Treatment of pancreatic fibrosis with siRNA against a collagen-specific chaperone in vitamin A-coupled liposomes

Hirotoshi Ishiwatari, Yasushi Sato, Kazuyuki Murase, Akihiro Yoneda, Ryosuke Fujita, Hiroki Nishita, Naoko Kubo Birukawa, Tsuyoshi Hayashi, Tsutomu Sato, Koji Miyanishi, Rishu Takimoto, Masayoshi Kobune, Shigenori Ota, Yasutoshi Kimura, Koichi Hirata, Junji Kato, Yoshiro Niitsu

Research output: Contribution to journalArticle

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Abstract

Background and 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 60-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 3H-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-lipsiRNAgp46- 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 VAlip- 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. Conclusion These data suggest the therapeutic potential of the present approach for reversing pancreatic fibrosis.

Original languageEnglish
Pages (from-to)1328-1339
Number of pages12
JournalGut
Volume62
Issue number9
DOIs
Publication statusPublished - Sep 1 2013

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Lip
Vitamin A
Liposomes
Pancreatic Stellate Cells
Small Interfering RNA
Fibrosis
Collagen
Ceruletide
Therapeutics
Hydroxyproline
Immunoblotting
Flow Cytometry
Chronic Pancreatitis
Liver Cirrhosis
Radioactivity
Microscopy
Histology
Coloring Agents
Fluorescence
dibutyldichlorotin

All Science Journal Classification (ASJC) codes

  • Gastroenterology

Cite this

Treatment of pancreatic fibrosis with siRNA against a collagen-specific chaperone in vitamin A-coupled liposomes. / Ishiwatari, Hirotoshi; Sato, Yasushi; Murase, Kazuyuki; Yoneda, Akihiro; Fujita, Ryosuke; Nishita, Hiroki; Birukawa, Naoko Kubo; Hayashi, Tsuyoshi; Sato, Tsutomu; Miyanishi, Koji; Takimoto, Rishu; Kobune, Masayoshi; Ota, Shigenori; Kimura, Yasutoshi; Hirata, Koichi; Kato, Junji; Niitsu, Yoshiro.

In: Gut, Vol. 62, No. 9, 01.09.2013, p. 1328-1339.

Research output: Contribution to journalArticle

Ishiwatari, H, Sato, Y, Murase, K, Yoneda, A, Fujita, R, Nishita, H, Birukawa, NK, Hayashi, T, Sato, T, Miyanishi, K, Takimoto, R, Kobune, M, Ota, S, Kimura, Y, Hirata, K, Kato, J & Niitsu, Y 2013, 'Treatment of pancreatic fibrosis with siRNA against a collagen-specific chaperone in vitamin A-coupled liposomes', Gut, vol. 62, no. 9, pp. 1328-1339. https://doi.org/10.1136/gutjnl-2011-301746
Ishiwatari, Hirotoshi ; Sato, Yasushi ; Murase, Kazuyuki ; Yoneda, Akihiro ; Fujita, Ryosuke ; Nishita, Hiroki ; Birukawa, Naoko Kubo ; Hayashi, Tsuyoshi ; Sato, Tsutomu ; Miyanishi, Koji ; Takimoto, Rishu ; Kobune, Masayoshi ; Ota, Shigenori ; Kimura, Yasutoshi ; Hirata, Koichi ; Kato, Junji ; Niitsu, Yoshiro. / Treatment of pancreatic fibrosis with siRNA against a collagen-specific chaperone in vitamin A-coupled liposomes. In: Gut. 2013 ; Vol. 62, No. 9. pp. 1328-1339.
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abstract = "Background and 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 60-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 3H-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-lipsiRNAgp46- 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 VAlip- 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. Conclusion These data suggest the therapeutic potential of the present approach for reversing pancreatic fibrosis.",
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T1 - Treatment of pancreatic fibrosis with siRNA against a collagen-specific chaperone in vitamin A-coupled liposomes

AU - Ishiwatari, Hirotoshi

AU - Sato, Yasushi

AU - Murase, Kazuyuki

AU - Yoneda, Akihiro

AU - Fujita, Ryosuke

AU - Nishita, Hiroki

AU - Birukawa, Naoko Kubo

AU - Hayashi, Tsuyoshi

AU - Sato, Tsutomu

AU - Miyanishi, Koji

AU - Takimoto, Rishu

AU - Kobune, Masayoshi

AU - Ota, Shigenori

AU - Kimura, Yasutoshi

AU - Hirata, Koichi

AU - Kato, Junji

AU - Niitsu, Yoshiro

PY - 2013/9/1

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N2 - Background and 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 60-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 3H-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-lipsiRNAgp46- 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 VAlip- 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. Conclusion These data suggest the therapeutic potential of the present approach for reversing pancreatic fibrosis.

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