Up–Regulation of the Multidrug Resistance Genes, Mrp1 And Mdr1b , and Down–Regulation of the Organic Anion Transporter, Mrp2, and the Bile Salt Transporter, Spgp, in Endotoxemic Rat Liver

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Abstract
Endotoxin–induced cholestasis is mainly caused by an impaired canalicular secretion. Mrp2, the canalicular multispecific organic anion transporter, is strongly down–regulated in this situation, and canalicular bile salt secretion is also reduced. We hypothesized that other adenosine triphosphate-binding cassette (ABC) transporters may compensate for the decreased transport activity to protect the cell from cytokine–induced oxidative damage. Therefore, we examined the expression of ABC–transport proteins in membrane fractions of whole liver and of isolated hepatocytes of endotoxin–treated rats and performed reverse–transcriptase polymerase chain reaction (RT–PCR) on mRNA isolated from these livers. In addition, the localization of these transporters was examined using confocal scanning laser microscopy. By 6 hours after endotoxin administration, we found a clear increase of mrp1 mRNA and protein, whereas mrp2 mRNA and protein were decreased. This was confirmed in isolated hepatocytes. In addition, mdr1b mRNA was strongly increased, whereas mdr1a and mdr2 mRNA did not change significantly. Both the mRNA and protein levels of the sister of P–glycoprotein (spgp), the recently cloned bile salt transporter, decreased. After endotoxin treatment, the normally sharply delineated canalicular staining of mrp2 and spgp had changed to a fuzzy pattern, suggesting localization in a subapical compartment. We conclude that endotoxin–induced cholestasis is caused by decreased mrp2 and spgp levels, as well as an abnormal localization of these proteins. The simultaneous up–regulation of mrp1 and mdr1b may confer resistance to hepatocytes against cytokine–induced metabolic stress.