Pathophysiology Of AD Research Articles

RTP801/REDD1 contributes to neuroinflammation severity and memory impairments in Alzheimer\u2019s disease

RTP801/REDD1 is a stress-regulated protein whose upregulation is necessary and sufficient to trigger neuronal death. Its downregulation in Parkinson’s and Huntington’s disease models ameliorates the pathological phenotypes. In the context of Alzheimer’s disease (AD), the coding gene for RTP801, DDIT4, is responsive to Aβ and modulates its cytotoxicity in vitro. Also, RTP801 mRNA levels are increased in AD patients’ lymphocytes. However, the involvement of RTP801 in the pathophysiology of AD has not been yet tested. Here, we demonstrate that RTP801 levels are increased in postmortem hippocampal samples from AD patients. Interestingly, RTP801 protein levels correlated with both Braak and Thal stages of the disease and with GFAP expression. RTP801 levels are also upregulated in hippocampal synaptosomal fractions obtained from murine 5xFAD and rTg4510 mice models of the disease. A local RTP801 knockdown in the 5xFAD hippocampal neurons with shRNA-containing AAV particles ameliorates cognitive deficits in 7-month-old animals. Upon RTP801 silencing in the 5xFAD mice, no major changes were detected in hippocampal synaptic markers or spine density. Importantly, we found an unanticipated recovery of several gliosis hallmarks and inflammasome key proteins upon neuronal RTP801 downregulation in the 5xFAD mice. Altogether our results suggest that RTP801 could be a potential future target for theranostic studies since it could be a biomarker of neuroinflammation and neurotoxicity severity of the disease and, at the same time, a promising therapeutic target in the treatment of AD.

Dynamic Amyloid PET: Relationships to 18F-Flortaucipir Tau PET Measures.

Measuring amyloid and predicting tau status using a single amyloid PET study would be valuable for assessing brain AD pathophysiology. We hypothesized that early-frame amyloid PET (efAP) correlates with the presence of tau pathology because the initial regional brain concentrations of radioactivity are determined primarily by blood flow, which is expected to be decreased in the setting of tau pathology. Methods: The study included 120 participants (63 amyloid-positive and 57 amyloid-negative) with dynamic 18F-florbetapir PET and static 18F-flortaucipir PET scans obtained within 6 mo of each other. These subjects were predominantly cognitively intact in both the amyloid-positive (63%) and the amyloid-negative (93%) groups. Parameters for efAP quantification were optimized for stratification of tau PET positivity, assessed by either a tauopathy score or Braak regions. The ability of efAP to stratify tau positivity was measured using receiver-operating-characteristic analysis of area under the curve (AUC). Pearson r and Spearman ρ were used for parametric and nonparametric comparisons between efAP and tau PET, respectively. Standardized net benefit was used to evaluate improvement in using efAP as an additional copredictor over hippocampal volume in predicting tau PET positivity. Results: Measuring efAP within the hippocampus and summing the first 3 min of brain activity after injection showed the strongest discriminative ability to stratify for tau positivity (AUC, 0.67-0.89 across tau PET Braak regions) in amyloid-positive individuals. Hippocampal efAP correlated significantly with a global tau PET tauopathy score in amyloid-positive participants (r = -0.57, P < 0.0001). Compared with hippocampal volume, hippocampal efAP showed a stronger association with tau PET Braak stage (ρ = -0.58 vs. -0.37) and superior stratification of tau PET tauopathy score (AUC, 0.86 vs. 0.66; P = 0.002). Conclusion: Hippocampal efAP can provide additional information to conventional amyloid PET, including estimation of the likelihood of tau positivity in amyloid-positive individuals.

Alzheimer's disease and gut microbiota: does trimethylamine N-oxide (TMAO) play a role?

Alzheimer's disease (AD) is a neurodegenerative disease that affects memory and cognitive function. Clinical evidence has put into question our current understanding of AD development, propelling researchers to look into further avenues. Gut microbiota has emerged as a potential player in AD pathophysiology. Lifestyle factors, such as diet, can have negative effects on the gut microbiota and thus host health. A Western-type diet has been highlighted as a risk factor for both gut microbiota alteration as well as AD development. The gut-derived trimethylamine N-oxide (TMAO) has been previously implied in the development of cardiovascular diseases with recent evidence suggesting a plausible role of TMAO in AD development. Therefore, the main goal of the present review is to provide the reader with potential mechanisms of action through which consumption of a Western-type diet could increase AD risk, by acting through microbiota-produced TMAO. Although a link between TMAO and AD is far from definitive, this review will serve as a call for research into this new area of research.

Amyloid precursor protein elevates fusion of promyelocytic leukemia nuclear bodies in human hippocampal areas with high plaque load

The amyloid precursor protein (APP) is a type I transmembrane protein with unknown physiological function but potential impact in neurodegeneration. The current study demonstrates that APP signals to the nucleus causing the generation of aggregates consisting of its adapter protein FE65, the histone acetyltransferase TIP60 and the tumour suppressor proteins p53 and PML. APP C-terminal (APP-CT50) complexes co-localize and co-precipitate with p53 and PML. The PML nuclear body generation is induced and fusion occurs over time depending on APP signalling and STED imaging revealed active gene expression within the complex. We further show that the nuclear aggregates of APP-CT50 fragments together with PML and FE65 are present in the aged human brain but not in cerebral organoids differentiated from iPS cells. Notably, human Alzheimer’s disease brains reveal a highly significant reduction of these nuclear aggregates in areas with high plaque load compared to plaque-free areas of the same individual. Based on these results we conclude that APP-CT50 signalling to the nucleus takes place in the aged human brain and is involved in the pathophysiology of AD.

Ferulic Acid Ameliorates Alzheimer\u2019s Disease-like Pathology and Repairs Cognitive Decline by Preventing Capillary Hypofunction in APP/PS1 Mice

Brain capillaries are crucial for cognitive functions by supplying oxygen and other nutrients to and removing metabolic wastes from the brain. Recent studies have demonstrated that constriction of brain capillaries is triggered by beta-amyloid (Aβ) oligomers via endothelin-1 (ET1)-mediated action on the ET1 receptor A (ETRA), potentially exacerbating Aβ plaque deposition, the primary pathophysiology of Alzheimer’s disease (AD). However, direct evidence is still lacking whether changes in brain capillaries are causally involved in the pathophysiology of AD. Using APP/PS1 mouse model of AD (AD mice) relative to age-matched negative littermates, we identified that reductions of density and diameter of hippocampal capillaries occurred from 4 to 7 months old while Aβ plaque deposition and spatial memory deficit developed at 7 months old. Notably, the injection of ET1 into the hippocampus induced early Aβ plaque deposition at 5 months old in AD mice. Conversely, treatment of ferulic acid against the ETRA to counteract the ET1-mediated vasoconstriction for 30 days prevented reductions of density and diameter of hippocampal capillaries as well as ameliorated Aβ plaque deposition and spatial memory deficit at 7 months old in AD mice. Thus, these data suggest that reductions of density and diameter of hippocampal capillaries are crucial for initiating Aβ plaque deposition and spatial memory deficit at the early stages, implicating the development of new therapies for halting or curing memory decline in AD.

Development of a novel, sensitive translational immunoassay to detect plasma glial fibrillary acidic protein (GFAP) after murine traumatic brain injury

BackgroundGlial fibrillary acidic protein (GFAP) has emerged as a promising fluid biomarker for several neurological indications including traumatic brain injury (TBI), a leading cause of death and disability worldwide. In humans, serum or plasma GFAP levels can predict brain abnormalities including hemorrhage on computed tomography (CT) scans and magnetic resonance imaging (MRI). However, assays to quantify plasma or serum GFAP in preclinical models are not yet available.MethodsWe developed and validated a novel sensitive GFAP immunoassay assay for mouse plasma on the Meso Scale Discovery immunoassay platform and validated assay performance for robustness, precision, limits of quantification, dilutional linearity, parallelism, recovery, stability, selectivity, and pre-analytical factors. To provide proof-of-concept data for this assay as a translational research tool for TBI and Alzheimer’s disease (AD), plasma GFAP was measured in mice exposed to TBI using the Closed Head Impact Model of Engineered Rotational Acceleration (CHIMERA) model and in APP/PS1 mice with normal or reduced levels of plasma high-density lipoprotein (HDL).ResultsWe performed a partial validation of our novel assay and found its performance by the parameters studied was similar to assays used to quantify human GFAP in clinical neurotrauma blood specimens and to assays used to measure murine GFAP in tissues. Specifically, we demonstrated an intra-assay CV of 5.0%, an inter-assay CV of 7.2%, a lower limit of detection (LLOD) of 9.0 pg/mL, a lower limit of quantification (LLOQ) of 24.8 pg/mL, an upper limit of quantification (ULOQ) of at least 16,533.9 pg/mL, dilution linearity of calibrators from 20 to 200,000 pg/mL with 90–123% recovery, dilution linearity of plasma specimens up to 32-fold with 96–112% recovery, spike recovery of 67–100%, and excellent analyte stability in specimens exposed to up to 7 freeze-thaw cycles, 168 h at 4 °C, 24 h at room temperature (RT), or 30 days at − 20 °C. We also observed elevated plasma GFAP in mice 6 h after TBI and in aged APP/PS1 mice with plasma HDL deficiency. This assay also detects GFAP in serum.ConclusionsThis novel assay is a valuable translational tool that may help to provide insights into the mechanistic pathophysiology of TBI and AD.

Association of CSF proteins with tau and amyloid \u03b2 levels in asymptomatic 70-year-olds

BackgroundIncreased knowledge of the evolution of molecular changes in neurodegenerative disorders such as Alzheimer’s disease (AD) is important for the understanding of disease pathophysiology and also crucial to be able to identify and validate disease biomarkers. While several biological changes that occur early in the disease development have already been recognized, the need for further characterization of the pathophysiological mechanisms behind AD still remains.MethodsIn this study, we investigated cerebrospinal fluid (CSF) levels of 104 proteins in 307 asymptomatic 70-year-olds from the H70 Gothenburg Birth Cohort Studies using a multiplexed antibody- and bead-based technology.ResultsThe protein levels were first correlated with the core AD CSF biomarker concentrations of total tau, phospho-tau and amyloid beta (Aβ42) in all individuals. Sixty-three proteins showed significant correlations to either total tau, phospho-tau or Aβ42. Thereafter, individuals were divided based on CSF Aβ42/Aβ40 ratio and Clinical Dementia Rating (CDR) score to determine if early changes in pathology and cognition had an effect on the correlations. We compared the associations of the analysed proteins with CSF markers between groups and found 33 proteins displaying significantly different associations for amyloid-positive individuals and amyloid-negative individuals, as defined by the CSF Aβ42/Aβ40 ratio. No differences in the associations could be seen for individuals divided by CDR score.ConclusionsWe identified a series of transmembrane proteins, proteins associated with or anchored to the plasma membrane, and proteins involved in or connected to synaptic vesicle transport to be associated with CSF biomarkers of amyloid and tau pathology in AD. Further studies are needed to explore these proteins’ role in AD pathophysiology.

AlzGPS: a genome-wide positioning systems platform to catalyze multi-omics for Alzheimer\u2019s drug discovery

BackgroundRecent DNA/RNA sequencing and other multi-omics technologies have advanced the understanding of the biology and pathophysiology of AD, yet there is still a lack of disease-modifying treatments for AD. A new approach to integration of the genome, transcriptome, proteome, and human interactome in the drug discovery and development process is essential for this endeavor.MethodsIn this study, we developed AlzGPS (Genome-wide Positioning Systems platform for Alzheimer’s Drug Discovery, https://alzgps.lerner.ccf.org), a comprehensive systems biology tool to enable searching, visualizing, and analyzing multi-omics, various types of heterogeneous biological networks, and clinical databases for target identification and development of effective prevention and treatment for AD.ResultsVia AlzGPS: (1) we curated more than 100 AD multi-omics data sets capturing DNA, RNA, protein, and small molecule profiles underlying AD pathogenesis (e.g., early vs. late stage and tau or amyloid endophenotype); (2) we constructed endophenotype disease modules by incorporating multi-omics findings and human protein-protein interactome networks; (3) we provided possible treatment information from ~ 3000 FDA approved/investigational drugs for AD using state-of-the-art network proximity analyses; (4) we curated nearly 300 literature references for high-confidence drug candidates; (5) we included information from over 1000 AD clinical trials noting drug’s mechanisms-of-action and primary drug targets, and linking them to our integrated multi-omics view for targets and network analysis results for the drugs; (6) we implemented a highly interactive web interface for database browsing and network visualization.ConclusionsNetwork visualization enabled by AlzGPS includes brain-specific neighborhood networks for genes-of-interest, endophenotype disease module networks for omics-of-interest, and mechanism-of-action networks for drugs targeting disease modules. By virtue of combining systems pharmacology and network-based integrative analysis of multi-omics data, AlzGPS offers actionable systems biology tools for accelerating therapeutic development in AD.

Low Cerebrospinal Fluid Levels of Hemopexin Are Associated With Increased Alzheimer's Pathology, Hippocampal Hypometabolism, and Cognitive Decline.

Brain iron dyshomeostasis is a feature of Alzheimer's disease. Conventionally, research has focused on non-heme iron although degradation of heme from hemoglobin subunits can generate iron to augment the redox-active iron pool. Hemopexin both detoxifies heme to maintain iron homeostasis and bolsters antioxidant capacity via catabolic products, biliverdin and carbon monoxide to combat iron-mediated lipid peroxidation. The aim of the present study was to examine the association of cerebrospinal fluid levels (CSF) hemopexin and hemoglobin subunits (α and β) to Alzheimer's pathological proteins (amyloid and tau), hippocampal volume and metabolism, and cognitive performance. We analyzed baseline CSF heme/iron proteins (multiplexed mass spectrometry-based assay), amyloid and tau (Luminex platform), baseline/longitudinal neuroimaging (MRI, FDG-PET) and cognitive outcomes in 86 cognitively normal, 135 mild-cognitive impairment and 66 Alzheimer's participants from the Alzheimer's Disease Neuroimaging Initiative-1 (ADNI-1) cohort. Multivariate regression analysis was performed to delineate differences in CSF proteins between diagnosis groups and evaluated their association to amyloid and tau, neuroimaging and cognition. A p-value ≤ 0.05 was considered significant. Higher hemopexin was associated with higher CSF amyloid (implying decreased brain amyloid deposition), improved hippocampal metabolism and cognitive performance. Meanwhile, hemoglobin subunits were associated with increased CSF tau (implying increased brain tau deposition). When dichotomizing individuals with mild-cognitive impairment into stable and converters to Alzheimer's disease, significantly higher baseline hemoglobin subunits were observed in the converters compared to non-converters. Heme/iron dyshomeostasis is an early and crucial event in AD pathophysiology, which warrants further investigation as a potential therapeutic target.

Omega6/omega3 ethanolamin‐containing plasmalogen (PlsEtn) ratio as a biomarker of Alzheimer's disease progression

AbstractBackgroundSingle‐cell transcriptomic analysis of AD has revealed that myelination‐related processes are perturbed in brain, suggesting a key role in AD pathophysiology. Oligodendrocytes support neurons and signal transmission in the central nervous system by wrapping axons with myelin, a lipid‐rich membrane structure that depend on fatty acid synthesis and metabolism. Beside galactolipids, the major lipids of myelin are cholesterol and ethanolamin‐containing plasmalogens (PlsEtn). Polyunsaturated fatty acids (PUFA) such as docosahexaenoic acid (DHA, 22:6, omega3), eicosapentaenoic acid (EPA, 20:5, omega3), arachidonic acid (20:4, omega6), and monounsaturated fatty acid oleic acid (18:1, omega9) are the major constituent FA at the sn‐2 position of Pls. Among them, DHA is the most efficient to confer very specific physicochemical properties and membrane fluidity (Pinot et al., 2014).MethodOur results in lipidomics reveal that brain PUFAs‐Pls levels decrease importantly in cortex T1 and particularly in the frontal white matter (FWM) of AD individuals.ResultWe hypothesized that blood and brain phospholipids deficits could be closely associated with disease progression in AD. This hypothesis is corroborated by our previous results showing statistical association between PUFA‐phospholipids (PC, PE, PS) blood concentrations and brain atrophy and cognition in the ADNI cohort (previous communication at AAIC 2018). In follow‐up analyses, we further demonstrated that blood PUFAs‐PlsEtn deficits specifically related to demyelination present the most significative statistical association between an increase of the ratio omega6/omega3 and the increase of AD severity (high disease stage).ConclusionBlood omega6/omega3 PUFAs‐PlsEtn ratio could be a biomarker of the demyelination process observed in AD progression.

New evidence reveals weakening of association between language and thought in aMCI versus in normal aging

AbstractBackgroundClinical manifestation of prodromal AD is typically evaluated through measures that assess individual cognitive domains with less focus on their relationship. Further, while decades of research identify deterioration in episodic memory as well as semantic aspects of language, there has been less focus on other aspects of linguistic computation. We provide evidence that advances our understanding of how language deficits in aMCI extend to sentence production, and how relations between performances in cognitive functions, specifically language and cognition, may differ in aMCI vs. Healthy Aging (HA).MethodWe tested 24 individuals diagnosed with aMCI based on neurologic and neuropsychological evaluations, and 22 older adults with normal cognition (Healthy Aging (HA). All subjects were administered a multi‐domain cognitive screen (ACE‐R) and a psycholinguistic assessment of complex sentence production. The ACE‐R provided a Total Score and scores on attention and orientation, memory, fluency, language, and visuospatial measures (Mioshi et al 2006, Hsieh et al 2013, Monsch et al 1994).) The psycholinguistic test assessed Elicited Imitation of complex sentences with relative clauses (RC) in an experimental design varying syntactic and semantic factors (Table 1) (Lust et al 2017).ResultACE‐R Total score was significantly lower for aMCI than HA. aMCI also performed significantly worse than HA on the psycholinguistic test (Table 2). For HA, not aMCI, ACE‐R Total score significantly predicted successful sentence production on the psycholinguistic test (Table 3) (Figure 1). Similarly, for HA, not aMCI, the ACE‐R language score significantly predicted psycholinguistic test performance. In contrast, ACE‐R memory performance, which was significantly lower for aMCI than HA, did not predict linguistic performance for either group. Only ACE‐R phonemic fluency predicted linguistic performance for both groups. (Table 3).ConclusionResults suggest a weakening of associations between overall cognitive performance and performance on a linguistic measure for aMCI relative to HA suggesting that evaluation of associations between, not only within, cognitive domains strengthen detection of early changes in AD. This finding may be further expanded by linking this clinical manifestation to neuroscientific evidence of functional connectivity in AD pathophysiology and in normal aging (e.g., Brier et al 2014, Spreng and Turner 2013).

Targeting isoaspartate-modified A\u03b2 rescues behavioral deficits in transgenic mice with Alzheimer\u2019s disease-like pathology

BackgroundAmyloid β (Aβ)-directed immunotherapy has shown promising results in preclinical and early clinical Alzheimer’s disease (AD) trials, but successful translation to late clinics has failed so far. Compelling evidence suggests that post-translationally modified Aβ peptides might play a decisive role in onset and progression of AD and first clinical trials targeting such Aβ variants have been initiated. Modified Aβ represents a small fraction of deposited material in plaques compared to pan-Aβ epitopes, opening up pathways for tailored approaches of immunotherapy. Here, we generated the first monoclonal antibodies that recognize l-isoaspartate-modified Aβ (isoD7-Aβ) and tested a lead antibody molecule in 5xFAD mice.MethodsThis work comprises a combination of chemical and biochemical techniques as well as behavioral analyses. Aβ peptides, containing l-isoaspartate at position 7, were chemically synthesized and used for immunization of mice and antibody screening methods. Biochemical methods included anti-isoD7-Aβ monoclonal antibody characterization by surface plasmon resonance, immunohistochemical staining of human and transgenic mouse brain, and the development and application of isoD7-Aβ ELISA as well as different non-modified Aβ ELISA. For antibody treatment studies, 12 mg/kg anti-isoD7-Aβ antibody K11_IgG2a was applied intraperitoneally to 5xFAD mice for 38 weeks. Treatment controls implemented were IgG2a isotype as negative and 3D6_IgG2a, the parent molecule of bapineuzumab, as positive control antibodies. Behavioral studies included elevated plus maze, pole test, and Morris water maze.ResultsOur advanced antibody K11 showed a KD in the low nM range and > 400fold selectivity for isoD7-Aβ compared to other Aβ variants. By using this antibody, we demonstrated that formation of isoD7-Aβ may occur after formation of aggregates; hence, the presence of the isoD7-modification differentiates aged Aβ from newly formed peptides. Importantly, we also show that the Tottori mutation responsible for early-onset AD in a Japanese pedigree is characterized by massively accelerated formation of isoD7-Aβ in cell culture. The presence of isoD7-Aβ was verified by K11 in post mortem human cortex and 5xFAD mouse brain tissue. Passive immunization of 5xFAD mice resulted in a significant reduction of isoD7-Aβ and total Aβ in brain. Amelioration of cognitive impairment was demonstrated by Morris water maze, elevated plus maze, pole, and contextual fear conditioning tests. Interestingly, despite the lower abundance of the isoD7-Aβ epitope, the application of anti-isoD7-Aβ antibodies showed comparable treatment efficacy in terms of reduction of brain amyloid and spatial learning but did not result in an increase of plasma Aβ concentration as observed with 3D6 treatment.ConclusionsThe present study demonstrates, for the first time, that the antibody-mediated targeting of isoD7-modified Aβ peptides leads to attenuation of AD-like amyloid pathology. In conjunction with previously published data on antibodies directed against pGlu-modified Aβ, the results highlight the crucial role of modified Aβ peptides in AD pathophysiology. Hence, the results also underscore the therapeutic potential of targeting modified amyloid species for defining tailored approaches in AD therapy.

Double hit viral parasitism, polymicrobial CNS residency and perturbed proteostasis in Alzheimer’s disease: A data driven, in silico analysis of gene expression data

The aim of this study was to determine the interaction of peripheral immunity vs. the CNS in the setting of AD pathogenesis at the transcriptomic level in a data driven manner. For this purpose, publicly available gene expression data from the GEO Datasets repository. We performed differential gene expression and functional enrichment analyses were performed on the five retrieved studies: (a) three hippocampal cortex (HC) studies (b) one study of peripheral blood mononuclear cells (PBMC) and (c) one involving neurofibrillary tangle – containing neurons of the entorhinal cortex (NFT EC). Subsequently, BLAST was used to determine protein conservation between human proteins vs. microbial, whereas putative protein / oligopeptide antigenicity were determined via RANKPep. Gene ontology and pathway analyses revealed significantly enriched viral parasitism pathways in both PBMC and NFT – EC datasets, mediated by ribosomal protein families and epigenetic regulators. Among these, a salient viral pathway referred to Influenza A infection. NFT – EC annotations included leukocyte chemotaxis and immune response pathways. All datasets were significantly enriched for infectious pathways, as well as pathways involved in impaired proteostasis and non – phagocytic cell phagosomal cascades. In conclusion, our in silico analysis outlined an ad hoc model of AD pathophysiology in which double hit (PBMC and NFT-EC) viral parasitism is mediated by eukaryotic translational hijacking, and may be further implicated by impaired immune responses. Overall, our results overlap with the antimicrobial protection hypothesis of AD pathogenesis and support the notion of a pathogen – driven etiology.

The Neuropeptide Kyotorphin as a Possible Biomarker and Neuroprotective Agent in Alzheimer’s Disease

Summary Alzheimer’s disease (AD) is an age-related neurodegenerative disorder clinically characterized by memory impairment, disorientation, cognitive deficits, and behavioral disturbances. The neuropathological features are amyloid plaques containing aggregated amyloid-beta (Aβ) peptide, neurofibrillary tangles composed of the hyperphosphorylated form of the microtubule protein tau (HP-tau), and loss of neurons and synapses in the brain. There are no effective strategies for the prevention or treatment of the disease, leading to an increased need for AD biomarkers to improve early detection, accurate diagnosis, and accelerate drug development in this field. Recently, increasing attention has been dedicated to neuropeptides in searching for new drug targets in the treatment of nervous system disorders. Available data suggest that many neuropeptides may be associated with the pathophysiology and potential therapy of AD because of their wide distribution in brain areas responsible for learning and memory processes and their predominately neuroprotective actions. This short review aimed to briefly describe the neuropathology of AD and summarize the data related to one of its recently proposed biomarker - kyotorphin (KTP) neuropeptide. Our previous experiments showed moderate and selective protective effects of KTP against the late consequences of the intracerebroventricular streptozotocin-induced AD model.

CSF levels of the BACE1 substrate NRG1 correlate with cognition in Alzheimer\u2019s disease

BackgroundThe presynaptic protein neuregulin1 (NRG1) is cleaved by beta-site APP cleaving enzyme 1 (BACE1) in a similar way as amyloid precursor protein (APP) NRG1 can activate post-synaptic receptor tyrosine-protein kinase erbB4 (ErbB4) and was linked to schizophrenia. The NRG1/ErbB4 complex is neuroprotective, can trigger synaptogenesis and plasticity, increases the expression of NMDA and GABA receptors, and can induce neuroinflammation. This complex can reduce memory formation. In Alzheimer’s disease (AD) brains, NRG1 accumulates in neuritic plaques. It is difficult to determine if NRG1 has beneficial and/or detrimental effects in AD. BACE1 levels are increased in AD brains and cerebrospinal fluid (CSF) and may lead to enhanced NRG1 secretion, but no study has assessed CSF NRG1 levels in AD and mild cognitive impairment (MCI) patients.MethodsThis retrospective study included 162 patients suffering from AD dementia (54), MCI with progression to AD dementia (MCI-AD) (27), non-AD MCI (30), non-AD dementias (30), and neurological controls (27). All patients had neurological examinations, brain MRI, and neuropsychological evaluations. After written informed consent and using enzyme-linked immunosorbent assays (ELISAs), CSF samples were evaluated for Aβ1–42, Aβ1–40, total tau (T-tau), phosphorylated tau on threonine 181 (P-tau), BACE1, growth-associated protein 43 (GAP 43), neurogranin (Ng), and NRG1.ResultsLevels of NRG1 were significantly increased in the CSF of AD (+ 36%) and MCI-AD (+ 28%) patients compared to neurological controls and also non-AD MCI and non-AD dementias. In addition, in AD and MCI-AD patients, NRG1 levels positively correlated with Aβ1–42 but not with T-tau, P-tau, and BACE1 levels and negatively correlated with MMSE scores. A longitudinal follow-up study of AD patients revealed a trend (p = 0.08) between CSF NRG1 levels and cognitive decline. In the overall population, NRG1 correlated with MMSE and the synaptic biomarkers GAP 43 and neurogranin.ConclusionsOur results showed that CSF NRG1 levels are increased in AD and MCI-AD as compared to controls and other dementias. CSF NRG1 levels are associated with cognitive evolution, and a major outcome of our findings is that synaptic NRG1 could be involved in the pathophysiology of AD. Modulating brain NRG1 activity may represent a new therapeutic target in AD.

CSF total tau/\u03b1-synuclein ratio improved the diagnostic performance for Alzheimer\u2019s disease as an indicator of tau phosphorylation

BackgroundRecently, several studies suggested potential involvements of α-synuclein in Alzheimer’s disease (AD) pathophysiology. Higher concentrations of α-synuclein were reported in cerebrospinal fluid (CSF) of AD patients with a positive correlation towards CSF tau, indicating its possible role in AD. We analyzed the CSF biomarkers to verify whether α-synuclein could be an additional supported biomarker in AD diagnosis.MethodsIn this cross-sectional study, CSF samples of 71 early-onset AD, 34 late-onset AD, 11 mild cognitive impairment, 17 subjective cognitive decline, 45 Parkinson’s disease, and 32 healthy control (HC) were collected. CSF amyloid-β1-42 (A), total tau (N), and phosphorylated tau181 (T) were measured by commercial ELISA kits, and in-house ELISA kit was developed to quantify α-synuclein. The cognitive assessments and amyloid-PET imaging were also performed.ResultsCSF α-synuclein manifested a tendency to increase in AD and to decreased in Parkinson’s disease compared to HC. The equilibrium states of total tau and α-synuclein concentrations were changed significantly in AD, and the ratio of total tau/α-synuclein (N/αS) was dramatically increased in AD than HC. Remarkably, N/αS revealed a strong positive correlation with tau phosphorylation rate. Also, the combination of N/αS with amyloid-β1-42/phosphorylated tau181 ratio had the best diagnosis performance (AUC = 0.956, sensitivity = 96%, specificity = 87%). In concordance analysis, N/αS showed the higher diagnostic agreement with amyloid-β1-42 and amyloid-PET. Analysis of biomarker profiling with N/αS had distinctive characteristics and clustering of each group. Especially, among the group of suspected non-Alzheimer’s disease pathophysiology, all A−T+N+ patients with N/αS+ were reintegrated into AD.ConclusionsThe high correlation of α-synuclein with tau and the elevated N/αS in AD supported the involvement of α-synuclein in AD pathophysiology. Importantly, N/αS improved the diagnostic performance, confirming the needs of incorporating α-synuclein as a biomarker for neurodegenerative disorders. The incorporation of a biomarker group [N/αS] could contribute to provide better understanding and diagnosis of neurodegenerative disorders.

25-Hydroxycholesterol amplifies microglial IL-1\u03b2 production in an apoE isoform-dependent manner

BackgroundGenome-wide association studies of Alzheimer’s disease (AD) have implicated pathways related to lipid homeostasis and innate immunity in AD pathophysiology. However, the exact cellular and chemical mediators of neuroinflammation in AD remain poorly understood. The oxysterol 25-hydroxycholesterol (25-HC) is an important immunomodulator produced by peripheral macrophages with wide-ranging effects on cell signaling and innate immunity. Cholesterol 25-hydroxylase (CH25H), the enzyme responsible for 25-HC production, has also been found to be one of the disease-associated microglial (DAM) genes that are upregulated in the brain of AD and AD transgenic mouse models.MethodsWe used real-time PCR and immunoblotting to examine CH25H expression in human AD brain tissue and in transgenic mouse brain tissue-bearing amyloid-β plaques or tau pathology. The innate immune response of primary mouse microglia under different treatment conditions or bearing different genetic backgrounds was analyzed using ELISA, western blotting, or immunocytochemistry.ResultsWe found that CH25H expression is upregulated in human AD brain tissue and in transgenic mouse brain tissue-bearing amyloid-β plaques or tau pathology. Treatment with the toll-like receptor 4 (TLR4) agonist lipopolysaccharide (LPS) markedly upregulates CH25H expression in the mouse brain and stimulates CH25H expression and 25-HC secretion in mouse primary microglia. We found that LPS-induced microglial production of the pro-inflammatory cytokine IL-1β is markedly potentiated by 25-HC and attenuated by the deletion of CH25H. Microglia expressing apolipoprotein E4 (apoE4), a genetic risk factor for AD, produce greater amounts of 25-HC than apoE3-expressing microglia following treatment with LPS. Remarkably, 25-HC treatment results in a greater level of IL-1β secretion in LPS-activated apoE4-expressing microglia than in apoE2- or apoE3-expressing microglia. Blocking potassium efflux or inhibiting caspase-1 prevents 25-HC-potentiated IL-1β release in apoE4-expressing microglia, indicating the involvement of caspase-1 inflammasome activity.Conclusion25-HC may function as a microglial-secreted inflammatory mediator in the brain, promoting IL-1β-mediated neuroinflammation in an apoE isoform-dependent manner (E4>>E2/E3) and thus may be an important mediator of neuroinflammation in AD.

Brain-specific repression of AMPKα1 alleviates pathophysiology in Alzheimer's model mice.

AMPK is a key regulator at the molecular level for maintaining energy metabolism homeostasis. Mammalian AMPK is a heterotrimeric complex, and its catalytic α subunit exists in 2 isoforms: AMPKα1 and AMPKα2. Recent studies suggest a role of AMPKα overactivation in Alzheimer's disease-associated (AD-associated) synaptic failure. However, whether AD-associated dementia can be improved by targeting AMPK remains unclear, and roles of AMPKα isoforms in AD pathophysiology are not understood. Here, we showed distinct disruption of hippocampal AMPKα isoform expression patterns in postmortem human AD patients and AD model mice. We further investigated the effects of brain- and isoform-specific AMPKα repression on AD pathophysiology. We found that repression of AMPKα1 alleviated cognitive deficits and synaptic failure displayed in 2 separate lines of AD model mice. In contrast, AMPKα2 suppression did not alter AD pathophysiology. Using unbiased mass spectrometry-based proteomics analysis, we identified distinct patterns of protein expression associated with specific AMPKα isoform suppression in AD model mice. Further, AD-associated hyperphosphorylation of eukaryotic elongation factor 2 (eEF2) was blunted with selective AMPKα1 inhibition. Our findings reveal isoform-specific roles of AMPKα in AD pathophysiology, thus providing insights into potential therapeutic strategies for AD and related dementia syndromes.

Axl receptor tyrosine kinase is a regulator of apolipoprotein E

Alzheimer’s disease (AD), the leading cause of dementia, is a chronic neurodegenerative disease. Apolipoprotein E (apoE), which carries lipids in the brain in the form of lipoproteins, plays an undisputed role in AD pathophysiology. A high-throughput phenotypic screen was conducted using a CCF-STTG1 human astrocytoma cell line to identify small molecules that could upregulate apoE secretion. AZ7235, a previously discovered Axl kinase inhibitor, was identified to have robust apoE activity in brain microglia, astrocytes and pericytes. AZ7235 also increased expression of ATP-binding cassette protein A1 (ABCA1), which is involved in the lipidation and secretion of apoE. Moreover, AZ7235 did not exhibit Liver-X-Receptor (LXR) activity and stimulated apoE and ABCA1 expression in the absence of LXR. Target validation studies using AXL−/− CCF-STTG1 cells showed that Axl is required to mediate AZ7235 upregulation of apoE and ABCA1. Intriguingly, apoE expression and secretion was significantly attenuated in AXL-deficient CCF-STTG1 cells and reconstitution of Axl or kinase-dead Axl significantly restored apoE baseline levels, demonstrating that Axl also plays a role in maintaining apoE homeostasis in astrocytes independent of its kinase activity. Lastly, these effects may require human apoE regulatory sequences, as AZ7235 exhibited little stimulatory activity toward apoE and ABCA1 in primary murine glia derived from neonatal human APOE3 targeted-replacement mice. Collectively, we identified a small molecule that exhibits robust apoE and ABCA1 activity independent of the LXR pathway in human cells and elucidated a novel relationship between Axl and apoE homeostasis in human astrocytes.

Nrf2 activation through the PI3K/GSK-3 axis protects neuronal cells from A\u03b2-mediated oxidative and metabolic damage

BackgroundMounting evidence points to a crucial role of amyloid-β (Aβ) in the pathophysiology of Alzheimer’s disease (AD), a disorder in which brain glucose hypometabolism, downregulation of central elements of phosphorylation pathways, reduced ATP levels, and enhanced oxidative damage coexist, and sometimes precede, synaptic alterations and clinical manifestations. Since the brain has limited energy storage capacity, mitochondria play essential roles in maintaining the high levels of energy demand, but, as major consumers of oxygen, these organelles are also the most important generators of reactive oxygen species (ROS). Thus, it is not surprising that mitochondrial dysfunction is tightly linked to synaptic loss and AD pathophysiology. In spite of their relevance, the mechanistic links among ROS homeostasis, metabolic alterations, and cell bioenergetics, particularly in relation to Aβ, still remain elusive.MethodsWe have used classic biochemical and immunocytochemical approaches together with the evaluation of real-time changes in global energy metabolism in a Seahorse Metabolic Analyzer to provide insights into the detrimental role of oligAβ in SH-SY5Y and primary neurons testing their pharmacologic protection by small molecules.ResultsOur findings indicate that oligomeric Aβ induces a dramatic increase in ROS production and severely affects neuronal metabolism and bioenergetics. Assessment of global energy metabolism in real time demonstrated Aβ-mediated reduction in oxygen consumption affecting basal and maximal respiration and causing decreased ATP production. Pharmacologic targeting of Aβ-challenged neurons with a set of small molecules of known antioxidant and cytoprotective activity prevented the metabolic/bioenergetic changes induced by the peptide, fully restoring mitochondrial function while inducing an antioxidant response that counterbalanced the ROS production. Search for a mechanistic link among the protective small molecules tested identified the transcription factor Nrf2—compromised by age and downregulated in AD and transgenic models—as their main target and the PI3K/GSK-3 axis as the central pathway through which the compounds elicit their Aβ protective action.ConclusionsOur study provides insights into the complex molecular mechanisms triggered by oligAβ which profoundly affect mitochondrial performance and argues for the inclusion of small molecules targeting the PI3K/GSK-3 axis and Nrf2-mediated pathways as part of the current or future combinatorial therapies.