Amyloid strains and Alzheimer disease : roles in the propagation of amyloid pathology and amyloid/tau interaction

Etablissement Université Grenoble Alpes
École doctorale CSV- Chimie et Sciences du Vivant
Spécialité Neurosciences - Neurobiologie
Unité de recherche Grenoble Institut des Neurosciences
Encadrement de la thèse Alain BUISSON
Financement du 01-10-2024 au 01-10-2027 origine ministère de l'enseignement supérieur et de la recherche
Début de la thèse le 1 octobre 2024
Date limite de candidature (à 23h59) 30 avril 2024

Keywords

Alzheimer's disease, amyloid beta peptides, synaptotoxicity, therapeutic strategies, electrophysiology, cellular imagery

Project description

Alzheimer's disease (AD) is characterized by the cerebral accumulation of aggregated amyloid ß peptides (Aβ, amyloid plaques) and abnormal tau protein aggregates (1,2). Native Aß peptides are constantly produced in the human brain where they usually remain in a soluble state with a α-helical conformation. However Aß peptides can adopt alternative misfolded aggregation-prone conformations rich in ß-sheets that favour their association into oligomers, fibrillar structures and amyloid. Distinct structural variants, also called strains of Aβ, bearing point mutation in the Ab peptide sequence have been identified. They may explain the variability of the pathology within and among the AD patients. For example, recent studies demonstrated that patients with rapidly progressive forms of AD exhibit distinct molecular structures of Aβ when compared with those with normally progressive AD (3). The impact of these structural variants on ADpathology evolution is however still partly unknown.
The aim of this proposal is to characterize the molecular mechanisms by which selected variants of Aβ promote and amplify cerebral amyloid and tau pathology. For this purpose, we will generate specific Aβ variants and use cellular and in vivo mouse models to evaluate how Aβ variants impact AD pathology (Fig 1). We will test whether Aβ variants:
i. induce different cellular pathology (LTP, synaptotoxicity, and tau pathology).
ii. transfer their biophysical and pathologic characteristics to mouse models of AD after intracerebral inoculation and induce different
long-term amyloid accumulation and spreading patterns.
iii. conserve their biological specificities after self-replication in mouse models.

Thematique / Domain / Context

We selected amyloid strains that exhibit specific oligomers profiles (Fig 2): i. Human (H) Aβ1-42 produces an amyloid strain rich in oligomers or fibrils depending of the re-suspension protocol; ii. H Aβ1-42 with the icelandic mutation A2T generates amyloid strain less prone to aggregation that forms dimers, tetramers but no dodecamers ((4); unpublished results); iii. H A􀀀 1-42 with the Osaka mutation called E22D produces a peptide highly resistant to degradation with an enhanced oligomerization and fibrillization profile (5); iv. H Aβ 1-42 with the mutation F19S/L34P that remains as a monomer in solution (6).
We evaluated the impact of the selected Aβ variants on synaptic density by transfecting specific APP in primary neuronal cultures. We showed that APPwt, APPswe and APPOsaka expression induced a drastic reduction of the synaptic density but no effect was observed when neurons were transfected with APPIcelandic (Fig. 3).
Further, we analysed the influence of oligomers of Aβ1-42 Osaka and Iceland (at 100 nM) on synaptic plasticity (LTP). While the peptide bearing the Osaka deletion abolishes LTP, Aβ1-42 with the Icelandic mutation does not alter the LTP (Fig 4) validating the differential effect of the selected strains on AD pathologies.
Finally, in two preliminary studies, APP/PS1 mouse models were inoculated in the CA1 region of the hippocampus with control, AD brains (Fig 5a,b), Aβ1-42 oligomers or PBS (Fig 5c). Amyloid load was evaluated at 1 to 4 months post-inoculation. We observed that an amplification of amyloid pathology in the mice that received the AD brains (Fig. 5a, b) or the Aβ1-42 oligomers (Fig 5c).
These results validate the impact of oligomers on spreading and evolution of AD pathology.

Our project relies on a procedure that we implemented to produce recombinant ABeta peptides in order to generate Ab peptides with specific disease-associated amino acid mutations. This procedure is based on the expression of plasmids containing Aβ1–42 cDNA in Escherichia coli and allowed us to produce 4 strains of Aβ peptides (H Aβ1-42, H Aβ1-42 with the Osaka (AβE22D), Icelandic (AβA2T) and F19S/L34P mutations)) bearing subtle modifications in their sequences. Our preliminary results and other publications leave no doubt on the ability of the selected A beta peptides to generate different oligomerization profiles (5).

This project involves a strong collaboration with the research group led by Dr Dhenain in Paris. Experiments will be performed in the GIN and complementary approaches will be performed in Paris in Dr Dhenain Group.

Objectives

We will 1) characterize the structural profiles of the selected Aβ variant 2) study their impacts on AD pathologies with a specific interest on APP processing; synaptic morphology and plasticity and tau pathology in primary neuronal cultures and acute brain slices exposed to Aβ variants. All the methods to perform these studies are mastered by Partner 1 (6). Next, we will evaluate whether intrahippocampal inoculation in APPswe/PS1dE9 mice of the selected Aβ strain promotes differences in Aβ misfolding, spreading and synaptotoxicity in this mice model of AD. We will also study amyloid load and spreading and behavioural outcomes in mice 4 months after injection of the selected strains. Partner 2 (Dhenain’s team) already has a large experience in Aβ inoculation in mice and in the evaluation of their impacts (7,8).

Method

This project involves technical approaches ranging from molecular biology, and cellular biology. We will be using neuronal cultures approaches to develop humain neuronal cultures from iPSCs. By transfecting these neurons, we will manipulate the expression of the major identified molecular actors of Alzheimer's disease physiopathology.

Expected results

There is currently no effective treatment for AD making the societal demand for a cure even stronger . The induction of AD lesions by  inoculation of Aβ strains is revolutionizing our vision of AD physiopathology (11,12). The fact that humans who have been accidentally contaminated with amyloid peptides developed a cerebral amyloidosis provided the first demonstration that AD can be caused by peripheral contaminations (13). These discoveries bring new concepts on mechanisms associated with AD such as seeding, spreading and Aβ strain effects. The strain effect is still poorly understood and the influence of selected amyloid strains on pathology evolution including on the induction of amyloid load and tau pathologies will bring important insights on the pathologic cascades involved in the spreading of AD. Further understanding of the concept of amyloid strains will greatly influence future treatments against AD and promotes the development of personalized medicine for AD patients.

Bibliography

[1] Sperling RA, et al. Toward defining the preclinical stages of Alzheimer’s disease: Recommendations from the National Institute on Aging and the Alzheimer's Association workgroup. Alzheimer's and Dementia. 2011;7:280-92.
[2] Frandemiche ML, et al. Activity-dependent tau protein translocation to excitatory synapse is disrupted by exposure to amyloid-beta oligomers. J Neurosci. 2014 Apr 23;34(17):6084-97.
(3) Cohen ML, et al. Rapidlly progressive Alzheimer's disease features distinct structures of amyloid-b. Brain 138:1009-1022.
[4] Zheng X, et al. Amyloid beta-Protein Assembly: Differential Effects of the Protective A2T Mutation and Recessive A2V Familial Alzheimer's Disease Mutation. ACS chemical neuroscience. 2015 Oct 21;6(10):1732-40.
[5] Tomiyama T, et al. A new amyloid beta variant favoring oligomerization in Alzheimer's-type dementia. Ann Neurol. 2008 Mar;63(3):377-87.
[6] Ahmed M, et al. Structural conversion of neurotoxic amyloid-beta(1-42) oligomers to fibrils. Nature structural & molecular biology. 2010 May;17(5):561-7.
[7] Amar F, et al. The amyloid-beta oligomer Abeta*56 induces specific alterations in neuronal signaling that lead to tau phosphorylation and aggregation. Science signaling. 2017 May 9;10(478).
(8] Dudeffant C, et al. Contrast-enhanced MR microscopy of amyloid plaques in five mouse models of amyloidosis and in human Alzheimer's disease brains. Sci Rep. 2017 Jul 10;7(1):4955.
[9] Dubois A, et al. Detection by voxel-wise statistical analysis of significant changes in regional cerebral glucose uptake in an APP/PS1 transgenic mouse model of Alzheimer's disease. Neuroimage. 2010 Jun;51(2):586-98.
[10] Vandenberghe ME, et al. High-throughput 3D whole-brain quantitative histopathology in rodents. Scientific Reports. 2016;6:20958.
[11] Lebenberg J, et al. Validation of MRI-based 3D digital atlas registration with histological and autoradiographic volumes: An anatomofunctional transgenic mouse brain imaging study. Neuroimage. 2010 Mar 10;51(3):1037-46.
[12] Gary C, et al. Experimental transmissibility of Alzheimer pathology in a non-human primate. Neurodegenerative Diseases. 2015;15(suppl 1):74.
[13] Jaunmuktane Z, et al. Evidence for human transmission of amyloid-beta pathology and cerebral amyloid angiopathy. Nature. 2015 Sep 10;525(7568):247-50.

Profile and skills required

The candidate will have a background in neuroscience, with a particular interest in neurovegetative processes. He/she will be trained in molecular and cellular biology approaches to neural pathophysiology in Alzheimer's disease. Training in electrophysiology will also be possible as part of the thesis.