Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.3 (collagenase)
 18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The identification and purification of human osteoclast precursors is essential to further our understanding of the mechanisms that control human osteoclast differentiation. Osteoclastoma tissue potentially provides a rich source of human osteoclast precursors, and in previous studies we have demonstrated the existence of a population of mononuclear cells within this tissue that is reactive with osteoclast-selective vitronectin receptor monoclonal antibodies. In this study, mononuclear cells expressing the vitronectin receptor, as defined by their ability to react with a murine monoclonal antibody to the beta 3 chain of the vitronectin receptor (87MEM1), were isolated from collagenase digests of osteoclastoma tissue using a fluorescence activated cell sorter. Based on their fluorescence signal and size, approximately 2-3% of the viable cells (typically 2 x 10(5)) were obtained and prepared for further phenotyping. The isolated cells demonstrated a number of phenotypic characteristics of osteoclasts: positive tartrate-resistant acid phosphatase (TRAP) activity, reactivity with human osteoclast-selective antibodies, expression of calcitonin receptors, cathepsin K (a novel osteoclast-selective cysteine proteinase) mRNA, and osteopontin mRNA and protein. These phenotypic characteristics were also detected in mononuclear cells within cryostat sections of the native osteoclastoma tissue as well as in resorption lacunae of sections of human bone. In contrast, isolated peripheral blood monocytes were negative for TRAP activity and osteopontin expression and, unlike the osteoclastoma-derived cells, demonstrated strong nonspecific esterase activity. Significantly, when the osteoclastoma-derived 87MEM1 positive cells were cocultured on whale dentine for 1-3 weeks with stromal cells, extensive resorption of the dentine surface was observed. This is the first demonstration of the purification of human osteoclast precursors. These cells provide an homogeneous cell population for studying cellular events that occur during human osteoclast differentiation.
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PMID:Purification and characterization of fully functional human osteoclast precursors. 891 68

A human in vitro resorption assay has been developed using osteoclastoma-derived osteoclasts and used to evaluate novel antiresorptive agents including antagonists of the alphavbeta3 integrin, and inhibitors of cathepsin K and the osteoclast ATPase. The potency of novel compounds in the in vitro resorption assay correlates with functional assays for each class of inhibitor: the human alphavbeta3-mediated cell adhesion assay for the vitronectin receptor antagonists (r2 = 0.82), the chick osteoclast vacuolar ATPase enzyme assay for the H+-ATPase inhibitors (r2 = 0.77) and the recombinant human cathepsin K enzyme assay for the cathepsin K inhibitors (r2 = 0.80). Cell suspensions, rich in osteoclasts, are prepared by collagenase digestion of the tumor tissue. These cells can be stored long-term in liquid nitrogen and upon thawing maintain their bone-resorbing phenotype. The cryopreserved cells can be cultured on bovine cortical bone for 24-48 h and resorption can be measured by either confocal microscopy or biochemical assays. The resorptive activity of osteoclasts derived from a number of tumors can be inhibited reproducibly using a number of mechanistically unique antiresorptive compounds. In addition, the measurement of resorption pits by laser confocal microscopy correlates with the release of type I collagen C-telopeptides or N-telopeptides, as measured by enzyme-linked immunosorbent assay. Resorption can be measured reproducibly using a 48-h incubation of osteoclasts on bone slices, or a 24-h incubation with bone particles. This in vitro human osteoclast resorption assay provides a robust system for the evaluation of inhibitors of osteoclastic function that may be developed for the treatment of metabolic bone diseases such as osteoporosis.
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PMID:Development and characterization of a human in vitro resorption assay: demonstration of utility using novel antiresorptive agents. 1046 85

We have previously demonstrated that a tartrate-resistant acid phosphatase (TRAP)-positive subpopulation of mononuclear cells isolated from collagenase digests of human osteoclastoma tissue exhibits an osteoclast phenotype and can be induced to resorb bone. Using these osteoclast precursors as a model system, we have assessed the chemotactic potential of 16 chemokines. Three CC chemokines, the recently described CKbeta-8, RANTES, and MIP-1alpha elicited significant chemotactic responses. In contrast, 10 other CC chemokines (MIP-1beta, MCP-1, MCP-2, MCP-3, MCP-4, HCC-1, eotaxin-2, PARC, SLC, ELC) and 3 CXC chemokines (IL-8, GROalpha, SDF-1) were inactive. None of these chemokines showed any chemotactic activity for either primary osteoblasts derived from human bone explants or the osteoblastic MG-63 cell line. The identity of the osteoclast receptor that mediates the chemotactic response remains to be established. However, all three active chemokines have been reported to bind to CCR1 and cross-desensitization studies demonstrate that RANTES and MIP-1alpha can partially inhibit the chemotactic response elicited by CKbeta-8. CKbeta-8, the most potent of the active CC chemokines (EC(max) 0.1-0.3 nM), was further characterized with regard to expression in human bone and cartilage. Although expression is not restricted to these tissues, CKbeta-8 mRNA was shown to be highly expressed in osteoblasts and chondrocytes in human fetal bone by in situ hybridization. In addition, CKbeta-8 protein was shown to be present in human osteophytic tissue by immunolocalization. These observations suggest that CKbeta-8, and perhaps other chemokines, may play a role in the recruitment of osteoclast precursors to sites of bone resorption.
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PMID:CKbeta-8 [CCL23], a novel CC chemokine, is chemotactic for human osteoclast precursors and is expressed in bone tissues. 1073 95