title: Treatment of pancreatic fibrosis with siRNA against a collagen-specific chaperone in vitamin A-coupled liposomes.
abstract: OBJECTIVE
  Fibrosis associated with chronic pancreatitis is an irreversible lesion that can disrupt pancreatic exocrine and endocrine function. Currently, there are no approved treatments for this disease. We previously showed that siRNA against collagen-specific chaperone protein gp46, encapsulated in vitamin A-coupled liposomes (VA-lip-siRNAgp46), resolved fibrosis in a model of liver cirrhosis. This treatment was investigated for pancreatic fibrosis induced by dibutyltin dichloride (DBTC) and cerulein in rats.
  
  
  METHODS
  Specific uptake of VA-lip-siRNAgp46, conjugated with 6'-carboxyfluorescein (FAM) by activated pancreatic stellate cells (aPSCs), was analysed by fluorescence activated cell sorting (FACS). Intracellular distribution of VA-lip-siRNAgp46-FAM was examined by fluorescent microscopy. Suppression of gp46 expression by VA-lip-siRNAgp46 was assessed by immunoblotting. Collagen synthesis in aPSCs was assayed by dye-binding. Specific delivery of VA-lip-siRNAgp46 to aPSCs in DBTC rats was verified following intravenous VA-lip-siRNA-FAM and (3)H-VA-lip-siRNAgp46. The effect of VA-lip-siRNA on pancreatic histology in DBTC- and cerulein-treated rats was determined by Azan-Mallory staining and hydroxyproline content.
  
  
  RESULTS
  FACS analysis revealed specific uptake of VA-lip-siRNAgp46-FAM through the retinol binding protein receptor by aPSCs in vitro. Immunoblotting and collagen assay verified knockdown of gp46 and suppression of collagen secretion, respectively, by aPSCs after transduction of VA-lip-siRNAgp46. Specific delivery of VA-lip-siRNAgp46 to aPSCs in fibrotic areas in DBTC rats was confirmed by fluorescence and radioactivity 24 h after the final injection. 10 systemic VA-lip-siRNAgp46 treatments resolved pancreatic fibrosis, and suppressed tissue hydroxyproline levels in DBTC- and cerulein-treated rats.
  
  
  CONCLUSIONS
  These data suggest the therapeutic potential of the present approach for reversing pancreatic fibrosis.
authors.affiliation: Fourth Department of Internal Medicine, Sapporo Medical University, Sapporo, Japan.
authors.forename: Hirotoshi
authors.initials: H
authors.lastname: Ishiwatari
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Sato
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Murase
authors.forename: Akihiro
authors.initials: A
authors.lastname: Yoneda
authors.forename: Ryosuke
authors.initials: R
authors.lastname: Fujita
authors.forename: Hiroki
authors.initials: H
authors.lastname: Nishita
authors.forename: Naoko Kubo
authors.initials: NK
authors.lastname: Birukawa
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Hayashi
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Sato
authors.forename: Koji
authors.initials: K
authors.lastname: Miyanishi
authors.forename: Rishu
authors.initials: R
authors.lastname: Takimoto
authors.forename: Masayoshi
authors.initials: M
authors.lastname: Kobune
authors.forename: Shigenori
authors.initials: S
authors.lastname: Ota
authors.forename: Yasutoshi
authors.initials: Y
authors.lastname: Kimura
authors.forename: Koichi
authors.initials: K
authors.lastname: Hirata
authors.forename: Junji
authors.initials: J
authors.lastname: Kato
authors.forename: Yoshiro
authors.initials: Y
authors.lastname: Niitsu
mesh_terms: D000818:Animals; D002108:Ceruletide / Q000494:pharmacology; D003094:Collagen / Q000096:biosynthesis*; D005355:Fibrosis / Q000188:drug therapy* / Q000209:etiology; D005765:Gastrointestinal Agents / Q000494:pharmacology; D050958:HSP47 Heat-Shock Proteins / Q000037:antagonists & inhibitors*; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology; D008081:Liposomes; D008297:Male; D023421:Models, Animal; D009947:Organotin Compounds / Q000494:pharmacology; D010179:Pancreas / Q000473:pathology*; D050500:Pancreatitis, Chronic / Q000150:complications*; D034741:RNA, Small Interfering / Q000008:administration & dosage*; D051381:Rats; D011917:Rats, Inbred Lew; D017208:Rats, Wistar; D016896:Treatment Outcome; D014801:Vitamin A / Q000494:pharmacology; D014815:Vitamins / Q000494:pharmacology
chemical_list: D005765:Gastrointestinal Agents; D050958:HSP47 Heat-Shock Proteins; D007166:Immunosuppressive Agents; D008081:Liposomes; D009947:Organotin Compounds; D034741:RNA, Small Interfering; D014815:Vitamins; D014801:Vitamin A; D002108:Ceruletide; D003094:Collagen; C020735:dibutyldichlorotin