Investigation of erythroid differentiation using transgene-free canine and feline induced pluripotent stem cells

• Project/Area Number: 15K07747
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Veterinary medical science
• Research Institution: Osaka Prefecture University
• Principal Investigator: Hatoya Shingo 大阪府立大学, 生命環境科学研究科, 准教授 (40453138)
• Co-Investigator(Kenkyū-buntansha): 杉浦 喜久弥 大阪府立大学, 生命環境科学研究科, 教授 (30171143) ; 竹中 重雄 大阪府立大学, 総合リハビリテーション学研究科, 教授 (10280067)
• Co-Investigator(Renkei-kenkyūsha): NAKANISHI Mahito 国立研究開発法人産業技術総合研究所, その他部局等, 研究員 (10172355) ; HISASUE Masaharu 麻布大学, 獣医学部, 准教授 (80333144)
• Project Period (FY): 2015-04-01 – 2018-03-31
• Project Status: Completed (Fiscal Year 2017)
• Budget Amount: ¥4,810,000 (Direct Cost: ¥3,700,000、Indirect Cost: ¥1,110,000); Fiscal Year 2017: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2016: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2015: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
• Keywords: iPS細胞 / 多能性幹細胞 / 再生医療 / 獣医学 / イヌ / ネコ / 血液 / センダイウイルスベクター / 獣医再生医療 / 再生獣医療

Outline of Final Research Achievements

The objective of this study was to establish genome integration-free canine and feline induced pluripotent stem cells (ciPSCs), and to induce hematopoietic cells from these stem cells. The following results were obtained.
①We confirmed that the Sendai virus vector (SeVdp) could infect canine and feline embryonic fibroblasts by detecting GFP expression. ②We could reprogram feline embryonic fibroblasts into iPS-like cells using SeVdp containing 6 transcription factors with TGF-beta inhibitor. These cells were positive for pluripotent markers. ③We transduced four transcriptional factors into canine fibroblast cells with SeVdp, and could established iPS-like cell line using TGF-beta inhibitor. Finally, we reprogrammed canine embryonic fibroblasts using the auto-erasable defective and persistent SeVdp, and canine iPS-like cells were formed. Further studies are required to induce hematopoietic cells from these canine or feline stem cells.

Research Products

• [Journal Article] Innate immunity mediated by dendritic cells/macrophages plays a central role in the early period in tumor treatment using gene of Mycobacterium tuberculosis antigen (2018)
  • Author(s): Takahiro Ushigusa, Yoshiyuki Koyama, Tomoko Ito, Kenichi Watanabe, James K. Chambers, Aya Hasegawa, Kazuyuki Uchida, Ryoji Kanegi, Shingo Hatoya, Toshio Inaba, Kikuya Sugiura
  • Journal Title: Journal of Veterinary Medical Science Volume: 80 Issue: 2 Pages: 190-196
  • DOI: 10.1292/jvms.17-0466
  • NAID: 130006339616
  • ISSN: 0916-7250, 1347-7439
  • Peer Reviewed / Open Access
• [Journal Article] Development of effective tumor immunotherapy using a novel dendritic cell?targeting Toll-like receptor ligand (2017)
  • Author(s): De Silva Nadeeka H.、Akazawa Takashi、Wijewardana Viskam、Inoue Norimitsu、Oyamada Maremichi、Ohta Atsuko、Tachibana Yuki、Wijesekera Daluthgamage Patsy H.、Kuwamura Mitsuru、Nishizawa Yasuko、Itoh Kazuyuki、Izawa Takeshi、Hatoya Shingo、Hasegawa Tetsuya、Yamate Jyoji、Inaba Toshio、Sugiura Kikuya
  • Journal Title: PLOS ONE Volume: 12 Issue: 11 Pages: 0188738-0188738
  • DOI: 10.1371/journal.pone.0188738
  • Peer Reviewed / Open Access
• [Journal Article] Generation of canine induced extraembryonic endoderm-like cell line that forms both extraembryonic and embryonic endoderm derivatives (2017)
  • Author(s): Nishimura T, Unezaki N, Kanegi R, Wijesekera DPH, Hatoya S, Sugiura K, Kawate N, Tamada H, Imai H, Inaba T.
  • Journal Title: Stem Cells and Development Volume: 印刷中 Issue: 15 Pages: 1111-1120
  • DOI: 10.1089/scd.2017.0026
  • Peer Reviewed / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Feeder-independent canine induced pluripotent stem cells maintained under serum-free conditions (2017)
  • Author(s): Nishimura T, Hatoya S, Kanegi R, Wijesekera DP, Sanno K, Tanaka E, Sugiura K, Hiromitsu Tamada NK, Imai H, Inaba T.
  • Journal Title: Molecular Reproduction and Development Volume: 印刷中 Issue: 4 Pages: 329-339
  • DOI: 10.1002/mrd.22789
  • Peer Reviewed / Open Access
• [Journal Article] Production of feline leukemia inhibitory factor with biological activity in Escherichia coli. (2016)
  • Author(s): Kanegi R, Hatoya S, Tsujimoto Y, Takenaka S, Nishimura T, Wijewardana V, Sugiura K, Takahashi M, Kawate N, Tamada H, Inaba T.
  • Journal Title: Theriogenology Volume: 印刷中 Issue: 2 Pages: 604-611
  • DOI: 10.1016/j.theriogenology.2016.02.013
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Positive correlation between patency and mRNA levels for cyclooxygenase-2 and prostaglandin E synthase in the uterine cervix of bitches with pyometra (2016)
  • Author(s): Tamada H, Adachi N, Kawate N, Inaba T, Hatoya S, Sawada T.
  • Journal Title: Journal of Veterinary Medical Science Volume: 78 Issue: 3 Pages: 525-528
  • DOI: 10.1292/jvms.15-0520
  • NAID: 130005140898
  • ISSN: 0916-7250, 1347-7439
  • Peer Reviewed
• [Journal Article] Expression and localization of epidermal growth factor, transforming growth factor-α and epidermal growth factor receptor in the canine testis. (2016)
  • Author(s): Tamada, H., Takemoto,K,, Tominaga, M., Kawate, N., Takahashi, M., Hatoya, S., Matsuyama, S., Inaba, T., Sawada, T.
  • Journal Title: J. Reprod. Dev. Volume: 62 Issue: 1 Pages: 59-64
  • DOI: 10.1262/jrd.2015-079
  • NAID: 130005126327
  • Peer Reviewed / Open Access
• [Journal Article] Feeding hydroalcoholic extract powder of Lepidium meyenii (maca) increases serum testosterone concentration and enhances steroidogenic ability of Leydig cells in male rats. (2016)
  • Author(s): Ohta, Y., Yoshida, K., Kamiya, S., Kawate, N., Takahashi, M., Inaba, T., Hatoya, S., Morii, H., Takahashi, K., Ito, M., Ogawa, H., Tamada, H.
  • Journal Title: Andrologia. Volume: 48 Issue: 3 Pages: 347-354
  • DOI: 10.1111/and.12453
  • Peer Reviewed / Open Access
• [Journal Article] Expression analyses of insulin-like peptide 3, RXFP2, LH receptor, and 3β-hydroxysteroid dehydrogenase in testes of normal and cryptorchid dogs. (2015)
  • Author(s): Hannan, MA., Kawate, N., Kubo, Y., Pathirana, IN., Büllesbach, EE., Hatoya, S., Inaba, T., Takahashi, M., Tamada, H.
  • Journal Title: Theriogenology. Volume: 84 Issue: 7 Pages: 1176-1184
  • DOI: 10.1016/j.theriogenology.2015.06.021
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Effect of ovarian hormones on maturation of dendritic cells from peripheral blood monocytes in dogs (2015)
  • Author(s): Wijewardana V, Sugiura K, Wijesekera DPH, Hatoya S, Nishimura T, Kanegi R, Ushigusa T, Inaba T.
  • Journal Title: Journal of Veterinary Medical Science Volume: 77 Issue: 7 Pages: 771-775
  • DOI: 10.1292/jvms.14-0558
  • NAID: 130005089954
  • ISSN: 0916-7250, 1347-7439
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Presentation] 医学・獣医学における幹細胞を用いた内科治療のエビデンス (2018)
  • Author(s): 鳩谷晋吾
  • Organizer: 日本獣医再生医療学会 第13回年次大会
  • Invited
• [Presentation] 獣医師が行う再生医療研究～小・大動物臨床への応用から畜産・動物保護～「ネコiPS細胞作製の試み」 (2018)
  • Author(s): 鳩谷晋吾
  • Organizer: 第17回日本再生医療学会総会
  • Invited
• [Presentation] 臨床応用へ向けたイヌiPS細胞の効率的な作製 (2018)
  • Author(s): 塚本雅也、鳩谷晋吾、金城綾二、大高真奈美、西村健、中西真人、稲葉俊夫、杉浦喜久弥
  • Organizer: 日本獣医再生医療学会 第13回年次大会
• [Presentation] 犬iPS細胞の基礎研究とその問題点 (2017)
  • Author(s): 鳩谷晋吾
  • Organizer: 日本再生医療学会
  • Place of Presentation: 仙台国際センター（宮城県仙台市）
  • Year and Date: 2017-03-07
  • Invited
• [Presentation] 獣医再生医療における幹細胞の役割とは (2017)
  • Author(s): 鳩谷晋吾
  • Organizer: 日本獣医再生医療学会
  • Place of Presentation: 名古屋プライムセントラルタワー（愛知県名古屋市）
  • Year and Date: 2017-02-11
  • Invited
• [Presentation] 幹細胞療法適応の展望と課題 (2017)
  • Author(s): 鳩谷晋吾
  • Organizer: 日本獣医再生医療学会
  • Place of Presentation: 名古屋プライムセントラルタワー（愛知県名古屋市）
  • Year and Date: 2017-02-11
  • Invited
• [Presentation] Primed型およびNaive型イヌiPS細胞の効率的な作製方法の検討 (2017)
  • Author(s): 塚本雅也、鳩谷晋吾、金城凌二、大高真奈美、西村健、中西真人、稲葉俊夫、杉浦喜久弥
  • Organizer: 第160回日本獣医学会学術集会
• [Presentation] ネコ精巣上体および卵巣における冷蔵保存法の検討 (2017)
  • Author(s): 花房佳祐、MD. EMTIAJ ALAM、鳩谷晋吾、辻本恭典、玉田尋通、川手憲俊、稲葉俊夫、杉浦喜久弥
  • Organizer: 第160回日本獣医学会学術集会
• [Presentation] ネコiPS細胞株のセンダイウイルスベクターを用いた作製 (2016)
  • Author(s): 沖田良太、鳩谷晋吾、杉浦喜久弥、大高真奈美、西村健、中西真人、稲葉俊夫
  • Organizer: 第159回日本獣医学会学術集会
  • Place of Presentation: 日本大学（神奈川県藤沢市）
  • Year and Date: 2016-09-06
• [Presentation] イヌ体細胞を用いた人工誘導胚体外内胚葉細胞の作製と肝細胞への分化誘導 (2016)
  • Author(s): 畦崎直哉、西村俊哉、金城綾二、Himali Wijesekera、鳩谷晋吾、杉浦喜久弥、川手憲俊、玉田尋通、今井裕、稲葉俊夫
  • Organizer: 第159回日本獣医学会学術集会
  • Place of Presentation: 日本大学（神奈川県藤沢市）
  • Year and Date: 2016-09-06
• [Presentation] GENERATION OF MULTIPOTENT CANINE EXTRAEMBRYONIC ENDODERM-LIKE CELLS VIA THE FORMATION OF INDUCED PLURIPOTENT STEM CELLS. (2016)
  • Author(s): Unezaki Naoya, Nishimura Toshiya, Kanegi Ryoji, Wijesekera Daluthgamage Patsy Himali, Sugiura, Kikuya, Hatoya Shingo, Kawate Noritoshi, Tamada Hiromichi, Imai Hiroshi and Inaba Toshio
  • Organizer: the International Society for Stem Cell Research（ISSCR） ２０１６ Annual Meeting (国際学会）
  • Place of Presentation: サンフランシスコ
  • Year and Date: 2016-06-22
  • Int'l Joint Research
• [Presentation] イヌiPS細胞のマトリゲル上での培養と間葉系幹細胞への分化誘導 (2016)
  • Author(s): 西村俊哉、田中恵里菜、鳩谷晋吾、金城綾二、Daluthgamage Pasty Himali Wijesekeral、杉浦喜久弥、玉田尋通、川手憲俊、今井裕、稲葉俊夫
  • Organizer: 第15回日本再生医療学会総会
  • Place of Presentation: 大阪国際会議場（大阪府大阪市）
  • Year and Date: 2016-03-17
• [Presentation] 再生医療 伴侶動物獣医療におけるESとiPS細胞の夢と現状 (2016)
  • Author(s): 鳩谷晋吾
  • Organizer: 第 12回 日本獣医内科学アカデミー学術大会(JCVIM 2016)
  • Place of Presentation: パシフィコ横浜（神奈川県横浜市）
  • Year and Date: 2016-02-21
  • Invited
• [Presentation] 獣医再生医療の未来（ES,iPS細胞の基礎,基礎研究,臨床応用へ） (2016)
  • Author(s): 鳩谷晋吾
  • Organizer: 第11回日本獣医再生医療学会年次大会
  • Place of Presentation: ホテルクライトン新大阪（大阪府大阪市）
  • Year and Date: 2016-02-06
  • Invited
• [Presentation] ネコの未成熟精子およびフリーズドライ精子による顕微授精 (2015)
  • Author(s): 辻本恭典、鳩谷晋吾、金子武人、杉浦喜久弥、稲葉俊夫
  • Organizer: 第158回日本獣医学会学術集会
  • Place of Presentation: 北里大学獣医学部（青森県十和田市）
  • Year and Date: 2015-09-07
• [Presentation] FEEDER-INDEPENDENT AND SERUM-FREE CANINE INDUCED PLURIPOTENT STEM CELLS REPROGRAMMED BY A DRUG-INDUCIBLE SYSTEM (2015)
  • Author(s): Nishimura Toshiya, Kanegi Ryoji, Wijesekera Daluthgamage Patsy Himali, Sanno Kousuke, Tanaka Erina, Sugiura Kikuya, Hatoya Shingo, Kawate Noritoshi, Tamada Hiromichi, Imai Hiroshi, Inaba Toshio
  • Organizer: The International Society for Stem Cell Research (ISSCR) 2015 Annual Meeting.
  • Place of Presentation: Stockholmsmassan Exhibition and Convention Center, Stockholm, Sweden
  • Year and Date: 2015-06-24
  • Int'l Joint Research
• [Remarks] 細胞病態学教室ホームページ
  • URL: http://www.vet.osakafu-u.ac.jp/cell/
• [Remarks] 研究室ホームページ
  • URL: http://www.vet.osakafu-u.ac.jp/cell/
• [Remarks] http://www.vet.osakafu-u.ac.jp/cell/