Modeling Inherited Arrhythmia Disorders Using Patient-Derived Induced Pluripotent Stem Cells

2018 Fiscal Year Annual Research Report

• Project/Area Number: 16K09499
• Research Institution: Kyoto University
• Principal Investigator: 牧山 武 京都大学, 医学研究科, 助教 (30528302)
• Project Period (FY): 2016-04-01 – 2019-03-31
• Keywords: 不整脈 / 遺伝子 / iPS細胞 / 心筋症 / 疾患モデル

Outline of Annual Research Achievements

１．カテコラミン誘発性多形性心室頻拍（CPVT）関連リアノジン受容体変異に関する3Dシミュレーションモデルを用いた検討
CPVTは、運動時に重症心室性不整脈による失神、突然死を来す遺伝性不整脈疾患であり、約70%に心臓リアノジン受容体（RyR2）をコードするRYR2遺伝子の変異を認めが、巨大な蛋白なため変異チャネルの機能解析は困難である。本研究では、CPVT症例にて検出されたRYR2変異93個に関して凍結電顕3Dシミュレーションモデルを用いた構造学的検討を行った。結果、アミノ酸の電荷・極性の変化を来す変異だけではなく、チャネルの重要な部位（ドメイン間のヒンジ部、チャネル孔、FKBP蛋白結合部位、Caセンサー）に近接する変異が多く認められ、これらの変異によりリアノジン受容体が容易にCaを放出し催不整脈性を来すと推察された。本研究により、3DモデルがRyR2チャネル機能異常の推察に有用である可能性が示唆された。
２．心臓電位依存性L型カルシウムチャネル（LTCC）のイオン選択性異常によるQT延長症候群、ブルガダ症候群オーバーラップ症例における患者由来iPS細胞研究
LTCCは、Caイオン選択性に関してイオン孔に”グルタミン酸リング（EEEE）”が非常に重要である。我々は、リングの一つのグルタミン酸がリシン（K）に置換される変異が検出されたQT延長症候群症例において、患者由来iPS細胞を作製し、分化心筋の解析を行った。パッチクランプ法を用いた電気生理学的機能解析では、患者由来iPS細胞分化心筋では、イオン選択性が低下したことによると思われる内向きCa電流の低下、異常な遅延内向きNa電流を認め、健常人由来分化心筋と比較し、有意な活動電位持続時間延長を認めた。これらの機能変化は、患者の表現型に合致する所見と考えら、今後、本iPS細胞モデルを用いた薬効評価を予定している。

Research Products

• [Journal Article] Association of Genetic and Clinical Aspects of Congenital Long QT Syndrome With Life-Threatening Arrhythmias in Japanese Patients (2019)
  • Author(s): Shimizu Wataru、Makimoto Hisaki、Yamagata Kenichiro、Kamakura Tsukasa、Wada Mitsuru、Miyamoto Koji、Inoue-Yamada Yuko、Okamura Hideo、Ishibashi Kohei、Noda Takashi、Nagase Satoshi、Miyazaki Aya、Sakaguchi Heima、Shiraishi Isao、Makiyama Takeru、Aiba Takeshi, et al.
  • Journal Title: JAMA Cardiology Volume: - Pages: -
  • DOI: 10.1001/jamacardio.2018.4925
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Complex aberrant splicing in the induced pluripotent stem cell?derived cardiomyocytes from a patient with long QT syndrome carrying KCNQ1-A344Aspl mutation (2018)
  • Author(s): Wuriyanghai Yimin、Makiyama Takeru、Sasaki Kenichi、Kamakura Tsukasa、Yamamoto Yuta、Hayano Mamoru、Harita Takeshi、Nishiuchi Suguru、Chen Jiarong、Kohjitani Hirohiko、Hirose Sayako、Yokoi Fumika、Gao Jingshan、Chonabayashi Kazuhisa、Watanabe Ken、Ohno Seiko、Yoshida Yoshinori、Kimura Takeshi、Horie Minoru
  • Journal Title: Heart Rhythm Volume: 15 Pages: 1566～1574
  • DOI: 10.1016/j.hrthm.2018.05.028
  • Peer Reviewed
• [Journal Article] Clinical Manifestations and Long-Term Mortality in <i>Lamin A/C</i> Mutation Carriers From a Japanese Multicenter Registry (2018)
  • Author(s): Nakajima Kenzaburo、Aiba Takeshi、Makiyama Takeru、Kusano Kengo, et al.
  • Journal Title: Circulation Journal Volume: 82 Pages: 2707～2714
  • DOI: 10.1253/circj.CJ-18-0339
  • Peer Reviewed
• [Journal Article] Differential Diagnosis Between Catecholaminergic Polymorphic Ventricular Tachycardia and Long QT Syndrome Type 1 ― Modified Schwartz Score ― (2018)
  • Author(s): Ozawa Junichi、Ohno Seiko、Fujii Yusuke、Makiyama Takeru、Suzuki Hiroshi、Saitoh Akihiko、Horie Minoru
  • Journal Title: Circulation Journal Volume: 82 Pages: 2269～2276
  • DOI: 10.1253/circj.CJ-17-1032
  • Peer Reviewed
• [Presentation] Modelling long-QT syndrome with human iPS cells and development of new pharmacotherapies (2018)
  • Author(s): Takeru Makiyama
  • Organizer: 23rd International Society of Cardiovascular Pharmacotherapy (ISCP)
  • Int'l Joint Research / Invited
• [Presentation] Human induced pluripotent stem cell models for inherited arrhythmias (2018)
  • Author(s): Takeru Makiyama
  • Organizer: The 2018 International Congress on Electrocardiology (ICE 2018)
  • Int'l Joint Research / Invited
• [Presentation] Modelling calmodulin-related long-QT syndrome using human iPS cells: therapeutic genome editing using CRISPR/Cas9 system (2018)
  • Author(s): Takeru Makiyama
  • Organizer: 第65回日本不整脈心電学会学術大会
  • Invited
• [Presentation] Can patient-derived iPSCs be applied to phenotypic cell-based high throughput screening assessment to distinguish between different types of long-QT syndrome? (2018)
  • Author(s): Daisuke Yoshinaga, Shiro Baba, Hirofumi Shibata, Takuya Hirata, Takeru Makiyama, Ryuta Nishikomori
  • Organizer: European Society of Cardiology Congress 2018
  • Int'l Joint Research
• [Presentation] Significance of cell-specific precise computer simulation using new mathematical models of human induced pluripotent stem cell derived cardiomyocyte in drug testing (2018)
  • Author(s): Hirohiko Kohjitani, Shigeya Kouda, Yukiko Himeno, Takeru Makiyama, Fumika Yokoi, Sayako Hirose, Yimin Wuriyanghai, Yuta Yamamoto, Minoru Horie, Takeshi Kimura, Akinori Noma, Akira Amano
  • Organizer: European Society of Cardiology Congress 2018
  • Int'l Joint Research
• [Presentation] Analysis of Disease-Causing Mechanism of Lamin A/C-Related Cardiomyopathy Using Homozygous Lamin A/C Knock out Human Induced Pluripotent Stem Cell Model (2018)
  • Author(s): Yimin Wuriyanghai, Takeru Makiyama, Yuta Yamamoto, Suguru, Nishiuchi, Sayako Hirose, Hirohiko Kohjitani, Jingshan Gao, Asami Kashiwa, Seiko Ohno, Minoru Horie, Takeshi Kimura
  • Organizer: The 11th Asia Pacific Heart Rhythm Society Scientific Session
  • Int'l Joint Research
• [Presentation] Three-dimensional Structural Analysis of Mutant Ryanodine Receptor Type 2 Channels Associated with Catecolaminergic Polymorphic Ventricular Tachycardia (2018)
  • Author(s): Jingshan Gao, Takeru Makiyama, Seiko Ohno, Yuta Yamamoto, Yimin Wuriyanghai, Hirohiko Kohjitani, Asami Kashiwa, Sayako Hirose, Minoru Horie, Takeshi Kimura
  • Organizer: The 11th Asia Pacific Heart Rhythm Society Scientific Session
  • Int'l Joint Research
• [Presentation] Overlapping Sick Sinus Syndrome Associated with SCN5A Mutations in Patients with Brugada Syndrome (2018)
  • Author(s): Asami Kashiwa, Takeru Makiyama, Seiko Ohno, Hisaaki Aoki, Hirohiko Kohjitani Yimin Wuriyanghai, Yuta Yamamoto, Jingshan Gao, Huang Hai, Minoru Horie, Takeshi Kimura
  • Organizer: 第83回日本循環器学会学術集会