A novel assay for analysis of the regulation of the function of human osteoclasts
<p>Abstract</p> <p>Background</p> <p>Very little is known of the regulation of the function of human osteoclasts, largely due to the virtual impossibility of obtaining human osteoclasts <it>ex vivo</it>. It has recently become possible to generate human oste...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
BMC
2006-11-01
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| Series: | Journal of Translational Medicine |
| Online Access: | http://www.translational-medicine.com/content/4/1/45 |
| _version_ | 1819027341695778816 |
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| author | Chambers Timothy J Shiroo Masahiro Samuelsson Bertil Grabowska Urszula Kirstein Barrie Fuller Karen |
| author_facet | Chambers Timothy J Shiroo Masahiro Samuelsson Bertil Grabowska Urszula Kirstein Barrie Fuller Karen |
| author_sort | Chambers Timothy J |
| collection | DOAJ |
| description | <p>Abstract</p> <p>Background</p> <p>Very little is known of the regulation of the function of human osteoclasts, largely due to the virtual impossibility of obtaining human osteoclasts <it>ex vivo</it>. It has recently become possible to generate human osteoclasts <it>in vitro</it>, by incubation of peripheral blood mononuclear cells (PBMCs) in macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL). However, the assays at present available do not distinguish clearly between the distinct effects of agents on differentiation and function.</p> <p>Materials and methods</p> <p>We developed a novel assay for resorptive function of human osteoclasts that minimizes inter-assay variability by using each culture as its own baseline, and that minimizes the confounding effects of agents on differentiation by assessing resorptive function over a short test period. In this assay, the development of resorptive activity is monitored in sample cultures. When resorption is underway, bone resorption (measured as the release of the C-terminal telopeptide degradation product of type I collagen (CTX-I) into the supernatant) is compared before <it>vs </it>after incubation for 1–24 h in test agent.</p> <p>Results</p> <p>Using this assay, we found that changes in bone resorption could be detected using substantially fewer cultures per variable. Moreover, we could detect effects of agents on resorption within 1 h of addition, a time sufficiently short that a change in release is likely to reflect an effect on function rather than on differentiation.</p> <p>Conclusion</p> <p>The assay makes it possible to distinguish the effects of agents on osteoclastic function, independent of their effects on differentiation.</p> |
| first_indexed | 2024-12-21T05:40:56Z |
| format | Article |
| id | doaj.art-a7a29563ae5148d088bc28c271ecfb16 |
| institution | Directory Open Access Journal |
| issn | 1479-5876 |
| language | English |
| last_indexed | 2024-12-21T05:40:56Z |
| publishDate | 2006-11-01 |
| publisher | BMC |
| record_format | Article |
| series | Journal of Translational Medicine |
| spelling | doaj.art-a7a29563ae5148d088bc28c271ecfb162022-12-21T19:14:16ZengBMCJournal of Translational Medicine1479-58762006-11-01414510.1186/1479-5876-4-45A novel assay for analysis of the regulation of the function of human osteoclastsChambers Timothy JShiroo MasahiroSamuelsson BertilGrabowska UrszulaKirstein BarrieFuller Karen<p>Abstract</p> <p>Background</p> <p>Very little is known of the regulation of the function of human osteoclasts, largely due to the virtual impossibility of obtaining human osteoclasts <it>ex vivo</it>. It has recently become possible to generate human osteoclasts <it>in vitro</it>, by incubation of peripheral blood mononuclear cells (PBMCs) in macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL). However, the assays at present available do not distinguish clearly between the distinct effects of agents on differentiation and function.</p> <p>Materials and methods</p> <p>We developed a novel assay for resorptive function of human osteoclasts that minimizes inter-assay variability by using each culture as its own baseline, and that minimizes the confounding effects of agents on differentiation by assessing resorptive function over a short test period. In this assay, the development of resorptive activity is monitored in sample cultures. When resorption is underway, bone resorption (measured as the release of the C-terminal telopeptide degradation product of type I collagen (CTX-I) into the supernatant) is compared before <it>vs </it>after incubation for 1–24 h in test agent.</p> <p>Results</p> <p>Using this assay, we found that changes in bone resorption could be detected using substantially fewer cultures per variable. Moreover, we could detect effects of agents on resorption within 1 h of addition, a time sufficiently short that a change in release is likely to reflect an effect on function rather than on differentiation.</p> <p>Conclusion</p> <p>The assay makes it possible to distinguish the effects of agents on osteoclastic function, independent of their effects on differentiation.</p>http://www.translational-medicine.com/content/4/1/45 |
| spellingShingle | Chambers Timothy J Shiroo Masahiro Samuelsson Bertil Grabowska Urszula Kirstein Barrie Fuller Karen A novel assay for analysis of the regulation of the function of human osteoclasts Journal of Translational Medicine |
| title | A novel assay for analysis of the regulation of the function of human osteoclasts |
| title_full | A novel assay for analysis of the regulation of the function of human osteoclasts |
| title_fullStr | A novel assay for analysis of the regulation of the function of human osteoclasts |
| title_full_unstemmed | A novel assay for analysis of the regulation of the function of human osteoclasts |
| title_short | A novel assay for analysis of the regulation of the function of human osteoclasts |
| title_sort | novel assay for analysis of the regulation of the function of human osteoclasts |
| url | http://www.translational-medicine.com/content/4/1/45 |
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