title: Ursodeoxycholic acid may inhibit deoxycholic acid-induced apoptosis by modulating mitochondrial transmembrane potential and reactive oxygen species production.
abstract: BACKGROUND
  The hydrophilic bile salt ursodeoxycholate (UDCA) inhibits injury by hydrophobic bile acids and is used to treat cholestatic liver diseases. Interestingly, hepatocyte cell death from bile acid-induced toxicity occurs more frequently from apoptosis than from necrosis. However, both processes appear to involve the mitochondrial membrane permeability transition (MPT). In this study, we determined the inhibitory effect of UDCA on deoxycholic acid (DCA)-induced MPT in isolated mitochondria by measuring changes in transmembrane potential (delta psi m) and production of reactive oxygen species (ROS). In addition, we examined the expression of apoptosis-associated proteins in mitochondria isolated from livers of bile acid-fed animals.
  
  
  METHODS
  Adult male rats were maintained on standard diet supplemented with DCA and/or UDCA for 10 days. Mitochondria were isolated from livers by sucrose/percoll gradient centrifugation and MPT was measured using spectrophotometric and fluorimetric assays. delta psi m and ROS generation were determined by FACScan analysis. Cytoplasmic and mitochondrial protein abundance were determined by Western blot analysis.
  
  
  RESULTS
  DCA increased mitochondrial swelling 25-fold over controls (p < 0.001); UDCA reduced the swelling by > 40% (p < 0.001). Similarly, UDCA inhibited DCA-mediated release of calcein-loaded mitochondria by 50% (p < 0.001). delta psi m was significantly decreased in mitochondria incubated with DCA but not with UDCA. delta psi m disruption was followed closely by increased superoxide anion and peroxides production (p < 0.01). Coincubation of mitochondria with UDCA significantly inhibited the changes associated with DCA (p < 0.05). In vivo, DCA feeding was associated with a 4.5-fold increase in mitochondria-associated Bax protein levels (p < 0.001); combination feeding with UDCA almost totally inhibited this increase (p < 0.001).
  
  
  CONCLUSIONS
  UDCA significantly reduces DCA-induced disruption of delta psi m, ROS production, and Bax protein abundance in mitochondria, suggesting both short- and long-term mechanisms in preventing MPT. The results suggest a possible role for UDCA as a therapeutic agent in the treatment of both hepatic and nonhepatic diseases associated with high levels of apoptosis.
authors.affiliation: Department of Medicine, University of Minnesota Medical School, Minneapolis 55455, USA.
authors.forename: C M
authors.initials: CM
authors.lastname: Rodrigues
authors.forename: G
authors.initials: G
authors.lastname: Fan
authors.forename: P Y
authors.initials: PY
authors.lastname: Wong
authors.forename: B T
authors.initials: BT
authors.lastname: Kren
authors.forename: C J
authors.initials: CJ
authors.lastname: Steer
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001152:Arsenicals / Q000494:pharmacology; D016572:Cyclosporine / Q000494:pharmacology; D003840:Deoxycholic Acid / Q000494:pharmacology*; D007425:Intracellular Membranes / Q000187:drug effects / Q000502:physiology; D008099:Liver / Q000737:chemistry; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D008930:Mitochondria, Liver / Q000201:enzymology / Q000378:metabolism / Q000502:physiology*; D010539:Permeability; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D014580:Ursodeoxycholic Acid / Q000494:pharmacology*
chemical_list: D001152:Arsenicals; D019253:Proto-Oncogene Proteins c-bcl-2; D017382:Reactive Oxygen Species; D003840:Deoxycholic Acid; C029341:oxophenylarsine; D014580:Ursodeoxycholic Acid; D016572:Cyclosporine