title: Trophic activity of human P2X7 receptor isoforms A and B in osteosarcoma.
abstract: The P2X7 receptor (P2X7R) is attracting increasing attention for its involvement in cancer. Several recent studies have shown a crucial role of P2X7R in tumour cell growth, angiogenesis and invasiveness. In this study, we investigated the role of the two known human P2X7R functional splice variants, the full length P2X7RA and the truncated P2X7RB, in osteosarcoma cell growth. Immunohistochemical analysis of a tissue array of human osteosarcomas showed that forty-four, of a total fifty-four tumours (81.4%), stained positive for both P2X7RA and B, thirty-one (57.4%) were positive using an anti-P2X7RA antibody, whereas fifteen of the total number (27.7%) expressed only P2X7RB. P2X7RB positive tumours showed increased cell density, at the expense of extracellular matrix. The human osteosarcoma cell line Te85, which lacks endogenous P2X7R expression, was stably transfected with either P2X7RA, P2X7RB, or both. Receptor expression was a powerful stimulus for cell growth, the most efficient growth-promoting isoform being P2X7RB alone. Growth stimulation was matched by increased Ca(2+) mobilization and enhanced NFATc1 activity. Te85 P2X7RA+B cells presented pore formation as well as spontaneous extracellular ATP release. The ATP release was sustained in all clones by P2X7R agonist (BzATP) and reduced following P2X7R antagonist (A740003) application. BzATP also increased cell growth and activated NFATc1 levels. On the other hand cyclosporin A (CSA) affected both NFATc1 activation and cell growth, definitively linking P2X7R stimulation to NFATc1 and cell proliferation. All transfected clones also showed reduced RANK-L expression, and an overall decreased RANK-L/OPG ratio. Mineralization was increased in Te85 P2X7RA+B cells while it was significantly diminished in Te85 P2X7RB clones, in agreement with immunohistochemical results. In summary, our data show that the majority of human osteosarcomas express P2X7RA and B and suggest that expression of either isoform is differently coupled to cell growth or activity.
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, Section of Experimental Pathology, Oncology and Biology, University of Ferrara, Ferrara, Italy.
authors.forename: Anna Lisa
authors.initials: AL
authors.lastname: Giuliani
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, Section of Experimental Pathology, Oncology and Biology, University of Ferrara, Ferrara, Italy.
authors.forename: Davide
authors.initials: D
authors.lastname: Colognesi
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, Section of Experimental Pathology, Oncology and Biology, University of Ferrara, Ferrara, Italy.
authors.forename: Tiziana
authors.initials: T
authors.lastname: Ricco
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, Section of Experimental Pathology, Oncology and Biology, University of Ferrara, Ferrara, Italy.
authors.forename: Carlotta
authors.initials: C
authors.lastname: Roncato
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, Section of Experimental Pathology, Oncology and Biology, University of Ferrara, Ferrara, Italy.
authors.forename: Marina
authors.initials: M
authors.lastname: Capece
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, Section of Experimental Pathology, Oncology and Biology, University of Ferrara, Ferrara, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Amoroso
authors.affiliation: Centre For Tissue Injury and Repair, The University of Manchester, Manchester, United Kingdom.
authors.forename: Qi Guang
authors.initials: QG
authors.lastname: Wang
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, Section of Experimental Pathology, Oncology and Biology, University of Ferrara, Ferrara, Italy.
authors.forename: Elena
authors.initials: E
authors.lastname: De Marchi
authors.affiliation: The Mellanby Center for Bone Research, the University of Sheffield, Sheffield, United Kingdom.
authors.forename: Allison
authors.initials: A
authors.lastname: Gartland
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, Section of Experimental Pathology, Oncology and Biology, University of Ferrara, Ferrara, Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Di Virgilio
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, Section of Experimental Pathology, Oncology and Biology, University of Ferrara, Ferrara, Italy.
authors.forename: Elena
authors.initials: E
authors.lastname: Adinolfi
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000293:Adolescent; D000328:Adult; D001859:Bone Neoplasms / Q000235:genetics / Q000378:metabolism*; D002113:Calcification, Physiologic; D045744:Cell Line, Tumor; D002648:Child; D002675:Child, Preschool; D005110:Extracellular Space / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D008875:Middle Aged; D050778:NFATC Transcription Factors / Q000378:metabolism; D053244:Osteoprotegerin / Q000378:metabolism; D012516:Osteosarcoma / Q000235:genetics / Q000378:metabolism*; D020033:Protein Isoforms; D053245:RANK Ligand / Q000378:metabolism; D058486:Receptors, Purinergic P2X7 / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D055815:Young Adult
chemical_list: D050778:NFATC Transcription Factors; D053244:Osteoprotegerin; D020033:Protein Isoforms; D053245:RANK Ligand; D058486:Receptors, Purinergic P2X7; D000255:Adenosine Triphosphate