, which is not diminished by dexamethasone. These results strongly suggest that the two factors modify MME mRNA level by stimulating (PMA) or inhibiting (dexamethasone) the transcription of the gene, rather than by modifying the transcript stability. Analysis of the cell-conditioned media by elastin zymography showed the MME as a lysis band in the 22-kDa region, the intensity of which varied with the treatments. The MME secretion is stimulated by PMA, inhibited by dexamethasone and does not show any variation after LPS treatment.
Monet-Kuntz
C
C
Laboratoire de Physiopathologie et Génétique Rénale et Pulmonaire, Institut National de la Santé et de la Recherche Médicale, Unité 295, St Etienne Du Rouvray, France.
Cuvelier
A
A
Sarafan
N
N
Martin
J P
JP
D000818:Animals; D002460:Cell Line; D017077:Culture Media, Conditioned; D003907:Dexamethasone / Q000494:pharmacology*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D017403:In Situ Hybridization; D007700:Kinetics; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008264:Macrophages / Q000201:enzymology*; D053508:Matrix Metalloproteinase 12; D008666:Metalloendopeptidases / Q000096:biosynthesis*; D051379:Mice; D010196:Pancreatic Elastase / Q000096:biosynthesis*; D012333:RNA, Messenger / Q000096:biosynthesis; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects*
D017077:Culture Media, Conditioned; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D003907:Dexamethasone; D010196:Pancreatic Elastase; D008666:Metalloendopeptidases; D053508:Matrix Metalloproteinase 12; D013755:Tetradecanoylphorbol Acetate