Cytokine-independent repression of rodent Ntcp in obstructive cholestasis.
Cholestatic liver injury is associated not only with accumulation of bile acids but also with activation of proinflammatory cytokines. Common bile duct ligation (CBDL) induces sustained downregulation of the Na(+)/taurocholate cotransporter (Ntcp) in rodent liver. Although repression of Ntcp during endotoxemia is cytokine mediated, it is unclear whether inflammatory cytokines contribute to this downregulation in obstructive cholestasis. Cytokine inactivation in CBDL rats and mice was either performed directly with tumor necrosis factor alpha (etanercept) or interleukin 1 beta inactivation (anakinra/AMG 719) or indirectly Kupffer cell depletion via intraperitoneal administration of liposome-encapsulated dichloromethylene bisphosphonate. Protein and messenger RNA (mRNA) expression of Ntcp and short heterodimer partner (SHP) were analyzed via Western and Northern blotting. Key regulators of Ntcp (hepatocyte nuclear factor 1 alpha [HNF-1alpha], HNF-4alpha, retinoid X receptor alpha [RXRalpha]:retinoic acid receptor alpha [RARalpha]) were studied via electrophoretic mobility shift analysis and nuclear Western blot analysis. Both methods of cytokine inactivation failed to maintain Ntcp protein or mRNA expression within 3 days after CBDL in either rats or mice (20%-40% of sham controls), while SHP mRNA expression increased three- to five-fold. Decreased nuclear HNF-1alpha and HNF-4alpha protein levels (45% and 60% of sham controls, respectively) and HNF-1alpha binding activity (32% of sham controls) were not restored during cytokine inactivation after CBDL, indicating cytokine-independent mechanisms of Ntcp regulation. RXRalpha:RARalpha binding remained unchanged in all experimental conditions. In conclusion, during obstructive cholestasis accumulating bile acids per se, without major contribution of cytokines, leads to downregulation of Ntcp via repression of HNF-1alpha and HNF-4alpha.
Geier
Andreas
A
Department of Internal Medicine III, Aachen University (RWTH), Aachen, Germany.
Zollner
Gernot
G
Dietrich
Christoph G
CG
Wagner
Martin
M
Fickert
Peter
P
Denk
Helmut
H
van Rooijen
Nico
N
Matern
Siegfried
S
Gartung
Carsten
C
Trauner
Michael
M
D000818:Animals; D002779:Cholestasis / Q000378:metabolism*; D016207:Cytokines / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics; D005786:Gene Expression Regulation; D051537:Hepatocyte Nuclear Factor 1; D051538:Hepatocyte Nuclear Factor 1-alpha; D051557:Hepatocyte Nuclear Factor 4; D007728:Kupffer Cells / Q000502:physiology; D008297:Male; D026901:Membrane Transport Proteins / Q000235:genetics*; D051379:Mice; D009687:Nuclear Proteins / Q000235:genetics; D029363:Organic Anion Transporters, Sodium-Dependent; D010750:Phosphoproteins / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics; D027981:Symporters; D014157:Transcription Factors / Q000235:genetics
D016207:Cytokines; D004268:DNA-Binding Proteins; D051538:Hepatocyte Nuclear Factor 1-alpha; D051557:Hepatocyte Nuclear Factor 4; C496488:Hnf1a protein, mouse; C496489:Hnf1a protein, rat; D026901:Membrane Transport Proteins; D009687:Nuclear Proteins; D029363:Organic Anion Transporters, Sodium-Dependent; D010750:Phosphoproteins; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D027981:Symporters; D014157:Transcription Factors; C099713:nuclear receptor subfamily 0, group B, member 2; D051537:Hepatocyte Nuclear Factor 1; C071908:sodium-bile acid cotransporter