title: Osteotropism of neuroendocrine tumors: role of the CXCL12/ CXCR4 pathway in promoting EMT in vitro.
abstract: Neuroendocrine tumors (NETs) metastasize to the skeleton in approximately 20% of patients. We have previously shown that the epithelial-mesenchymal transition (EMT) regulates the NET osteotropism and that CXCR4 overexpression predicts bone spreading. Here, we unravel the molecular mechanisms linking the activation of the CXCL12/CXCR4 axis to the bone colonization of NETs using cell lines representative of pancreatic (BON1, CM, QGP1), intestinal (CNDT 2.5), and bronchial origin (H727). By combining flow cytometry and ELISA, BON1, CM and QGP1 cells were defined as CXCR4high/CXCL12low, while H727 and CNDT 2.5 were CXCR4low/CXCL12high. CXCL12 was inert on cell proliferation, but significantly increased the in vitro osteotropism of CXCR4high/CXCL12low cells, as assessed by transwell assays with or without Matrigel membranes. In these cells, CXCL12 induced in vitro a marked EMT-like transcriptional shift with acquirement of a mesenchymal shape. The nuclei of CXCR4high/CXCL12low NET cells were typically enriched in non-phosphorylated CXCR4, particularly upon agonist stimulation. Silencing of CXCR4 via siRNA prevented the CXCL12-induced EMT in CXCR4high/CXCL12low NET cell lines resulting in the abrogation of both migration and transcriptional mesenchymal patterns. Our data suggest that CXCL12 conveys EMT-promoting signals in NET cells through CXCR4, which in turn regulates transcriptional, morphologic and functional modifications resulting in enhanced in vitro osteotropism of NET cells. Unique functions of CXCR4 may be segregated in relation to its subcellular localization and may acquire potential relevance in future in vivo studies.
authors.affiliation: Department of Biomedical Sciences and Human Oncology, University of Bari "A. Moro", Bari, Italy.
authors.forename: Mauro
authors.initials: M
authors.lastname: Cives
authors.affiliation: Department of Biomedical Sciences and Human Oncology, University of Bari "A. Moro", Bari, Italy.
authors.forename: Davide
authors.initials: D
authors.lastname: Quaresmini
authors.affiliation: Department of Biomedical Sciences and Human Oncology, University of Bari "A. Moro", Bari, Italy.
authors.forename: Francesca Maria
authors.initials: FM
authors.lastname: Rizzo
authors.affiliation: Department of Biomedical Sciences and Human Oncology, University of Bari "A. Moro", Bari, Italy.
authors.forename: Claudia
authors.initials: C
authors.lastname: Felici
authors.affiliation: Department of Biomedical Sciences and Human Oncology, University of Bari "A. Moro", Bari, Italy.
authors.forename: Stella
authors.initials: S
authors.lastname: D'Oronzo
authors.affiliation: Department of Biomedical Sciences and Human Oncology, University of Bari "A. Moro", Bari, Italy.
authors.forename: Valeria
authors.initials: V
authors.lastname: Simone
authors.affiliation: Department of Biomedical Sciences and Human Oncology, University of Bari "A. Moro", Bari, Italy.
authors.forename: Franco
authors.initials: F
authors.lastname: Silvestris
mesh_terms: D001859:Bone Neoplasms / Q000276:immunology* / Q000556:secondary; D001842:Bone and Bones / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement; D054377:Chemokine CXCL12 / Q000378:metabolism*; D058750:Epithelial-Mesenchymal Transition; D006801:Humans; D008648:Mesoderm / Q000502:physiology; D009362:Neoplasm Metastasis; D018358:Neuroendocrine Tumors / Q000276:immunology* / Q000473:pathology; D010766:Phosphorylation; D019718:Receptors, CXCR4 / Q000378:metabolism*; D015398:Signal Transduction; D059467:Transcriptome; D018523:Tropism
chemical_list: D054377:Chemokine CXCL12; D019718:Receptors, CXCR4
keywords: GPCR nuclear translocation; SDF-1; bone metastasis; carcinoid; epithelial-mesenchymal transition