title: Medicinal value of asiaticoside for Alzheimer's disease as assessed using single-molecule-detection fluorescence correlation spectroscopy, laser-scanning microscopy, transmission electron microscopy, and in silico docking.
abstract: BACKGROUND
  Identifying agents that inhibit amyloid beta peptide (Aβ) aggregation is the ultimate goal for slowing Alzheimer's disease (AD) progression. This study investigated whether the glycoside asiaticoside inhibits Aβ1-42 fibrillation in vitro.
  
  
  METHODS
  Fluorescence correlation spectroscopy (FCS), evaluating the Brownian diffusion times of moving particles in a small confocal volume at the single-molecule level, was used. If asiaticoside inhibits early Aβ1-42 fibrillation steps, more Aβs would remain free and rapidly diffuse in the confocal volume. In contrast, "weaker or no inhibition" permits a greater number of Aβs to polymerize into oligomers, leading to fibers and gives rise to slow diffusion times in the solution. Trace amounts of 5-carboxytetramethylrhodamine (TAMRA)-labeled Aβ1-42 in the presence of excess unlabeled Aβ1-42 (10 μM) was used as a fluorescent probe. Steady-state and kinetic-Thioflavin T (ThT) fluorospectroscopy, laser-scanning fluorescence microscopy (LSM), and transmission electron microscopy (TEM) were also used to monitor fibrillation. Binding of asiaticoside with Aβ1-42 at the atomic level was computationally examined using the Molegro Virtual Docker and PatchDock.
  
  
  RESULTS
  With 1 h of incubation time for aggregation, FCS data analysis revealed that the diffusion time of TAMRA-Aβ1-42 was 208 ± 4 μs, which decreased to 164 ± 8.0 μs in the presence of asiaticoside, clearly indicating that asiaticoside inhibited the early stages Aβ1-42 of fibrillation, leaving more free Aβs in the solution and permitting their rapid diffusion in the confocal volume. The inhibitory effects were also evidenced by reduced fiber formation as assessed by steady-state and kinetic ThT fluorospectroscopy, LSM, and TEM. Asiaticoside elongated the lag phase of Aβ1-42 fibrillation, indicating the formation of smaller amyloid species were impaired in the presence of asiaticoside. Molecular docking revealed that asiaticoside binds with amyloid intra- and inter-molecular amino acid residues, which are responsible for β-sheet formation and longitudinal extension of fibrils.
  
  
  CONCLUSIONS
  Finally, asiaticoside prevents amyloidogenesis that precedes neurodegeneration in patients with Alzheimer's disease.
authors.affiliation: Department of Environmental Physiology, Shimane University Faculty of Medicine, Izumo, 693-8501, Japan. shahdat@dhaka.net.|Department of Biochemistry and Molecular Biology, Jahangirnagar University, Savar, Dhaka, Bangladesh. shahdat@dhaka.net.
authors.forename: Shahdat
authors.initials: S
authors.lastname: Hossain
authors.affiliation: Department of Environmental Physiology, Shimane University Faculty of Medicine, Izumo, 693-8501, Japan. michio1@med.shimane-u.ac.jp.
authors.forename: Michio
authors.initials: M
authors.lastname: Hashimoto
authors.affiliation: Department of Environmental Physiology, Shimane University Faculty of Medicine, Izumo, 693-8501, Japan. katakura@med.shimane-u.ac.jp.
authors.forename: Masanori
authors.initials: M
authors.lastname: Katakura
authors.affiliation: Department of Environmental Physiology, Shimane University Faculty of Medicine, Izumo, 693-8501, Japan. mamun104@gmail.com.
authors.forename: Abdullah
authors.initials: A
authors.lastname: Al Mamun
authors.affiliation: Department of Environmental Physiology, Shimane University Faculty of Medicine, Izumo, 693-8501, Japan. o-shido@med.shimane-u.ac.jp.
authors.forename: Osamu
authors.initials: O
authors.lastname: Shido
mesh_terms: D000544:Alzheimer Disease / Q000378:metabolism* / Q000517:prevention & control; D000682:Amyloid / Q000378:metabolism*; D016229:Amyloid beta-Peptides / Q000378:metabolism*; D028041:Centella / Q000737:chemistry*; D005453:Fluorescence; D006801:Humans; D008853:Microscopy / Q000379:methods; D062105:Molecular Docking Simulation / Q000379:methods; D010446:Peptide Fragments / Q000378:metabolism*; D008517:Phytotherapy; D010936:Plant Extracts / Q000494:pharmacology* / Q000627:therapeutic use; D012235:Rhodamines / Q000378:metabolism; D013057:Spectrum Analysis / Q000379:methods; D014315:Triterpenes / Q000494:pharmacology* / Q000627:therapeutic use
chemical_list: C437523:5-carboxytetramethylrhodamine; D000682:Amyloid; D016229:Amyloid beta-Peptides; D010446:Peptide Fragments; D010936:Plant Extracts; D012235:Rhodamines; D014315:Triterpenes; C075222:amyloid beta-protein (1-42); C004446:asiaticoside