Variability in the type and layer distribution of cortical Aβ pathology in familial Alzheimer's disease.

Familial Alzheimer's disease (FAD) is caused by autosomal dominant mutations in the PSEN1, PSEN2 or APP genes, giving rise to considerable clinical and pathological heterogeneity in FAD. Here we investigate variability in clinical data and the type and distribution of Aβ pathologies throughout the cortical layers of different FAD mutation cases. Brain tissue from 20 FAD cases [PSEN1 pre-codon 200 (n = 10), PSEN1 post-codon 200 (n = 6), APP (n = 4)] were investigated. Frontal cortex sections were stained immunohistochemically for Aβ, and Nissl to define the cortical layers. The frequency of different amyloid-beta plaque types was graded for each cortical layer and the severity of cerebral amyloid angiopathy (CAA) was determined in cortical and leptomeningeal blood vessels. Comparisons were made between FAD mutations and APOE4 status, with associations between pathology, clinical and genetic data investigated. In this cohort, possession of an APOE4 allele was associated with increased disease duration but not with age at onset, after adjusting for mutation sub-group and sex. We found Aβ pathology to be heterogeneous between cases although Aβ load was highest in cortical layer 3 for all mutation groups and a higher Aβ load was associated with APOE4. The PSEN1 post-codon 200 group had a higher Aβ load in lower cortical layers, with a small number of this group having increased cotton wool plaque pathology in lower layers. Cotton wool plaque frequency was positively associated with the severity of CAA in the whole cohort and in the PSEN1 post-codon 200 group. Carriers of the same PSEN1 mutation can have differing patterns of Aβ deposition, potentially because of differences in risk factors. Our results highlight possible influences of APOE4 genotype, and PSEN1 mutation type on Aβ deposition, which may have effects on the clinical heterogeneity of FAD

The PSEN1 E280G mutation leads to increased amyloid-β43 production in induced pluripotent stem cell neurons and deposition in brain tissue

Mutations in the presenilin 1 gene, PSEN1, which cause familial Alzheimer’s disease alter processing of amyloid precursor protein, leading to the generation of various amyloid-β peptide species. These species differ in their potential for aggregation. Mutation-specific amyloid-β peptide profiles may thereby influence pathogenicity and clinical heterogeneity. There is particular interest in comparing mutations with typical and atypical clinical presentations, such as E280G. We generated PSEN1 E280G mutation induced pluripotent stem cells from two patients and differentiated them into cortical neurons, along with previously reported PSEN1 M146I, PSEN1 R278I and two control lines. We assessed both the amyloid-β peptide profiles and presenilin 1 protein maturity. We also compared amyloid-β peptide profiles in human post-mortem brain tissue from cases with matched mutations. Amyloid-β ratios significantly differed compared with controls and between different patients, implicating mutation-specific alterations in amyloid-β ratios. Amyloid-β42:40 was increased in the M146I and both E280G lines compared with controls. Amyloid-β42:40 was not increased in the R278I line compared with controls. The amyloid-β43:40 ratio was increased in R278I and both E280G lines compared with controls, but not in M146I cells. Distinct amyloid-β peptide patterns were also observed in human brain tissue from individuals with these mutations, showing some similar patterns to cell line observations. Reduced presenilin 1 maturation was observed in neurons with the PSEN1 R278I and E280G mutations, but not the M146I mutation. These results suggest that mutation location can differentially alter the presenilin 1 protein and affect its autoendoproteolysis and processivity, contributing to the pathological phenotype. Investigating differences in underlying molecular mechanisms of familial Alzheimer’s disease may inform our understanding of clinical heterogeneity

Association of MAPT haplotype-tagging polymorphisms with cerebrospinal fluid biomarkers of Alzheimer's disease: a preliminary study in a Croatian cohort

INTRODUCTION: Alzheimer's disease (AD) is the world leading cause of dementia. Early detection of AD is essential for faster and more efficacious usage of therapeutics and preventive measures. Even though it is well known that one ε4 allele of apolipoprotein E gene increases the risk for sporadic AD five times, and that two ε4 alleles increase the risk 20 times, reliable genetic markers for AD are not yet available. Previous studies have shown that microtubule-associated protein tau (MAPT) gene polymorphisms could be associated with increased risk for AD. ----- METHODS: The present study included 113 AD patients and 53 patients with mild cognitive impairment (MCI), as well as nine healthy controls (HC) and 53 patients with other primary causes of dementia. The study assessed whether six MAPT haplotype-tagging polymorphisms (rs1467967, rs242557, rs3785883, rs2471738, del-In9, and rs7521) and MAPT haplotypes are associated with AD pathology, as measured by cerebrospinal fluid (CSF) AD biomarkers amyloid β1-42 (Aβ1-42 ), total tau (t-tau), tau phosphorylated at epitopes 181 (p-tau181 ), 199 (p-tau199 ), and 231 (p-tau231 ), and visinin-like protein 1 (VILIP-1). ----- RESULTS: Significant increases in t-tau and p-tau CSF levels were found in patients with AG and AA MAPT rs1467967 genotype, CC MAPT rs2471738 genotype and in patients with H1H2 or H2H2 MAPT haplotype. ----- CONCLUSIONS: These results indicate that MAPT haplotype-tagging polymorphisms and MAPT haplotypes should be further tested as potential genetic biomarkers of AD

A single nuclear transcriptomic characterisation of mechanisms responsible for impaired angiogenesis and blood-brain barrier function in Alzheimer's disease

Brain perfusion and blood-brain barrier (BBB) integrity are reduced early in Alzheimer's disease (AD). We performed single nucleus RNA sequencing of vascular cells isolated from AD and non-diseased control brains to characterise pathological transcriptional signatures responsible for this. We show that endothelial cells (EC) are enriched for expression of genes associated with susceptibility to AD. Increased β-amyloid is associated with BBB impairment and a dysfunctional angiogenic response related to a failure of increased pro-angiogenic HIF1A to increased VEGFA signalling to EC. This is associated with vascular inflammatory activation, EC senescence and apoptosis. Our genomic dissection of vascular cell risk gene enrichment provides evidence for a role of EC pathology in AD and suggests that reducing vascular inflammatory activation and restoring effective angiogenesis could reduce vascular dysfunction contributing to the genesis or progression of early AD.</p

Plasma amyloid-β ratios in autosomal dominant Alzheimer's disease: the influence of genotype.

In vitro studies of autosomal dominant Alzheimer's disease implicate longer amyloid-β peptides in disease pathogenesis; however, less is known about the behaviour of these mutations in vivo. In this cross-sectional cohort study, we used liquid chromatography-tandem mass spectrometry to analyse 66 plasma samples from individuals who were at risk of inheriting a mutation or were symptomatic. We tested for differences in amyloid-β (Aβ)42:38, Aβ42:40 and Aβ38:40 ratios between presenilin 1 (PSEN1) and amyloid precursor protein (APP) carriers. We examined the relationship between plasma and in vitro models of amyloid-β processing and tested for associations with parental age at onset. Thirty-nine participants were mutation carriers (28 PSEN1 and 11 APP). Age- and sex-adjusted models showed marked differences in plasma amyloid-β between genotypes: higher Aβ42:38 in PSEN1 versus APP (P < 0.001) and non-carriers (P < 0.001); higher Aβ38:40 in APP versus PSEN1 (P < 0.001) and non-carriers (P < 0.001); while Aβ42:40 was higher in both mutation groups compared to non-carriers (both P < 0.001). Amyloid-β profiles were reasonably consistent in plasma and cell lines. Within the PSEN1 group, models demonstrated associations between Aβ42:38, Aβ42:40 and Aβ38:40 ratios and parental age at onset. In vivo differences in amyloid-β processing between PSEN1 and APP carriers provide insights into disease pathophysiology, which can inform therapy development

A single nuclear transcriptomic characterisation of mechanisms responsible for impaired angiogenesis and blood-brain barrier function in Alzheimer’s disease

Brain perfusion and blood-brain barrier (BBB) integrity are reduced early in Alzheimer’s disease (AD). We performed single nucleus RNA sequencing of vascular cells isolated from AD and non-diseased control brains to characterise pathological transcriptional signatures responsible for this. We show that endothelial cells (EC) are enriched for expression of genes associated with susceptibility to AD. Increased β-amyloid is associated with BBB impairment and a dysfunctional angiogenic response related to a failure of increased pro-angiogenic HIF1A to increased VEGFA signalling to EC. This is associated with vascular inflammatory activation, EC senescence and apoptosis. Our genomic dissection of vascular cell risk gene enrichment provides evidence for a role of EC pathology in AD and suggests that reducing vascular inflammatory activation and restoring effective angiogenesis could reduce vascular dysfunction contributing to the genesis or progression of early AD

Deciphering pathological heterogeneity in familial Alzheimer’s disease

Introduction: Alzheimer’s disease (AD) is the most common neurodegenerative dementia and its cause is unknown. In rare cases AD can be caused by mutations in the PSEN1, PSEN2 or APP gene and this form of AD is termed familial Alzheimer’s disease (FAD). While the genetic cause is determined, there is considerable heterogeneity in terms of clinical presentation and pathological appearance at post-mortem. Previously it has been suggested pathological features of FAD may influence clinical features. It has also been suggested that FAD mutations may influence both pathological and clinical features. Common features of AD may also play a role in FAD, such as microglial activation and the influence of genetic modifiers of disease, such as APOE. The aims of this thesis were to investigate the associations of Aβ pathology (including CAA) and microglial load to age at onset and disease duration. Investigate histological profiles of Aβ pathologies (including CAA) and microglial load and the associations between these pathologies in genetic causes of FAD and APOE genotypes. Observe the contribution of specific Aβ peptide species to the histological profiles of Aβ pathology, and the association of these peptide with FAD and APOE genotypes. Generate and differentiate FAD patient derived iPSC to neuronal cultures to assess the association of FAD mutation with Aβ peptide profiles and PSEN1 maturity in a neuronal model of FAD. We hypothesise that histological features and Aβ peptide profiles will segregate with FAD mutation location, while microglial phenotype will associate with specific Aβ pathologies. Additionally, we predict the Aβ profiles observed in FAD cell lines will reflect histological profiles of Aβ aggregation. Materials and Methods: Nissl staining was performed on the frontal cortex of 20 FAD cases from the Queen Square Brain Bank (QSBB) (PSEN1 mutation carriers n=16, 10 pre-codon 200, 6 post-codon 200 and APP mutation carriers n=4). Cortical layers were delineated and serial sections immunohistochemically stained with antibodies against Aβ, Iba1, CD68 and CR3/43 and a subset were also stained for Tau. Aβ plaque type, load (% area stained), proportion of Aβ positive cerebral amyloid angiopathy (CAA), and microglial load were analysed per cortical layer. Additionally, in frontal and occipital cortex tissue from the 20 QSBB cases and an additional 21 FAD cases from the Institute of Psychiatry, Psychology and Neuroscience (combined total n=41, PSEN1 mutation carriers n=31, 20 pre-codon 200, 11 post-codon 200 and APP mutation carriers n=10) the proportion and severity of cortical and leptomeningeal CAA were investigated via vessel counts. In the temporal and occipital cortex of the 20 QSBB cases, IHC with Aβ isoform specific antibodies was conducted to investigate genetic contribution to isoform specific pathology. Finally, 5 induced pluripotent stem cell (iPSC) lines from FAD patients were generated. Four FAD lines and two control lines were differentiated into cortical neurons to investigate Aβ isoform production via ELISA and PSEN1 protein levels and maturity were assessed. Results: Clinical features in this FAD cohort were influenced by both FAD mutation and APOE status. Pathological Aβ deposits showed variability across cortical layers, and specific features were more associated with distinct mutations. Microglial phenotype did not differ by FAD mutation group or APOE status however associations with Aβ pathologies were observed. CAA differed between mutations groups, while APOE4 genotype had a non-significant effect of CAA pathology in FAD. Analysis of Aβ production from FAD iPSC derived cortical neurons showed that Aβ production differed not only compared to control cell lines but compared to other independent PSEN1 mutations. This could be the result of changes to PSEN1 maturation. Conclusions: It was shown that there is pathological heterogeneity in FAD of which some aspects associate with specific FAD mutation subgroup. For instance, greater Aβ and CWP frequency in the lower layers in the PSEN1 post-codon 200 group as well as greater proportion and severity of CAA. Correlations between plaques, CAA and microglia indicate contribution of clearance mechanisms to histological features observed in FAD, which can differ by mutation group. Specifically, CD68 was generally associated with greater CAA and reduced Aβ pathology. In the cellular models, specific FAD mutations, particularly those post-codon 200, affect Aβ peptide ratios, which associates with observed pathological heterogeneity and suggests that differences in the aggregation and clearance of these peptides modifies the histological appearance of Aβ pathology. Combined with the observed effect of APOE genotype on disease duration, peptide profiles and CAA severity, this may contribute to the differences in clinical aspects of FAD

Characterisation of premature cell senescence in Alzheimer’s disease using single nuclear transcriptomics

Aging is associated with cell senescence and is the major risk factor for AD. We characterized premature cell senescence in postmortem brains from non-diseased controls (NDC) and donors with Alzheimer’s disease (AD) using imaging mass cytometry (IMC) and single nuclear RNA (snRNA) sequencing (&gt; 200,000 nuclei). We found increases in numbers of glia immunostaining for galactosidase beta (&gt; fourfold) and p16 INK4A (up to twofold) with AD relative to NDC. Increased glial expression of genes related to senescence was associated with greater β-amyloid load. Prematurely senescent microglia downregulated phagocytic pathways suggesting reduced capacity for β-amyloid clearance. Gene set enrichment and pseudo-time trajectories described extensive DNA double-strand breaks (DSBs), mitochondrial dysfunction and ER stress associated with increased β-amyloid leading to premature senescence in microglia. We replicated these observations with independent AD snRNA-seq datasets. Our results describe a burden of senescent glia with AD that is sufficiently high to contribute to disease progression. These findings support the hypothesis that microglia are a primary target for senolytic treatments in AD