Three-dimensional real time imaging of amyloid β aggregation on living cells

Abstract

Alzheimer’s disease (AD) is a progressive disorder of the brain that gradually decreases thinking, memory, and language abilities. The aggregation process of amyloid β (Aβ) is a key step in the expression of its neurocytotoxicity and development of AD because Aβ aggregation and accumulation around neuronal cells induces cell death. However, the molecular mechanism underlying the neurocytotoxicity and cell death by Aβ aggregation has not been clearly elucidated. In this study, we successfully visualized real-time process of Aβ42 aggregation around living cells by applying our established QD imaging method. 3D observations using confocal laser microscopy revealed that Aβ42 preferentially started to aggregate at the region where membrane protrusions frequently formed. Furthermore, we found that inhibition of actin polymerization using cytochalasin D reduced aggregation of Aβ42 on the cell surface. These results indicate that actin polymerization-dependent cell motility is responsible for the promotion of Aβ42 aggregation at the cell periphery. 3D observation also revealed that the aggregates around the cell remained in that location even if cell death occurred, implying that amyloid plaques found in the AD brain grew from the debris of dead cells that accumulated Aβ42 aggregates.

Enhanced neprilysin-mediated degradation of hippocampal Aβ42 with a somatostatin peptide that enters the brain

“…A number of previous studies indicate that the intermediate soluble species are the most toxic forms of Aβ 6 - 9 . Interaction of Aβ with the membrane of the neurons can induce cell toxicity and affect its aggregation cascade 10 , 11 . These point towards the significance of targeting membrane-bound Aβ over the soluble one.…”

Dietary Intake of Rosmarinic Acid Increases Serum Inhibitory Activity in Amyloid A Aggregation and Suppresses Deposition in the Organs of Mice

“…Previously, we reported a real-time imaging method of amyloid β (Aβ 42 ) and tau aggregation using quantum-dot (QD) nanoprobes and developed a microliter-scale high-throughput screening system (MSHTS) for aggregation inhibitors that applied this imaging method [ 27 , 28 , 29 , 30 , 31 ]. This screening system can be used to analyze a 5 µL sample volume when a 1536-well plate is used, and inhibitory activity can be estimated as half-maximal effective concentration (EC 50 ).…”

Functional Switching of a Native Neuropeptide Through Interactions with Pathogenic Factors in Dementia

“…To identify a relationship between the decrease in the cytotoxicity of Cu(II)-Ab42 and Zn(II)-Ab42 by SST and their interactions with cell membranes, we carried out immunocytochemistry studies. After treatment of metal-Ab and metal-free Ab added with or without SST to living cells, cytoplasmic membranes were stained by a fluorescent membrane dye, CellBrite green 45,52 , followed by immunoblotting Ab species using an anti-Ab antibody (6E10) and the Alexa Fluor 647goat anti-mouse secondary antibody, as illustrated in Fig. 6c and Supplementary Figs.…”

“…When Ab species are positioned on the cellular surface, the green and red fluorescence signals are overlapped 52 . To quantitatively analyze the interaction between Ab species and cell membranes, the Pearson's correlation coefficient (PCC), indicative of the degree of the overlap between the two fluorophores, was calculated.…”