Galactosamine (GAL) prevents ethinylestradiol (EE)-induced cholestasis.

SÁNCHEZ POZZI, ENRIQUE J; CROCENZI, FERNANDO A; PELLEGRINO, JOSÉ M; CATANIA, VIVIANA A.; LUQUITA, MARCELO G.

Salvador

Congreso; Biennial Scientific Meeting of the International Association for the Study of the Liver (IASL); 2004

International Association for the Study of the Liver (IASL)

EE-17ß-glucuronide (EE17G), a potent cholestatic agent, is postulated to mediate EE cholestasis. Previously, we demonstrated that ursodeoxycholate prevention of EE cholestasis involves a decrease in EE17G formation (JPET 306:279-286, 2003) and in this study we further investigate the role of glucuronidation, evaluating the effect of gal, a drug that depletes cells from UDP-glucuronic acid (UDPGA), on this model of cholestasis. Uridine (Ur) was used to restore UDPGA levels. Methods: In preliminary experiments, gal in a dose of 0.2 g/kg bw reduced hepatic UDPGA levels (µmol/g liver) from 0.42±0.03 to 0.06±0.01, 2 hs after injection whereas Ur 0.9 g/kg bw prevented this decrease (0.76±0.11). Male Wistar rats were divided into 6 groups: 3 groups received for 5 days EE (5 mg/kg bw, sc), one of them was co-administered with gal (0.2 g/kg bw, ip, EG), another received gal+Ur (0.9 g/kg bw, ip, EGU), and the third received solvent ip (E). The other 3 groups received solvent sc plus an ip administration of either solvent (C), gal (G) or Ur (U). Eighteen hs after the last injection, bile flow (BF) and biliary excretion of bile salt (BSE) and gluthathione (GSHE) were assessed. Plasma was obtained to determine ASAT and ALAT. Western blot analysis of UGT2B1, isozyme that converts EE into EE17G, was performed in liver microsomes. In a set of animals from all groups, tauroursodeoxycholate (TUDC) was infused at increasing rates to evaluate its canalicular transport maximum (TUDC TM). The data from this last experiment were plotted as BF vs. BSE to estimate the bile salt-independent fraction of bile flow (BSIBF). Results: Gal and Ur per se did not affect any of the parameters studied. Gal did not increase serum levels of ASAT and ALAT indicating that the dose was not hepatotoxic neither alone nor co-administered with EE. Western blot of UGT2B1: EE decreased enzyme protein content by a 45% but this was not modified by gal or gal+Ur. Biliary studies: BF and BSE declines produced by EE were significantly prevented by gal and were not modified by gal+Ur (BF (µl/min.g liver): C: 2.29±0.14; EE: 0.94±0.05a; EG: 1.27±0.08a,b, EGU: 0.99±0.07a,c; BSE (nmol/min.g liver): C: 52±4; E: 28±2a; EG: 42±2b; EGU: 37±4a). GSHE (nmol/min.g liver) was not restored by gal (C: 8.7±0.3; E: 0.3±0.2a; EG: 0.4±0.4a; EGU: 0.4±0.4a). EE decreased TUDC TM (nmol/min.g liver), gal partially restored it, and Ur reverted gal effect (C: 898±28; E: 112±16a; EG: 249±18a,b; EGU: 139±11a,c). Finally, BSIBF estimation (nmol/min g liver) showed a significant alteration by EE that was prevented by gal (C: 1.93±0.15; E: 0.61±0.09a; EG: 1.01±0.05a,b; EGU: 0.73±0.05a,c). a, b and  c: significantly different from C, E and EG, respectively (p<0.05). Conclusion: Glucuronidation of EE represents a key step in cholestasis. EE17G seems to be partially responsible for the decrease in BF, in the canalicular transport of bile salts and in the BSIBF, in spite of the lack of improvement of GSH excretion.