title: In vitro spontaneous osteoclastogenesis of human peripheral blood mononuclear cells is not crucially dependent on T lymphocytes.
abstract: OBJECTIVE
  In vitro spontaneous osteoclastogenesis from peripheral blood mononuclear cells (PBMCs) is increased in diseases with excessive bone loss. The purpose of this study was to reassess the role of T lymphocytes in this process.
  
  
  METHODS
  Fresh or cryopreserved PBMCs obtained from healthy subjects and from patients with rheumatoid arthritis, psoriatic arthritis, and non-psoriatic spondylarthritis were cultured at high density and stained for tartrate-resistant acid phosphatase (TRAP). Resorption of mineralized matrix was assessed by a dentin disc assay. CD14+ monocytes and CD3+ T cells were selected using magnetically labeled antibodies.
  
  
  RESULTS
  Numerous multinucleated, TRAP+, dentin-resorbing osteoclasts developed spontaneously from fresh PBMCs from healthy individuals. This process was abrogated by T cell depletion and was restored by exogenous macrophage colony-stimulating factor (M-CSF) and RANKL, indicating the important role of T cells in spontaneous osteoclastogenesis in vitro. Using physiologic freezing and thawing as a model for the activation of PBMCs, spontaneous osteoclastogenesis was significantly increased in cryopreserved versus fresh cells. Under these conditions, spontaneous osteoclastogenesis was not dependent on T lymphocytes, since it was not influenced by T cell depletion and persisted in purified CD14+ cell cultures supplemented with M-CSF and RANKL. In contrast to studies with fresh PBMCs, spontaneous osteoclastogenesis under these conditions did not appear to be clearly different between healthy subjects and patients with arthritis.
  
  
  CONCLUSIONS
  Spontaneous osteoclastogenesis in vitro is dependent on T lymphocytes or on the direct activation of monocytic cells, depending on the test conditions. This variability warrants better validation of the relevance of this functional test for in vivo osteoclastogenesis.
authors.affiliation: Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Bernard
authors.initials: B
authors.lastname: Vandooren
authors.forename: Lode
authors.initials: L
authors.lastname: Melis
authors.forename: Eric M
authors.initials: EM
authors.lastname: Veys
authors.forename: Paul P
authors.initials: PP
authors.lastname: Tak
authors.forename: Dominique
authors.initials: D
authors.lastname: Baeten