Visualizing mineral binding and uptake of bisphosphonate by osteoclasts and non-resorbing cells

Fraser Coxon, Keith Thompson, Anke J. Roelofs, F. Hal Ebetino, Michael J. Rogers

Research output: Contribution to journalArticle

151 Citations (Scopus)

Abstract

Bisphosphonates (BPs) target bone due to their high affinity for calcium ions. During osteoclastic resorption, these drugs are released from the acidified bone surface and taken up by osteoclasts, where they act by inhibiting the pretrylation of small GTPases essential for osteoclast function. However, it remains unclear exactly how osteoclasts intemalise BPs from bone and whether other cells in the bone microenvironment can also take up BPs from the bone surface. We have investigated. this using a novel fluorescently-labelled alendronate analogue (FL-ALN), and by examining changes in protein prenylation following treatment of cells with risedronate (RIS). Confocal microscopic analysis showed that FL-ALN was efficiently internalised from solution or from the surface of dentine by resorbing osteoclasts into intracellular vesicles. Accordingly, unprenylated Rap] A accumulated to the same extent whether osteoclasts were cultured on RIS-coated dentine or with RIS in solution. By contrast, J774 macrophages intemalised FL-ALN and RIS from solution, but took up comparatively little from dentine, due to their inability to resorb the mineral. Calvarial osteoblasts and MCF-7 tumour cells internalised even less FL-ALN and RIS, both from solution and from the surface of dentine. Accordingly, the viability of J774 and MCF-7 cells was drastically reduced when cultured with RIS in solution, but not when cultured on dentine pre-coated with RIS. However, when J774 macrophages were co-cultured with rabbit osteoclasts, J774 cells that were adjacent to resorbing osteoclasts frequently internalised more FL-ALN than J774 cells more distant from osteoclasts. This was possibly a result of increased availability of BP to these J774 cells due to transcytosis through osteoclasts, since FL-ALN partially co-localised with trancytosed, resorbed matrix protein within osteoclasts. In addition, J774 cells occupying resorption pits internalised more FL-ALN than those on unresorbed surfaces. These data demonstrate that osteoclasts are able to take up large amounts of BP, due to their ability to release the BP from the dentine surface during resorption. By contrast, non-resorbing cells take up only small amounts of BP that becomes available due to natural desorption from the dentine surface. However, BP uptake by non-resorbing cells can be increased when cultured in the presence of resorbing osteoclasts. (c) 2008 Elsevier Inc. All rights reserved.

Original languageEnglish
Pages (from-to)848-860
Number of pages13
JournalBone
Volume42
Issue number5
Early online date26 Jan 2008
DOIs
Publication statusPublished - May 2008

Keywords

  • bisphosphonate
  • osteoclast
  • bone resorption
  • bone mineral
  • transcyrosis
  • RAB geranylgeranyl transferase
  • bone-resorption
  • in-vitro
  • molecular-mechanisms
  • zoledronic acid
  • alendronate
  • apoptosis
  • localization
  • osteoblasts
  • prenylation

Cite this