Impact of Four Common Hydrogels on Amyloid-β (Aβ) Aggregation and Cytotoxicity: Implications for 3D Models of Alzheimer’s Disease

[Image: see text] The physiochemical properties of hydrogels utilized in 3D culture can be used to modulate cell phenotype and morphology with a striking resemblance to cellular processes that occur in vivo. Indeed, research areas including regenerative medicine, tissue engineering, in vitro cancer models, and stem cell differentiation have readily utilized 3D biomaterials to investigate cell biological questions. However, cells are only one component of this biomimetic milieu. In many models of disease such as Alzheimer’s disease (AD) that could benefit from the in vivo-like cell morphology associated with 3D culture, other aspects of the disease such as protein aggregation have yet to be methodically considered in this 3D context. A hallmark of AD is the accumulation of the peptide amyloid-β (Aβ), whose aggregation is associated with neurotoxicity. We have previously demonstrated the attenuation of Aβ cytotoxicity when cells were cultured within type I collagen hydrogels versus on 2D substrates. In this work, we investigated the extent to which this phenomenon is conserved when Aβ is confined within hydrogels of varying physiochemical properties, notably mesh size and bioactivity. We investigated the Aβ structure and aggregation kinetics in solution and hydrogels composed of type I collagen, agarose, hyaluronic acid, and polyethylene glycol using fluorescence correlation spectroscopy and thioflavin T assays. Our results reveal that all hydrogels tested were associated with enhanced Aβ aggregation and Aβ cytotoxicity attenuation. We suggest that confinement itself imparts a profound effect, possibly by stabilizing Aβ structures and shifting the aggregate equilibrium toward larger species. If this phenomenon of altered protein aggregation in 3D hydrogels can be generalized to other contexts including the in vivo environment, it may be necessary to reevaluate aspects of protein aggregation disease models used for drug discovery.