Postmortem Alzheimer's Disease Research Articles

Premortem perivascular space morphology is a predictor of postmortem glia tau pathology in Alzheimer's disease.

Alzheimer's Disease (AD) is the most common form of dementia in the United States, nearly 5 million people have the disease. Perivascular space (PVS) changes were previously considered an inconsequential finding of AD, but there is recent interest regarding its role in AD progression 1,2 . Here, we investigate the association between several PVS morphological features (from in vivo MRI) and AD pathology (from postmortem histology). Data were derived from Alzheimer's Disease Neuroimaging Initiative (ADNI)3 . Demographic information (sex, race, age) and ADNI pathologic scores were collected. Scores were for Amyloid-B, Tau, and neurofibrillary tangle status from histological images. We also assessed scores for neuronal loss and cerebral amyloid angiopathy (CAA). Using the last premortem MRI, manual segmentation of basal ganglia PVS of each individual was conducted. PVS of the white matter was also automatically mapped using our recently developed technique 1,4 . We then used linear regression to assess if premortem PVS features are predictors of postmortem AD pathology. Basal ganglia PVS count was a predictor of glial Tau pathology in the superior and middle temporal gyri (t=4.1, p=0.00003) and the parahippocampal gyrus (t=3.5, p=0.0005). Basal ganglia PVS volume was also a predictor of glial Tau pathology in the Entorhinal Cortex (t=3.14, p=0.002). Interestingly, PVS volume fraction in white matter centrum semiovale was a predictor of vascular pathology, including CAA in the Amygdala (t=2.6, p=0.008) and Entorhinal cortex (t=2.9, p=0.003). We found that PVS morphology in the basal ganglia was a predictor of glial Tau pathology but not of neurofibrillary tangle pathology. Furthermore, PVS morphology in the basal ganglia was not a predictor of Amyloid-Bdiffuse or cored plaques but interestingly was a predictor for vascular damage.

Microglia across the spectrum from homeostatic to disease phenotypes display enhanced inflammatory responses in human postmortem AD brain.

Microglia-mediated neuroinflammation is hypothesized to contribute to disease progression in neurodegenerative diseases such as Alzheimer's Disease (AD). Recent single-cell RNA-seq studies highlight the heterogeneity of microglial subtypes. Since microglia are a relatively small proportion of CNS cells, standard single-cell omics approaches do not fully capture the diversity of microglia in human subjects. Understanding the gene expression and regulatory networks which define the spectrum of microglia phenotypes is critical to identifying specific targets for neuroinflammation modulating therapies. Our study utilized post-mortem brain tissue from 22 total (7 male) participants with an average age of 86.23 years and post-mortem interval of 5.42 hours. Of those individuals, 12 (3 male) had significant AD neuropathic change present in their brains upon postmortem analysis. Nuclei isolated from prefrontal cortex were sorted for PU.1 expression using fluorescence activated nucleus sorting (FANS). The FANS approach yields larger numbers of nuclei annotated as microglia with high quality sequence from each individual. We performed single-nucleus RNA-seq using the 10X Genomics Chromium platform. We isolated more than 4,000 microglia nuclei from each of the 22 participants, facilitating the comparison of profiles at an individual level in addition to group comparisons based on disease state. Unbiased clustering revealed 8 microglia clusters and improved resolution of microglia heterogeneity compared to standard single-cell approaches. Previously identified mouse disease-associated microglia (DAM) genes are not located in a single cluster of human microglia. "DAM" genes are also present in control aged brains. We identify clusters of microglia enriched for biological pathways implicating defined myeloid roles including a cluster of interferon-stimulated microglia as well as a cluster of autophagic/phagocytic microglia. In many, but not all, of our annotated clusters, including that with high expression of homeostatic genes, individuals with AD have greater enrichment of inflammatory pathways using pseudobulk analysis. We demonstrate significant heterogeneity in microglia subtypes in human brain. Regulation of expression in inflammation-related networks is detected in microglia from both control and AD individuals, though enrichment of specific inflammatory pathways differs. This is important as the field looks to target microglia mechanisms for pharmacological intervention while limiting off-target effects.

Apolipoprotein E4 drives tau propagation through astrocyte-secreted glypican-4.

Apolipoprotein E4 (APOE4) is the most crucial genetic risk factor of late-onset Alzheimer's disease (AD). However, the mechanism through which APOE4 induces AD risk remains unknown. We have utilized post-mortem AD brain tissues of APOE variants, biochemical assays and a series of in vitro and in vivo experiments with transgenic mouse models to study the role of GPC-4 in APOE4 induced tau pathology. Here, we report the astrocyte-secreted protein glypican-4 (GPC-4), as a novel binding partner of APOE4, drives tau pathology. APOE4-carrying post-mortem AD brain tissue display more tau accumulation compared to APOE4-noncarring AD patients. GPC-4 is highly expressed in APOE4 post-mortem AD brains and regulated by microglial factors via NF-κB signaling pathway. The astrocyte-secreted GPC-4 induced both tau accumulation and spreading in vitro and in vivo animal models. Further, GPC-4 is required for APOE4-mediated surface trafficking of low-density lipoprotein receptor-related protein 1 (LRP1) and tau propagation. GPC-4 activates unfolded protein response (UPR) pathway IRE1α, and pharmacological inhibition of IRE1α with KIRA6 blocks GPC-4 induced tau propagation in in vivo animal models. Together, our data demonstrate that one of the APOE4-induced AD risk is directly mediated by GPC-4, and that perturbing GPC-4 induced IRE1α pathway has therapeutic opportunities.

Targeted microRNA characterization in various brain cell‐derived exosome subtypes to identify novel biomarkers for Alzheimer’s disease

AbstractBackgroundMicroRNAs (miRs) offer exciting diagnostic and therapeutic opportunities for Alzheimer’s disease (AD). However, most information about human miRs is from either plasma or CSF of uncertain cellular origin or from postmortem AD brain, which suffers significantly for miR integrity and expression. The discovery of brain cells‐derived exosome (BCDEs) in plasma has led to studies examining their role as ‘liquid biopsies’ for AD. Here, we simultaneously isolated 6 BCDE subtypes from plasma and characterized them for specific miRNAs regulating synaptic plasticity, memory, Aβ, tau, ischemia, and neuroinflammation related signaling pathways.MethodTE were isolated from plasma samples obtained from the Wake Forest Alzheimer’s Disease Research Center: normal cognition (NC; n=8), mild‐cognitive impairment (MCI; n=3), MCI to AD (n=6), and AD (n=11). Next, following 6 BCDE subtypes were extracted using streptavidin‐coated magnetic beads and specific biotin‐tagged antibodies: neuron‐derived exosomes (L1CAM‐biotin antibody), astrocytes‐derived exosomes (GLAST‐biotin antibody), microglia‐derived exosome (TMEM119‐biotin antibody), oligodendrocyte‐derived exosomes (PDGFRα‐biotin antibody), endothelial‐derived exosomes (PECAM1‐biotin antibody), and pericyte‐derived exosomes (PDGFRβ‐biotin antibody). The purity, concentration and size of TE and each BCDE subtype was assessed by flow cytometry, nanoparticle tracking, immunogold labeling, and electron microscopy. The expression of 8 specific miRs (miR‐9a‐5p, miR‐29‐5p, miR‐106‐5p, miR‐107, miR‐125b‐5p, miR‐132‐5p, miR‐135b‐5p, and miR‐210) was measured by TaqMan assays on real‐time PCR.ResultWe observed statistically significant changes in the expression of all 8 miRs in various BCDE subtypes at MCI, MCI‐AD and AD stages. Importantly, we did not observe significant change for most miRNAs in TE, except for miR‐107 and miR‐135b‐5p, highlighting the importance of studying BCDE. Moreover, basal expression of several miRNAs was significantly different between TE and BCDEs. Interestingly, miR‐106b‐5p, miR‐135b‐5p, miR‐107, and miR‐9a‐5b consistently showed significantly higher expression at MCI, MCI‐AD and AD stages for most BCDE subtypes. While the expression of miR‐125b‐5p, miR‐29a‐5p, and miR‐132‐5p showed statistically significant differences based upon BCDE subtype (either an increase or a decrease); for hypoxiamir ‘miR‐210’, expression was higher at MCI stage but decreased with AD progression.ConclusionBCDEs characterization in plasma offers a less invasive opportunity to better understand AD pathogenesis at the cellular level, and to discover novel theranostic targets.

TNF-mediated neuroinflammation is linked to neuronal necroptosis in Alzheimer's disease hippocampus

The pathogenetic mechanisms underlying neuronal death and dysfunction in Alzheimer’s disease (AD) remain unclear. However, chronic neuroinflammation has been implicated in stimulating or exacerbating neuronal damage. The tumor necrosis factor (TNF) superfamily of cytokines are involved in many systemic chronic inflammatory and degenerative conditions and are amongst the key mediators of neuroinflammation. TNF binds to the TNFR1 and TNFR2 receptors to activate diverse cellular responses that can be either neuroprotective or neurodegenerative. In particular, TNF can induce programmed necrosis or necroptosis in an inflammatory environment. Although activation of necroptosis has recently been demonstrated in the AD brain, its significance in AD neuron loss and the role of TNF signaling is unclear. We demonstrate an increase in expression of multiple proteins in the TNF/TNF receptor-1-mediated necroptosis pathway in the AD post-mortem brain, as indicated by the phosphorylation of RIPK3 and MLKL, predominantly observed in the CA1 pyramidal neurons. The density of phosphoRIPK3 + and phosphoMLKL + neurons correlated inversely with total neuron density and showed significant sexual dimorphism within the AD cohort. In addition, apoptotic signaling was not significantly activated in the AD brain compared to the control brain. Exposure of human iPSC-derived glutamatergic neurons to TNF increased necroptotic cell death when apoptosis was inhibited, which was significantly reversed by small molecule inhibitors of RIPK1, RIPK3, and MLKL. In the post-mortem AD brain and in human iPSC neurons, in response to TNF, we show evidence of altered expression of proteins of the ESCRT III complex, which has been recently suggested as an antagonist of necroptosis and a possible mechanism by which cells can survive after necroptosis has been triggered. Taken together, our results suggest that neuronal loss in AD is due to TNF-mediated necroptosis rather than apoptosis, which is amenable to therapeutic intervention at several points in the signaling pathway.

A nexus of miR-1271, PAX4 and ALK/RYK influences the cytoskeletal architectures in Alzheimer's Disease and Type 2 Diabetes.

Alzheimer's Disease (AD) and Type 2 Diabetes (T2D) share a common hallmark of insulin resistance. Reportedly, two non-canonical Receptor Tyrosine Kinases (RTKs), ALK and RYK, both targets of the same micro RNA miR-1271, exhibit significant and consistent functional down-regulation in post-mortem AD and T2D tissues. Incidentally, both have Grb2 as a common downstream adapter and NOX4 as a common ROS producing factor. Here we show that Grb2 and NOX4 play critical roles in reducing the severity of both the diseases. The study demonstrates that the abundance of Grb2 in degenerative conditions, in conjunction with NOX4, reverse cytoskeletal degradation by counterbalancing the network of small GTPases. PAX4, a transcription factor for both Grb2 and NOX4, emerges as the key link between the common pathways of AD and T2D. Down-regulation of both ALK and RYK through miR-1271, elevates the PAX4 level by reducing its suppressor ARX via Wnt/β-Catenin signaling. For the first time, this study brings together RTKs beyond Insulin Receptor (IR) family, transcription factor PAX4 and both AD and T2D pathologies on a common regulatory platform.

Citrullination of Amyloid-β Peptides in Alzheimer's Disease.

Protein citrullination (deimination of arginine residue) is a well-known biomarker of inflammation. Elevated protein citrullination has been shown to colocalize with extracellular amyloid plaques in postmortem AD patient brains. Amyloid-β (Aβ) peptides which aggregate and accumulate in the plaques of Alzheimer's disease (AD) have sequential N-terminal truncations and multiple post-translational modifications (PTM) such as isomerization, pyroglutamate formation, phosphorylation, nitration, and dityrosine cross-linking. However, no conclusive biochemical evidence exists whether citrullinated Aβ is present in AD brains. In this study, using high-resolution mass spectrometry, we have identified citrullination of Aβ in sporadic and familial AD brains by characterizing the tandem mass spectra of endogenous N-truncated citrullinated Aβ peptides. Our quantitative estimations demonstrate that ∼ 35% of pyroglutamate3-Aβ pool was citrullinated in plaques in the sporadic AD temporal cortex and ∼ 22% in the detergent-insoluble frontal cortex fractions. Similarly, hypercitrullinated pyroglutamate3-Aβ (∼ 30%) was observed in both the detergent-soluble as well as insoluble Aβ pool in familial AD cases. Our results indicate that a common mechanism for citrullination of Aβ exists in both the sporadic and familial AD. We establish that citrullination of Aβ is a remarkably common PTM, closely associated with pyroglutamate3-Aβ formation and its accumulation in AD. This may have implications for Aβ toxicity, autoantigenicity of Aβ, and may be relevant for the design of diagnostic assays and therapeutic targeting.

LC–MS/MS assay for the investigation of acetylated Alpha-synuclein in serum from postmortem Alzheimer’s disease pathology

Alpha-synuclein (α-Syn), a neuronal protein, has been linked to the inflammation and development of neurodegenerative diseases. In a number of neurodegenerations, α-Syn has been investigated in the central nervous system and cerebrospinal fluid. However, there are few studies concerning the variations in peripheral α-Syn in postmortem Alzheimer’s disease (AD) pathology. In this study, the quantitative procedure for the determination of peripheral acetylated α-Syn regarding N-terminal amino acid’s site (α-Syn1-6; MDVFMK and Ac-α-Syn1-6; Ac-MDVFMK) was developed using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) and tryptic digestion without antibody. Serum samples were selected from postmortem specimens based on autopsy pathological examination of AD remark. The LC-MS/MS assay with ACQUITY UPLC BEH C18 column was applied on the basis of electrospray positive ionization. When subjected to N-terminal α-Syn peptides using MonoSpin Typsin HP preparation, doubly- and singly-charged α-Syn1-6 and Ac-α-Syn1-6 ions were observed at m/z 386 > 104 and m/z 813 > 72, respectively, which correspond to quantitative profiling with internal standards. In the calibration, the range of 10–1000 nmol/L showed r2 = 0.999 and recovery from 86.0% to 115.0% (RSD < 9.0%). Using this procedure, peripheral α-Syn1-6 from serum samples could not be detected. On the other hand, Ac-α-Syn1-6 levels were measured from 106.9 to 319.8 nmol/L (AD; n = 10) and 147.1–292.0 nmol/L (control; n = 10) with an insignificant difference. From these preliminary results, individual Ac-α-Syn levels in serum were inferred with nonspecific biomarker regarding to AD pathology.

Systematic review of human post-mortem immunohistochemical studies and bioinformatics analyses unveil the complexity of astrocyte reaction in Alzheimer's disease.

AimsReactive astrocytes in Alzheimer's disease (AD) have traditionally been demonstrated by increased glial fibrillary acidic protein (GFAP) immunoreactivity; however, astrocyte reaction is a complex and heterogeneous phenomenon involving multiple astrocyte functions beyond cytoskeletal remodelling. To better understand astrocyte reaction in AD, we conducted a systematic review of astrocyte immunohistochemical studies in post‐mortem AD brains followed by bioinformatics analyses on the extracted reactive astrocyte markers.MethodsNCBI PubMed, APA PsycInfo and WoS‐SCIE databases were interrogated for original English research articles with the search terms ‘Alzheimer's disease’ AND ‘astrocytes.’ Bioinformatics analyses included protein–protein interaction network analysis, pathway enrichment, and transcription factor enrichment, as well as comparison with public human ‐omics datasets.ResultsA total of 306 articles meeting eligibility criteria rendered 196 proteins, most of which were reported to be upregulated in AD vs control brains. Besides cytoskeletal remodelling (e.g., GFAP), bioinformatics analyses revealed a wide range of functional alterations including neuroinflammation (e.g., IL6, MAPK1/3/8 and TNF), oxidative stress and antioxidant defence (e.g., MT1A/2A, NFE2L2, NOS1/2/3, PRDX6 and SOD1/2), lipid metabolism (e.g., APOE, CLU and LRP1), proteostasis (e.g., cathepsins, CRYAB and HSPB1/2/6/8), extracellular matrix organisation (e.g., CD44, MMP1/3 and SERPINA3), and neurotransmission (e.g., CHRNA7, GABA, GLUL, GRM5, MAOB and SLC1A2), among others. CTCF and ESR1 emerged as potential transcription factors driving these changes. Comparison with published ‐omics datasets validated our results, demonstrating a significant overlap with reported transcriptomic and proteomic changes in AD brains and/or CSF.ConclusionsOur systematic review of the neuropathological literature reveals the complexity of AD reactive astrogliosis. We have shared these findings as an online resource available at www.astrocyteatlas.org.

Elevated Expression of MiR-17 in Microglia of Alzheimer's Disease Patients Abrogates Autophagy-Mediated Amyloid-β Degradation.

Autophagy is a proposed route of amyloid-β (Aβ) clearance by microglia that is halted in Alzheimer’s Disease (AD), though mechanisms underlying this dysfunction remain elusive. Here, primary microglia from adult AD (5xFAD) mice were utilized to demonstrate that 5xFAD microglia fail to degrade Aβ and express low levels of autophagy cargo receptor NBR1. In 5xFAD mouse brains, we show for the first time that AD microglia express elevated levels of microRNA cluster Mirc1/Mir17-92a, which is known to downregulate autophagy proteins. By in situ hybridization in post-mortem AD human tissue sections, we observed that the Mirc1/Mir17-92a cluster member miR-17 is also elevated in human AD microglia, specifically in the vicinity of Aβ deposits, compared to non-disease controls. We show that NBR1 expression is negatively correlated with expression of miR-17 in human AD microglia via immunohistopathologic staining in human AD brain tissue sections. We demonstrate in healthy microglia that autophagy cargo receptor NBR1 is required for Aβ degradation. Inhibiting elevated miR-17 in 5xFAD mouse microglia improves Aβ degradation, autophagy, and NBR1 puncta formation in vitro and improves NBR1 expression in vivo. These findings offer a mechanism behind dysfunctional autophagy in AD microglia which may be useful for therapeutic interventions aiming to improve autophagy function in AD.

Astrocyte reactivity with late-onset cognitive impairment assessed in vivo using 11C-BU99008 PET and its relationship with amyloid load

11C-BU99008 is a novel positron emission tomography (PET) tracer that enables selective imaging of astrocyte reactivity in vivo. To explore astrocyte reactivity associated with Alzheimer’s disease, 11 older, cognitively impaired (CI) subjects and 9 age-matched healthy controls (HC) underwent 3T magnetic resonance imaging (MRI), 18F-florbetaben and 11C-BU99008 PET. The 8 amyloid (Aβ)-positive CI subjects had higher 11C-BU99008 uptake relative to HC across the whole brain, but particularly in frontal, temporal, medial temporal and occipital lobes. Biological parametric mapping demonstrated a positive voxel-wise neuroanatomical correlation between 11C-BU99008 and 18F-florbetaben. Autoradiography using 3H-BU99008 with post-mortem Alzheimer’s brains confirmed through visual assessment that increased 3H-BU99008 binding localised with the astrocyte protein glial fibrillary acid protein and was not displaced by PiB or florbetaben. This proof-of-concept study provides direct evidence that 11C-BU99008 can measure in vivo astrocyte reactivity in people with late-life cognitive impairment and Alzheimer’s disease. Our results confirm that increased astrocyte reactivity is found particularly in cortical regions with high Aβ load. Future studies now can explore how clinical expression of disease varies with astrocyte reactivity.

ER stress associated TXNIP-NLRP3 inflammasome activation in hippocampus of human Alzheimer's disease

Although the exact etiology of Alzheimer's disease (AD) is poorly understood, experimental and clinical evidences suggest the contribution of neuroinflammation in the pathogenesis of AD. Pathologically, AD brain is characterized by an imbalance in redox status, elevated endoplasmic reticulum (ER) stress, synaptic dysfunction, inflammation, and progressive neurodegeneration. It has been noted that continuous accumulation of amyloid-beta (Aβ) and intracellular neurofibrillary tangles (NFTs) in AD brain trigger ER stress, which contributes to neurodegeneration. Similarly, experimental evidences supports the hypothesis that thioredoxin-interacting protein (TXNIP), an endogenous regulator of redox regulator thioredoxin (TRX), is activated by ER stress and contributes to activation of NLRP3 (NOD-like receptor protein 3) inflammatory cascade in hippocampus of the AD brain. Hippocampus of postmortem human AD and aged matched non-AD controls were analyzed for the expression ER stress markers and TXNIP-NLRP3 inflammasome at cellular and molecular levels. We found higher expression of TXNIP at protein and transcript levels in close association with pathological markers of AD such as Aβ and NFTs in AD hippocampus. In addition, our results demonstrated that TXNIP was co-localized in neurons and microglia. Moreover, expression of binding immunoglobulin protein (BiP), activated eukaryotic initiation factor-2α (eIf2α) and C/EBP homology protein (CHOP), proteins involved the development of ER stress, were elevated in AD hippocampus. Further, elevated expression of effector molecules of NLRP3 inflammasome activation such as apoptosis associated speck-like protein (ASC), cleaved caspase-1 and cleaved interleukin-1β were observed in the AD hippocampus. The study suggests that TXNIP could be a link that connect ER stress with neuroinflammation. Thus, TXNIP can be a possible therapeutic target to mitigate the progression of neuroinflammation in the pathogenesis of AD.

Enhancing myelin renewal reverses cognitive dysfunction in a murine model of Alzheimer’s disease

Severe cognitive decline is a hallmark of Alzheimer's disease (AD). In addition to gray matter loss, significant white matter pathology has been identified in AD patients. Here, we characterized the dynamics of myelin generation and loss in the APP/PS1 mouse model of AD. Unexpectedly, we observed a dramatic increase in the rate of new myelin formation in APP/PS1 mice, reminiscent of the robust oligodendroglial response to demyelination. Despite this increase, overall levels of myelination are decreased in the cortex and hippocampus of APP/PS1 mice and postmortem AD tissue. Genetically or pharmacologically enhancing myelin renewal, by oligodendroglial deletion of the muscarinic M1 receptor or systemic administration of the pro-myelinating drug clemastine, improved the performance of APP/PS1 mice in memory-related tasks and increased hippocampal sharp wave ripples. Taken together, these results demonstrate the potential of enhancing myelination as a therapeutic strategy to alleviate AD-related cognitive impairment.

ARCAM-1 Facilitates Fluorescence Detection of Amyloid-Containing Deposits in the Retina

PurposeTo investigate the use of an amyloid-targeting fluorescent probe, ARCAM-1, to identify amyloid-containing deposits in the retina of a transgenic mouse model of Alzheimer's disease (AD) and in human postmortem AD patients.MethodsAged APP/PS1 transgenic AD and wild-type (WT) mice were given an intraperitoneal (IP) injection of ARCAM-1 and their retinas imaged in vivo using a fluorescence ophthalmoscope. Eyes were enucleated and dissected for ex vivo inspection of retinal amyloid deposits. Additionally, formalin-fixed eyes from human AD and control patients were dissected, and the retinas were stained using ARCAM-1 or with an anti-amyloid-β antibody. Confocal microscopy was used to image amyloid-containing deposits stained with ARCAM-1 or with immunostaining.ResultsFour out of eight APP/PS1 mice showed the presence of amyloid aggregates in the retina during antemortem imaging. Retinas from three human AD patients stained with ARCAM-1 showed an apparent increased density of fluorescently labeled amyloid-containing deposits compared to the retinas from two healthy, cognitively normal (CN) patients. Immunolabeling confirmed the presence of amyloid deposits in both the retinal neuronal layers and in retinal vasculature.ConclusionsARCAM-1 facilitates antemortem detection of amyloid aggregates in the retina of a mouse model for AD, and postmortem detection of amyloid-containing deposits in human retinal tissues from AD patients. These results support the hypothesis of AD pathology manifesting in the eye and highlight a novel area for fluorophore development for the optical detection of retinal amyloid in AD patients.Translational RelevanceThis paper represents an initial examination for potential translation of an amyloid-targeting fluorescent probe to a retinal imaging agent for aiding in the diagnosis of Alzheimer's disease.

Replicative senescence dictates the emergence of disease-associated microglia and contributes to Aβ pathology.

SummaryThe sustained proliferation of microglia is a key hallmark of Alzheimer’s disease (AD), accelerating its progression. Here, we aim to understand the long-term impact of the early and prolonged microglial proliferation observed in AD, hypothesizing that extensive and repeated cycling would engender a distinct transcriptional and phenotypic trajectory. We show that the early and sustained microglial proliferation seen in an AD-like model promotes replicative senescence, characterized by increased βgal activity, a senescence-associated transcriptional signature, and telomere shortening, correlating with the appearance of disease-associated microglia (DAM) and senescent microglial profiles in human post-mortem AD cases. The prevention of early microglial proliferation hinders the development of senescence and DAM, impairing the accumulation of Aβ, as well as associated neuritic and synaptic damage. Overall, our results indicate that excessive microglial proliferation leads to the generation of senescent DAM, which contributes to early Aβ pathology in AD.

Introducing ADNP and SIRT1 as new partners regulating microtubules and histone methylation.

Activity-dependent neuroprotective protein (ADNP) is essential for brain formation and function. As such, de novo mutations in ADNP lead to the autistic ADNP syndrome and somatic ADNP mutations may drive Alzheimer's disease (AD) tauopathy. Sirtuin 1 (SIRT1) is positively associated with aging, the major risk for AD. Here, we revealed two key interaction sites for ADNP and SIRT1. One, at the microtubule end-binding protein (EB1 and EB3) Tau level, with EB1/EB3 serving as amplifiers for microtubule dynamics, synapse formation, axonal transport, and protection against tauopathy. Two, on the DNA/chromatin site, with yin yang 1, histone deacetylase 2, and ADNP, sharing a DNA binding motif and regulating SIRT1, ADNP, and EB1 (MAPRE1). This interaction was linked to sex- and age-dependent altered histone modification, associated with ADNP/SIRT1/WD repeat-containing protein 5, which mediates the assembly of histone modification complexes. Single-cell RNA and protein expression analyses as well as gene expression correlations placed SIRT1-ADNP and either MAPRE1 (EB1), MAPRE3 (EB3), or both in the same mouse and human cell; however, while MAPRE1 seemed to be similarly regulated to ADNP and SIRT1, MAPRE3 seemed to deviate. Finally, we demonstrated an extremely tight correlation for the gene transcripts described above, including related gene products. This correlation was specifically abolished in affected postmortem AD and Parkinson's disease brain select areas compared to matched controls, while being maintained in blood samples. Thus, we identified an ADNP-SIRT1 complex that may serve as a new target for the understanding of brain degeneration.

246 Maintenance of Circadian/Daily Activity Patterns and Cognitive Resilience to Alzheimer’s Pathology in Late Life

Abstract Introduction Many individuals remain free from dementia despite substantial plaque and tangle deposits, the hallmarks of Alzheimer’s disease (AD). Understanding of this cognitive resilience is poor. Evidence suggests that circadian disturbances predict increased risk of incident AD, and that circadian regulation deteriorates as clinical AD progresses. We hypothesize that circadian robustness protects against dementia, and the effect is stronger in individuals with AD pathology than those without. Methods We studied 575 deceased participants (age at death: 91.1□6.2; female: 414) in the Rush Memory and Aging Project who underwent brain autopsy at death, had clinical diagnostic opinion of dementia and motor activity assessments with actigraphy of ~10 days before death. Using actigraphy proximate to death, we calculated four circadian metrics: amplitude, acrophase, interdaily stability, and intradaily variability. Logistic regressions, stratified by postmortem pathologic AD diagnosis, were used to examine associations of circadian metrics with odds of dementia, and separately cognitive impairment (CI, including both dementia and mild cognitive impairment [MCI]), adjusting for age at death, sex, education, and time-lag between actigraphy and death. Results Based on postmortem assessment, 378 participants met the NIA-Reagan criteria for high/intermediate likelihood AD (AD group), including 197 clinically diagnosed with dementia, 86 MCI, and 85 cognitively intact. Non-AD group consisted of the remaining 197 participants, including 36 with clinical dementia, 47 MCI, and 114 cognitively intact. In the AD group, greater amplitude, greater interdaily stability, and lower intradaily variability were associated with lower odds of CI and dementia, i.e., odds ratios [OR] for CI corresponding to 1-SD changes were 0.54 (95% CI: 0.40-0.71), 0.70 (0.54-0.91), and 0.63 (0.47-0.84), and were 0.46 (0.35-0.60), 0.55 (0.43-0.70), and 0.61 (0.48-0.78) for dementia. In the non-AD group, only amplitude was associated with the odds of CI or dementia, i.e., the ORs corresponding to 1-SD increase was 0.61 (0.42-0.88) and 0.50 (0.31-0.82), respectively. Conclusion Better preserved circadian function, as characterized by more pronounced, more stable and less fragmented rest-activity rhythms, links to lower risk of CI or dementia in older people, especially those with pathological AD. Support (if any) NIH RF1AG064312, RF1AG059867, R01AG017917, R01AG56352; and the BrightFocus Foundation A2020886S.

Bioenergetic and inflammatory systemic phenotypes in Alzheimer's disease APOE ε4-carriers.

We examined the impact of an APOE ε4 genotype on Alzheimer's disease (AD) subject platelet and lymphocyte metabolism. Mean platelet mitochondrial cytochrome oxidase Vmax activity was lower in APOE ε4 carriers and lymphocyte Annexin V, a marker of apoptosis, was significantly higher. Proteins that mediate mitophagy and energy sensing were higher in APOE ε4 lymphocytes which could represent compensatory changes and recapitulate phenomena observed in post‐mortem AD brains. Analysis of the lipid synthesis pathway found higher AceCSI, ATP CL, and phosphorylated ACC levels in APOE ε4 lymphocytes. Lymphocyte ACC changes were also observed in post‐mortem brain tissue. Lymphocyte RNAseq showed lower APOE ε4 carrier sphingolipid Transporter 3 (SPNS3) and integrin Subunit Alpha 1 (ITGA1) expression. RNAseq pathway analysis revealed APOE ε4 alleles activated inflammatory pathways and modulated bioenergetic signaling. These findings support a relationship between APOE genotype and bioenergetic pathways and indicate platelets and lymphocytes from APOE ε4 carriers exist in a state of bioenergetic stress. Neither medication use nor brain‐localized AD histopathology can account for these findings, which define an APOE ε4‐determined molecular and systemic phenotype that informs AD etiology.

Alzheimer's Disease Alters Oligodendrocytic Glycolytic and Ketolytic Gene Expression

Sporadic Alzheimer's disease (AD) is the most common form of senescence-related dementia, and is clinically characterized by neurodegeneration that progressively impairs memory and behavior. It is widely accepted that AD is strongly correlated with impaired brain glucose metabolism, which may contribute to disease onset and progression. Ketolysis has been suggested as an alternative pathway to fuel the brain. In the present study, RNA-seq profiles of postmortem AD brains were used to determine whether dysfunctional AD brain metabolism can be determined by impairments in glycolytic and ketolytic gene expression. Data were obtained from the Knight Alzheimer's Disease Research Center (62 cases; 13 controls), Mount Sinai Brain Bank (110 cases; 44 controls), and the Mayo Clinic Brain Bank (80 cases; 76 controls), and were normalized to cell type: astrocytes, microglia, neurons, oligodendrocytes. In oligodendrocytes, both glycolytic and ketolytic pathways were significantly impaired in regions of the brain relevant to AD. However, ketolytic gene expression was not significantly altered in neurons, astrocytes, and microglia. These results indicate that oligodendrocytes may contribute to brain hypometabolism and AD pathology more than previously thought. These findings are also suggestive of a potential link between AD hypometabolism and dysmyelination, and suggest that ketones may be therapeutic in AD due to their ability to fuel neurons despite impairments in glycolytic metabolism.

Pro-inflammatory interleukin-6 signaling links cognitive impairments and peripheral metabolic alterations in Alzheimer\u2019s disease

Alzheimer’s disease (AD) is associated with memory impairment and altered peripheral metabolism. Mounting evidence indicates that abnormal signaling in a brain-periphery metabolic axis plays a role in AD pathophysiology. The activation of pro-inflammatory pathways in the brain, including the interleukin-6 (IL-6) pathway, comprises a potential point of convergence between memory dysfunction and metabolic alterations in AD that remains to be better explored. Using T2-weighted magnetic resonance imaging (MRI), we observed signs of probable inflammation in the hypothalamus and in the hippocampus of AD patients when compared to cognitively healthy control subjects. Pathological examination of post-mortem AD hypothalamus revealed the presence of hyperphosphorylated tau and tangle-like structures, as well as parenchymal and vascular amyloid deposits surrounded by astrocytes. T2 hyperintensities on MRI positively correlated with plasma IL-6, and both correlated inversely with cognitive performance and hypothalamic/hippocampal volumes in AD patients. Increased IL-6 and suppressor of cytokine signaling 3 (SOCS3) were observed in post-mortem AD brains. Moreover, activation of the IL-6 pathway was observed in the hypothalamus and hippocampus of AD mice. Neutralization of IL-6 and inhibition of the signal transducer and activator of transcription 3 (STAT3) signaling in the brains of AD mouse models alleviated memory impairment and peripheral glucose intolerance, and normalized plasma IL-6 levels. Collectively, these results point to IL-6 as a link between cognitive impairment and peripheral metabolic alterations in AD. Targeting pro-inflammatory IL-6 signaling may be a strategy to alleviate memory impairment and metabolic alterations in the disease.