Modulation of organic anion transporting polypeptide 1 and multidrug resistance protein 3 expression in the liver and kidney of Gunn rats.

Background: Gunn rat is an animal model of Crigler-Najjar syndrome (CNS) type I that develops jaundice due to defect of bilirubin conjugation. Bilirubin UDP-glucuronosyltransferase (UGT1A1), which plays a critical role in bilirubin glucuronidation, has been reported to be deficient in CNS type 1. On the other hand, little is known about the expression of organic anion transporters in Gunn rats. In the present study, we evaluated expressions of organic anion transporting polypeptide (oatp) 1 and 2, multidrug resistance-associated protein (mrp) 2 and mrp3 in the liver and kidney of Gunn rats. Methods: Serum samples, liver and kidney tissues were obtained from Gunn rats and normal SD rats ( [Formula: see text], in each group). Semi-quantitative mRNA expression of oatp1, oatp2, mrp2, and mrp3 mRNA was evaluated by constructed reverse transcription-polymerase chain reaction (RT-PCR). Protein expressions were determined by Western blotting and by immunohistochemistry. Results: Marked elevation of serum unconjugated bilirubin concentration ( [Formula: see text] micromol/l) was observed in Gunn rats. Hepatic expression of oatp1 and oatp2 mRNA was 44% ( [Formula: see text] ) and 35% ( [Formula: see text] ) lower in Gunn rats than in SD rats, respectively. Hepatic oatp1 protein expression was 37% ( [Formula: see text] ) lower in Gunn rats than in SD rats. In contrast to oatp1, hepatic expression of mrp3 mRNA and protein was 76% ( [Formula: see text] ) and 557% ( [Formula: see text] ) higher in Gunn rats than in SD rats, respectively. Hepatic expression of oatp2 and mrp2 protein was not significantly different between Gunn rats and SD rats. Like the Western blot analysis, immunohistochemical staining disclosed decrease of oatp1 and increase of mrp3 protein expressions in the liver of Gunn rats. Decrease of oatp1 and increase of mrp3 expressions were also observed in the kidney of Gunn rats. Conclusion: Decreased expression of oatp1 and increased expression of mrp3 were observed in the liver and kidney of Gunn rats. Deficient UGT1A1 activity-associated retention of unconjugated bilirubin in the hepatocytes may modulate the expressions of these transporters in Gunn rats.

Dietary iron restriction improves aminotransferase levels in chronic hepatitis C patients.

BACKGROUND/AIMS: It is generally accepted that iron overload plays an important role in the pathogenesis of liver cell injury in chronic hepatitis C. The present study was undertaken to evaluate whether low-iron diet improves liver function tests in patients with chronic hepatitis C. METHODOLOGY: Seventeen patients with chronic hepatitis C (13 men and 4 women, 54 +/- 14 years old) that did not respond to, or were unsuitable for interferon therapy, were enrolled in this study. All patients had been pretreated with ursodeoxycholic acid for more than 12 months before the beginning of the study. Dietary iron intake was restricted to less than 7 mg/day, and the patients were followed up for 18 months. RESULTS: Mean daily iron intakes, calculated from food records, were 5.9 and 6.4 mg after 6 and 12 months, respectively. The mean serum ferritin decreased significantly from 362 ng/mL at entry to 179 ng/mL after 18 months. The serum unsaturated iron binding capacity level increased significantly from 163 micrograms/dL at entry to 203 micrograms/dL after 18 months. The serum aspartate aminotransferase decreased significantly from 62 IU/L at entry to 47 IU/L after 18 months, and serum alanine aminotransferase from 68 IU/L at entry to 53 IU/L after 18 months. Serum iron, hepatitis C virus-RNA titer and platelet count remained unchanged throughout the study. CONCLUSIONS: These results suggest that iron-restricted diet may be an important therapeutic modality for improving liver injury in patients with chronic hepatitis C.

Increased renal expression of bilirubin glucuronide transporters in a rat model of obstructive jaundice.

Regulation of bilirubin glucuronide transporters during hyperbilirubinemia in hepatic and extrahepatic tissues is not completely clear. In the present study, we evaluated the regulation of the bilirubin glucuronide transporters, multidrug resistance-associated proteins (MRP)2 and 3, in rats with obstructive jaundice. Bile duct ligation (BDL) or sham operation was performed in Wistar rats. Liver and kidneys were removed 1, 3, and 5 days after BDL (n = 4, in each group). Serum and urine were collected to measure bilirubin levels just before animal killing. MRP2 And MRP3 mRNA expressions were determined by real-time RT-PCR. Protein expression of MRP2 and MRP3 was determined by Western blotting. Renal MRP2 function was evaluated by para-aminohippurate (PAH) clearance. The effect of conjugated bilirubin, unconjugated bilirubin, human bile, and sulfate-conjugated bile acid on MRP2 gene expression was also evaluated in renal and hepatocyte cell lines. Serum bilirubin and urinary bilirubin excretion increased significantly after BDL. In the liver, the mRNA expression of MRP2 decreased 59, 86, and 82%, and its protein expression decreased 25, 74, and 93% compared with sham-operated animals after 24, 72, and 120 h of BDL, respectively. In contrast, the liver expression of MRP3 mRNA increased 138, 2,137, and 3,295%, and its protein expression increased 560, 634, and 612% compared with sham-operated animals after 24, 72, and 120 h of BDL, respectively. On the other hand, in the kidneys, the mRNA expression of MRP2 increased 162, 73, and 21%, and its protein expression increased 387, 558, and 472% compared with sham-operated animals after 24, 72, and 120 h of BDL, respectively. PAH clearance was significantly increased after BDL. The mRNA expression of MRP2 increased in renal proximal tubular epithelial cells after treatment with conjugated bilirubin, sulfate-conjugated bile acid or human bile. Upregulation of MRP2 in the kidneys and MRP3 in the liver may be a compensatory mechanism to improve bilirubin clearance during obstructive jaundice.

Increased expression of multidrug resistance-associated protein 1 (mrp1) in hepatocyte basolateral membrane and renal tubular epithelia after bile duct ligation in rats.

Components of the multidrug resistance-associated protein (mrp) family mediate the adenosine triphosphate (ATP)-dependent transport of conjugated organic anions in the liver. Of these, mrp1 and mrp2 have been shown to have similar substrate specificity and nucleotide sequence. The intracellular localization and distribution of mrp1 under normal condition and cholestasis have not been as yet completely elucidated. To clarify this point, in the present study we evaluated the intracellular localization of mrp1 in rat liver and kidney after bile duct ligation (BDL). Bile duct was ligated in Wistar rats. Sequential staining of mrp1 by immunofluorescence was carried out in rat liver and kidneys 1, 3, and 5 days after bile duct ligation using confocal laser scanning microscopy. Weak granular staining of mrp1 was observed in cytoplasm of control rat hepatocytes. In addition to increased cytoplasm staining of mrp1, belt-and granule-like staining of mrp1 in basolateral membrane of hepatocytes was also shown after BDL. Furthermore, mrp1 immunofluorescence increased over time after BDL. No specific immunoflurescence of mrp1 was detected in control rat kidney. However, mrp1-positive staining was observed in epithelia of some renal tubules after BDL. This study showed that mrp1 immunofluorescence increased in hepatocyte basolateral membrane and cytoplasm and epithelia of some renal tubules after BDL. This increased mrp1 expression may be an adaptive response to impairment of hepato-biliary organic anion transport during obstructive cholestasis.