Analysis of the effect of Aβ fibril polymorphism on toxicity, degradation and activation of an inflammatory response

Alzheimer’s disease (AD) is the most common form of dementia and has a large social, medical and economic impact. One of the main pathological hallmarks of AD is the presence of extracellular plaques consisting primarily of β-amyloid (Aβ). Aβ fibrils share a characteristic cross-β structure, however structural models for Aβ fibrils have revealed polymorphism at a molecular level in vitro and there is also evidence that distinct structural variants could be linked to the presentation of different subtypes of AD.

Working at the interface between structural and cellular biology, this research investigates differences in the cellular responses to different Aβ fibril preparations. Distinct populations of Aβ fibrils were formed from recombinantly purified Aβ monomers, reproducing fibrillation conditions previously used in the production of fibrils from which structures have been published. Namely, 2A, 3Q and de novo Aβ40 fibril populations were produced, in addition to fibrils formed from a familial mutant form of the peptide, E22∆ Aβ40. Further to this, two populations of Aβ42 fibrils were compared; one formed at pH 8 and the other formed at pH 2. Fibrils were characterised using transmission electron microscopy (TEM), Thioflavin T (ThT) monitoring, oligothiophene probes and fibril yield analysis.

With neuroinflammation being another major hallmark of AD, this research focuses specifically on the effects of these different Aβ fibril populations on monocytic and microglial cells. A combination of MTT, ATP and LDH viability assays were performed which revealed differences in the toxicity of the different Aβ fibrils towards BV-2 microglial cells, RAW 264.7 macrophage cells and monocytic THP-1 cells. Aβ42 fibrils that were formed at pH 8 were found to be significantly more toxic than the population of Aβ42 fibrils that were formed at pH 2. E22∆ Aβ40 fibrils were also shown to be more toxic towards these cells than the fibril populations that were formed from wild-type Aβ40 peptide.

Further to this, differences in the levels of the pro-inflammatory cytokines IL-6, IL-1β and TNF-α released from cells in response to the different fibrils were identified, with the population of Aβ42 fibrils produced at pH 8 resulting in the greatest release compared to other fibril populations. These fibrils were also found to associate more with cells, possibly helping to explain this increased toxicity and inflammatory response. Furthermore, differences in the efficiency of clearance by microglial cells of Aβ40 fibrils were also identified, with reduced 2A Aβ40 fibril clearance compared to other fibril populations. Differences identified between these Aβ fibrils could help to explain the huge amount of variability in symptoms and disease severity that exists between Alzheimer’s patients.