Aims

There are three main clinical presentations in Alzheimer disease (AD) with diversity in phenotype, onset and progression of clinical symptoms: autosomal dominant (ADAD), early onset (EOAD) and late onset (LOAD); yet, they share the common AD pathology: deposition of Aβ and ptau protein aggregates in the brain. This work aims to identify and characterize common disrupted pathways across AD etiologies.

Methods

We examined bulk transcriptomic data from brain donors to the DIAN (ADAD, N=19) and Knight-ADRC (EOAD, N=13; LOAD, N=55; controls (CO), N=16). Through differential gene expression (DGE) analyses, we identified common signals across AD etiologies. We used three independent datasets to replicate our findings and followed our top gene using functional assays that include iPSC and N2A cells.

Results

DGE analysis in our discovery dataset identified AGFG2, a gene under the AD GWAs signal for NYAP1, to have higher expression ((p=7×10^-4) in ADAD, EOAD or LOAD compared to CO. This effect was replicated in Mount Sinai (p=8.63×10^-3), Mayo (p=5.88×10^-12) and ROSMAP (p=3.96×10^-05). We have overexpressed human AGFG2 on N2a APP695wt and observed, via ELISA, an increase in extracellular Aβ40 and Aβ42. Using brain single cell nuclei data, we identified that astrocytes are the main cell type expressing AGFG2. We have cultured human iPSC-derived astrocytes and knockdown this gene using siRNA.

Conclusions

AGFG2 is a gene primarily expressed in astrocytes and we have observed that higher AGFG2 leads to higher extracellular Aβ40 and Aβ42. Upcoming experiments will use iPSC-derived astrocytes to further identify the relationship between AGFG2 and Aβ.

Aims

The relationship between aging and Alzheimer’s disease (AD) phenotypes is imperfect, which challenges the long-standing belief that dementia is an inevitable consequence of aging and underscores the need for investigations of resilience factors. We examined whether the functionally advantageous KL-VS variant of the aging suppressor KLOTHO gene confers resilience against the deleterious effects of aging on cerebrospinal fluid (CSF) markers of neuroinflammation (sTREM2, YKL-40, GFAP, IL-6) and synaptic dysfunction (α-synuclein, S100, neurogranin) in a middle-aged and older adult cohort enriched for AD risk.

Methods

The sample included non-demented adults (N=454; M_age=62±8) from the Wisconsin Registry for Alzheimer's Prevention and the Wisconsin Alzheimer's Disease Research Center who were genotyped for KLOTHO and underwent CSF sampling. Covariate-adjusted multivariate regression examined relationships between age (mean split: Younger (age<62) vs. Older (age≥62)) and CSF biomarkers (Roche NeuroToolKit), and whether those relationships differed between KL-VS non-carriers (N=332) and heterozygotes (N=122).

Results

Older age was associated with poorer CSF biomarker profiles among KL-VS non-carriers (P’s ≤ 0.05). These effects were attenuated in heterozygotes for α-synuclein, neurogranin, S100, and IL-6 (P’s ≥ 0.13). Although significant differences in GFAP, sTREM2, and YKL40 between Younger and Older groups were evident in both non-carriers (P’s ≤ 0.001) and heterozygotes (P’s ≤ 0.01), the effects within the non-carriers were markedly stronger (2- to 5-fold) compared to heterozygotes.

Conclusions

Higher levels of neuroinflammatory and synaptic dysfunction markers in older age were attenuated in carriers of a functionally advantageous KLOTHO variant. KL-VS heterozygosity appears to protect against age-related biomolecular alterations that confer risk for AD.

Aims

Non-canonical functions of autophagy proteins have been implicated in neurodegenerative conditions, including Alzheimer’s Disease (AD). We previously identified a novel role for the autophagy machinery in regulating the conjugation of LC3 to endosomes, a process we termed LC3-associated endocytosis (LANDO). Here we investigate the mechanism responsible for exacerbated neuroinflammation downstream of LANDO-deficiency.

Methods

We identified a robust increase in the expression of necroptotic regulators including MLKL in AD patient brain samples and further found increases in both expression and activation of MLKL and RIPK1/3 in the brains of LANDO-deficient mice on the 5xFAD background, predominantly in microglia. To investigate the contribution of necroptosis and the role of Rubicon and the LANDO pathway in mitigating neuroinflammation, we have used multiple approaches in mice and human microglia.

Results

Abrogation of necroptosis reversed neuroinflammation, tau hyperphosphorylation, neurodegeneration, and memory impairment in the LANDO-deficient AD mouse model. Necroptotic activation in the 5xFAD/LANDO-deficient mouse was restricted to microglia, with no observable activation of necroptosis in neurons or astrocytes. Moreover, we identified that the LANDO regulator Rubicon interacts with/restricts the plasma membrane localization of MLKL, protecting microglia from necroptosis and limiting inflammatory cytokine release.

Conclusions

Taken together our results demonstrate a novel mechanism for the regulation of necroptosis and highlight a putative pathological setting of necroptotic activation in microglia. Furthermore, it provides a direct pathway of regulating inflammation downstream of LANDO, which to date has remained elusive. Our findings in mice are correlative to human expression data and brain slide analysis as will be presented.

Aims

The presence of reactive astrocytes in Alzheimer’s disease (AD) brain was demonstrated to correlate with neuronal loss and cognitive deficits. Evidence indeed supports the role of reactive astrocytes as mediators of changes in neurons, including synapses.

Oxysterols (cholesterol oxidation products) are crucial for brain cholesterol homeostasis and we previously demonstrated that changes in the brain levels of various oxysterols correlate with AD progression.

In order to deepen the role of oxysterols in AD, we investigated whether they could contribute to astrocyte reactivity, and consequently impact on neuronal health.

Methods

Two oxysterol mixtures were used, whose compositions represent respectively mild and severe AD brain samples. Mouse primary astrocytes and neurons were used as in vitro models. Lcn2 gene silencing was performed in astrocytes.

Results

Oxysterols induce a clear morphological change in astrocytes and the upregulation of some reactive astrocyte markers, including Lcn2. Astrocyte conditioned media analysis revealed an increase in the release of Lcn2, cytokines, and chemokines in response to oxysterols. A reduction of PSD95 and an increase in cleaved caspase-3 protein levels have been demonstrated in neurons co-cultured with oxysterol-treated astrocytes, pointing out the impact of mediators released by astrocytes; among them, Lcn2 has been demonstrated to play a major role on synapses, affecting neurite morphology and decreasing dendritic spine density.

Conclusions

Results demonstrated that oxysterols present in the AD brain promote astrocyte reactivity, determining the release of several mediators that affect neuronal health and synapses. Lcn2 has been shown to exert a key role in mediating the synaptotoxic effect of oxysterol-treated astrocytes.

Aims

There is a large variation in the clinical presentations and progression between Parkinson’s disease patients (PD). The generation of deeply and longitudinally phenotyped patient cohorts offers an enormous potential to identify disease subtypes with prognostic and therapeutic utility.

Methods

Using three large cohorts of PD patients with extensive clinical observational data repeatedly collected over many years, we developed a Bayesian multiple phenotypes mixed model incorporating the genetic relationships between individuals, able to reduce a large number of diverse clinical measurements into a smaller number of continuous underlying factors (“phenotypic axes”).

Results

We identify three principal axes of PD patient phenotypic variation, which are reproducibly found across three independent, deeply and diversely phenotyped cohorts. The most influential axis was specifically associated with the genetic risk of Alzheimer's disease (AD) and CSF Aβ1-42. As for AD onset risk but different to PD onset risk, the genetics influencing this phenotypic axis were in genomic regions enriched for microglia-expressed genes. These results suggest that neuroinflammation plays a key role in the development of a more aggressive form of PD, but that neuroinflammation does not affect the risk of PD itself. Finally, we show that by focussing our method specifically on AD genetic risk we can more accurately predict the clinical progression of PD.

Conclusions

PD patients with a high genetic risk for AD are more likely to develop a more severe and rapid form of PD including, but not limited to, dementia.

Aims

WM abnormalities constitute an early hallmark in AD pathogenesis, presenting many years prior to cortical neuronal loss and dementia. Yet, the underlying pathways and mechanisms linking subcortical and cortical pathogenic changes remain widely unexplored. We aimed to evaluate glial expression profiles in WM of individuals with differing cognitive statuses and address possible links with cortical markers of early cellular damage in AD.

Methods

We performed immunohistochemical analyses using artificial-intelligence based cell-quantifications of WM of demented and non-demented individuals with identical tau burden (Braak III/IV), and healthy controls. We evaluated brain regions temporally matched (temporal pole) and posterior (occipital cortex) to the tau deposition stage of these study subjects at postmortem, assessing diverging glial expression patterns.

Results

We found a differing WM expression profile with increased activated and reduced homeostatic glia, which was present in demented individuals in brain regions with and without overt tau deposition. In addition, we found an association between dysfunctional glia burden in subcortical-WM and presence of early cellular vulnerability markers in the cortex.

Conclusions

Our results suggest that WM dysfunction occurs early in AD and prior to overt cortical tau deposition. Moreover, dysfunctional glial profiles with an increase in activated and loss of homeostatic astroglia markedly differ between demented and non-demented individuals. Lastly, the burden of subcortical glial patterns shows association with cortical cellular vulnerability markers. Overall, these results suggest that WM could be an early site of vulnerability in AD and that its dysfunction could be key in subsequent cortical cellular loss, tau deposition and dementia in AD.

Abstract Body

Parkinson's disease (PD) , a neurodegenerative disease, is associated with impairment with motor activity and rigidity. The genetic risks of the disease is reported to be between 5 and 10%. It was suggested that activated microglia may play a role in neurotoxicity to dopaminergic neurons (Lazdon et al JNC 2019). We found that reduction in DJ-1 protein ( found in PD patients) in microglia increase both their pro-inflammatory profile and neurotoxicity. Autophagy is an important mechanism for the degradation of intracellular proteins and organelles. We discovered that impaired DJ-1 microglia exhibit an impaired autophagy-dependent degradation of p62 and LC3 proteins, an important protein in the autophagy flux. We discovered that impaired autophagy affects the ability of DJ-1 microglia to uptake αSyn. Further research suggests that αSyn affects microglia metabolism both in vitro and in vivo, leading to their neurotoxic properties. Targeting those metabolic changes in microglia may lead to new therapeutic avenues in the disease.