Optimal blood tau species for the detection of Alzheimer's disease neuropathology: an immunoprecipitation mass spectrometry and autopsy study.

Plasma-to-autopsy studies are essential for validation of blood biomarkers and understanding their relation to Alzheimer's disease (AD) pathology. Few such studies have been done on phosphorylated tau (p-tau) and those that exist have made limited or no comparison of the different p-tau variants. This study is the first to use immunoprecipitation mass spectrometry (IP-MS) to compare the accuracy of eight different plasma tau species in predicting autopsy-confirmed AD. The sample included 123 participants (AD = 69, non-AD = 54) from the Boston University Alzheimer's disease Research Center who had an available ante-mortem plasma sample and donated their brain. Plasma samples proximate to death were analyzed by targeted IP-MS for six different tryptic phosphorylated (p-tau-181, 199, 202, 205, 217, 231), and two non-phosphorylated tau (195-205, 212-221) peptides. NIA-Reagan Institute criteria were used for the neuropathological diagnosis of AD. Binary logistic regressions tested the association between each plasma peptide and autopsy-confirmed AD status. Area under the receiver operating curve (AUC) statistics were generated using predicted probabilities from the logistic regression models. Odds Ratio (OR) was used to study associations between the different plasma tau species and CERAD and Braak classifications. All tau species were increased in AD compared to non-AD, but p-tau217, p-tau205 and p-tau231 showed the highest fold-changes. Plasma p-tau217 (AUC = 89.8), p-tau231 (AUC = 83.4), and p-tau205 (AUC = 81.3) all had excellent accuracy in discriminating AD from non-AD brain donors, even among those with CDR < 1). Furthermore, p-tau217, p-tau205 and p-tau231 showed the highest ORs with both CERAD (ORp-tau217 = 15.29, ORp-tau205 = 5.05 and ORp-tau231 = 3.86) and Braak staging (ORp-tau217 = 14.29, ORp-tau205 = 5.27 and ORp-tau231 = 4.02) but presented increased levels at different amyloid and tau stages determined by neuropathological examination. Our findings support plasma p-tau217 as the most promising p-tau species for detecting AD brain pathology. Plasma p-tau231 and p-tau205 may additionally function as markers for different stages of the disease.

Ante-mortem plasma phosphorylated tau (181) predicts Alzheimer's disease neuropathology and regional tau at autopsy.

Blood-based biomarkers such as tau phosphorylated at threonine 181 (phosphorylated-tau181) represent an accessible, cost-effective and scalable approach for the in vivo detection of Alzheimer's disease pathophysiology. Plasma-pathological correlation studies are needed to validate plasma phosphorylated-tau181 as an accurate and reliable biomarker of Alzheimer's disease neuropathological changes. This plasma-to-autopsy correlation study included participants from the Boston University Alzheimer's Disease Research Center who had a plasma sample analysed for phosphorylated-tau181 between 2008 and 2018 and donated their brain for neuropathological examination. Plasma phosphorelated-tau181 was measured with single molecule array technology. Of 103 participants, 62 (60.2%) had autopsy-confirmed Alzheimer's disease. Average time between blood draw and death was 5.6 years (standard deviation = 3.1 years). Multivariable analyses showed higher plasma phosphorylated-tau181 concentrations were associated with increased odds for having autopsy-confirmed Alzheimer's disease [AUC = 0.82, OR = 1.07, 95% CI = 1.03-1.11, P < 0.01; phosphorylated-tau standardized (z-transformed): OR = 2.98, 95% CI = 1.50-5.93, P < 0.01]. Higher plasma phosphorylated-tau181 levels were associated with increased odds for having a higher Braak stage (OR = 1.06, 95% CI = 1.02-1.09, P < 0.01) and more severe phosphorylated-tau across six cortical and subcortical brain regions (ORs = 1.03-1.06, P < 0.05). The association between plasma phosphorylated-tau181 and Alzheimer's disease was strongest in those who were demented at time of blood draw (OR = 1.25, 95%CI = 1.02-1.53), but an effect existed among the non-demented (OR = 1.05, 95% CI = 1.01-1.10). There was higher discrimination accuracy for Alzheimer's disease when blood draw occurred in years closer to death; however, higher plasma phosphorylated-tau181 levels were associated with Alzheimer's disease even when blood draw occurred >5 years from death. Ante-mortem plasma phosphorylated-tau181 concentrations were associated with Alzheimer's disease neuropathology and accurately differentiated brain donors with and without autopsy-confirmed Alzheimer's disease. These findings support plasma phosphorylated-tau181 as a scalable biomarker for the detection of Alzheimer's disease.

Alzheimer's disease heterogeneity explained by polygenic risk scores derived from brain transcriptomic profiles.

INTRODUCTION: Alzheimer's disease (AD) is heterogeneous, both clinically and neuropathologically. We investigated whether polygenic risk scores (PRSs) integrated with transcriptome profiles from AD brains can explain AD clinical heterogeneity. METHODS: We conducted co-expression network analysis and identified gene sets (modules) that were preserved in three AD transcriptome datasets and associated with AD-related neuropathological traits including neuritic plaques (NPs) and neurofibrillary tangles (NFTs). We computed the module-based PRSs (mbPRSs) for each module and tested associations with mbPRSs for cognitive test scores, cognitively defined AD subgroups, and brain imaging data. RESULTS: Of the modules significantly associated with NPs and/or NFTs, the mbPRSs from two modules (M6 and M9) showed distinct associations with language and visuospatial functioning, respectively. They matched clinical subtypes and brain atrophy at specific regions. DISCUSSION: Our findings demonstrate that polygenic profiling based on co-expressed gene sets can explain heterogeneity in AD patients, enabling genetically informed patient stratification and precision medicine in AD. HIGHLIGHTS: Co-expression gene-network analysis in Alzheimer's disease (AD) brains identified gene sets (modules) associated with AD heterogeneity. AD-associated modules were selected when genes in each module were enriched for neuritic plaques and neurofibrillary tangles. Polygenic risk scores from two selected modules were linked to the matching cognitively defined AD subgroups (language and visuospatial subgroups). Polygenic risk scores from the two modules were associated with cognitive performance in language and visuospatial domains and the associations were confirmed in regional-specific brain atrophy data.

Integrative brain transcriptome analysis links complement component 4 and HSPA2 to the APOE ε2 protective effect in Alzheimer disease.

Mechanisms underlying the protective effect of apolipoprotein E (APOE) ε2 against Alzheimer disease (AD) are not well understood. We analyzed gene expression data derived from autopsied brains donated by 982 individuals including 135 APOE ɛ2/ɛ3 carriers. Complement pathway genes C4A and C4B were among the most significantly differentially expressed genes between ɛ2/ɛ3 AD cases and controls. We also identified an APOE ε2/ε3 AD-specific co-expression network enriched for astrocytes, oligodendrocytes and oligodendrocyte progenitor cells containing the genes C4A, C4B, and HSPA2. These genes were significantly associated with the ratio of phosphorylated tau at position 231 to total Tau but not with amyloid-ß 42 level, suggesting this APOE ɛ2 related co-expression network may primarily be involved with tau pathology. HSPA2 expression was oligodendrocyte-specific and significantly associated with C4B protein. Our findings provide the first evidence of a crucial role of the complement pathway in the protective effect of APOE ε2 for AD.

Protein phosphatase 2A and complement component 4 are linked to the protective effect of APOE ɛ2 for Alzheimer's disease.

INTRODUCTION: The apolipoprotein E (APOE) ɛ2 allele reduces risk against Alzheimer's disease (AD) but mechanisms underlying this effect are largely unknown. METHODS: We conducted a genome-wide association study for AD among 2096 ɛ2 carriers. The potential role of the top-ranked gene and complement 4 (C4) proteins, which were previously linked to AD in ɛ2 carriers, was investigated using human isogenic APOE allele-specific induced pluripotent stem cell (iPSC)-derived neurons and astrocytes and in 224 neuropathologically examined human brains. RESULTS: PPP2CB rs117296832 was the second most significantly associated single nucleotide polymorphism among ɛ2 carriers (P = 1.1 × 10-7 ) and the AD risk allele increased PPP2CB expression in blood (P = 6.6 × 10-27 ). PPP2CB expression was correlated with phosphorylated tau231/total tau ratio (P = .01) and expression of C4 protein subunits C4A/B (P = 2.0 × 10-4 ) in the iPSCs. PPP2CB (subunit of protein phosphatase 2A) and C4b protein levels were correlated in brain (P = 3.3 × 10-7 ). DISCUSSION: PP2A may be linked to classical complement activation leading to AD-related tau pathology.

Genome-wide association and multi-omics studies identify MGMT as a novel risk gene for Alzheimer's disease among women.

INTRODUCTION: Variants in the tau gene (MAPT) region are associated with breast cancer in women and Alzheimer's disease (AD) among persons lacking apolipoprotein E ε4 (ε4-). METHODS: To identify novel genes associated with tau-related pathology, we conducted two genome-wide association studies (GWAS) for AD, one among 10,340 ε4- women in the Alzheimer's Disease Genetics Consortium (ADGC) and another in 31 members (22 women) of a consanguineous Hutterite kindred. RESULTS: We identified novel associations of AD with MGMT variants in the ADGC (rs12775171, odds ratio [OR] = 1.4, P = 4.9 × 10-8 ) and Hutterite (rs12256016 and rs2803456, OR = 2.0, P = 1.9 × 10-14 ) datasets. Multi-omics analyses showed that the most significant and largest number of associations among the single nucleotide polymorphisms (SNPs), DNA-methylated CpGs, MGMT expression, and AD-related neuropathological traits were observed among women. Furthermore, promoter capture Hi-C analyses revealed long-range interactions of the MGMT promoter with MGMT SNPs and CpG sites. DISCUSSION: These findings suggest that epigenetically regulated MGMT expression is involved in AD pathogenesis, especially in women.

Plasma p-tau181 shows stronger network association to Alzheimer's disease dementia than neurofilament light and total tau.

INTRODUCTION: We examined the ability of plasma hyperphosphorylated tau (p-tau)181 to detect cognitive impairment due to Alzheimer's disease (AD) independently and in combination with plasma total tau (t-tau) and neurofilament light (NfL). METHODS: Plasma samples were analyzed using the Simoa platform for 235 participants with normal cognition (NC), 181 with mild cognitive impairment due to AD (MCI), and 153 with AD dementia. Statistical approaches included multinomial regression and Gaussian graphical models (GGMs) to assess a network of plasma biomarkers, neuropsychological tests, and demographic variables. RESULTS: Plasma p-tau181 discriminated AD dementia from NC, but not MCI, and correlated with dementia severity and worse neuropsychological test performance. Plasma NfL similarly discriminated diagnostic groups. Unlike plasma NfL or t-tau, p-tau181 had a direct association with cognitive diagnosis in a bootstrapped GGM. DISCUSSION: These results support plasma p-tau181 for the detection of AD dementia and the use of blood-based biomarkers for optimal disease detection.

Genome-wide association study of Alzheimer's disease endophenotypes at prediagnosis stages.

INTRODUCTION: Genetic associations for endophenotypes of Alzheimer's disease (AD) in cognitive stages preceding AD have not been thoroughly evaluated. METHODS: We conducted genome-wide association studies for AD-related endophenotypes including hippocampal volume, logical memory scores, and cerebrospinal fluid Aß42 and total/phosphorylated tau in cognitively normal (CN), mild cognitive impairment, and AD dementia subjects from the Alzheimer's Disease Neuroimaging Initiative study. RESULTS: In CN subjects, study-wide significant (P < 8.3 × 10-9) loci were identified for total tau near SRRM4 and C14orf79 and for hippocampal volume near MTUS1. In mild cognitive impairment subjects, study-wide significant association was found with single nucleotide polymorphisms (SNPs) near ZNF804B for logical memory test of delayed recall scores. We found consistent expression patterns of C14orf40 and MTUS1 in carriers with risk alleles of expression SNPs and in brains of AD patients, compared with in the noncarriers and in brains of controls. DISCUSSION: Our findings for AD-related brain changes before AD provide insight about early AD-related biological processes.

Transethnic genome-wide scan identifies novel Alzheimer's disease loci.

INTRODUCTION: Genetic loci for Alzheimer's disease (AD) have been identified in whites of European ancestry, but the genetic architecture of AD among other populations is less understood. METHODS: We conducted a transethnic genome-wide association study (GWAS) for late-onset AD in Stage 1 sample including whites of European Ancestry, African-Americans, Japanese, and Israeli-Arabs assembled by the Alzheimer's Disease Genetics Consortium. Suggestive results from Stage 1 from novel loci were followed up using summarized results in the International Genomics Alzheimer's Project GWAS dataset. RESULTS: Genome-wide significant (GWS) associations in single-nucleotide polymorphism (SNP)-based tests (P < 5 × 10-8) were identified for SNPs in PFDN1/HBEGF, USP6NL/ECHDC3, and BZRAP1-AS1 and for the interaction of the (apolipoprotein E) APOE ε4 allele with NFIC SNP. We also obtained GWS evidence (P < 2.7 × 10-6) for gene-based association in the total sample with a novel locus, TPBG (P = 1.8 × 10-6). DISCUSSION: Our findings highlight the value of transethnic studies for identifying novel AD susceptibility loci.

APOE genotype-specific methylation patterns are linked to Alzheimer disease pathology and estrogen response.

The joint effects of APOE genotype and DNA methylation on Alzheimer disease (AD) risk is relatively unknown. We conducted genome-wide methylation analyses using 2,021 samples in blood (91 AD cases, 329 mild cognitive impairment, 1,391 controls) and 697 samples in brain (417 AD cases, 280 controls). We identified differentially methylated levels in AD compared to controls in an APOE genotype-specific manner at 25 cytosine-phosphate-guanine (CpG) sites in brain and 36 CpG sites in blood. Additionally, we identified seven CpG sites in the APOE region containing TOMM40, APOE, and APOC1 genes with P < 5 × 10-8 between APOE ε4 carriers and non-carriers in brain or blood. In brain, the most significant CpG site hypomethylated in ε4 carriers compared to non-carriers was from the TOMM40 in the total sample, while most of the evidence was derived from AD cases. However, the CpG site was not significantly modulating expression of these three genes in brain. Three CpG sites from the APOE were hypermethylated in APOE ε4 carriers in brain or blood compared in ε4 non-carriers and nominally significant with APOE expression in brain. Three CpG sites from the APOC1 were hypermethylated in blood, which one of the 3 CpG sites significantly lowered APOC1 expression in blood using all subjects or ε4 non-carriers. Co-methylation network analysis in blood and brain detected eight methylation networks associated with AD and APOE ε4 status. Five of the eight networks included genes containing network CpGs that were significantly enriched for estradiol perturbation, where four of the five networks were enriched for the estrogen response pathway. Our findings provide further evidence of the role of APOE genotype on methylation levels associated with AD, especially linked to estrogen response pathway.

Brain Cell-based Genetic Subtyping and Drug Repositioning for Alzheimer Disease.

Alzheimer's Disease (AD) is characterized by its complex and heterogeneous etiology and gradual progression, leading to high drug failure rates in late-stage clinical trials. In order to better stratify individuals at risk for AD and discern potential therapeutic targets we employed a novel procedure utilizing cell-based co-regulated gene networks and polygenic risk scores (cbPRSs). After defining genetic subtypes using extremes of cbPRS distributions, we evaluated correlations of the genetic subtypes with previously defined AD subtypes defined on the basis of domain-specific cognitive functioning and neuroimaging biomarkers. Employing a PageRank algorithm, we identified priority gene targets for the genetic subtypes. Pathway analysis of priority genes demonstrated associations with neurodegeneration and suggested candidate drugs currently utilized in diabetes, hypertension, and epilepsy for repositioning in AD. Experimental validation utilizing human induced pluripotent stem cell (hiPSC)-derived astrocytes demonstrated the modifying effects of estradiol, levetiracetam, and pioglitazone on expression of APOE and complement C4 genes, suggesting potential repositioning for AD.

Cross-sectional and longitudinal evaluation of plasma glial fibrillary acidic protein to detect and predict clinical syndromes of Alzheimer's disease.

Introduction: This study examined plasma glial fibrillary acidic protein (GFAP) as a biomarker of cognitive impairment due to Alzheimer's disease (AD) with and against plasma neurofilament light chain (NfL), and phosphorylated tau (p-tau)181+231. Methods: Plasma samples were analyzed using Simoa platform for 567 participants spanning the AD continuum. Cognitive diagnosis, neuropsychological testing, and dementia severity were examined for cross-sectional and longitudinal outcomes. Results: Plasma GFAP discriminated AD dementia from normal cognition (adjusted mean difference = 0.90 standard deviation [SD]) and mild cognitive impairment (adjusted mean difference = 0.72 SD), and demonstrated superior discrimination compared to alternative plasma biomarkers. Higher GFAP was associated with worse dementia severity and worse performance on 11 of 12 neuropsychological tests. Longitudinally, GFAP predicted decline in memory, but did not predict conversion to mild cognitive impairment or dementia. Discussion: Plasma GFAP was associated with clinical outcomes related to suspected AD and could be of assistance in a plasma biomarker panel to detect in vivo AD.

Blood and brain transcriptome analysis reveals APOE genotype-mediated and immune-related pathways involved in Alzheimer disease.

BACKGROUND: While Alzheimer disease (AD) is generally considered as a brain disorder, blood biomarkers may be useful for the diagnosis and prediction of AD brain pathology. The APOE ε4 allele has shown cerebrovascular effects including acceleration of blood-brain barrier (BBB) breakdown. METHODS: We evaluated the differential expression of previously established AD genes in brains from 344 pathologically confirmed AD cases and 232 controls and in blood from 112 pathologically confirmed AD cases and 67 controls from the Religious Orders Study and Memory and Aging Project. Differential gene expression between AD cases and controls was analyzed in the blood and brain jointly using a multivariate approach in the total sample and within APOE genotype groups. Gene set enrichment analysis was performed within APOE genotype groups using the results from the combined blood and brain analyses to identify biologically important pathways. Gene co-expression networks in brain and blood samples were investigated using weighted correlation network analysis. Top-ranked genes from networks and pathways were further evaluated with vascular injury traits. RESULTS: We observed differentially expressed genes with P < 0.05 in both brain and blood for established AD genes INPP5D (upregulated) and HLA-DQA1 (downregulated). PIGHP1 and FRAS1 were differentially expressed at the transcriptome-wide level (P < 3.3 × 10-6) within ε2/ε3 and ε3/ε4 groups, respectively. Gene set enrichment analysis revealed 21 significant pathways (false discovery rate P < 0.05) in at least one APOE genotype group. Ten pathways were significantly enriched in the ε3/ε4 group, and six of these were unique to these subjects. Four pathways (allograft rejection, interferon gamma response, peroxisome, and TNFA signaling via NFKB) were enriched for AD upregulated genes in the ε3/ε4 group and AD downregulated genes in subjects lacking ε4. We identified a co-expressed gene network in the brain that reproduced in blood and showed higher average expression in ε4 carriers. Twenty-three genes from pathway and network analyses were significantly associated with at least one vascular injury trait. CONCLUSION: These results suggest that the APOE genotype contributes to unique expression network profiles in both blood and brain. Several genes in these networks are associated with measures of vascular injury and potentially contribute to ε4's effect on the BBB.

Cytokine Levels in Human Vitreous in Proliferative Diabetic Retinopathy.

In this study, we compare the vitreous cytokine profile in patients with proliferative diabetic retinopathy (PDR) to that of patients without PDR. The identification of novel cytokines involved in the pathogenesis of PDR provides candidate therapeutic targets that may stand alone or work synergistically with current therapies in the management of diabetic retinopathy. Undiluted vitreous humor specimens were collected from 74 patients undergoing vitrectomy for various vitreoretinal disorders. Quantitative immunoassay was performed for a panel of 36 neuroinflammatory cytokines in each specimen and assessed to identify differences between PDR (n = 35) and non-PDR (n = 39) patients. Levels of interleukin-8 (IL-8), IL-15, IL-16, vascular endothelial growth factor (VEGF), VEGF-D, c-reactive protein (CRP), serum amyloid-A (SAA), and intracellular adhesion molecule-1 (ICAM1) were significantly increased in the vitreous of PDR patients compared to non-PDR patients (p < 0.05). We report novel increases in IL-15 and IL-16, in addition to the expected VEGF, in the human vitreous humor of patients with PDR. Additionally, we confirm the elevation of ICAM-1, VCAM-1, SAA, IL-8 and CRP in the vitreous of patients with PDR, which has previously been described.

A missense variant in SHARPIN mediates Alzheimer's disease-specific brain damages.

Established genetic risk factors for Alzheimer's disease (AD) account for only a portion of AD heritability. The aim of this study was to identify novel associations between genetic variants and AD-specific brain atrophy. We conducted genome-wide association studies for brain magnetic resonance imaging measures of hippocampal volume and entorhinal cortical thickness in 2643 Koreans meeting the clinical criteria for AD (n = 209), mild cognitive impairment (n = 1449) or normal cognition (n = 985). A missense variant, rs77359862 (R274W), in the SHANK-associated RH Domain Interactor (SHARPIN) gene was associated with entorhinal cortical thickness (p = 5.0 × 10-9) and hippocampal volume (p = 5.1 × 10-12). It revealed an increased risk of developing AD in the mediation analyses. This variant was also associated with amyloid-ß accumulation (p = 0.03) and measures of memory (p = 1.0 × 10-4) and executive function (p = 0.04). We also found significant association of other SHARPIN variants with hippocampal volume in the Alzheimer's Disease Neuroimaging Initiative (rs3417062, p = 4.1 × 10-6) and AddNeuroMed (rs138412600, p = 5.9 × 10-5) cohorts. Further, molecular dynamics simulations and co-immunoprecipitation indicated that the variant significantly reduced the binding of linear ubiquitination assembly complex proteins, SHPARIN and HOIL-1 Interacting Protein (HOIP), altering the downstream NF-κB signaling pathway. These findings suggest that SHARPIN plays an important role in the pathogenesis of AD.

Novel Alzheimer Disease Risk Loci and Pathways in African American Individuals Using the African Genome Resources Panel: A Meta-analysis.

Importance: Compared with non-Hispanic White individuals, African American individuals from the same community are approximately twice as likely to develop Alzheimer disease. Despite this disparity, the largest Alzheimer disease genome-wide association studies to date have been conducted in non-Hispanic White individuals. In the largest association analyses of Alzheimer disease in African American individuals, ABCA7, TREM2, and an intergenic locus at 5q35 were previously implicated. Objective: To identify additional risk loci in African American individuals by increasing the sample size and using the African Genome Resource panel. Design, Setting, and Participants: This genome-wide association meta-analysis used case-control and family-based data sets from the Alzheimer Disease Genetics Consortium. There were multiple recruitment sites throughout the United States that included individuals with Alzheimer disease and controls of African American ancestry. Analysis began October 2018 and ended September 2019. Main Outcomes and Measures: Diagnosis of Alzheimer disease. Results: A total of 2784 individuals with Alzheimer disease (1944 female [69.8%]) and 5222 controls (3743 female [71.7%]) were analyzed (mean [SD] age at last evaluation, 74.2 [13.6] years). Associations with 4 novel common loci centered near the intracellular glycoprotein trafficking gene EDEM1 (3p26; P = 8.9 × 10-7), near the immune response gene ALCAM (3q13; P = 9.3 × 10-7), within GPC6 (13q31; P = 4.1 × 10-7), a gene critical for recruitment of glutamatergic receptors to the neuronal membrane, and within VRK3 (19q13.33; P = 3.5 × 10-7), a gene involved in glutamate neurotoxicity, were identified. In addition, several loci associated with rare variants, including a genome-wide significant intergenic locus near IGF1R at 15q26 (P = 1.7 × 10-9) and 6 additional loci with suggestive significance (P ≤ 5 × 10-7) such as API5 at 11p12 (P = 8.8 × 10-8) and RBFOX1 at 16p13 (P = 5.4 × 10-7) were identified. Gene expression data from brain tissue demonstrate association of ALCAM, ARAP1, GPC6, and RBFOX1 with brain ß-amyloid load. Of 25 known loci associated with Alzheimer disease in non-Hispanic White individuals, only APOE, ABCA7, TREM2, BIN1, CD2AP, FERMT2, and WWOX were implicated at a nominal significance level or stronger in African American individuals. Pathway analyses strongly support the notion that immunity, lipid processing, and intracellular trafficking pathways underlying Alzheimer disease in African American individuals overlap with those observed in non-Hispanic White individuals. A new pathway emerging from these analyses is the kidney system, suggesting a novel mechanism for Alzheimer disease that needs further exploration. Conclusions and Relevance: While the major pathways involved in Alzheimer disease etiology in African American individuals are similar to those in non-Hispanic White individuals, the disease-associated loci within these pathways differ.

Neurofilament light chain in the vitreous humor of the eye.

BACKGROUND: Neurofilament light chain (NfL) is a promising biomarker of neurodegeneration in the cerebrospinal fluid and blood. This study investigated the presence of NfL in the vitreous humor and its associations with amyloid beta, tau, inflammatory cytokines and vascular proteins, apolipoprotein E (APOE) genotypes, Mini-Mental State Examination (MMSE) scores, systemic disease, and ophthalmic diseases. METHODS: This is a single-site, prospective, cross-sectional cohort study. Undiluted vitreous fluid (0.5-1.0 mL) was aspirated during vitrectomy, and whole blood was drawn for APOE genotyping. NfL, amyloid beta (Aß), total Tau (t-Tau), phosphorylated Tau (p-Tau181), inflammatory cytokines, chemokines, and vascular proteins in the vitreous were quantitatively measured by immunoassay. The main outcome measures were the detection of NfL levels in the vitreous humor and its associations with the aforementioned proteins. Linear regression was used to test the associations of NfL with other proteins, APOE genotypes, MMSE scores, and ophthalmic and systemic diseases after adjustment for age, sex, education level, and other eye diseases. RESULTS: NfL was detected in all 77 vitreous samples. NfL was not found to be associated with ophthalmic conditions, APOE genotypes, MMSE scores, or systemic disease (p > 0.05). NfL levels were positively associated with increased vitreous levels of Aß40 (p = 7.7 × 10-5), Aß42 (p = 2.8 × 10-4), and t-tau (p = 5.5 × 10-7), but not with p-tau181 (p = 0.53). NfL also had significant associations with inflammatory cytokines such as interleukin-15 (IL-15, p = 5.3 × 10-4), IL-16 (p = 2.2 × 10-4), monocyte chemoattractant protein-1 (MCP1, p = 4.1 × 10-4), and vascular proteins such as vascular endothelial growth factor receptor-1 (VEGFR1, p = 2.9 × 10-6), Vegf-C (p = 8.6 × 10-6), vascular cell adhesion molecule-1 (VCAM-1, p = 5.0 × 10-4), Tie-2 (p = 6.3 × 10-4), and intracellular adhesion molecular-1 (ICAM-1, p = 1.6 × 10-4). CONCLUSION: NfL is detectable in the vitreous humor of the eye and significantly associated with amyloid beta, t-tau, and select inflammatory and vascular proteins in the vitreous. Additionally, NfL was not associated with patients' clinical eye condition. Our results serve as a foundation for further investigation of NfL in the ocular fluids to inform us about the potential utility of its presence in the eye.

Exceptionally low likelihood of Alzheimer's dementia in APOE2 homozygotes from a 5,000-person neuropathological study.

Each additional copy of the apolipoprotein E4 (APOE4) allele is associated with a higher risk of Alzheimer's dementia, while the APOE2 allele is associated with a lower risk of Alzheimer's dementia, it is not yet known whether APOE2 homozygotes have a particularly low risk. We generated Alzheimer's dementia odds ratios and other findings in more than 5,000 clinically characterized and neuropathologically characterized Alzheimer's dementia cases and controls. APOE2/2 was associated with a low Alzheimer's dementia odds ratios compared to APOE2/3 and 3/3, and an exceptionally low odds ratio compared to APOE4/4, and the impact of APOE2 and APOE4 gene dose was significantly greater in the neuropathologically confirmed group than in more than 24,000 neuropathologically unconfirmed cases and controls. Finding and targeting the factors by which APOE and its variants influence Alzheimer's disease could have a major impact on the understanding, treatment and prevention of the disease.

Common variants in SOX-2 and congenital cataract genes contribute to age-related nuclear cataract.

Nuclear cataract is the most common type of age-related cataract and a leading cause of blindness worldwide. Age-related nuclear cataract is heritable (h2 = 0.48), but little is known about specific genetic factors underlying this condition. Here we report findings from the largest to date multi-ethnic meta-analysis of genome-wide association studies (discovery cohort N = 14,151 and replication N = 5299) of the International Cataract Genetics Consortium. We confirmed the known genetic association of CRYAA (rs7278468, P = 2.8 × 10-16) with nuclear cataract and identified five new loci associated with this disease: SOX2-OT (rs9842371, P = 1.7 × 10-19), TMPRSS5 (rs4936279, P = 2.5 × 10-10), LINC01412 (rs16823886, P = 1.3 × 10-9), GLTSCR1 (rs1005911, P = 9.8 × 10-9), and COMMD1 (rs62149908, P = 1.2 × 10-8). The results suggest a strong link of age-related nuclear cataract with congenital cataract and eye development genes, and the importance of common genetic variants in maintaining crystalline lens integrity in the aging eye.

Comparison of methods for multivariate gene-based association tests for complex diseases using common variants.

Complex diseases are usually associated with multiple correlated phenotypes, and the analysis of composite scores or disease status may not fully capture the complexity (or multidimensionality). Joint analysis of multiple disease-related phenotypes in genetic tests could potentially increase power to detect association of a disease with common SNPs (or genes). Gene-based tests are designed to identify genes containing multiple risk variants that individually are weakly associated with a univariate trait. We combined three multivariate association tests (O'Brien method, TATES, and MultiPhen) with two gene-based association tests (GATES and VEGAS) and compared performance (type I error and power) of six multivariate gene-based methods using simulated data. Data (n = 2000) for genetic sequence and correlated phenotypes were simulated by varying causal variant proportions and phenotype correlations for various scenarios. These simulations showed that two multivariate association tests (TATES and MultiPhen, but not O'Brien) paired with VEGAS have inflated type I error in all scenarios, while the three multivariate association tests paired with GATES have correct type I error. MultiPhen paired with GATES has higher power than competing methods if the correlations among phenotypes are low (r < 0.57). We applied these gene-based association methods to a GWAS dataset from the Alzheimer's Disease Genetics Consortium containing three neuropathological traits related to Alzheimer disease (neuritic plaque, neurofibrillary tangles, and cerebral amyloid angiopathy) measured in 3500 autopsied brains. Gene-level significant evidence (P < 2.7 × 10-6) was identified in a region containing three contiguous genes (TRAPPC12, TRAPPC12-AS1, ADI1) using O'Brien and VEGAS. Gene-wide significant associations were not observed in univariate gene-based tests.