The synthesis of collagenase, gelatinase-A (72 kDa) and -B (95 kDa), and TIMP-1 and -2 by human osteoblasts from normal and arthritic bone.
Bone resorption is a complex multistep process that involves removal of both the organic and mineral constituents of bone matrix by proteolytic enzymes synthesized by osteoblasts and osteoclasts. To further understand the role of matrix metalloproteinases (MMPs) and their specific inhibitors TIMPs (tissue inhibitor of metalloproteinases) in this process, human osteoblasts were obtained by sequential enzymatic digestion from samples of bone from normal donors and patients with various forms of arthritis; first passage cells were used in all experiments and cultured on a type I collagen substratum. Collagenase was detected by an ELISA in supernatants from unstimulated osteoblasts (range 12-730 ng/mL), although the levels did not appear to bear any relationship to the age or clinical status of the patient; treatment with parathyroid hormone (PTH; 2 units/mL) and 1,25-dihydroxyvitamin D3 [1,25(OH)2D3, 10 ng/mL] had no added effect, but mononuclear cell conditioned medium (MCM; 5% v/v) and interleukin-1 alpha (IL-1 alpha; 1 ng/mL) both stimulated collagenase synthesis, in the case of MCM by two orders of magnitude. TIMP-1 was detected in unstimulated cultures by an ELISA (range 320-590 ng/mL), the mean level being three-fold greater than for collagenase and was stimulated by 1,25(OH)2D3 and MCM treatment. Degradation studies showed that, over a 120 h culture period, one third of the collagen substratum was degraded by unstimulated cells. PTH and 1,25(OH)2D3 had no effect on this endogenous level of lysis, but addition of MCM and IL-1 alpha resulted in a significant increase in collagen degradation.(ABSTRACT TRUNCATED AT 250 WORDS)
Cell and Molecular Biology Department, Strangeways Research Laboratory, Cambridge, UK.
M C
MC
Meikle
S
S
Bord
R M
RM
Hembry
J J
JJ
Reynolds
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001168:Arthritis / Q000378:metabolism*; D016723:Bone Remodeling; D002478:Cells, Cultured; D017364:Collagenases / Q000096:biosynthesis*; D005260:Female; D018093:Gelatinases / Q000096:biosynthesis*; D006023:Glycoproteins / Q000096:biosynthesis*; D006801:Humans; D008297:Male; D008875:Middle Aged; D010006:Osteoblasts / Q000378:metabolism*; D014176:Protein Biosynthesis; D019716:Tissue Inhibitor of Metalloproteinase-2; D019714:Tissue Inhibitor of Metalloproteinases
D006023:Glycoproteins; D019714:Tissue Inhibitor of Metalloproteinases; D019716:Tissue Inhibitor of Metalloproteinase-2; D017364:Collagenases; D018093:Gelatinases