INVESTIGADORES
ESPELT Maria Victoria
congresos y reuniones científicas
Título:
ATP induce ATP release in hepatoma cells
Autor/es:
ESPELT MV; SANCHEZ ALBERTI G; CHARA O; SCHWARZBAUM PJ
Lugar:
Tarragona, Espana
Reunión:
Congreso; Purines 2010; 2010
Resumen:
<!-- /* Style Definitions */ p.MsoNormal, li.MsoNormal, div.MsoNormal {mso-style-parent:""; margin:0cm; margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman"; mso-fareast-font-family:"Times New Roman"; mso-ansi-language:EN-US; mso-fareast-language:ES-AR;} @page Section1 {size:595.3pt 841.9pt; margin:70.85pt 3.0cm 70.85pt 3.0cm; mso-header-margin:35.4pt; mso-footer-margin:35.4pt; mso-paper-source:0;} div.Section1 {page:Section1;} --> Background: In hepatic cells extracellular nucleotides act via P receptors regulating different physiological responses as: glucogenolysis, gluconeogenesis, canalicular contraction, etc. The aim of our study was to analyze how extracellular ATP (ATPe) activates ATP release in HepG2 cells. Methods: ATPe was quantified using the luciferin-luciferase method. Intracellular calcium was measured by fluorescence microscopy. Results: After addition of exogenous ATP, ATPe rised to a maximum followed by an exponential decay. The maximum reached was 43.1±3.01 nM (with 50 nM ATP added; n=6), 135.0±19.25 nM (100 nM ATP; n=7), 269.0±26.78 nM (200 nM ATP; n=7), and 555.8±39.70 nM (400 nM, n=5). When cells were preincubated with cibacron blue and suramin (P2 blockers), 400 nM ATP induced 426.0±33.02 nM ATPe. ATP release increased to 823.4±36.16 nM when cells were stimulated with 400 nM ATP (n=4) and preincubated with PI3K inhibitors, being this release inhibited with P2 inhibitors (640.2±71.45 nM). Regarding potential 2nd messengers activating ATP release, we determined the percentage of cells showing at least one spike in Ca2+ after incubation with different ATPe concentrations. The spike frequency augmented hyperbolically with ATPe concentration with a K0.5=3.8 uM. Conclusion: ATP induce ATP release in HepG2 cells in concentrations between 100-400 nM. The release is further stimulated with PI3K inhibitors and inhibited with cibacron blue and suramin. ATPe increases Ca2+ spike frequency, which in principle could lead to ATP secretion.