title: Type I collagen degradation by mouse calvarial osteoblasts stimulated with 1,25-dihydroxyvitamin D-3: evidence for a plasminogen-plasmin-metalloproteinase activation cascade.
abstract: To understand the mechanisms regulating osteoid removal by osteoblasts, mouse calvarial osteoblasts were grown on 14C-labelled type I collagen films and stimulated with 1,25-dihydroxyvitamin D-3 (2.5.10(-8) M) for 48-72 h. In the presence of 5% non-inhibitory rabbit serum this resulted in a 2-3-fold increase in collagen degradation and a dramatic change in osteoblast morphology, when compared with untreated osteoblasts. Collagenolysis was accompanied by increased synthesis and release of latent collagenase, gelatinase and stromelysin and a concomitant decrease in their specific inhibitor, TIMP (tissue inhibitor of metalloproteinases). In serum-free medium, osteoblasts failed to degrade collagen, but their ability to lyse collagen could be restored by adding plasminogen (5 micrograms/ml) to the cultures. Plasminogen-dependent collagenolysis was inhibited by human recombinant TIMP (5 units/ml), demonstrating that plasmin, derived from plasminogen, activated latent collagenase and did not itself degrade collagen. Plasminogen activator production was confirmed by culturing osteoblasts on 125I-labelled fibrin plates. Comparison with urokinase-type and tissue-type plasminogen activator standards suggested that osteoblast plasminogen activator was predominantly cell-associated and likely to be of the urokinase type. Immunocytochemistry indicated that osteoblasts also constitutively produce plasminogen activator inhibitor-1. These findings provide evidence for the involvement of a plasminogen-plasmin-latent metalloproteinase activation cascade in type I collagen degradation by osteoblasts, and for its regulation by TIMP and plasminogen activator inhibitor-1.
authors.affiliation: Cell Physiology Department, Strangeways Research Laboratory, Worts Causeway, Cambridge, U.K.
authors.forename: B M
authors.initials: BM
authors.lastname: Thomson
authors.forename: S J
authors.initials: SJ
authors.lastname: Atkinson
authors.forename: A M
authors.initials: AM
authors.lastname: McGarrity
authors.forename: R M
authors.initials: RM
authors.lastname: Hembry
authors.forename: J J
authors.initials: JJ
authors.lastname: Reynolds
authors.forename: M C
authors.initials: MC
authors.lastname: Meikle
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D002117:Calcitriol / Q000494:pharmacology*; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism*; D004789:Enzyme Activation; D005341:Fibrinolysin / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D018093:Gelatinases; D006023:Glycoproteins / Q000494:pharmacology; D007150:Immunohistochemistry; D019278:Matrix Metalloproteinase 3; D008666:Metalloendopeptidases / Q000378:metabolism*; D051379:Mice; D003012:Microbial Collagenase / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D010434:Pepsin A / Q000378:metabolism; D010958:Plasminogen / Q000378:metabolism*; D015849:Plasminogen Inactivators / Q000378:metabolism; D011994:Recombinant Proteins; D019714:Tissue Inhibitor of Metalloproteinases
chemical_list: D006023:Glycoproteins; D015849:Plasminogen Inactivators; D011994:Recombinant Proteins; D019714:Tissue Inhibitor of Metalloproteinases; D010958:Plasminogen; D003094:Collagen; D005341:Fibrinolysin; D010434:Pepsin A; D018093:Gelatinases; D008666:Metalloendopeptidases; D019278:Matrix Metalloproteinase 3; D003012:Microbial Collagenase; D002117:Calcitriol