Cellular Internalization of Monomeric and Oligomeric Amyloid-Beta 42 Peptide

Abstract

Amyloid beta (Abeta) is the major component of extracellular plaques found in Alzheimer's disease (AD). Uptake of Abeta from extracellular to intracellular space, which is likely by endocytosis, appears to be an important process for understanding AD pathology (Friedrich et al., PNAS 2010; Hu et al., PNAS 2009). It may also be a promising target for treatment and prevention.We aimed to study biophysical details of Abeta uptake, and to determine the uptake kinetics for different Abeta forms: monomers, oligomers, higher molecular mass aggregates of Abeta in and small fibrils. We used neuroblastoma (Sh-EP) cells as a model for neurons. Uptakes kinetic were followed by using the fluorescent labelled Abeta 42 that was monomerized and was purified by gel filtration. Abeta 42 peptide was added to the cell culture medium and the amount of intracellular aggregates was quantified using automated fluorescence microscopy.We compared uptake kinetics and aggregation kinetics in buffer and in cell culture medium at different Abeta 42 concentrations to test whether aggregation precedes uptake or vice versa. To compare the uptake of different Abeta species, pre-aggregated Abeta oligomers or small fibrils were added to the cells. We found that pre-aggregation accelerated the formation of intracellular aggregates, which suggests that Abeta oligomers and / or small fibrils may be taken up more rapidly than Abeta monomer.In the future the form and location of intracellular Abeta will be monitored by high resolution fluorescence nanoscopy combined with atomic force microscopy.