title: Autocrine/paracrine stimulation of purinergic receptors in osteoblasts: contribution of vesicular ATP release.
abstract: Extracellular nucleotides such as ATP and UTP are released in response to mechanical stimulation in different cell systems. It is becoming increasingly evident that ATP release plays a role in autocrine and paracrine stimulation of osteoblasts. Mechanical stimulation, as shear stress, membrane stretch or hypo-osmotic swelling, as well as oscillatory fluid flow, stimulates ATP release from different osteoblastic cell lines. Human osteoblast-like initial transfectant (HOBIT) cells release ATP in response to mechanical stimulation. In the present study, we show that HOBIT cells are activated by nanomolar levels of extracellular ATP, concentrations that can be detected under resting conditions and increase following hypotonic shock. Cell activation by hypotonic medium induced intracellular Ca2+ oscillations, and Egr-1 synthesis and DNA-binding activity. Quinacrine staining of living, resting cells revealed a granular fluorescence, typical of ATP-storing vesicles. Monensin prevented quinacrine staining and considerably inhibited hypotonic-induced ATP release. Finally, elevated levels of cytosolic Ca2+ activated massive ATP release and a dose-dependent loss of quinacrine granules. The contribution of a vesicular mechanism for ATP release is proposed to sustain paracrine osteoblast activation.
authors.affiliation: Department of Biochemistry, University of Trieste, via L. Giorgeri 1, 34100 Trieste, Italy.
authors.forename: Milena
authors.initials: M
authors.lastname: Romanello
authors.forename: Andrea
authors.initials: A
authors.lastname: Codognotto
authors.forename: Massimiliano
authors.initials: M
authors.lastname: Bicego
authors.forename: Alex
authors.initials: A
authors.lastname: Pines
authors.forename: Gianluca
authors.initials: G
authors.lastname: Tell
authors.forename: Paola
authors.initials: P
authors.lastname: D'Andrea
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D010006:Osteoblasts / Q000378:metabolism*; D011796:Quinacrine / Q000378:metabolism; D011983:Receptors, Purinergic / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D011983:Receptors, Purinergic; D000255:Adenosine Triphosphate; D011796:Quinacrine