title: High calcium concentration in bones promotes bone metastasis in renal cell carcinomas expressing calcium-sensing receptor.
abstract: BACKGROUND
  The prognosis for renal cell carcinoma (RCC) is related to a high rate of metastasis, including 30% of bone metastasis. Characteristic for bone tissue is a high concentration of calcium ions. In this study, we show a promoting effect of an enhanced extracellular calcium concentration on mechanisms of bone metastasis via the calcium-sensing receptor (CaSR) and its downstream signaling molecules.
  
  
  METHODS
  Our analyses were performed using 33 (11/category) matched specimens of normal and tumor tissue and 9 (3/category) primary cells derived from RCC patients of the 3 categories: non-metastasized, metastasized into the lung and metastasized into bones during a five-year period after nephrectomy. Expression of CaSR was determined by RT-PCR, Western blot analyses and flow cytometry, respectively. Cells were treated by calcium and the CaSR inhibitor NPS 2143. Cell migration was measured in a Boyden chamber with calcium (10 μM) as chemotaxin and proliferation by BrdU incorporation. The activity of intracellular signaling mediators was quantified by a phospho-kinase array and Western blot.
  
  
  RESULTS
  The expression of CaSR was highest in specimens and cells of patients with bone metastases. Calcium treatment induced an increased migration (19-fold) and proliferation (2.3-fold) exclusively in RCC cells from patients with bone metastases. The CaSR inhibitor NPS 2143 elucidated the role of CaSR on the calcium-dependent effects. After treatment with calcium, the activity of AKT, PLCγ-1, p38α and JNK was clearly enhanced and PTEN expression was almost completely abolished in bone metastasizing RCC cells.
  
  
  CONCLUSIONS
  Our results indicate a promoting effect of extracellular calcium on cell migration and proliferation of bone metastasizing RCC cells via highly expressed CaSR and its downstream signaling pathways. Consequently, CaSR may be regarded as a new prognostic marker predicting RCC bone metastasis.
authors.forename: Elke
authors.initials: E
authors.lastname: Joeckel
authors.forename: Tobias
authors.initials: T
authors.lastname: Haber
authors.forename: Dirk
authors.initials: D
authors.lastname: Prawitt
authors.forename: Kerstin
authors.initials: K
authors.lastname: Junker
authors.forename: Christian
authors.initials: C
authors.lastname: Hampel
authors.forename: Joachim W
authors.initials: JW
authors.lastname: Thüroff
authors.forename: Frederik C
authors.initials: FC
authors.lastname: Roos
authors.affiliation: Department of Urology, Johannes Gutenberg University Medical Center, Langenbeckstr 1, Mainz 55131, Germany. brenner@uni-mainz.de.
authors.forename: Walburgis
authors.initials: W
authors.lastname: Brenner
mesh_terms: D015153:Blotting, Western; D001859:Bone Neoplasms / Q000556:secondary*; D001842:Bone and Bones / Q000737:chemistry*; D002118:Calcium / Q000378:metabolism*; D002292:Carcinoma, Renal Cell / Q000378:metabolism* / Q000556:secondary; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D007680:Kidney Neoplasms / Q000378:metabolism* / Q000473:pathology; D044169:Receptors, Calcium-Sensing / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D044169:Receptors, Calcium-Sensing; D002118:Calcium