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:
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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.