Phosphatidylinositide 3-kinase (PI3K) modulates the canalicular transporter internalization and the secretory failure induced by estradiol 17ß-glucuronide (E17G): Possible role for PKB

BOAGLIO, A.C.; ZUCCHETTI A.E.; SÁNCHEZ POZZI E.J.; MOTTINO A.D.; CROCENZI F.A.; ROMA M.G.

Copenhague, Dinamara

Congreso; 44th Annual Meeting of the European Association for the Study of the Liver (EASL); 2009

European Association for the Study of the Liver (EASL)

<!-- /* Style Definitions */ p.MsoNormal, li.MsoNormal, div.MsoNormal {mso-style-parent:""; margin:0cm; margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:"Times New Roman"; mso-fareast-font-family:"Times New Roman";} @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;} --> E17G is an endogenous, cholestatic metabolite that induces endocytic internalization of the canalicular transporters relevant to bile secretion Bsep and Mrp2; this may justify its causal association with pregnancy-induced cholestasis. PI3K mediates cholestatic events, e.g. taurolithocholate-induced cholestasis (JBC 278: 17810, 2003). Since common pathomechanisms exist between both kinds of cholestasis, we assessed here whether PI3K is involved in E17G cholestatic effects. E17G (200 µM) activated PI3K from 20 min onwards, as assessed by the phosphorylation of the final PI3K effector Akt in primary cultured hepatocytes. When preadministered to isolated rat hepatocyte couplets (IRHCs), the PI3K inhibitor wortmannin (WM; 100 nM) prevented partially the reduction induced by E17G (12.5-800 µM) in the proportion of IRHCs accumulating apically the fluorescent Bsep and Mrp2 substrates cholyllysylfluorescein and glutathionylmethylfluorescein, respectively. The alternative PI3K inhibitor LY294002 (50 µM) and the ?Akt inhibitor? (Calbiochem® #124005; 20 µM) showed similar protective effects. Immunostaining of Bsep and Mrp2 in IRHC followed by confocal microscopy and image analysis revealed that E17G induces endocytic internalization of the transporters. This redistribution was extensively prevented by WM. To evaluate this phenomenon in a more physiological model, we assessed the anticholestatic effect of WM in perfused rat livers. A bolus, intraportal injection of E17G (2 µmol) induced an acute decrease in bile flow within 10 min, which did not recover during the remaining perfusion period (Figure 1). WM (200 nM) did not prevent this initial decay, but greatly accelerated the recovery to normality of bile flow. A similar behaviour was observed for the biliary excretion of the Bsep and Mrp2 substrates [3H]taurocholate and glutathione, respectively. We conclude that PI3K/Akt signalling pathway is involved in the the biliary secretory failure induced by E17G by producing a sustained internalization of canalicular transporters. PI3K/Akt may therefore be a potential therapeutic target in cholestatic phenomena associated to estrogens.