Hormonal modulation of hepatic cAMP prevents estradiol 17β-D-glucuronide-induced cholestasis in perfused rat liver.
BACKGROUND
Estradiol-17β-D-glucuronide (E17G) induces cholestasis in vivo, endocytic internalization of the canalicular transporters multidrug resistance-associated protein 2 (Abcc2) and bile salt export pump (Abcb11) being a key pathomechanism. Cyclic AMP (cAMP) prevents cholestasis by targeting these transporters back to the canalicular membrane. In hepatocyte couplets, glucagon and salbutamol, both of which increase cAMP, prevented E17G action by stimulating the trafficking of these transporters by different mechanisms, namely: glucagon activates a protein kinase A-dependent pathway, whereas salbutamol activates an exchange-protein activated by cAMP (Epac)-mediated, microtubule-dependent pathway.


METHODS
The present study evaluated whether glucagon and salbutamol prevent E17G-induced cholestasis in a more physiological model, i.e., the perfused rat liver (PRL). Additionally, the preventive effect of in vivo alanine administration, which induces pancreatic glucagon secretion, was evaluated.


RESULTS
In PRLs, glucagon and salbutamol prevented E17G-induced decrease in both bile flow and the secretory activity of Abcc2 and Abcb11. Salbutamol prevention fully depended on microtubule integrity. On the other hand, glucagon prevention was microtubule-independent only at early time periods after E17G administration, but it was ultimately affected by the microtubule disrupter colchicine. Cholestasis was associated with endocytic internalization of Abcb11 and Abcc2, the intracellular carriers being partially colocalized with the endosomal marker Rab11a. This effect was completely prevented by salbutamol, whereas some transporter-containing vesicles remained colocalized with Rab11a after glucagon treatment. In vivo, alanine administration increased hepatic cAMP and accelerated the recovery of bile flow and Abcb11/Abcc2 transport function after E17G administration. The initial recovery afforded by alanine was microtubule-independent, but microtubule integrity was required to sustain this protective effect.


CONCLUSIONS
We conclude that modulation of cAMP levels either by direct administration of cAMP modulators or by physiological manipulations leadings to hormone-mediated increase of cAMP levels (alanine administration), prevents estrogen-induced cholestasis in models with preserved liver architecture, through mechanisms similar to those arisen from in vitro studies.
Instituto de Fisiología Experimental (IFISE), Facultad de Ciencias Bioquímicas y Farmacéuticas (CONICET, U.N.R.), Suipacha 570, S2002LRL, Rosario, Argentina.
Andrés E
AE
Zucchetti
Ismael R
IR
Barosso
Andrea C
AC
Boaglio
Marcelo G
MG
Luquita
Marcelo G
MG
Roma
Fernando A
FA
Crocenzi
Enrique J
EJ
Sánchez Pozzi
D000074020:ATP Binding Cassette Transporter, Subfamily B, Member 11; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D058666:Adrenergic beta-2 Receptor Agonists / Q000627:therapeutic use*; D000409:Alanine / Q000627:therapeutic use; D000420:Albuterol / Q000627:therapeutic use*; D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002779:Cholestasis / Q000209:etiology / Q000378:metabolism / Q000517:prevention & control*; D000242:Cyclic AMP / Q000819:agonists* / Q000378:metabolism; D004958:Estradiol; D005260:Female; D005934:Glucagon / Q000627:therapeutic use*; D006728:Hormones / Q000627:therapeutic use*; D008099:Liver / Q000378:metabolism / Q000503:physiopathology; D051381:Rats; D017208:Rats, Wistar; D016896:Treatment Outcome; D020691:rab GTP-Binding Proteins / Q000378:metabolism; D000099193:rab11 GTP-Binding Proteins
D000074020:ATP Binding Cassette Transporter, Subfamily B, Member 11; D018528:ATP-Binding Cassette Transporters; D058666:Adrenergic beta-2 Receptor Agonists; D000409:Alanine; D000420:Albuterol; D015415:Biomarkers; D000242:Cyclic AMP; D004958:Estradiol; D005934:Glucagon; D006728:Hormones; D020691:rab GTP-Binding Proteins; D000099193:rab11 GTP-Binding Proteins; C490548:Abcb11 protein, rat; C423013:Abcc2 protein, rat