Fibronectin fragments induce the expression of stromelysin-1 mRNA and protein in bovine chondrocytes in monolayer culture.
Addition of proteolytically generated fibronectin fragments (Fn-f) to cultured cartilage tissue causes greatly enhanced release of metalloproteinases (MMPs), such as pro-stromelysin-1 (proSln-1), and suppression of proteoglycan (PG) synthesis, through release of catabolic cytokines, while native fibronectin is ineffective. We have investigated whether enhanced release of proSln-1 was due to up-regulation of pro-Sln-1 mRNA. We report the addition of a 29-kDa (amino-terminal heparin-binding Fn-f) or a 140-kDa (central cell-binding Fn-f) to bovine chondrocytes in monolayer culture causes a dose dependent increase in the expression of pro-Sln-1 mRNA and the greatly enhanced release of pro-Sln-1 protein into the culture media. Up to 700 nM pro-Sln-1 was found in the conditioned media and metabolic labeling showed that it constituted a major portion of newly synthesized protein. A potential activator of pro-Sln-1, urokinase (u-PA), was released at elevated levels in the presence of the Fn-f while other activators, tissue plasminogen activator (t-PA) and plasmin activities were not detected. Addition of these activators to conditioned media did not allow conversion of pro-Sln-1 to active Sln-1. However, aminophenyl mercuric acid activated pro-Sln-1 to a 48-kDa Sln-1 form capable of degrading PG when added to cartilage suspensions. Gelatinase A mRNA was also enhanced, suggesting that the Fn-f may induce MMPs in general. However, the major regulator of Sln-1 activity, tissue inhibitor of MMPs form 1 (TIMP-1), was not induced at the gene level. Thus, a major effect of Fn-f on chondrocytes is to up-regulate pro-Sln-1 expression at the gene level, resulting in pro-Sln-1 as a major protein product.
Bewsey
K E
KE
Department of Biochemistry, Rush Medical College at Rush-Presbyterian-St. Luke's Medical Center, Chicago, IL 60612-3864, USA.
Wen
C
C
Purple
C
C
Homandberg
G A
GA
D000818:Animals; D002356:Cartilage / Q000166:cytology / Q000201:enzymology*; D002364:Caseins / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D017077:Culture Media, Conditioned; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D005353:Fibronectins / Q000494:pharmacology*; D018093:Gelatinases / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism; D019278:Matrix Metalloproteinase 3 / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D008666:Metalloendopeptidases / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D008970:Molecular Weight; D010446:Peptide Fragments / Q000494:pharmacology*; D010662:Phenylmercuric Acetate / Q000031:analogs & derivatives / Q000494:pharmacology; D010960:Plasminogen Activators / Q000378:metabolism / Q000494:pharmacology; D011509:Proteoglycans / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D019714:Tissue Inhibitor of Metalloproteinases; D015854:Up-Regulation
D002364:Caseins; D017077:Culture Media, Conditioned; D004792:Enzyme Precursors; D005353:Fibronectins; D006023:Glycoproteins; D010446:Peptide Fragments; D011509:Proteoglycans; D012333:RNA, Messenger; D019714:Tissue Inhibitor of Metalloproteinases; C010024:4-aminophenylmercuriacetate; D010960:Plasminogen Activators; D018093:Gelatinases; D008666:Metalloendopeptidases; C048047:prostromelysin; D019278:Matrix Metalloproteinase 3; D010662:Phenylmercuric Acetate