The association between neighborhood deprivation and DNA methylation in an autopsy cohort.

Previous research has found that living in a disadvantaged neighborhood is associated with poor health outcomes. Living in disadvantaged neighborhoods may alter inflammation and immune response in the body, which could be reflected in epigenetic mechanisms such as DNA methylation (DNAm). We used robust linear regression models to conduct an epigenome-wide association study examining the association between neighborhood deprivation (Area Deprivation Index; ADI), and DNAm in brain tissue from 159 donors enrolled in the Emory Goizueta Alzheimer's Disease Research Center (Georgia, USA). We found one CpG site (cg26514961, gene PLXNC1) significantly associated with ADI after controlling for covariates and multiple testing (p-value=5.0e-8). Effect modification by APOE ε4 was statistically significant for the top ten CpG sites from the EWAS of ADI, indicating that the observed associations between ADI and DNAm were mainly driven by donors who carried at least one APOE ε4 allele. Four of the top ten CpG sites showed a significant concordance between brain tissue and tissues that are easily accessible in living individuals (blood, buccal cells, saliva), including DNAm in cg26514961 (PLXNC1). Our study identified one CpG site (cg26514961, PLXNC1 gene) that was significantly associated with neighborhood deprivation in brain tissue. PLXNC1 is related to immune response, which may be one biological pathway how neighborhood conditions affect health. The concordance between brain and other tissues for our top CpG sites could make them potential candidates for biomarkers in living individuals.

MAPT H2 haplotype and risk of Pick's disease in the Pick's disease International Consortium: a genetic association study.

BACKGROUND: Pick's disease is a rare and predominantly sporadic form of frontotemporal dementia that is classified as a primary tauopathy. Pick's disease is pathologically defined by the presence in the frontal and temporal lobes of Pick bodies, composed of hyperphosphorylated, three-repeat tau protein, encoded by the MAPT gene. MAPT has two distinct haplotypes, H1 and H2; the MAPT H1 haplotype is the major genetic risk factor for four-repeat tauopathies (eg, progressive supranuclear palsy and corticobasal degeneration), and the MAPT H2 haplotype is protective for these disorders. The primary aim of this study was to evaluate the association of MAPT H2 with Pick's disease risk, age at onset, and disease duration. METHODS: In this genetic association study, we used data from the Pick's disease International Consortium, which we established to enable collection of data from individuals with pathologically confirmed Pick's disease worldwide. For this analysis, we collected brain samples from individuals with pathologically confirmed Pick's disease from 35 sites (brainbanks and hospitals) in North America, Europe, and Australia between Jan 1, 2020, and Jan 31, 2023. Neurologically healthy controls were recruited from the Mayo Clinic (FL, USA, or MN, USA between March 1, 1998, and Sept 1, 2019). For the primary analysis, individuals were directly genotyped for the MAPT H1-H2 haplotype-defining variant rs8070723. In a secondary analysis, we genotyped and constructed the six-variant-defined (rs1467967-rs242557-rs3785883-rs2471738-rs8070723-rs7521) MAPT H1 subhaplotypes. Associations of MAPT variants and MAPT haplotypes with Pick's disease risk, age at onset, and disease duration were examined using logistic and linear regression models; odds ratios (ORs) and ß coefficients were estimated and correspond to each additional minor allele or each additional copy of the given haplotype. FINDINGS: We obtained brain samples from 338 people with pathologically confirmed Pick's disease (205 [61%] male and 133 [39%] female; 338 [100%] White) and 1312 neurologically healthy controls (611 [47%] male and 701 [53%] female; 1312 [100%] White). The MAPT H2 haplotype was associated with increased risk of Pick's disease compared with the H1 haplotype (OR 1·35 [95% CI 1·12 to 1·64], p=0·0021). MAPT H2 was not associated with age at onset (ß -0·54 [95% CI -1·94 to 0·87], p=0·45) or disease duration (ß 0·05 [-0·06 to 0·16], p=0·35). Although not significant after correcting for multiple testing, associations were observed at p less than 0·05: with risk of Pick's disease for the H1f subhaplotype (OR 0·11 [0·01 to 0·99], p=0·049); with age at onset for H1b (ß 2·66 [0·63 to 4·70], p=0·011), H1i (ß -3·66 [-6·83 to -0·48], p=0·025), and H1u (ß -5·25 [-10·42 to -0·07], p=0·048); and with disease duration for H1x (ß -0·57 [-1·07 to -0·07], p=0·026). INTERPRETATION: The Pick's disease International Consortium provides an opportunity to do large studies to enhance our understanding of the pathobiology of Pick's disease. This study shows that, in contrast to the decreased risk of four-repeat tauopathies, the MAPT H2 haplotype is associated with an increased risk of Pick's disease in people of European ancestry. This finding could inform development of isoform-related therapeutics for tauopathies. FUNDING: Wellcome Trust, Rotha Abraham Trust, Brain Research UK, the Dolby Fund, Dementia Research Institute (Medical Research Council), US National Institutes of Health, and the Mayo Clinic Foundation.

Bridging the Gap: Multi-Omics Profiling of Brain Tissue in Alzheimer's Disease and Older Controls in Multi-Ethnic Populations.

INTRODUCTION: Multi-omics studies in Alzheimer's disease (AD) revealed many potential disease pathways and therapeutic targets. Despite their promise of precision medicine, these studies lacked African Americans (AA) and Latin Americans (LA), who are disproportionately affected by AD. METHODS: To bridge this gap, Accelerating Medicines Partnership in AD (AMP-AD) expanded brain multi-omics profiling to multi-ethnic donors. RESULTS: We generated multi-omics data and curated and harmonized phenotypic data from AA (n=306), LA (n=326), or AA and LA (n=4) brain donors plus Non-Hispanic White (n=252) and other (n=20) ethnic groups, to establish a foundational dataset enriched for AA and LA participants. This study describes the data available to the research community, including transcriptome from three brain regions, whole genome sequence, and proteome measures. DISCUSSION: Inclusion of traditionally underrepresented groups in multi-omics studies is essential to discover the full spectrum of precision medicine targets that will be pertinent to all populations affected with AD.

Vascular Heparan Sulfate and Amyloid-ß in Alzheimer's Disease Patients.

Alzheimer's disease (AD) is a debilitating neurodegenerative disease characterized by the accumulation of extracellular amyloid-ß peptides (Aß) within the cerebral parenchyma and vasculature, which is known as cerebral amyloid angiopathy (CAA). This study utilized confocal imaging to investigate heparan sulfate (HS) expression within the cerebrovasculature and its associations with Aß, gender, and ApoE4 genotype in AD. Our investigation revealed elevated levels of HS in the cerebrovasculature of AD patients with severe CAA. Additionally, these patients exhibited higher HS colocalization with Aß in the cerebrovasculature, including both endothelial and vascular smooth muscle cell compartments. Intriguingly, a reversal in the polarized expression of HS within the cerebrovasculature was detected in AD patients with severe CAA. Furthermore, male patients exhibited lower levels of both parenchymal and cerebrovascular HS. Additionally, ApoE4 carriers displayed heightened cerebrovascular Aß expression and a tendency of elevated cerebrovascular HS levels in AD patients with severe CAA. Overall, these findings reveal potential intricate interplay between HS, Aß, ApoE, and vascular pathology in AD, thereby underscoring the potential roles of cerebrovascular HS in CAA development and AD pathology. Further study of the underlying mechanisms may present novel therapeutic avenues for AD treatment.

Association of Structural Forms of 17q21.31 with the Risk of Progressive Supranuclear Palsy and MAPT Sub-haplotypes.

Importance: The chromosome 17q21.31 region, containing a 900 Kb inversion that defines H1 and H2 haplotypes, represents the strongest genetic risk locus in progressive supranuclear palsy (PSP). In addition to H1 and H2, various structural forms of 17q21.31, characterized by the copy number of α, ß, and γ duplications, have been identified. However, the specific effect of each structural form on the risk of PSP has never been evaluated in a large cohort study. Objective: To assess the association of different structural forms of 17q.21.31, defined by the copy numbers of α, ß, and γ duplications, with the risk of PSP and MAPT sub-haplotypes. Design setting and participants: Utilizing whole genome sequencing data of 1,684 (1,386 autopsy confirmed) individuals with PSP and 2,392 control subjects, a case-control study was conducted to investigate the association of copy numbers of α, ß, and γ duplications and structural forms of 17q21.31 with the risk of PSP. All study subjects were selected from the Alzheimer's Disease Sequencing Project (ADSP) Umbrella NG00067.v7. Data were analyzed between March 2022 and November 2023. Main outcomes and measures: The main outcomes were the risk (odds ratios [ORs]) for PSP with 95% CIs. Risks for PSP were evaluated by logistic regression models. Results: The copy numbers of α and ß were associated with the risk of PSP only due to their correlation with H1 and H2, while the copy number of γ was independently associated with the increased risk of PSP. Each additional duplication of γ was associated with 1.10 (95% CI, 1.04-1.17; P = 0.0018) fold of increased risk of PSP when conditioning H1 and H2. For the H1 haplotype, addition γ duplications displayed a higher odds ratio for PSP: the odds ratio increases from 1.21 (95%CI 1.10-1.33, P = 5.47 × 10-5) for H1ß1γ1 to 1.29 (95%CI 1.16-1.43, P = 1.35 × 10-6) for H1ß1γ2, 1.45 (95%CI 1.27-1.65, P = 3.94 × 10-8) for H1ß1γ3, and 1.57 (95%CI 1.10-2.26, P = 1.35 × 10-2) for H1ß1γ4. Moreover, H1ß1γ3 is in linkage disequilibrium with H1c (R2 = 0.31), a widely recognized MAPT sub-haplotype associated with increased risk of PSP. The proportion of MAPT sub-haplotypes associated with increased risk of PSP (i.e., H1c, H1d, H1g, H1o, and H1h) increased from 34% in H1ß1γ1 to 77% in H1ß1γ4. Conclusions and relevance: This study revealed that the copy number of γ was associated with the risk of PSP independently from H1 and H2. The H1 haplotype with more γ duplications showed a higher odds ratio for PSP and were associated with MAPT sub-haplotypes with increased risk of PSP. These findings expand our understanding of how the complex structure at 17q21.31 affect the risk of PSP.

Differential DNA methylation in the brain as potential mediator of the association between traffic-related PM2.5 and neuropathology markers of Alzheimer's disease.

INTRODUCTION: Growing evidence indicates that fine particulate matter (PM2.5) is a risk factor for Alzheimer's disease (AD), but the underlying mechanisms have been insufficiently investigated. We hypothesized differential DNA methylation (DNAm) in brain tissue as a potential mediator of this association. METHODS: We assessed genome-wide DNAm (Illumina EPIC BeadChips) in prefrontal cortex tissue and three AD-related neuropathological markers (Braak stage, CERAD, ABC score) for 159 donors, and estimated donors' residential traffic-related PM2.5 exposure 1, 3, and 5 years prior to death. We used a combination of the Meet-in-the-Middle approach, high-dimensional mediation analysis, and causal mediation analysis to identify potential mediating CpGs. RESULTS: PM2.5 was significantly associated with differential DNAm at cg25433380 and cg10495669. Twenty-four CpG sites were identified as mediators of the association between PM2.5 exposure and neuropathology markers, several located in genes related to neuroinflammation. DISCUSSION: Our findings suggest differential DNAm related to neuroinflammation mediates the association between traffic-related PM2.5 and AD. HIGHLIGHTS: First study to evaluate the potential mediation effect of DNA methylation for the association between PM2.5 exposure and neuropathological changes of Alzheimer's disease. Study was based on brain tissues rarely investigated in previous air pollution research. Cg10495669, assigned to RBCK1 gene playing a role in inflammation, was associated consistently with 1-year, 3-year, and 5-year traffic-related PM2.5 exposures prior to death. Meet-in-the-middle approach and high-dimensional mediation analysis were used simultaneously to increase the potential of identifying the differentially methylated CpGs. Differential DNAm related to neuroinflammation was found to mediate the association between traffic-related PM2.5 and Alzheimer's disease.

Network Proteomics of the Lewy Body Dementia Brain Reveals Presynaptic Signatures Distinct from Alzheimer's Disease.

Lewy body dementia (LBD), a class of disorders comprising Parkinson's disease dementia (PDD) and dementia with Lewy bodies (DLB), features substantial clinical and pathological overlap with Alzheimer's disease (AD). The identification of biomarkers unique to LBD pathophysiology could meaningfully advance its diagnosis, monitoring, and treatment. Using quantitative mass spectrometry (MS), we measured over 9,000 proteins across 138 dorsolateral prefrontal cortex (DLPFC) tissues from a University of Pennsylvania autopsy collection comprising control, Parkinson's disease (PD), PDD, and DLB diagnoses. We then analyzed co-expression network protein alterations in those with LBD, validated these disease signatures in two independent LBD datasets, and compared these findings to those observed in network analyses of AD cases. The LBD network revealed numerous groups or "modules" of co-expressed proteins significantly altered in PDD and DLB, representing synaptic, metabolic, and inflammatory pathophysiology. A comparison of validated LBD signatures to those of AD identified distinct differences between the two diseases. Notably, synuclein-associated presynaptic modules were elevated in LBD but decreased in AD relative to controls. We also found that glial-associated matrisome signatures consistently elevated in AD were more variably altered in LBD, ultimately stratifying those LBD cases with low versus high burdens of concurrent beta-amyloid deposition. In conclusion, unbiased network proteomic analysis revealed diverse pathophysiological changes in the LBD frontal cortex distinct from alterations in AD. These results highlight the LBD brain network proteome as a promising source of biomarkers that could enhance clinical recognition and management.

Association of PM2.5 Exposure and Alzheimer Disease Pathology in Brain Bank Donors-Effect Modification by APOE Genotype.

BACKGROUND AND OBJECTIVES: Fine particulate matter (PM2.5) exposure has been found to be associated with Alzheimer disease (AD) and is hypothesized to cause inflammation and oxidative stress in the brain, contributing to neuropathology. The APOE gene, a major genetic risk factor of AD, has been hypothesized to modify the association between PM2.5 and AD. However, little prior research exists to support these hypotheses. This study investigates the association between traffic-related PM2.5 and AD hallmark pathology, including effect modification by APOE genotype, in an autopsy cohort. METHODS: A cross-sectional study was conducted using brain tissue donors enrolled in the Emory Goizueta AD Research Center who died before 2020 (n = 224). Donors were assessed for AD pathology including the Braak stage, Consortium to Establish a Registry for AD (CERAD) score, and combined AD neuropathologic change (ABC) score. Traffic-related PM2.5 concentrations were modeled for the metro-Atlanta area during 2002-2019 with a spatial resolution of 200-250 m. One-year, 3-year, and 5-year average PM2.5 concentrations before death were matched to participants' home address. We assessed the association between traffic-related PM2.5 and AD hallmark pathology and effect modification by APOE genotype, using adjusted ordinal logistic regression models. RESULTS: Among the 224 participants, the mean age of death was 76 years, and 57% had at least 1 APOE ε4 copy. Traffic-related PM2.5 was significantly associated with the CERAD score for the 1-year exposure window (odds ratio [OR] 1.92; 95% CI 1.12-3.30) and the 3-year exposure window (OR 1.87; 95% CI 1.01-3.17). PM2.5 was also associated with higher Braak stage and ABC score albeit nonsignificantly. The strongest associations between PM2.5 and neuropathology markers were among those without APOE ε4 alleles (e.g., for the CERAD score and 1-year exposure window, OR 2.31; 95% CI 1.36-3.94), though interaction between PM2.5 and APOE genotype was not statistically significant. DISCUSSION: Our study found traffic-related PM2.5 exposure was associated with the CERAD score in an autopsy cohort, contributing to epidemiologic evidence that PM2.5 affects ß-amyloid deposition in the brain. This association was particularly strong among donors without APOE ε4 alleles. Future studies should further investigate the biological mechanisms behind this association.

Whole-Genome Sequencing Analysis Reveals New Susceptibility Loci and Structural Variants Associated with Progressive Supranuclear Palsy.

Background: Progressive supranuclear palsy (PSP) is a rare neurodegenerative disease characterized by the accumulation of aggregated tau proteins in astrocytes, neurons, and oligodendrocytes. Previous genome-wide association studies for PSP were based on genotype array, therefore, were inadequate for the analysis of rare variants as well as larger mutations, such as small insertions/deletions (indels) and structural variants (SVs). Method: In this study, we performed whole genome sequencing (WGS) and conducted association analysis for single nucleotide variants (SNVs), indels, and SVs, in a cohort of 1,718 cases and 2,944 controls of European ancestry. Of the 1,718 PSP individuals, 1,441 were autopsy-confirmed and 277 were clinically diagnosed. Results: Our analysis of common SNVs and indels confirmed known genetic loci at MAPT, MOBP, STX6, SLCO1A2, DUSP10, and SP1, and further uncovered novel signals in APOE, FCHO1/MAP1S, KIF13A, TRIM24, TNXB, and ELOVL1. Notably, in contrast to Alzheimer's disease (AD), we observed the APOE ε2 allele to be the risk allele in PSP. Analysis of rare SNVs and indels identified significant association in ZNF592 and further gene network analysis identified a module of neuronal genes dysregulated in PSP. Moreover, seven common SVs associated with PSP were observed in the H1/H2 haplotype region (17q21.31) and other loci, including IGH, PCMT1, CYP2A13, and SMCP. In the H1/H2 haplotype region, there is a burden of rare deletions and duplications (P = 6.73×10-3) in PSP. Conclusions: Through WGS, we significantly enhanced our understanding of the genetic basis of PSP, providing new targets for exploring disease mechanisms and therapeutic interventions.

Toward a generalizable machine learning workflow for neurodegenerative disease staging with focus on neurofibrillary tangles.

Machine learning (ML) has increasingly been used to assist and expand current practices in neuropathology. However, generating large imaging datasets with quality labels is challenging in fields which demand high levels of expertise. Further complicating matters is the often seen disagreement between experts in neuropathology-related tasks, both at the case level and at a more granular level. Neurofibrillary tangles (NFTs) are a hallmark pathological feature of Alzheimer disease, and are associated with disease progression which warrants further investigation and granular quantification at a scale not currently accessible in routine human assessment. In this work, we first provide a baseline of annotator/rater agreement for the tasks of Braak NFT staging between experts and NFT detection using both experts and novices in neuropathology. We use a whole-slide-image (WSI) cohort of neuropathology cases from Emory University Hospital immunohistochemically stained for Tau. We develop a workflow for gathering annotations of the early stage formation of NFTs (Pre-NFTs) and mature intracellular (iNFTs) and show ML models can be trained to learn annotator nuances for the task of NFT detection in WSIs. We utilize a model-assisted-labeling approach and demonstrate ML models can be used to aid in labeling large datasets efficiently. We also show these models can be used to extract case-level features, which predict Braak NFT stages comparable to expert human raters, and do so at scale. This study provides a generalizable workflow for various pathology and related fields, and also provides a technique for accomplishing a high-level neuropathology task with limited human annotations.

Differential DNA Methylation in the Brain as Potential Mediator of the Association between Traffic-related PM2.5 and Neuropathology Markers of Alzheimer's Disease.

INTRODUCTION: Growing evidence indicates fine particulate matter (PM2.5) as risk factor for Alzheimer's' disease (AD), but the underlying mechanisms have been insufficiently investigated. We hypothesized differential DNA methylation (DNAm) in brain tissue as potential mediator of this association. METHODS: We assessed genome-wide DNAm (Illumina EPIC BeadChips) in prefrontal cortex tissue and three AD-related neuropathological markers (Braak stage, CERAD, ABC score) for 159 donors, and estimated donors' residential traffic-related PM2.5 exposure 1, 3 and 5 years prior to death. We used a combination of the Meet-in-the-Middle approach, high-dimensional mediation analysis, and causal mediation analysis to identify potential mediating CpGs. RESULTS: PM2.5 was significantly associated with differential DNAm at cg25433380 and cg10495669. Twenty-six CpG sites were identified as mediators of the association between PM2.5 exposure and neuropathology markers, several located in genes related to neuroinflammation. DISCUSSION: Our findings suggest differential DNAm related to neuroinflammation mediates the association between traffic-related PM2.5 and AD.

Increased 3-O-sulfated heparan sulfate in Alzheimer's disease brain is associated with genetic risk gene HS3ST1.

HS3ST1 is a genetic risk gene associated with Alzheimer's disease (AD) and overexpressed in patients, but how it contributes to the disease progression is unknown. We report the analysis of brain heparan sulfate (HS) from AD and other tauopathies using a LC-MS/MS method. A specific 3-O-sulfated HS displayed sevenfold increase in the AD group (n = 14, P < 0.0005). Analysis of the HS modified by recombinant sulfotransferases and HS from genetic knockout mice revealed that the specific 3-O-sulfated HS is made by 3-O-sulfotransferase isoform 1 (3-OST-1), which is encoded by the HS3ST1 gene. A synthetic tetradecasaccharide (14-mer) carrying the specific 3-O-sulfated domain displayed stronger inhibition for tau internalization than a 14-mer without the domain, suggesting that the 3-O-sulfated HS is used in tau cellular uptake. Our findings suggest that the overexpression of HS3ST1 gene may enhance the spread of tau pathology, uncovering a previously unidentified therapeutic target for AD.

Ancestry-related differences in chromatin accessibility and gene expression of APOE ε4 are associated with Alzheimer's disease risk.

INTRODUCTION: European local ancestry (ELA) surrounding apolipoprotein E (APOE) ε4 confers higher risk for Alzheimer's disease (AD) compared to African local ancestry (ALA). We demonstrated significantly higher APOE ε4  expression in ELA versus ALA in AD brains from APOE ε4/ε4 carriers. Chromatin accessibility differences could contribute to these expression changes. METHODS: We performed single nuclei assays for transposase accessible chromatin sequencing from the frontal cortex of six ALA and six ELA AD brains, homozygous for local ancestry and APOE ε4. RESULTS: Our results showed an increased chromatin accessibility at the APOE ε4  promoter area in ELA versus ALA astrocytes. This increased accessibility in ELA astrocytes extended genome wide. Genes with increased accessibility in ELA in astrocytes were enriched for synapsis, cholesterol processing, and astrocyte reactivity. DISCUSSION: Our results suggest that increased chromatin accessibility of APOE ε4  in ELA astrocytes contributes to the observed elevated APOE ε4  expression, corresponding to the increased AD risk in ELA versus ALA APOE ε4/ε4 carriers.

Fine particulate air pollution and neuropathology markers of Alzheimer's disease in donors with and without APOE ε4 alleles - results from an autopsy cohort.

Introduction: Higher fine particulate matter (PM2.5) exposure has been found to be associated with Alzheimer's disease (AD). PM2.5 has been hypothesized to cause inflammation and oxidative stress in the brain, contributing to neuropathology. A major genetic risk factor of AD, the apolipoprotein E (APOE) gene, has also been hypothesized to modify the association between PM2.5 and AD. However, little prior research exisits to support these hypotheses. Therefore, this paper aims to investigate the association between traffic-related PM2.5 and AD hallmark pathology, including effect modification by APOE genotype, in an autopsy cohort. Methods: Brain tissue donors enrolled in the Emory Goizueta Alzheimer's Disease Research Center (ADRC) who died before 2020 (n=224) were assessed for AD pathology including Braak Stage, Consortium to Establish a Registry for AD (CERAD) score, and the combined AD neuropathologic change (ABC score). Traffic-related PM2.5 concentrations were modeled for the metro-Atlanta area during 2002-2019 with a spatial resolution of 200-250m. One-, 3-, and 5-year average PM2.5 concentrations prior to death were matched to participants home address. We assessed the association between traffic-related PM2.5 and AD hallmark pathology, as well as effect modification by APOE genotype, using adjusted ordinal logistic regression models. Results: Traffic-related PM2.5 was significantly associated with CERAD score for the 1-year exposure window (OR: 1.92; 95% CI: 1.12, 3.30), and the 3-year exposure window (OR: 1.87; 95%-CI: 1.01, 3.17). PM2.5 had harmful, but non-significant associations on Braak Stage and ABC score. The strongest associations between PM2.5 and neuropathology markers were among those without APOE ε4 alleles (e.g., for CERAD and 1-year exposure window, OR: 2.31; 95% CI: 1.36, 3.94), though interaction between PM2.5 and APOE genotype was not statistically significant. Conclusions: Our study found traffic-related PM2.5 exposure was associated with CERAD score in an autopsy cohort, contributing to epidemiologic evidence that PM2.5 affects Aß deposition in the brain. This association was particularly strong among donors without APOE ε4 alleles. Future studies should further investigate the biological mechanisms behind this assocation.

Cross-Platform Synaptic Network Analysis of Human Entorhinal Cortex Identifies TWF2 as a Modulator of Dendritic Spine Length.

Proteomic studies using postmortem human brain tissue samples have yielded robust assessments of the aging and neurodegenerative disease(s) proteomes. While these analyses provide lists of molecular alterations in human conditions, like Alzheimer's disease (AD), identifying individual proteins that affect biological processes remains a challenge. To complicate matters, protein targets may be highly understudied and have limited information on their function. To address these hurdles, we sought to establish a blueprint to aid selection and functional validation of targets from proteomic datasets. A cross-platform pipeline was engineered to focus on synaptic processes in the entorhinal cortex (EC) of human patients, including controls, preclinical AD, and AD cases. Label-free quantification mass spectrometry (MS) data (n = 2260 proteins) was generated on synaptosome fractionated tissue from Brodmann area 28 (BA28; n = 58 samples). In parallel, dendritic spine density and morphology was measured in the same individuals. Weighted gene co-expression network analysis was used to construct a network of protein co-expression modules that were correlated with dendritic spine metrics. Module-trait correlations were used to guide unbiased selection of Twinfilin-2 (TWF2), which was the top hub protein of a module that positively correlated with thin spine length. Using CRISPR-dCas9 activation strategies, we demonstrated that boosting endogenous TWF2 protein levels in primary hippocampal neurons increased thin spine length, thus providing experimental validation for the human network analysis. Collectively, this study describes alterations in dendritic spine density and morphology as well as synaptic proteins and phosphorylated tau from the entorhinal cortex of preclinical and advanced stage AD patients.SIGNIFICANCE STATEMENT Proteomic studies can yield vast lists of molecules that are altered under various experimental or disease conditions. Here, we provide a blueprint to facilitate mechanistic validation of protein targets from human brain proteomic datasets. We conducted a proteomic analysis of human entorhinal cortex (EC) samples spanning cognitively normal and Alzheimer's disease (AD) cases with a comparison of dendritic spine morphology in the same samples. Network integration of proteomics with dendritic spine measurements allowed for unbiased discovery of Twinfilin-2 (TWF2) as a regulator of dendritic spine length. A proof-of-concept experiment in cultured neurons demonstrated that altering Twinfilin-2 protein level induced corresponding changes in dendritic spine length, thus providing experimental validation for the computational framework.

MicroRNA-650 Regulates the Pathogenesis of Alzheimer's Disease Through Targeting Cyclin-Dependent Kinase 5.

Alzheimer's disease (AD) pathogenesis feature progressive neurodegeneration, amyloid-ß plaque formation, and neurofibrillary tangles. Ample evidence has indicated the involvement of epigenetic pathways in AD pathogenesis. Here, we show that the expression of microRNA 650 (miR-650) is altered in brains from AD patients. Furthermore, we found that the processing of primary miR-650 to mature miR-650 is misregulated. Bioinformatic analysis predicted that miR-650 targets the expression of three AD-associated components: Apolipoprotein E (APOE), Presenilin 1 (PSEN1), and Cyclin-Dependent Kinase 5 (CDK5), and we have experimentally confirmed that miR-650 is able to significantly reduce the expression of APOE, PSEN1, and CDK5 in vitro. Importantly, the overexpression of miR-650 was further shown to significantly alter the CDK5 level and ameliorate AD pathologies in APP-PSEN1 transgenic mice. Overall, our results indicate that miR-650 influences AD pathogenesis through regulation of CDK5.

Single nucleus multiome analysis of the prefrontal cortex from C9orf72 ALS/FTD patients illuminates pathways affected during disease progression.

Repeat expansions in the C9orf72 gene are the most common genetic cause of amyotrophic lateral sclerosis and familial frontotemporal dementia (ALS/FTD). To identify molecular defects that take place in the dorsolateral frontal cortex of patients with C9orf72 ALS/FTD, we compared healthy controls with C9orf72 ALS/FTD donor samples staged based on the levels of cortical phosphorylated TAR DNA binding protein (pTDP-43), a neuropathological hallmark of disease progression. We identified distinct molecular changes in different cell types that take place during disease progression. These alterations include downregulation of nuclear and mitochondrial ribosomal protein genes in early disease stages that become upregulated as the disease progresses. High ratios of premature oligodendrocytes expressing low levels of genes encoding major myelin protein components are characteristic of late disease stages and may represent a unique signature of C9orf72 ALS/FTD. Microglia with increased reactivity and astrocyte specific transcriptome changes in genes involved in glucose/glycogen metabolism are also associated with disease progression. Late stages of C9orf72 ALS/FTD correlate with sequential changes in the regulatory landscape of several genes in glial cells, namely MBP/MAG/MOG in oligodendrocytes, CD83/IRF8 in microglia, and GLUT1/GYS2/AGL in astrocytes. Only layer 2-3 cortical projection neurons with high expression of CUX2/LAMP5 are significantly reduced in C9orf72 ALS/FTD patients with respect to controls. Our findings reveal previously unknown progressive functional changes in cortical cells of C9orf72 ALS/FTD patients that shed light on the mechanisms underlying the pathology of this disease.

Medin co-aggregates with vascular amyloid-ß in Alzheimer's disease.

Aggregates of medin amyloid (a fragment of the protein MFG-E8, also known as lactadherin) are found in the vasculature of almost all humans over 50 years of age1,2, making it the most common amyloid currently known. We recently reported that medin also aggregates in blood vessels of ageing wild-type mice, causing cerebrovascular dysfunction3. Here we demonstrate in amyloid-ß precursor protein (APP) transgenic mice and in patients with Alzheimer's disease that medin co-localizes with vascular amyloid-ß deposits, and that in mice, medin deficiency reduces vascular amyloid-ß deposition by half. Moreover, in both the mouse and human brain, MFG-E8 is highly enriched in the vasculature and both MFG-E8 and medin levels increase with the severity of vascular amyloid-ß burden. Additionally, analysing data from 566 individuals in the ROSMAP cohort, we find that patients with Alzheimer's disease have higher MFGE8 expression levels, which are attributable to vascular cells and are associated with increased measures of cognitive decline, independent of plaque and tau pathology. Mechanistically, we demonstrate that medin interacts directly with amyloid-ß to promote its aggregation, as medin forms heterologous fibrils with amyloid-ß, affects amyloid-ß fibril structure, and cross-seeds amyloid-ß aggregation both in vitro and in vivo. Thus, medin could be a therapeutic target for prevention of vascular damage and cognitive decline resulting from amyloid-ß deposition in the blood vessels of the brain.

Common Variants Near ZIC1 and ZIC4 in Autopsy-Confirmed Multiple System Atrophy.

BACKGROUND: Multiple System Atrophy is a rare neurodegenerative disease with alpha-synuclein aggregation in glial cytoplasmic inclusions and either predominant olivopontocerebellar atrophy or striatonigral degeneration, leading to dysautonomia, parkinsonism, and cerebellar ataxia. One prior genome-wide association study in mainly clinically diagnosed patients with Multiple System Atrophy failed to identify genetic variants predisposing for the disease. OBJECTIVE: Since the clinical diagnosis of Multiple System Atrophy yields a high rate of misdiagnosis when compared to the neuropathological gold standard, we studied only autopsy-confirmed cases. METHODS: We studied common genetic variations in Multiple System Atrophy cases (N = 731) and controls (N = 2898). RESULTS: The most strongly disease-associated markers were rs16859966 on chromosome 3, rs7013955 on chromosome 8, and rs116607983 on chromosome 4 with P-values below 5 × 10-6 , all of which were supported by at least one additional genotyped and several imputed single nucleotide polymorphisms. The genes closest to the chromosome 3 locus are ZIC1 and ZIC4 encoding the zinc finger proteins of cerebellum 1 and 4 (ZIC1 and ZIC4). INTERPRETATION: Since mutations of ZIC1 and ZIC4 and paraneoplastic autoantibodies directed against ZIC4 are associated with severe cerebellar dysfunction, we conducted immunohistochemical analyses in brain tissue of the frontal cortex and the cerebellum from 24 Multiple System Atrophy patients. Strong immunohistochemical expression of ZIC4 was detected in a subset of neurons of the dentate nucleus in all healthy controls and in patients with striatonigral degeneration, whereas ZIC4-immunoreactive neurons were significantly reduced inpatients with olivopontocerebellar atrophy. These findings point to a potential ZIC4-mediated vulnerability of neurons in Multiple System Atrophy. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Large-scale deep multi-layer analysis of Alzheimer's disease brain reveals strong proteomic disease-related changes not observed at the RNA level.

The biological processes that are disrupted in the Alzheimer's disease (AD) brain remain incompletely understood. In this study, we analyzed the proteomes of more than 1,000 brain tissues to reveal new AD-related protein co-expression modules that were highly preserved across cohorts and brain regions. Nearly half of the protein co-expression modules, including modules significantly altered in AD, were not observed in RNA networks from the same cohorts and brain regions, highlighting the proteopathic nature of AD. Two such AD-associated modules unique to the proteomic network included a module related to MAPK signaling and metabolism and a module related to the matrisome. The matrisome module was influenced by the APOE ε4 allele but was not related to the rate of cognitive decline after adjustment for neuropathology. By contrast, the MAPK/metabolism module was strongly associated with the rate of cognitive decline. Disease-associated modules unique to the proteome are sources of promising therapeutic targets and biomarkers for AD.