Estradiol 17-glucuronide (E217G)-induced internalization of the canalicular bile salt export pump, Bsep, in the rat: Its role in E217G-induced cholestasis and its prevention by dibutyryl cyclic AMP (DB-cAMP)

CROCENZI, FERNANDO A.; VEGGI, LUIS M.; CAO, JINGSONG; VORE, MARY; COLEMAN, ROGER; MOTTINO, ALDO D.; ROMA, MARCELO G

Congreso; 37th Annual Meeting of the European Association for the Study of the Liver (EASL); 2002

This study was aimed to establish the role of changes in the cellular localization of Bsep in the acute cholestasis induced by E217G, and the ability of the vesicular-transport stimulator, DBcAMP, to prevent these alterations. We analyzed in Wistar rats both bile flow (BF) and subcellular localization of Bsep (by Western Blotting and immunohistochemistry, followed by confocal microscopy), at the peak of E217G (15 micromol/kg, i.v.)-induced cholestasis, i.e. 20 min after its administration. We also evaluated the effect of DBcAMP (20 micromol/kg, i.v.), administered 30 min before E217G injection. E217G diminished BF by 87±17% (p<0.05 vs controls), and induced relocalization of Bsep from the canalicular membrane to intracellular vesicles, as assessed by the increase in Bsep content in microsomal membranes (+55±18%; p<0.05). This relocalization was further confirmed by confocal microscopy, showing loss of Bsep in the canalicular membrane and its simultaneous appearance in intracellular vesicles. In DBcAMP-pretreated rats, E217G diminished BF by only 44±22% (p<0.05 vs either E217G or controls), and substantially prevented Bsep internalization. In "in vitro" studies using isolated rat hepatocyte couplets, E217G (50 microM, 20 min) significantly diminished the percentage of couplets accumulating the fluorescent bile-salt-analogue, cholyl-lysyl-fluorescein, into their canalicular vacuoles (-57±3%; p<0.05), and Bsep retrieval from the canalicular membrane; DBcAMP (10 microM) almost completely prevented both deleterious effects. This suggests that changes in Bsep localization may be involved in E217G-induced impairment of BF and bile salt transport, and that DBcAMP prevents this effect, probably by stimulating insertion of transporter-containing vesicles.