Resveratrol modifies biliary secretion of cholephilic compounds in sham-operated and cholestatic rats

AIM To investigate the effect of resveratrol on biliary secretion of cholephilic compounds in healthy and bile duct-obstructed rats. METHODS Resveratrol(RSV) or saline were administered to rats by daily oral gavage for 28 d after sham operation or reversible bile duct obstruction(BDO). Bile was collected 24 h after the last gavage during an intravenous bolus dose of the Mdr1/Mrp2 substrate azithromycin. Bile acids,glutathione and azithromycin were measured in bile to quantify their level of biliary secretion. Liver expression of enzymes and transporters relevant for bile production and biliary secretion of major bile constituents and drugs were analyzed at the m RNA and protein levels using q RT-PCR and Western blot analysis,respectively. The TR-FRET PXR Competitive Binding Assay kit was used to determine the agonism of RSV at the pregnane X receptor. RESULTS RSV increased bile flow in sham-operated rats due to increased biliary secretion of bile acids(BA) and glutathione. This effect was accompanied by the induction of the hepatic rate-limiting transporters for bile acids and glutathione,Bsep and Mrp2,respectively. RSV also induced Cyp7 a1,an enzyme that is crucial for bile acid synthesis; Mrp4,a transporter important for BA secretion from hepatocytes to blood; and Mdr1,the major apical transporter for xenobiotics. The findings were supported by increased biliary secretion of azithromycin. The TR-FRET PXR competitive binding assay confirmed RSV as a weak agonist of the human nuclear receptor PXR,which is a transcriptional regulator of Mdr1/Mrp2. RSV demonstrated significant hepatoprotective properties against BDO-induced cirrhosis. RSV also reduced bile flow in BDO rats without any corresponding change in the levels of the transporters and enzymes involved in RSV-mediated hepatoprotection. CONCLUSION Resveratrol administration for 28 d has a distinct effect on bile flow and biliary secretion of cholephilic compounds in healthy and bile duct-obstructed rats.

The 3’-untranslated region contributes to the pregnane X receptor(PXR) expression downregulation by PXR ligands and up-regulation by glucocorticoids

Pregnane X receptor(PXR)is the major regulator of xenobiotic metabolism.PXR itself is controlled by various signaling molecules including glucocorticoids.Moreover,negative feed-back regulation has been proposed at the transcriptional level.We examined the involvement of the 3’-untranslated region(3’-UTR)of NR1I2 mRNA and microRNAs in PXR-and glucocorticoid receptor(GR)-mediated regulation of NR1I2 gene expression.PXR ligands were found to significantly downregulate NR1I2 mRNA expression in a set of 14 human hepatocyte cultures.Similarly,PXR was downregulated by PCN in the C57/BL6 mice liver.In mechanistic studies with the full-length 3’-UTR cloned into luciferase reporter or expression vectors,we showed that the 3’-UTR reduces PXR expression.From the miRNAs tested,miR-18a-5p inhibited both NR 112 expression and CYP3A4 gene induction.Importantly,we observed significant upregulation of miR-18a-5p expression 6 h after treatment with the PXR ligand rifampicin,which indicates a putative mechanism underlying NR1I2 negative feed-back regulation in hepatic cells.Additionally,glucocorticoids upregulated NR1I2 expression not only through the promoter region but also via 3’-UTR regulation,which likely involves downregulation of miR-18a-5p.We conclude that miR-18a-5p is involved in the down-regulation of NR1I2expression by its ligands and in the upregulation of NR1I2 mRNA expression by glucocorticoids in hepatic cells.