Development of a new method for obtaining osteoclasts from endosteal surfaces |
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Authors: | Lisa G. May Carol V. Gay |
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Affiliation: | (1) Present address: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, 469 North Frear Building, 16802 University Park, Pennsylvania;(2) Present address: Department of Poultry Science, The Pennsylvania State University, 469 North Frear Building, 16802 University Park, Pennsylvania |
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Abstract: | Summary Techniques for the isolation of ahhighly pure population of viable osteoclasts are limited. For this reason, we developed an isolation procedure that results in a high yield of osteoclast-like cells, up to 92% pure, from 3-wk-old chicken tibias. The unique feature of the method is the migration of cells from marrow-free endosteal surfaces to vitronectin-coated plates. The cells retain the osteoclast phenotype and remain viable in culture for a minimum of 1 wk. The cells were characterized and compared to two populations of authentic avian osteoclasts, which were isolated on the basis of association with fibronectin-coated plates. The cells contained substantial amounts of tartrate-resistant acid phosphatase. Alkaline phosphatase levels were negligible, suggesting little contamination by osteoblasts. Response to parathyroid hormone, dibutyryl cyclic adenosine monophosphate, calcitonin, acetazolamide, 17β-estradiol, and prostaglandin E2 was evident, as detected by measuring acid production. The vitronectin-associating cells contained numerous mitochondria, had the ability to resorb bone in anin vitro bone slice assay, and specifically bound biotinylated vitronectin. At 5 d of culture, the cells demonstrated marginal multinuclearity, having two to three nuclei. A large number (∼1×106 cells/tibia) of viable cells that exhibit characteristics of authentic osteoclasts can be obtained by the method described. Potentially, this method could be applied to other species. |
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Keywords: | osteoclast isolation migratory osteoclasts vitronectin-coated surfaces osteoclast-like cell characterization |
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