Establishment of mouse-directed HCV infection models

2011 Fiscal Year Final Research Report

• Project/Area Number: 22659145
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Single-year Grants
• Research Field: Gastroenterology
• Research Institution: Tokyo Medical and Dental University
• Principal Investigator: SAKAMOTO Naoya 東京医科歯科大学, 大学院・医歯学総合研究科, 寄附講座教員 (10334418)
• Co-Investigator(Kenkyū-buntansha): KAKINUMA Sei 東京医科歯科大学, 大学院・医歯学総合研究科, 寄付講座教員 (30372444) ; NAKAGAWA Mina 東京医科歯科大学, 医学部附属病院, 助教 (30401342)
• Project Period (FY): 2010 – 2011
• Keywords: HCV / CD81

Research Abstract

In our present study, we constructed two fluorescence protein-tagged recombinant JFH1 virus clones, JFH1-EYFP and JFH1-AsRed, as well as two corresponding clones with adaptive mutations, JFH1-EYFP mutant and JFH1-AsRed mutant, that and were as effective as JFH1 in producing infectious virus particles, and investigated their viral infection life cycles. After infection of the fluorescence-tagged mutant viruses, infected cells increased exponentially. In cells, EYFP or AsRed and NS5A were expressed as a fusion protein and co-localized in core proteins. The rate of the cell. cell spread was dependent on the cell densities with a maximum of 102. 5/day. Treatment of cells with interferon or a protease inhibitor suppressed expansion of virus-positive cells. Taken together, these results indicate that fluorescence-tagged HCV is a useful tool to study virus infection life cycles and to assist in the search for novel antiviral compounds.

Research Products

• [Journal Article] Suppression of hepatitis C virus replicon by adenovirus vector-mediated expression of tough decoy RNA against miR-122a (2012)
  • Author(s): Sakurai F, Furukawa N, Higuchi M, Okamoto S, Ono K, Yoshida T, Kondoh M, Yagi K, Sakamoto N, Katayama K, Mizuguchi H
  • Journal Title: Virus Research
  • DOI: DOI:10.1016/j.virusres.2012.02.003
• [Journal Article] Identification of novel N-(morpholine-4-carbonyloxy) amidine compounds as potent inhibitors against hepatitis C virus replication (2012)
  • Author(s): Kusano-Kitazume A, Sakamoto N(equalcontribution), Okuno Y, Sekine-Osajima Y, Nakagawa M, Kakinuma S, Kiyonashi K, Nitta S, Murakawa M, Azuma S, Nishimura-Sakurai Y, Hagiwara M, Watanabe M
  • Journal Title: Antimicrob Agent Chemother Volume: 56(3) Pages: 1315-1323
• [Journal Article] Use of human hepatocyte-like cells derived from induced pluripotent stem cells as a model for hepatocytes in hepatitis C virus infection (2011)
  • Author(s): Yoshida T, Takayama K, Kondoh M, Sakurai F, Tani H, Sakamoto N, Matsuura Y, Mizuguchi H, Yagi K
  • Journal Title: Biochem Biophys Res Commun Volume: 416(1-2) Pages: 119-124
• [Journal Article] Adenovirus vector-mediated assay system for hepatitis C virus replication (2011)
  • Author(s): Yoshida T, Kondoh M, Ojima M, Mizuguchi H, Yamagishi Y, Sakamoto N, Yagi K.
  • Journal Title: Nucleic Acids Res Volume: 39(10) Pages: e64
• [Journal Article] Analysis of interferon signaling by infectious hepatitis C virus clones with substitutions of core amino acids 70 and 91 (2011)
  • Author(s): Funaoka Y, Sakamoto N(equal contribution), Nakagawa M, Kakinuma S, Suda S, Watanabe T, Nitta S, Kitazume A, Kiyohashi K, Murakawa M, Azuma S, Tsuchiya K, and Watanabe M
  • Journal Title: J Virol Volume: 85(12) Pages: 5986-5994
• [Journal Article] Studies on virus kinetics using infectious fluorescence-tagged hepatitis C virus cell culture (2011)
  • Author(s): Yamamoto M, Sakamoto N, Nakamura T, Itsui Y, Nakagawa M, Nishimura-Sakurai Y, Kakinuma S, Azuma S, Tsuchiya K, Kato T, Wakita T, Watanabe M
  • Journal Title: Hepatol Res Volume: 41 Pages: 258-269
• [Journal Article] Association between lipid accumulation and the cannabinoid system in Huh7 cells expressing HCV genes (2011)
  • Author(s): Toyoda M, Kitaoka A, Machida K, Nishinakagawa T, Yada R, Kohjima M, Kato M, Kotoh K, Sakamoto N, Shiota G, Nakamuta M, Nakashima M, Enjoji M
  • Journal Title: Int J Mol Med Volume: 27 Pages: 619-624
• [Journal Article] シクロフィリン阻害剤 (2011)
  • Author(s): 中川美奈、坂本直哉、渡辺守
  • Journal Title: 肝胆膵 Volume: 62 Pages: 403-412
• [Journal Article] IL-6-mediated intersubgenotypic variation of interferon sensitivity in hepatitis C virus genotype 2a/2b chimeric clones (2010)
  • Author(s): Suda G, Sakamoto N, Itsui Y, Nakagawa M, Tasaka-Fujita M, Funaoka Y, Watanabe T, Nitta S, Kiyohashi K, Azuma S, Kakinuma S, Tsuchiya K, Imamura M, Hiraga N, Chayama K, Watanabe M
  • Journal Title: Virology Volume: 407 Pages: 80-90
• [Journal Article] Cell culture and in vivo analyses of cytopathic hepatitis C virus mutants (2010)
  • Author(s): Mishima K, Sakamoto N, Sekine-Osajima Y, Nakagawa M, Itsui Y, Azuma S, Kakinuma S, Kiyohashi K, Kitazume A, Tsuchiya K, Imamura M, Hiraga N, Chayama K, Wakita T, Watanabe M
  • Journal Title: Virology Volume: 405 Pages: 361-369
• [Journal Article] HCVの複製・増殖機構と遺伝子型・遺伝子変異 (2010)
  • Author(s): 筬島裕子、中川美奈、坂本直哉
  • Journal Title: 日本臨床 Volume: 69 Pages: 52-58
• [Presentation] A high-content screening assay using infectious fluorescence-tagged hepatitis C virus reveals candidates for small molecule inhibitors of viral entry (2011)
  • Author(s): Kusano-Kitazume A, Sakamoto N, Okuno Y, Yamamoto M, Sekine-Osajima Y, Nakagawa M, Kakinuma S, Kiyohashi K, Nitta S, Murakawa M, Hagiwara M, Watanabe M
  • Organizer: 62th. Annual Meeting of American Association for the Study of Liver Diseases
  • Place of Presentation: San Francisco, CA.(Poster# 383)
  • Year and Date: 2011-11-04
• [Presentation] EYFP発現HCV培養系を用いたHigh-content screening assayによるウイルス侵入阻害剤の同定.(ポスターP26) (2011)
  • Author(s): 北詰晶子、坂本直哉、奥野友紀子、中川美奈、柿沼晴、山本満千、幾世橋佳、新田沙由梨、村川美也子、萩原正敏、渡辺守
  • Organizer: 第15回日本肝臓学会大会
  • Place of Presentation: 福岡
  • Year and Date: 2011-10-20
• [Presentation] Cell based high-throughput screeningによる新規抗ウイルス小分子の同定及びその作用機構の解析 (2011)
  • Author(s): 北詰晶子、坂本直哉、奥野友紀子、筬島裕子、中川美奈、柿沼晴、植山真由美、小野塚泉、船岡祐介、渡辺貴子、幾世橋佳、新田沙由梨、村川美也子、萩原正敏、渡辺守
  • Organizer: 第47回日本肝臓学会総会
  • Place of Presentation: 東京(一般口演, O-156)
  • Year and Date: 2011-06-03
• [Presentation] 蛍光蛋白発現HCV培養系を用いたHigh-content screening assayの樹立及びHCVエントリー阻害剤探索への応用 (2011)
  • Author(s): 北詰晶子、坂本直哉、奥野友紀子、筬島裕子、中川美奈、柿沼晴、植山真由美、小野塚泉、船岡祐介、渡辺貴子、幾世橋佳、新田沙由梨、村川美也子、萩原正敏、渡辺守
  • Organizer: 第47回日本肝臓学会総会
  • Place of Presentation: 東京(一般口演, O-164)
  • Year and Date: 2011-06-03
• [Presentation] Ezetimibe and interferon synergistically suppress subgenomic replication of hepatitis C virus via the acceleration of IFN-mediated ISG15 conjugation system (2010)
  • Author(s): Masayoshi Yada, Ryoko Yada, Motoyuki Kojima, Kazuto Goto, Naoya Sasamoto, Akihide Masumoto and Makoto Nakamuta
  • Organizer: Asian Pacific Association for the Study of the Liver(APASL) The 7th Single Topic Conference
  • Place of Presentation: Chiba, Japan
  • Year and Date: 2010-12-17
• [Presentation] 蛍光タグ付加HCV培養系を用いたウイルスライフサイクルの解析 (2010)
  • Author(s): 山本満千、坂本直哉、中川美奈、柿沼晴、東正新、加藤孝宣、脇田隆字、渡辺守
  • Organizer: JDDW2010
  • Place of Presentation: 横浜(ポスター)
  • Year and Date: 2010-10-13
• [Presentation] 細胞障害性HCV-JFH1subcloneの細胞培養系および感染動物モデルにおける機能解析 (2010)
  • Author(s): 三島果子、坂本直哉、筬島裕子、中川美奈、柿沼晴、東正新、脇田隆字、今村道雄、茶山一彰、渡辺守
  • Organizer: JDDW2010
  • Place of Presentation: 横浜(ポスター)
  • Year and Date: 2010-10-13
• [Presentation] Identification of Novel Small Molecules Inhibitors of Hepatitis C Virus Replication Using a Cell-Based High-Throughput Screen. (2010)
  • Author(s): Akiko Kusano-Kitazume, Naoya Sakamoto, Yuko Sekine-Osajima, Seishin Azuma, Mina Nakagawa, Sei Kakinuma, Yuki Nishimura-Sakurai, Yukiko Okuno, Masatoshi Hagiwara, and Mamoru Watanabe
  • Organizer: 17th International Meeting on Hepatitis C Virus & Related Viruses
  • Place of Presentation: Yokohama, Japan
  • Year and Date: 2010-09-09
• [Presentation] 蛍光タグ付加HCV培養系を用いたウイルスライフサイクルの解析.(一般口演) (2010)
  • Author(s): 山本満千、坂本直哉、中川美奈、柿沼晴、東正新、加藤孝宣、脇田隆字、渡辺守
  • Organizer: 第46回日本肝臓学会総会
  • Place of Presentation: 山形
  • Year and Date: 2010-05-27
• [Presentation] HCVレプリコン系を用いた抗ウイルス小分子の大規模スクリーニング.(ポスター) (2010)
  • Author(s): 北詰晶子、坂本直哉、筬島裕子、中川美奈、東正新、柿沼晴、渡辺守
  • Organizer: 第46回日本肝臓学会総会
  • Place of Presentation: 山形
  • Year and Date: 2010-05-27