Single-cell analysis of autoimmune encephalitis of unknown etiology for treatment stratification

• Project/Area Number: 20K16577
• Research Category: Grant-in-Aid for Early-Career Scientists
• Allocation Type: Multi-year Fund
• Review Section: Basic Section 52020:Neurology-related
• Research Institution: Osaka University
• Principal Investigator: Ishikura Teruyuki 大阪大学, 大学院医学系研究科, 招へい教員 (80838524)
• Project Period (FY): 2020-04-01 – 2023-03-31
• Project Status: Completed (Fiscal Year 2022)
• Budget Amount: ¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000); Fiscal Year 2022: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000); Fiscal Year 2021: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000); Fiscal Year 2020: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
• Keywords: 自己免疫性脳炎 / 視神経脊髄炎スペクトラム障害 / アクアポリン4抗体 / 神経障害性疼痛 / シングルセル解析 / CyTOF / NMOSD / NORSE

Outline of Research at the Start

自己免疫介在性脳炎(Autoimmne-mediated encephalitis:AIE)は幅広い年齢層に発症し、高度の神経障害が残存する難治性疾患である。近年は抗NMDA受容体抗体を代表とする様々な疾患特異的自己抗体が同定され、分離不能であった疾患群が層別化され、各種特異的脳炎スペクトラムの確定診断と個別化治療が可能となってきた。しかし未だ約半数のAIEで特異的自己抗体は陰性であり、病態機序が不明であり、経験的治療に委ねられている。本研究ではAIEにおいて、革新的免疫解析法であるsingle-cell解析を駆使し、新規バイオマーカーの確立、免疫動態解明による治療層別化を目指す。

Outline of Final Research Achievements

We collected clinical information, cerebral spinal fluid, serum, and peripheral blood mononuclear cells from autoimmune encephalitis patients. We generated patient-derived aquaporin 4 antibody by Single-cell sorting of cerebral spinal fluid plasmablasts from neuromyelitis optica spectrum disorder (NMOSD) patients. By injecting the antibody directly into the rat spinal cord, we created a NMOSD pain animal model and clarified the pain mechanism focusing on ATP. Currently, we are still screening for anti-neuronal antibodies in cerebral spinal fluid and serum of autoimmune encephalitis patients using rat brain sections, and some samples containing anti-neuronal antibodies have been collected. We hope to create an animal model in the future by generating pathogenic autoantibodies from cerebral spinal fluid cells and peripheral blood mononuclear cells from the autoimmune encephalitis patients.

Academic Significance and Societal Importance of the Research Achievements

NMOSD患者の疼痛を呈する割合は高く、薬剤抵抗性であり、QOLを大きく低下させるため臨床的に大きな問題となっている。今回動物モデルを用いてNMOSD疼痛にATPが重要であることを示し、治療方法に結び付くと考えられる。またシングルセルソーティングによって得られた患者由来自己抗体から動物モデルを作成する手法は、自己免疫性脳炎にも今後適応することができると考えている。

Research Products

• [Journal Article] Cryptogenic new-onset refractory status epilepticus responded to anti-interleukin-6 treatment (2022)
  • Author(s): Tomoya Wadayama, Mikito Shimizu, Tomohiro Yata, Teruyuki Ishikura, Yuta Kajiyama, Daisuke Hirozawa, Tatsusada Okuno, Hideki Mochizuki
  • Journal Title: Journal of Neuroimmunology Volume: 363 Pages: 577789-577789
  • DOI: 10.1016/j.jneuroim.2021.577789
  • Peer Reviewed
• [Journal Article] High cell surface expression and peptide binding affinity of HLA-DQA1*05:03, a susceptible allele of neuromyelitis optica spectrum disorders (NMOSD) (2022)
  • Author(s): Beppu Shohei、Kinoshita Makoto、Wilamowski Jan、Suenaga Tadahiro、Yasumizu Yoshiaki、Ogawa Kotaro、Ishikura Teruyuki、Tada Satoru、Koda Toru、Murata Hisashi、Shiraishi Naoyuki、Sugiyama Yasuko、Kihara Keigo、Sugimoto Tomoyuki、Arase Hisashi、Standley Daron M.、Okuno Tatsusada、Mochizuki Hideki
  • Journal Title: Scientific Reports Volume: 12 Issue: 1 Pages: 1-12
  • DOI: 10.1038/s41598-021-04074-1
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Anti-AQP4 autoantibodies promote ATP release from astrocytes and induce mechanical pain in rats (2021)
  • Author(s): Ishikura Teruyuki、Kinoshita Makoto、Shimizu Mikito、Murata Hisashi、Beppu Shohei、Koda Toru、Tada Satoru、Miyamoto Katsuichi、Kusunoki Susumu、Sugimoto Tomoyuki、Kumanogoh Atsushi、Okuno Tatsusada、Mochizuki Hideki
  • Journal Title: Journal of Neuroinflammation Volume: 18 Issue: 1 Pages: 1-12
  • DOI: 10.1186/s12974-021-02232-w
  • Peer Reviewed / Open Access
• [Journal Article] Prenatal clinical manifestations in individuals with COL4A1/2 variants (2020)
  • Author(s): Itai T, Miyatake S, Taguri M, Nakashima M, Saitsu H, Matsumoto N et al.
  • Journal Title: J Med Genet Volume: 0 Issue: 8 Pages: 106896-106896
  • DOI: 10.1136/jmedgenet-2020-106896
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Sema4A is implicated in the acceleration of Th17 cell-mediated neuroinflammation in the effector phase (2020)
  • Author(s): Koda Toru、Namba Akiko、Kinoshita Makoto、Nakatsuji Yuji、Sugimoto Tomoyuki、Sakakibara Kaori、Tada Satoru、Shimizu Mikito、Yamashita Kazuya、Takata Kazushiro、Ishikura Teruyuki、Murata Syo、Beppu Shohei、Kumanogoh Atsushi、Mochizuki Hideki、Okuno Tatsusada
  • Journal Title: Journal of Neuroinflammation Volume: 17 Issue: 1 Pages: 1-9
  • DOI: 10.1186/s12974-020-01757-w
  • Peer Reviewed / Open Access
• [Journal Article] Mitochondrial DNA enhance innate immune responses in neuromyelitis optica by monocyte recruitment and activation (2020)
  • Author(s): Mikito Shimizu, Tatsusada Okuno, Makoto Kinoshita, Hisae Sumi, Harutoshi Fujimura, Kazuya Yamashita, Tomoyuki Sugimoto, Shuhei Sakakibara, Kaori Sakakibara, Toru Koda, Satoru Tada, Teruyuki Ishikura, Hisashi Murata, Shohei Beppu, Naoyuki Shiraishi, Yasuko Sugiyama, Yuji Nakatsuji, Atsushi Kumanogoh & Hideki Mochizuki
  • Journal Title: Scientific Reports Volume: 10 Issue: 1 Pages: 13274-13274
  • DOI: 10.1038/s41598-020-70203-x
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Presentation] ATP mediates neuropathic pain in neuromyelitis optica via microglial activation (2021)
  • Author(s): Teruyuki Ishikura, Makoto Kinoshita, Tatsusada Okuno, Hideki Mochiduki
  • Organizer: 15th ISNI Congress - International Society of Neuroimmunology
• [Presentation] 新規NMOSD疼痛動物モデルとATP、ミクログリア、IL-1βによる疼痛機序 (2021)
  • Author(s): 石倉照之、木下允、甲田亨、宮本勝一、奥野龍禎、望月秀樹
  • Organizer: 第39回日本神経治療学会
• [Presentation] ATP induces neuropathic pain in neuromyelitis optica spectrum disorder through microglial activation (2021)
  • Author(s): Teruyuki Ishikura
  • Organizer: 第62回日本神経学会学術大会
• [Presentation] 新規NMOSD疼痛モデル動物樹立とATPシグナル経路解明 (2020)
  • Author(s): 石倉照之
  • Organizer: 日本神経免疫学会