In vivo inhibition of bilirubin conjugation does not induce a loss of its protective effect in oxidative stress-induced cholestasis

MARTÍN P.L.; TAURIZANO D.; RAZORI M.V.; ARRIAGA S.M.; PISANI G.B.; SÁNCHEZ POZZI E.J.; ROMA M.G.; BASIGLIO C.L.

Rosario, Santa Fe

Congreso; Reunión Anual de la Sociedad Argentina de Fisiología (SAFIS) 2019; 2019

Sociedad Argentina de Fisiología (SAFIS) 2019

Our group proved that induction of hemeoxygenase 1 (HO1) and consequent elevation of bilirubin (BR) protect the liver from oxidative injury occurring in cholestatic diseases. To assess whether BR thus generated needs to be conjugated to exert its protective effect, we aimed to impair BR conjugation by depleting the endogenous pool of UDP, cofactor of UDP-glucuronosyl transferase (UGT), key enzyme in BR conjugation. Male Wistar rats were used throughout. Oxidative cholestatic injury was induced by tert-butyl hydroperoxide (T, 440 μmol/kg b.w., i.p.). Induction of HO1 and inhibition of UGT were achieved by administration of hemin (H, 20 mg/kg b.w., i.p.) and galactosamine (G, 200 mg/kg b.w., i.p.), respectively. Results are expressed as media±SD. Bile flow (μl/min/g liver) decreased 4-6 h post T injection (1.65±0.73 vs 3.12±0.51 for control -T vehicle-, p<0.05, n=6); pre-treatment with H completely prevented this reduction (2.89±0.84, p<0.05 vs T, n=6). G administration to HT animals did not impair the protective effect of H on bile flow (2.33±0.83, p>0.05 vs HT, n=6), thus evidencing that BR conjugation is not crucial for its protective effect on biliary function. Biliary excretion of BR (mg/min/g liver) decreased after T (0.48±0.31 vs 1.10±0.16 for control, p<0.05, n=6) and this decrease was prevented by H (0.95±0.21, p<0.05 vs T, n=6). As expected, pre-treatment with G induced a decrease in biliary excretion of BR (0.53±0.13, p<0.05 vs HT, n=6), since BR conjugation is impaired and, consequently, BR cannot be excreted into bile. Lipid peroxidation (pmol MDA/mg protein) increased after T treatment (3.53±0.38 vs 1.78±0.44 for control, p<0.05, n=6) while H prevented that increase (1.82±0.61, p<0.05 vs T, n=6), likely due to the elevated levels of antioxidant BR generated by HO1 induction. G did not produce a decline in the antioxidant protective effect of BR (2.23±0.36 p>0.05 vs HT, n=6), likely as a consequence of its accumulation in liver tissue. We conclude that BR conjugation is not essential for BR to exert its antioxidant properties. These results are in line with previous findings of our group as regards the in vitro antioxidant action of unconjugated BR.