title: Breast cancer increases osteoclastogenesis by secreting M-CSF and upregulating RANKL in stromal cells.
abstract: BACKGROUND
  Breast cancer metastasis to bone causes resorption of the mineralized matrix by osteoclasts. Macrophage colony stimulating factor (M-CSF)and receptor activator of the NF-kappaB ligand (RANKL) are produced by stromal cells and are essential for osteoclast formation. The human breast cancer cell line, MDA-MB-231, reliably forms bone metastases in a murine model and stimulates osteoclast formation in culture. We hypothesized that MDA-MB-231 stimulates osteoclast formation through secretion of M-CSF and/or RANKL.
  
  
  METHODS
  We cocultured MDA-MB-231 and a bone marrow derived cell line, UAMS-33, and evaluated the expression of M-CSF and RANKL mRNA. Osteoclast formation was assessed using these cells added to hematopoietic cell cultures.
  
  
  RESULTS
  MDA-MB-231 exhibited constitutive expression of M-CSF mRNA. As expected, addition of recombinant M-CSF (30 ng/ml) and RANKL (30 ng/ml) to hematopoietic osteoclast precursors supported osteoclast formation, while the addition of soluble RANKL alone or MDA-231 without added RANKL did not. Notably, coculture of MDA-231 with hematopoietic cells and added soluble RANKL stimulated significant osteoclast formation, indicating that MDA-231 served as an effective source for M-CSF. MDA-231 did not express RANKL. However, when cocultured with the murine bone marrow stromal cell line UAMS-33, RANKL expression was significantly increased in the latter cells. MDA-231 also stimulated osteoclast formation in coculture with UAMS-33 and hematopoietic cells.
  
  
  CONCLUSIONS
  We conclude that MDA-MB-231 increases osteoclast formation by secreting adequate amounts of M-CSF protein and enhancing the expression of RANKL by stromal support cells. The ability to stimulate osteoclasts may explain the ability to metastasize to bone.
authors.affiliation: Department of Surgery, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA.
authors.forename: A T
authors.initials: AT
authors.lastname: Mancino
authors.forename: V S
authors.initials: VS
authors.lastname: Klimberg
authors.forename: M
authors.initials: M
authors.lastname: Yamamoto
authors.forename: S C
authors.initials: SC
authors.lastname: Manolagas
authors.forename: E
authors.initials: E
authors.lastname: Abe
mesh_terms: D000818:Animals; D001859:Bone Neoplasms / Q000503:physiopathology / Q000556:secondary*; D001943:Breast Neoplasms / Q000473:pathology* / Q000503:physiopathology; D002117:Calcitriol / Q000494:pharmacology; D002120:Calcium Channel Agonists / Q000494:pharmacology; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D018920:Coculture Techniques; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects / Q000502:physiology; D006412:Hematopoietic Stem Cells / Q000166:cytology; D006801:Humans; D016173:Macrophage Colony-Stimulating Factor / Q000235:genetics / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008806:Mice, Inbred AKR; D008819:Mice, Nude; D010010:Osteoclasts / Q000473:pathology*; D010012:Osteogenesis / Q000502:physiology; D053245:RANK Ligand; D012333:RNA, Messenger / Q000032:analysis; D053246:Receptor Activator of Nuclear Factor-kappa B; D017154:Stromal Cells / Q000166:cytology / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D002120:Calcium Channel Agonists; D002352:Carrier Proteins; D008562:Membrane Glycoproteins; D053245:RANK Ligand; D012333:RNA, Messenger; D053246:Receptor Activator of Nuclear Factor-kappa B; C505686:TNFRSF11A protein, human; C505688:TNFSF11 protein, human; C505687:Tnfrsf11a protein, mouse; C505689:Tnfsf11 protein, mouse; D016173:Macrophage Colony-Stimulating Factor; D002117:Calcitriol