Cytoprotective properties of rifampicin are related to the regulation of detoxification system and bile acid transporter expression during hepatocellular injury induced by hydrophobic bile acids

Abstract Treatment with high dose human GH, although an effective anabolic agent, has been associated with increased incidence of sepsis, inflammation, multiple organ failure, and death in critically ill patients. We hypothesized that GH might increase mortality by exacerbating cholestasis through modulation of bile acid transporter expression. High dose GH was continuously infused over 4 d into rats, and on the final day lipopolysaccharides were injected. Hepatic bile acid transporter expression was measured by Northern analysis and immunoblotting and compared with serum markers of cholestasis and endotoxinemia. Compared with non-GH-treated controls, GH increased endotoxin-induced markers of cholestasis and liver damage as well as augmented IL-6 induction. In endotoxinemia, GH treatment significantly induced multidrug resistance-associated protein 1 mRNA and protein and suppressed organic anion transporting polypeptides, Oatp1 and Oatp4, mRNA, suggesting impaired uptake of bilirubin and bile acids at the basolateral surface of the hepatocyte, which could contribute to the observed worsening of cholestasis by GH. This study of endotoxinemia may thus provide a mechanistic link between GH treatment and exacerbation of cholestasis through modulation of basolateral bile acid transporter expression in the rat hepatocyte.

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Electrostatic and potential cation-π forces may guide the interaction of extracellular loop III with Na+ and bile acids for human apical Na+-dependent bile acid transporter

Biochemical Journal ◽

10.1042/bj20071300 ◽

2008 ◽

Vol 410 (2) ◽

pp. 391-400 ◽

Cited By ~ 14

Author(s):

Antara Banerjee ◽

Naissan Hussainzada ◽

Akash Khandelwal ◽

Peter W. Swaan

Keyword(s):

Amino Acids ◽

Bile Acid ◽

Bile Acids ◽

Conformational Changes ◽

Electrostatic Interactions ◽

Extracellular Loop ◽

Membrane Expression ◽

Ligand Interaction ◽

Bile Acid Transporter ◽

Acid Transporter

The hASBT (human apical Na+-dependent bile acid transporter) constitutes a key target of anti-hypercholesterolaemic therapies and pro-drug approaches; physiologically, hASBT actively reclaims bile acids along the terminal ileum via Na+ co-transport. Previously, TM (transmembrane segment) 7 was identified as part of the putative substrate permeation pathway using SCAM (substitute cysteine accessibility mutagenesis). In the present study, SCAM was extended through EL3 (extracellular loop 3; residues Arg254–Val286) that leads into TM7 from the exofacial matrix. Activity of most EL3 mutants was significantly hampered upon cysteine substitution, whereas ten (out of 31) were functionally inactive (<10% activity). Since only E282C lacked plasma membrane expression, EL3 amino acids predominantly fulfill critical functional roles during transport. Oppositely charged membrane-impermeant MTS (methanethiosulfonate) reagents {MTSET [(2-trimethylammonium) ethyl MTS] and MTSES [(2-sulfonatoethyl) MTS]} produced mostly similar inhibition profiles wherein only middle and descending loop segments (residues Thr267–Val286) displayed significant MTS sensitivity. The presence of bile acid substrate significantly reduced the rates of MTS modification for all MTS-sensitive mutants, suggesting a functional association between EL3 residues and bile acids. Activity assessments at equilibrative [Na+] revealed numerous Na+-sensitive residues, possibly performing auxiliary functions during transport such as transduction of protein conformational changes during translocation. Integration of these data suggests ligand interaction points along EL3 via electrostatic interactions with Arg256, Glu261 and probably Glu282 and a potential cation-π interaction with Phe278. We conclude that EL3 amino acids are essential for hASBT activity, probably as primary substrate interaction points using long-range electrostatic attractive forces.

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