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

Abstract

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.

Mrp4−/− mice have an impaired cytoprotective response in obstructive cholestasis

“…Although Mrp2 protein was increased significantly in the kidneys of Mrp4Ϫ/Ϫ mice, it did not reach statistical significance in wild-type mice after CBDL, unlike in the rat 30,31 (Fig. 5B-C).…”

“…These findings differ from those in rats where renal Mrp2 protein is significantly increased after CBDL, presumably by posttranscriptional mechanisms. 30,31 However, cholic acid feeding in Fxr ϩ/ϩ and FxrϪ/Ϫ mice increases Mrp2 mRNA in the kidneys. 11 Again, the lack of significant increases in Mrp2 protein in the kidneys of wild-type CBDL mice in the present study may reflect a less severe degree of cholestasis.…”

Expression and Function of Renal Organic Anion Transporters in Cholestasis

“…In this model, Oat1 up regulation might be found in order to enhance renal secretion of toxic compounds that may be harmful in the presence of the pathological state. Connected to this, Tanaka et al (2002) found that bilirubin ditaurate, sulfate conjugated bile acids, and some components of the human bile up regulate the expression of Mrp2 in human renal tubular cells. On the other hand, we found an increased 3 H-PAH uptake by S2 cells expressing Oat1 in the presence of bilirubin ditaurate (Brandoni et al, 2006c).…”

Gene expression of ATP-binding cassette transporters during liver regeneration after 90% hepatectomy in rats

“…Consequently, the direction of bilirubin transport changes from toward the bile canaliculi to toward the sinusoidal space, which leads to conjugated hyperbilirubinemia (13,14). As for the expression of the ABC transporters during liver regeneration of the small liver, we previously demonstrated the same patterns of MRP2 and MRP3 expression in rat hepatocytes by a western blot analysis (20).…”

“…Physiologically, conjugated bilirubin is evacuated from the cell into the bile canaliculi by a bilirubin-excreting transporter, an ATP-binding cassette (ABC) transporter called multidrug resistance protein (MRP) 2, which localizes to the canalicular membrane of the hepatocytes (10)(11)(12). In some pathological conditions, such as bile duct ligation (13)(14)(15), or under an endotoxin load (15)(16)(17)(18)(19), however, the expression of MRP2 is reported to be downregulated. As for liver regeneration, we previously reported that the MRP2 protein decreased and that MRP3, an isoform of MRP2, is expressed on the sinusoidal membrane of the hepatocytes, which leads to changes in the direction of the bilirubin transport from the bile canaliculi to the sinusoidal space (20).…”