Human osteoblasts in vitro secrete tissue inhibitor of metalloproteinases and gelatinase but not interstitial collagenase as major cellular products

L. Rifas, L. R. Halstead, W. A. Peck, L. V. Avioli, H. G. Welgus

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    Abstract

    Human osteoblast cultures (hOB) were examined for the production of interstitial collagenase, tissue inhibitor of metalloproteinases (TIMP), adn gelatinolytic enzymes, Cells were isolated by bacterial collagenase digestion of trabecular bone (vertebra, rib, tibia, and femur) from 11 subjects (neonatal to adult). Confluent cultures were exposed to phorbol 12-myristate 13-acetate, PTH, PGE2, epidermal growth factors, 1,25(OH)2 vitamin D3, recombinant human IL-1β, and dexamethasone. Collagenase and TIMP were assayed immunologically and also by measurements of functional activity. Collagenase was not secreted in significant quantities by human bone cells under any tested condition. Furthermore, collagenase mRNA could not be detected in hOB. However, hOB spontaneously secreted large amounts of TIMP for at least 72 h in culture. hOB TIMP was found to be identical to human fibroblast TIMP by double immunodiffusion, metabolic labeling and immunoprecipitation, Northern blot analysis, and stoichiometry of collagenase inhibition. SDS-substrate gel electrophoresis of hOB-contioned media revealed a prominent band of gelatinolytic activity at 68 kD, and specific polyclonal antisera established its identity with the major gelatinolytic protease of human fibroblasts. Abundant secretion of gelatinolytic, but no collagenolytic, enzymes by hOB may indicate that human osteoblasts do not initiate and direct the cleavage of osteoid collagen on the bone surface, but may participate in the preparation of the bone surface for osteoclast attachment by removal of denatured collagen peptides. The constitutive secretion of TIMP may function to regulate metalloproteinase activity.

    Original languageEnglish
    Pages (from-to)686-694
    Number of pages9
    JournalJournal of Clinical Investigation
    Volume84
    Issue number2
    DOIs
    StatePublished - Jan 1 1989

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