title: Liver Bid suppression for treatment of fibrosis associated with non-alcoholic steatohepatitis.
abstract: OBJECTIVE
  Liver fibrosis is the most worrisome feature of non-alcoholic steatohepatitis (NASH). Growing evidence supports a link between hepatocyte apoptosis and liver fibrogenesis. Our aim was to determine the therapeutic efficacy and safety of liver Bid, a key pro-apoptotic molecule, suppression using RNA interference (RNAi) for the treatment of fibrosis.
  
  
  METHODS
  First, we optimized the delivery system for Bid siRNA in mice using ten different stealth RNAi siRNAs and two lipid formulations -Invivofectamine2.0 and a newly developed Invivofectamine3.0 - that have been designed for high efficacy accumulation in the liver, assessed via real-time PCR of Bid mRNA. Next, C57BL/6 mice were placed on a choline-deficient L-amino acid defined (CDAA) diet. After 19weeks of the CDAA diet, a time point that results in severe fibrotic NASH, mice were injected with the selected Bid siRNA-Invivofectamine3.0 biweekly for three weeks. Additionally hepatocyte-specific Bid deficient (Bid(Δhep)) mice were placed on CDAA diet for 20weeks.
  
  
  RESULTS
  A maximum Bid knockdown was achieved at 1.5mg/kg siRNA with Invivofectamine3.0, whereas it was at 7mg/kg with Invivofectamine2.0. In NASH mice, after 3weeks of treatment, BID protein was reduced to 10% and this was associated with an improvement in liver fibrosis and inflammation associated with a marked reduction in TUNEL positive cells, caspase 3 activation, and a reduction in mitochondrial BAX and BAK. Bid(Δhep) mice showed similar protection from fibrotic changes.
  
  
  CONCLUSIONS
  Our data demonstrate that liver Bid suppression by RNAi technology, as well as hepatocyte-specific Bid deficiency, improves liver fibrosis coupled with a reduction of inflammation in experimental NASH. These findings are consistent with existing evidence that hepatocyte apoptosis triggers hepatic stellate cell activation and liver fibrosis and suggest that Bid inhibition may be useful as an antifibrotic NASH therapy.
authors.affiliation: Department of Pediatrics, University of California - San Diego, 9500 Gilman Drive, La Jolla, USA.
authors.forename: Akiko
authors.initials: A
authors.lastname: Eguchi
authors.affiliation: Life Technologies Corporation, Carlsbad, USA.
authors.forename: Xavier
authors.initials: X
authors.lastname: De Mollerat Du Jeu
authors.affiliation: Department of Pediatrics, University of California - San Diego, 9500 Gilman Drive, La Jolla, USA.
authors.forename: Casey D
authors.initials: CD
authors.lastname: Johnson
authors.affiliation: Life Technologies Corporation, Carlsbad, USA.
authors.forename: Andronikou
authors.initials: A
authors.lastname: Nektaria
authors.affiliation: Department of Pediatrics, University of California - San Diego, 9500 Gilman Drive, La Jolla, USA. Electronic address: afeldstein@ucsd.edu.
authors.forename: Ariel E
authors.initials: AE
authors.lastname: Feldstein
mesh_terms: D000818:Animals; D017209:Apoptosis; D051023:BH3 Interacting Domain Death Agonist Protein / Q000037:antagonists & inhibitors* / Q000235:genetics; D000067128:Extracellular Vesicles / Q000502:physiology; D056945:Hep G2 Cells; D006801:Humans; D008099:Liver / Q000473:pathology; D008106:Liver Cirrhosis, Experimental / Q000628:therapy*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008928:Mitochondria / Q000502:physiology; D065626:Non-alcoholic Fatty Liver Disease / Q000150:complications*; D034622:RNA Interference
chemical_list: D051023:BH3 Interacting Domain Death Agonist Protein; C494815:Bid protein, mouse
keywords: Apoptosis; Bid; Liver fibrosis; Liver inflammation; Mitochondrial dysfunction