Tauroursodeoxycholic acid protects rat hepatocytes from bile acid-induced apoptosis via activation of survival pathways.
Ursodeoxycholic acid (UDCA) is used in the treatment of cholestatic liver diseases, but its mechanism of action is not yet well defined. The aim of this study was to explore the protective mechanisms of the taurine-conjugate of UDCA (tauroursodeoxycholic acid [TUDCA]) against glycochenodeoxycholic acid (GCDCA)-induced apoptosis in primary cultures of rat hepatocytes. Hepatocytes were exposed to GCDCA, TUDCA, the glyco-conjugate of UDCA (GUDCA), and TCDCA. The phosphatidylinositol-3 kinase pathway (PI3K) and nuclear factor-kappaB were inhibited using LY 294002 and adenoviral overexpression of dominant-negative IkappaB, respectively. The role of p38 and extracellular signal-regulated protein kinase mitogen-activated protein kinase (MAPK) pathways were investigated using the inhibitors SB 203580 and U0 126 and Western blot analysis. Transcription was blocked by actinomycin-D. Apoptosis was determined by measuring caspase-3, -9, and -8 activity using fluorimetric enzyme detection, Western blot analysis, immunocytochemistry, and nuclear morphological analysis. Our results demonstrated that uptake of GCDCA is needed for apoptosis induction. TUDCA, but not TCDCA and GUDCA, rapidly inhibited, but did not delay, apoptosis at all time points tested. However, the protective effect of TUDCA was independent of its inhibition of caspase-8. Up to 6 hours of preincubation with TUDCA before addition of GCDCA clearly decreased GCDCA-induced apoptosis. At up to 1.5 hours after exposure with GCDCA, the addition of TUDCA was still protective. This protection was dependent on activation of p38, ERK MAPK, and PI3K pathways, but independent of competition on the cell membrane, NF-kappaB activation, and transcription. In conclusion, TUDCA contributes to the protection against GCDCA-induced mitochondria-controlled apoptosis by activating survival pathways.
Schoemaker
Marieke H
MH
Center for Liver, Digestive and Metabolic Diseases, Groningen University Institute for Drug Exploration, Groningen, The Netherlands.
Conde de la Rosa
Laura
L
Buist-Homan
Manon
M
Vrenken
Titia E
TE
Havinga
Rick
R
Poelstra
Klaas
K
Haisma
Hidde J
HJ
Jansen
Peter L M
PL
Moshage
Han
H
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001647:Bile Acids and Salts / Q000502:physiology*; D002352:Carrier Proteins / Q000494:pharmacology; D053148:Caspase 3; D053453:Caspase 9; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D005999:Glycochenodeoxycholic Acid / Q000494:pharmacology; D022781:Hepatocytes / Q000187:drug effects*; D008297:Male; D026901:Membrane Transport Proteins; D008928:Mitochondria / Q000187:drug effects / Q000502:physiology; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology; D029363:Organic Anion Transporters, Sodium-Dependent; D051381:Rats; D017208:Rats, Wistar; D027981:Symporters; D013655:Taurochenodeoxycholic Acid / Q000494:pharmacology*; D048051:p38 Mitogen-Activated Protein Kinases
D001647:Bile Acids and Salts; D002352:Carrier Proteins; D061945:Caspase Inhibitors; D026901:Membrane Transport Proteins; D029363:Organic Anion Transporters, Sodium-Dependent; D027981:Symporters; C071908:sodium-bile acid cotransporter; D013655:Taurochenodeoxycholic Acid; C031655:ursodoxicoltaurine; D005999:Glycochenodeoxycholic Acid; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; C505410:Casp3 protein, rat; C506435:Casp9 protein, rat; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases