Development of in vitro amyloid fibril formation systems that mimic the physiological fibrillogenesis conditions in vivo and their application to the analysis of fibril formation mechanism

• Project/Area Number: 15K06965
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Structural biochemistry
• Research Institution: University of Fukui
• Principal Investigator: Hasegawa Kazuhiro 福井大学, 学術研究院医学系部門, 助教 (60324159)
• Co-Investigator(Kenkyū-buntansha): 内木 宏延 福井大学, 学術研究院医学系部門, 教授 (10227704)
• Research Collaborator: Endo Yoshinori
• Project Period (FY): 2015-04-01 – 2019-03-31
• Project Status: Completed (Fiscal Year 2018)
• Budget Amount: ¥5,070,000 (Direct Cost: ¥3,900,000、Indirect Cost: ¥1,170,000); Fiscal Year 2017: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000); Fiscal Year 2016: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000); Fiscal Year 2015: ¥2,990,000 (Direct Cost: ¥2,300,000、Indirect Cost: ¥690,000)
• Keywords: 蛋白質 / 変性とフォールディング / 病理学 / アミロイド / アルツハイマー病

Outline of Final Research Achievements

We evaluate the roles of the various biological molecules or reaction conditions in the amyloid fibril formation using amyloid fibril formation systems in vitro. (1) We examined the effects of apoE and clusterin on the fibril formation of Alzheimer’s disease amyloid β-peptide (Aβ) using the previously developed near-physiological Aβ fibril formation system for evaluating the efficiency of protein components to induce the nucleation of Aβ peptide. We found that physiological concentrations of apoE and clusterin delayed the initiation time of amyloid growth kinetics in a concentration-dependent manner. (2) To further decrease the concentration of Aβ peptide in the reaction, we developed a high-sensitive system, in which the elongation of amyloid fibrils by sub-micromolar concentrations of Aβ peptide can be detected. (3) We tried to identify the proteins which induce the nucleation of beta2-microglobulin amyloid, but the reaction system was not sufficient to detect them.

Academic Significance and Societal Importance of the Research Achievements

アルツハイマー病(AD)患者が急増しており、根本的な予防法・治療法の開発が急務である。Aβ蛋白質(Aβ)は、ADの病態の最上流部にあるとされている。アミロイド線維形成など、Aβの脳内での挙動を解明する為に、試験管内で正確に再現することが必要。現状では、脳内の生理濃度(nM)よりかなり高濃度のAβを用いて解析しており、反応機構の再現が不完全な可能性がある。より低濃度で反応する試験管内反応系を構築し、重合、オリゴマー形成、共存蛋白質群との相互作用などを正確に再現し、予防・治療法の開発に役立てる。β2-mアミロイドでも生体における重合機序が完全には解明されておらず、試験管内重合反応系の構築が必要。

Research Products

• [Journal Article] Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy (2019)
  • Author(s): Endo Y, Hasegawa K, Nomura R, Arishima H, Kikuta K, Yamashita T, Inoue Y, Ueda M, Ando Y, Wilson MR, Hamano T, Nakamoto Y, Naiki H.
  • Journal Title: Acta Neuropathologica Communications Volume: 7 Issue: 1 Pages: 12-12
  • DOI: 10.1186/s40478-019-0662-1
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Antiamyloidogenic and proamyloidogenic chaperone effects of C-reactive protein and serum amyloid P component. (2017)
  • Author(s): Ozawa D, Nomura R, Mangione PP, Hasegawa K, Okoshi T, Porcari R, Bellotti V, Naiki H.
  • Journal Title: Amyloid Volume: 24(sup1) Issue: sup1 Pages: 28-29
  • DOI: 10.1080/13506129.2017.1295943
  • Int'l Joint Research
• [Journal Article] Multifaceted anti-amyloidogenic and pro-amyloidogenic effects of C-reactive protein and serum amyloid P component in vitro. (2016)
  • Author(s): Ozawa D, Nomura R, Mangione PP, Hasegawa K, Okoshi T, Porcari R, Bellotti V, Naiki H
  • Journal Title: Sci Rep Volume: 6 Issue: 1 Pages: 29077-29077
  • DOI: 10.1038/srep29077
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Molecular pathogenesis ofhuman amyloidosis: Lessons from β2 -microglobulin-related amyloidosis. (2016)
  • Author(s): Naiki H, Okoshi T, Ozawa D, Yamaguchi I, Hasegawa K.
  • Journal Title: Pathol Int. Volume: ;66(4) Issue: 4 Pages: 193-201
  • DOI: 10.1111/pin.12394
  • Peer Reviewed / Open Access
• [Journal Article] Endocytosed β2-Microglobulin Amyloid Fibrils Induce Necrosis and Apoptosis of Rabbit Synovial Fibroblasts by Disrupting Endosomal/Lysosomal Membranes: A Novel Mechanism on the Cytotoxicity of Amyloid Fibrils. (2015)
  • Author(s): Okoshi T, Yamaguchi I, Ozawa D, Hasegawa K, Naiki H
  • Journal Title: PLoS One Volume: 10 Issue: 9 Pages: e0139330-e0139330
  • DOI: 10.1371/journal.pone.0139330
  • Peer Reviewed / Open Access
• [Presentation] C反応性蛋白質と血清アミロイドP成分の多面的シャペロン効果 (2016)
  • Author(s): 小澤 大作, 野村 寮, Patrizia P. Mangione, 長谷川 一浩, 大越 忠和, Riccardo Porcari, Vittorio Bellotti, 内木 宏延
  • Organizer: 第16回日本蛋白質科学会年会
  • Place of Presentation: 福岡国際会議場, 福岡市
  • Year and Date: 2016-06-07