Journal of Bone and Mineral Metabolism

, Volume 29, Issue 1, pp 111–122

Modeled microgravity and hindlimb unloading sensitize osteoclast precursors to RANKL-mediated osteoclastogenesis

  • Ritu Saxena
  • George Pan
  • Erik D. Dohm
  • Jay M. McDonald
Original Article

DOI: 10.1007/s00774-010-0201-4

Cite this article as:
Saxena, R., Pan, G., Dohm, E.D. et al. J Bone Miner Metab (2011) 29: 111. doi:10.1007/s00774-010-0201-4

Abstract

Mechanical forces are essential to maintain skeletal integrity, and microgravity exposure leads to bone loss. The underlying molecular mechanisms leading to the changes in osteoblasts and osteoclast differentiation and function remain to be fully elucidated. Because of the infrequency of spaceflights and payload constraints, establishing in vitro and in vivo systems that mimic microgravity conditions becomes necessary. We have established a simulated microgravity (modeled microgravity, MMG) system to study the changes induced in osteoclast precursors. We observed that MMG, on its own, was unable to induce osteoclastogenesis of osteoclast precursors; however, 24 h of MMG activates osteoclastogenesis-related signaling molecules ERK, p38, PLCγ2, and NFATc1. Receptor activator of NFkB ligand (RANKL) (with or without M-CSF) stimulation for 3–4 days in gravity of cells that had been exposed to MMG for 24 h enhanced the formation of very large tartrate-resistant acid phosphatase (TRAP)-positive multinucleated (>30 nuclei) osteoclasts accompanied by an upregulation of the osteoclast marker genes TRAP and cathepsin K. To validate the in vitro system, we studied the hindlimb unloading (HLU) system using BALB/c mice and observed a decrease in BMD of femurs and a loss of 3D microstructure of both cortical and trabecular bone as determined by micro-CT. There was a marked stimulation of osteoclastogenesis as determined by the total number of TRAP-positive multinucleated osteoclasts formed and also an increase in RANKL-stimulated osteoclastogenesis from precursors removed from the tibias of mice after 28 days of HLU. In contrast to earlier reported findings, we did not observe any histomorphometric changes in the bone formation parameters. Thus, the foregoing observations indicate that microgravity sensitizes osteoclast precursors for increased differentiation. The in vitro model system described here is potentially a valid system for testing drugs for preventing microgravity-induced bone loss by targeting the molecular events occurring in microgravity-induced enhanced osteoclastogenesis.

Keywords

Modeled microgravityOsteoblastsOsteoclastsHindlimb unloadingRotary cell culture system

Supplementary material

774_2010_201_MOESM1_ESM.tif (519 kb)
Supplementary figure: RAW264.7 cells were incubated overnight in low-adhesion plates. Cells attached around the beads which were then incubated in gravity (G) and modeled microgravity (MMG) for 24h. Figure shows that the cells remain attached around the beads after incubation (10X magnification) (TIFF 518 kb)

Copyright information

© The Japanese Society for Bone and Mineral Research and Springer 2010

Authors and Affiliations

  • Ritu Saxena
    • 1
  • George Pan
    • 2
  • Erik D. Dohm
    • 3
  • Jay M. McDonald
    • 4
    • 5
  1. 1.Department of Cancer Biology and PharmacologyUniversity of Illinois College of Medicine at PeoriaPeoriaUSA
  2. 2.Department of PediatricsEmory University Medical SchoolAtlantaUSA
  3. 3.Animal Resources ProgramThe University of Alabama at BirminghamBirminghamUSA
  4. 4.Departments of Pathology and Cell BiologyThe University of Alabama at BirminghamBirminghamUSA
  5. 5.Veterans Administration Medical CenterBirminghamUSA