Aggregated amyloid-β(1-42) is internalized more efficiently than monomers by HEK293T cells.

An early and continual event in Alzheimer's disease (AD) is upregulation of the immune response concomitant with accumulation of aggregated amyloid-β peptide (Aβ). The mechanisms by which Aβ can trigger an immune response involve stimulation of cell surface receptors and intracellular pathways, as well as internalization of Aβ into the cell cytosol. The HEK293T cell line has been a valuable model system for exploring a variety of biological processes, including internalization of protein aggregates such as Aβ. There have been some questions regarding the role of Aβ conformation in the HEK293T internalization process. The current study investigated the influence of Aβ42 conformation on HEK293T internalization using ELISA, confocal microscopy and flow cytometry. Analysis of HEK293T cell lysates by ELISA after exposure to Aβ42 conformational species indicated that protofibrillar and fibrillar Aβ42 were internalized to a much greater extent compared to purified Aβ42 monomers. The same trend was observed in HEK293T whole cells by confocal fluorescence imaging. HEK293T cell internalization occurred by 4 h for aggregated Aβ42 at concentrations in the low micromolar range. Fluorescent labeling of Aβ42 did not impact cell internalization and facilitated the use of flow cytometry as a third strategy to show significantly greater internalization of Aβ42 protofibrils compared to monomers, further confirming the Aβ42 conformational influence on HEK293T cell internalization. HEK293T cell models are valuable tools for mechanistic investigations of human disease and characteristically generate detailed information about cellular processes or pathways. These include Aβ cellular internalization and intracellular Aβ, which are key aspects of AD pathogenesis.