title: Combination of tumor necrosis factor α and interleukin-6 induces mouse osteoclast-like cells with bone resorption activity both in vitro and in vivo.
abstract: OBJECTIVE
  To clarify the function of osteoclast-like multinuclear cells differentiated from bone marrow-derived macrophages (BMMs) by a combination of tumor necrosis factor α (TNFα) and interleukin-6 (IL-6), and to investigate the molecular mechanisms underlying the differentiation.
  
  
  METHODS
  BMMs were stimulated by TNFα and/or IL-6. The cells were then compared with conventional osteoclasts differentiated in vitro by RANKL. An in vitro pit formation assay on dentine slices and an in vivo resorption assay of calvarial bones were performed. We also evaluated the activities and expression levels of NF-κB, c-Fos, and NF-ATc1, which are essential to the differentiation of conventional osteoclasts. Small interfering RNA was used to knock down c-Fos. The effects of genetic ablation of STAT-3 and pharmacologic inhibitors of NF-AT, JAK, and ERK were also studied.
  
  
  RESULTS
  Osteoclast-like cell differentiation depended on TNFα and IL-6 and was not inhibited by osteoprotegerin. These differentiated cells were associated with both in vitro and in vivo bone resorption activity. TNFα and IL-6 had a synergistic effect on the activity and expression of c-Fos. Knockdown of c-Fos inhibited the expression of NF-ATc1 and the differentiation of osteoclast-like cells. All of these inhibitors blocked differentiation of the cells in vitro, but surprisingly, the conditional knockout of STAT-3 did not. Tofacitinib also inhibited the bone destruction caused by TNFα and IL-6 in vivo.
  
  
  CONCLUSIONS
  Our results demonstrate that a combination of the inflammatory cytokines TNFα and IL-6 can induce osteoclast-like cells that have in vitro and in vivo bone-resorptive activity.
authors.affiliation: Saitama Medical University, Saitama, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Yokota
authors.forename: Kojiro
authors.initials: K
authors.lastname: Sato
authors.forename: Takashi
authors.initials: T
authors.lastname: Miyazaki
authors.forename: Hideki
authors.initials: H
authors.lastname: Kitaura
authors.forename: Hisako
authors.initials: H
authors.lastname: Kayama
authors.forename: Fumihiko
authors.initials: F
authors.lastname: Miyoshi
authors.forename: Yasuto
authors.initials: Y
authors.lastname: Araki
authors.forename: Yuji
authors.initials: Y
authors.lastname: Akiyama
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Takeda
authors.forename: Toshihide
authors.initials: T
authors.lastname: Mimura
mesh_terms: D000818:Animals; D001862:Bone Resorption / Q000378:metabolism / Q000503:physiopathology*; D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D015850:Interleukin-6 / Q000494:pharmacology* / Q000502:physiology; D008264:Macrophages / Q000166:cytology / Q000187:drug effects*; D051379:Mice; D016328:NF-kappa B / Q000187:drug effects / Q000378:metabolism; D050778:NFATC Transcription Factors / Q000187:drug effects* / Q000378:metabolism; D010010:Osteoclasts / Q000166:cytology / Q000187:drug effects*; D053244:Osteoprotegerin / Q000494:pharmacology / Q000502:physiology; D010880:Piperidines / Q000494:pharmacology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D016760:Proto-Oncogene Proteins c-fos / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D011743:Pyrimidines / Q000494:pharmacology; D011758:Pyrroles / Q000494:pharmacology; D053245:RANK Ligand / Q000494:pharmacology / Q000502:physiology; D050796:STAT3 Transcription Factor / Q000235:genetics / Q000502:physiology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology* / Q000502:physiology
chemical_list: D015850:Interleukin-6; D016328:NF-kappa B; D050778:NFATC Transcription Factors; D053244:Osteoprotegerin; D010880:Piperidines; D047428:Protein Kinase Inhibitors; D016760:Proto-Oncogene Proteins c-fos; D011743:Pyrimidines; D011758:Pyrroles; D053245:RANK Ligand; D050796:STAT3 Transcription Factor; C494087:Stat3 protein, mouse; C506031:Tnfrsf11b protein, mouse; C505689:Tnfsf11 protein, mouse; D014409:Tumor Necrosis Factor-alpha; C479163:tofacitinib