Amyloid Pathogenesis Research Articles

Activation of the muscle-to-brain axis ameliorates neurocognitive deficits in an Alzheimer's disease mouse model via enhancing neurotrophic and synaptic signaling.

Skeletal muscle regulates central nervous system (CNS) function and health, activating the muscle-to-brain axis through the secretion of skeletal muscle-originating factors ("myokines") with neuroprotective properties. However, the precise mechanisms underlying these benefits in the context of Alzheimer's disease (AD) remain poorly understood. To investigate muscle-to-brain axis signaling in response to amyloid β (Aβ)-induced toxicity, we generated 5xFAD transgenic female mice with enhanced skeletal muscle function (5xFAD;cTFEB;HSACre) at prodromal (4-months old) and late (8-months old) symptomatic stages. Skeletal muscle TFEB overexpression reduced Aβ plaque accumulation in the cortex and hippocampus at both ages and rescued behavioral neurocognitive deficits in 8-month-old 5xFAD mice. These changes were associated with transcriptional and protein remodeling of neurotrophic signaling and synaptic integrity, partially due to the CNS-targeting myokine prosaposin (PSAP). Our findings implicate the muscle-to-brain axis as a novel neuroprotective pathway against amyloid pathogenesis in AD.

Genetic diversity differentially regulates molecular mechanisms of brain aging under normal and pathological conditions

AbstractBackgroundGenetically diverse mouse strains have varying life‐spans and varying brain‐aging hallmarks under normal conditions. Studies show that AD mouse models on diverse genomic backgrounds better recapitulate AD‐relevant neuropathological and molecular phenotypes. Mouse models carrying mutant APP and PS1 alleles (APPswe,PS1de9) were generated on eight diverse mouse strains, and significant strain‐specific differences were observed in amyloid deposition, vascular phenotypes, neurodegeneration and transcriptomic signatures. Among those, wild‐derived strains WSB/Eij‐(WSB) and PWK/Phj‐(PWK) gained further attention in AD research. Compared to the APP/PS1 model on standard C57BL/6J‐(B6) mice, WSB.APP/PS1 was associated with elevated neuronal and vascular impairments, whereas, PWK.APP/PS1 was shown to be resilient.MethodWild‐derived strains were originally generated via a backcross strategy, and all PWK.APP/PS1 and WSB.APP/PS1 used in this study were congenic. Cohorts of male and female PWK.APP/PS1 and WSB.APP/PS1 mice and wild‐type controls were aged to 4‐ and 14‐months, and brain hemispheres were processed for RNA‐Seq transcriptomics and neuropathological assessment. In both strains, 14‐months‐old wild‐type mice were compared to 4‐months‐old wild‐type controls to assess brain aging under normal conditions, and 14‐months‐old APP/PS1 mice were compared to 4‐months‐old wild‐type group to assess aging in presence of amyloid pathology (n = 12). Differentially expressed genes were identified and gene set enrichment analysis were performed to determine gene modules and biological pathways associated with brain aging in health and disease.ResultWild‐type aging signatures were mainly strain‐specific with a strong up‐regulation of immune response in WSB, and up‐regulation of metabolic and signaling pathways in PWK. Varying levels of amyloid plaques deposition was observed in the brain of both WSB.APP/PS1 and PWK.APP/PS1 mice. In presence of amyloid, consistent up‐regulation of microglia genes and an elevated immune response was observed. Down‐regulated genes were enriched in synaptic function in WSB, and in pathways of neurodegeneration in PWK.ConclusionThis study suggests that molecular mechanisms of brain aging under normal conditions is modulated by genetic background, and is associated with the response to amyloid pathogenesis. Our findings suggest a potential link between aging‐associated inflammatory state in wild‐type WSB mice and elevated neuronal damage in amyloid pathogenesis, and between preservation of cellular communication in the brain of aged PWK mice and neuronal resilience.

Night-to-night variability in sleep and amyloid beta burden in normal aging.

Alzheimer's disease is associated with sleep disturbances and accumulation of cerebral amyloid beta. The objective was to examine whether actigraphy-detected sleep parameters might be biomarkers for early amyloid burden. Participants underwent a week of actigraphy and an amyloid positron emission tomography (PET) scan. Sleep duration and continuity disruption (sleep fragmentation and nocturnal awakenings) were extracted and compared between amyloid-positive and amyloid-negative participants. Then multiple linear regressions were used between mean or night-to-night intra-individual variability (standard deviation) of sleep parameters and brain amyloid burden in a voxel-wise analysis. Eighty-six subjects were included (80.3±5.4 years; 48.8% of women). Amyloid-positive participants had a higher variability of sleep fragmentation compared to amyloid-negative participants. This parameter was associated with a higher amyloid burden in the frontal and parietal regions, and in the precuneus, in the whole sample. This study highlights the relevance of using variability in sleep continuity as a potential biomarker of early amyloid pathogenesis.

#4868 MASS-SPECTROMETRY ANALYSIS OF CONGOPHILIC URINE AND ITS DETERGENT RESISTANT FRACTION IN PATIENTS WITH AMYLOID AND NON-AMYLOID NEPHROPATHIES

Abstract Background and Aims Being a well-known intrinsic property of amyloid, сongophilia was recently demonstrated in urine of patients with proteinuria of different etiology including renal amyloidosis (RA) and non-amyloid nephropathies (NANP) [1]. Urine proteins (UPs) responsible for congophilia in RA and NANP are supposed to have another property of amyloid such as resistance to ionic detergents and present in urine as detergent-resistant aggregates (DRA). In the pilot study we performed mass-spectrometry (MS) analysis of congophilic urine samples and its detergent-resistant fraction in patients with RA and NANP to investigate UPs and their specificity to the particular kidney disease. Method We collected first morning void urine samples from patients with RA (n = 4) and NANP (n = 4). Urine congophilia was assessed by Congo red Dot test as described previously [1]. To analyze bulk urine (BU) proteins, 30 μl of supernatant was precipitated with 80% acetone and boiled in 2% sodium dodecyl sulfate (SDS) for 30 minutes. Then 5 μg of UPs was separated in 10% polyacrylamide gel electrophoresis (PAGE) followed by Coomassie blue staining. The ultracentrifugation of 0.5-3 ml of urine at 300,000x g for 16h following by the treatment of precipitate with 3% sarcosyl in phosphate-buffered saline (PBS) for 10 min with subsequent washing of resistant aggregates in PBS were applied to obtain DRA. After boiling in SDS 8 μl of the sample was separated in 10% PAGE. UPs concentration in the DRA was estimated by densitometry regarding to 2 μg of bovine serum albumin. Then 6 μg of UPs was digested with trypsin, followed by purification on silicate (CDS Empore™ C18 Extraction Disks). Prepared samples of BU and DRA were analyzed by electrospray ionization tandem MS. We used MsFragger software to obtain lists of UPs for each sample. Most representative UPs in the sample were selected by the unique spectral count (USC). We considered the protein as having diagnosis-specific potential (DSP) if it was present in every sample of the particular disease group, i.e. AA, AL, NANP in BU or DRA, and was absent in any sample of another disease group. Results The patients had following characteristics: age 51±13 years, 3 male/ 5 female, eGFR = 45 (19; 95) ml/min/1.73 m2; 24 h proteinuria = 6.5 (4.5; 7.9) g. In PAGE analysis, BU proteins appeared to be similar in all 8 samples with 2 major bends in 70 kDa and 50 kDa regions, corresponding to albumin and immunoglobulin (Ig) heavy chains, respectively (Fig. 1A). The amount of protein in DRA was small and comprised 1.02 (0.71; 1.61) % of the total protein in the sample (4.7 (2.3; 5,1) g/l). Compared with BU PAGE analysis of DRA proteins revealed other bends with trace albumin bend, predominance of 45 kDA region bend and more apparent 30 kDa bend (Ig light chains) in the majority of patients (Fig. 1B). Results of MS analysis are shown on the Figure 2. There were more DSP revealed in DRA vs BU: 14 vs 5, 46 vs 2 and 4 vs 2 UPs in samples of AL, AA and NANP, respectively. When compare by a particular disease group, there were no DSP found either in BU or DRA in RA samples. One protein (aminopeptidase N) and 3 proteins (isoform 3 of unconventional myosin-LC, protein S100-A8, elongation factor 1-α 1) were detected as DSP in BU and DRA in AL, respectively. Alpha-1-acid glycoprotein 1 was only DSP for AA in DRA as well as serum paraoxonase 1 for NANP in BU. Venn-diagrams of shared and divergent UPs are present on the Figures 2D-G. Conclusion Although DRA represented a small portion of UPs, its composition significantly differed from BU and could contain more specific disease markers that makes urine detergent-resistant fraction promising for the future research. Understanding the role of DRA proteins in the pathogenesis of amyloid and non-amyloid renal disease and their diagnostic utility requires further studies.

INPP5D deficiency attenuates amyloid pathology in a mouse model of Alzheimer's disease.

Inositol polyphosphate-5-phosphatase (INPP5D) is a microglia-enriched lipid phosphatase in the central nervous system. A non-coding variant (rs35349669) in INPP5D increases the risk for Alzheimer's disease (AD), and elevated INPP5D expression is associated with increased plaque deposition. INPP5D negatively regulates signaling via several microglial cell surface receptors, including triggering receptor expressed on myeloid cells 2 (TREM2); however, the impact of INPP5D inhibition on AD pathology remains unclear. We used the 5xFAD mouse model of amyloidosis to assess how Inpp5d haplodeficiency regulates amyloid pathogenesis. Inpp5d haplodeficiency perturbs the microglial intracellular signaling pathways regulating the immune response, including phagocytosis and clearing of amyloid beta (Aβ). It is important to note that Inpp5d haploinsufficiency leads to the preservation of cognitive function. Spatial transcriptomic analysis revealed that pathways altered by Inpp5d haploinsufficiency are related to synaptic regulation and immune cell activation. These data demonstrate that Inpp5d haplodeficiency enhances microglial functions by increasing plaque clearance and preserves cognitive abilities in 5xFAD mice. Inhibition of INPP5D is a potential therapeutic strategy for AD.

Genetically diverse mouse models of Alzheimer’s Disease reveal an association between amyloid deposition and glycan metabolism in the brain

AbstractBackgroundVarying degrees of cerebral amyloid angiopathy (CAA) co‐occur with parenchymal amyloid deposition in majority of Alzheimer’s Disease (AD) cases. However, common mouse models of familial AD generated on a single inbred strain do not recapitulate the spectrum of parenchymal and vascular amyloid pathogenesis in the brain. To better represent phenotypic diversity in AD, we generated a panel of genetically diverse Collaborative Cross (CC) mouse strains harboring APOE4 allele and amyloidogenic mutations.MethodFive CC strains (CC002, CC006, CC013, CC037 and CC041) were selected for maximal genetic and transcriptional variation in AD‐relevant genes. Selected CC strains were crossed to C57BL/6J (B6J) mice homozygous for humanized APOE4 allele and carrying mutant APP and PS1 alleles (APPswe , PS1de9 ). Cohorts of male and female B6JCCnF1.APOE4.APP/PS1 mice and controls were aged to 8 months, and brain hemispheres were processed for RNA‐Seq transcriptomics and neuropathological assessment. Weighted Gene Co‐Expression Networks Analysis (WGCNA) and gene set enrichment analysis were performed to identify gene modules and biological pathways associated with plaque deposition and CAA in CC strains.ResultNeuropathological assessment revealed significant strain and sex‐dependent differences in both parenchymal and vascular amyloid pathology. Highest levels of plaque deposition in cortex and hippocampus was observed in female CC002 mice. Female mice had higher plaque load compared to male mice in all strains, whereas CAA score was higher in males and in CC037 strain. CC006 represented lowest levels of plaque deposition and CAA at comparable levels with B6J mice. RNA‐Seq data revealed a common microglia/neuroinflammation related transcriptional response observed in all strains and both sexes. On the other hand, gene expression signatures associated with varying levels of plaque deposition and CAA revealed alterations in metabolic pathways, in particular glucose, lipid and glycan metabolisms.ConclusionOur results suggest that major genetic diversity modulates distinctive parenchymal and vascular amyloid phenotypes, and differentially impacts transcriptomic signatures in the mouse brain. Overall, CC lines mouse models better represent the spectrum of neuropathological and molecular phenotypes associated with AD.

APOE ε4-dependent effects on the early amyloid pathology in induced neurons of patients with Alzheimer’s disease

BackgroundThe ε4 allele of apolipoprotein E (APOE ε4) is the strongest known genetic risk factor for late-onset Alzheimer’s disease (AD), associated with amyloid pathogenesis. However, it is not clear how APOE ε4 accelerates amyloid-beta (Aβ) deposition during the seeding stage of amyloid development in AD patient neurons.MethodsAD patient induced neurons (iNs) with an APOE ε4 inducible system were prepared from skin fibroblasts of AD patients. Transcriptome analysis was performed using RNA isolated from the AD patient iNs expressing APOE ε4 at amyloid-seeding and amyloid-aggregation stages. Knockdown of IGFBP3 was applied in the iNs to investigate the role of IGFBP3 in the APOE ε4-mediated amyloidosis.ResultsWe optimized amyloid seeding stage in the iNs of AD patients that transiently expressed APOE ε4. Remarkably, we demonstrated that Aβ pathology was aggravated by the induction of APOE ε4 gene expression at the amyloid early-seeding stage in the iNs of AD patients. Moreover, transcriptome analysis in the early-seeding stage revealed that IGFBP3 was functionally important in the molecular pathology of APOE ε4-associated AD.ConclusionsOur findings suggest that the presence of APOE ε4 at the early Aβ-seeding stage in patient iNs is critical for aggravation of sporadic AD pathology. These results provide insights into the importance of APOE ε4 expression for the progression and pathogenesis of sporadic AD.

Association of Plasma Oligomerized Amyloid-β and Cerebral White Matter Lesions in a Health Screening Population.

Cerebral white matter lesions (WML) are related to a higher risk of vascular and Alzheimer's dementia. Moreover, oligomerized amyloid-β (OAβ) can be measured from blood for dementia screening. We aimed to investigate the relationship of plasma OAβ levels with clinical and radiological variables in a health screening population. WML, other volumetric parameters of magnetic resonance images, cognitive assessment, and plasma OAβ level were evaluated. Ninety-two participants were analyzed. The majority of participants' clinical dementia rating was 0 or 0.5 (96.7%). White matter hyperintensities (WMH) increased with age, but OAβ levels did not (r2 = 0.19, p < 0.001, r2 = 0.03, p = 0.10, respectively). No volumetric data, including cortical thickness/hippocampal volume, showed any significant correlation with OAβ. Log-WMH volume was positively correlated with OAβ (r = 0.24, p = 0.02), and this association was significant in the periventricular area. White matter signal abnormalities from 3D-T1 images were also correlated with the OAβ in the periventricular area (p = 0.039). Multivariate linear regression showed that log-WMH values were independently associated with OAβ (B = 0.879 (95% confidence interval 0.098 -1.660, p = 0.028)). Higher tertiles of WMH showed higher OAβ levels than lower tertiles showed (p = 0.044). Using a cutoff of 0.78 ng/mL, the high OAβ group had a larger WMH volume, especially in the periventricular area, than the low OAβ group (p = 0.036). Both WML and plasma OAβ levels can be early markers for neurodegeneration in the healthcare population. The lesions, especially in the periventricular area, might be related to amyloid pathogenesis, which strengthens the importance of WML in the predementia stage.

Alzheimer’s disease genetic risk variants show brain cell type‐specific associations with protein levels in cerebrospinal fluid

Neuronal dysfunction is central to the clinical manifestation of Alzheimer's disease (AD). However, genome-wide studies also suggest important roles for non-neuronal brain cell-types such as microglia and astrocytes. Our objective was to study whether brain cell type-specific polygenic risk scores (PGRS) for AD, including single nucleotide polymorphisms (SNPs) of genes expressed in one brain cell type, showed relationships with levels of cerebrospinal fluid (CSF) AD markers in individuals across the clinical spectrum of AD. We selected 1,535 subjects (552 controls/709 mild cognitive impairment/274 AD-dementia, age 71±8 years, 48%female) from the ADNI (N=617) and EMIF-AD MBD (N=918) study, who had genetic data available. We labelled AD risk genes as specific for neurons, astrocytes, microglia, oligodendrocytes or endothelial cells when more than 50% of the gene expression was produced by one cell type (otherwise as 'non-specific') according to the BRAIN RNASeq database (Zhang et al., 2014). We calculated cell type-specific-PGRS with cell type-corresponding SNPs (P<5e-8 ) using weights from De Rojas et al., (2020) AD case-control summary statistics. Associations between cell type-specific-PGRS and CSF biomarkers (amyloid-beta, total tau (t-tau), phosphorylated tau (p-tau), neurofilament light, neurogranin and YKL-40) were examined using linear regressions, adjusted for population structure, study, age and sex. Of 40 AD risk genes, 26 had detectable RNA levels in at least one brain cell type. Of these, sixteen were cell type-specific: 11 were microglia-specific, 4 astrocyte-specific (including APOE) and 1 neuron-specific (Figure 1). Astrocyte-PGRS (P=1.4e-6 , Pwithout_APOE =.1) and microglia-PGRS (P=6.3e-3 ) were increased in AD-type dementia compared to controls, whereas other PGRS were not (Figure 2). Astrocyte-PGRS (including APOE) were most strongly associated with CSF amyloid-beta (P=1.4e-31 ), t-tau (P=6.3e-10 ), p-tau (P=4.9e-9 ), and neurogranin (P=0.01). Astrocyte-specific (excluding APOE) and microglia-specific PGRS-AD also associated with CSF amyloid-beta (P<0.05), and with p-tau at trend-level (Figure 2). Apart from the tentative association between neuron-PGRS and CSF YKL-40, no other associations were observed. Findings indicate that AD risk variants have cell type-specific associations with amyloid and tau pathology, which seems mostly expressed by astrocytes (also without APOE) and microglia, suggesting that these cell types play a role in amyloid pathogenesis.

Incorporating genetic diversity to improve alignment of mouse models to human Alzheimer’s disease

Previous work by our labs, and others, reveal the benefits of incorporating diverse mouse strains to better model human Alzheimer's disease (AD). For instance, mouse models of AD on genetically distinct wild-derived mouse strains demonstrated robust differences in immune response to amyloid and neurodegeneration, better representing the phenotypic spectrum of AD than previous models. To further investigate the importance of genetic diversity, as part of MODEL-AD, we generated AD-relevant mouse models on Collaborative Cross (CC) lines background, a recombinant inbred mouse panel created from eight highly diverse founder strains.C57BL/6J (B6J) mice homozygous for humanized APOE4 allele and carrying mutant APP and PS1 alleles (APPswe , PS1de9 ) were crossed to five CC strains that were selected based on variation in AD-relevant genes. Cohorts of male and female B6JCCnF1.APOE4.APP/PS1 mice and controls were aged to 8 months (a mid-point for amyloid deposition in B6.APP/PS1 mice) and brain hemispheres processed for RNA-Seq and neuropathological assessment. Expectation Maximization for Allele Specific Expression (EMASE) tool was used to quantify transcriptome-wide expression at allelic level. Differential expression and KEGG pathway enrichment analysis were performed to determine altered molecular processes in CC strains. Strain specific transcriptional signatures were mapped to co-expression gene modules of human Late-Onset AD (LOAD) from the AMP-AD consortium. Neuropathological assessment included characterization of plaque deposition, Tau pathology, glial activation, neurodegeneration and cerebrovascular health.RNA-Seq analysis revealed up-regulation of neuroinflammation related pathways (lysosome, complement and coagulation cascades) in all CC strains as an effect of mutant APP and PS1 transgenes in presence of APOE4 allele. On the other hand, we observed significant CC strain and sex specific variation in immune (Fc gamma R-mediated phagocytosis) and metabolic (glycosphingolipid biosynthesis) response to amyloid pathogenesis. Alignment with human LOAD signatures revealed high strain and sex specific variation in correlation with synaptic signaling, cell cycle and DNA repair related gene modules. Overall, novel AD mouse models that incorporated CC lines demonstrated increased alignment to human AD-like transcriptomic signatures compared to B6J.This work builds on previous work highlighting the strengths of including multiple diverse mouse strains to better model complex neurodegenerative disorders such as AD and related dementias.

MicroRNA-425 loss mediates amyloid plaque microenvironment heterogeneity and promotes neurodegenerative pathologies.

Different cellular and molecular changes underlie the pathogenesis of Alzheimer's disease (AD). Among these, neuron‐specific dysregulation is a necessary event for accumulation of classic pathologies including amyloid plaques. Here, we show that AD‐associated pathophysiology including neuronal cell death, inflammatory signaling, and endolysosomal dysfunction is spatially colocalized to amyloid plaques in regions with abnormal microRNA‐425 (miR‐425) levels and this change leads to focal brain microenvironment heterogeneity, that is, an amyloid plaque‐associated microenvironment (APAM). APAM consists of multiple specific neurodegenerative signature pathologies associated with senile plaques that contribute to the heterogeneity and complexity of AD. Remarkably, miR‐425, a neuronal‐specific regulator decreased in AD brain, maintains a normal spatial transcriptome within brain neurons. We tested the hypothesis that miR‐425 loss correlates with enhanced levels of mRNA targets downstream, supporting APAM and AD progression. A miR‐425‐deficient mouse model has enhanced APP amyloidogenic processing, neuroinflammation, neuron loss, and cognitive impairment. In the APP/PS1 mouse model, intervening with miR‐425 supplementation ameliorated APAM changes and memory deficits. This study reveals a novel mechanism of dysregulation of spatial transcriptomic changes in AD brain, identifying a probable neuronal‐specific microRNA regulator capable of staving off amyloid pathogenesis. Moreover, our findings provide new insights for developing AD treatment strategies with miRNA oligonucleotide(s).

(-)-Oleocanthal Nutraceuticals for Alzheimer's Disease Amyloid Pathology: Novel Oral Formulations, Therapeutic, and Molecular Insights in 5xFAD Transgenic Mice Model.

Alzheimer’s disease (AD) is a complex progressive neurodegenerative disorder affecting humans mainly through the deposition of Aβ-amyloid (Aβ) fibrils and accumulation of neurofibrillary tangles in the brain. Currently available AD treatments only exhibit symptomatic relief but do not generally intervene with the amyloid and tau pathologies. The extra-virgin olive oil (EVOO) monophenolic secoiridoid S-(–)-oleocanthal (OC) showed anti-inflammatory activity through COX system inhibition with potency comparable to the standard non-steroidal anti-inflammatory drug (NSAID) like ibuprofen. OC also showed positive in vitro, in vivo, and clinical therapeutic effects against cardiovascular diseases, many malignancies, and AD. Due to its pungent, astringent, and irritant taste, OC should be formulated in acceptable dosage form before its oral use as a potential nutraceutical. The objective of this study is to develop new OC oral formulations, assess whether they maintained OC activity on the attenuation of β-amyloid pathology in a 5xFAD mouse model upon 4-month oral dosing use. Exploration of potential OC formulations underlying molecular mechanism is also within this study scope. OC powder formulation (OC-PF) and OC-solid dispersion formulation with erythritol (OC-SD) were prepared and characterized using FT-IR spectroscopy, powder X-ray diffraction, and scanning electron microscopy (ScEM) analyses. Both formulations showed an improved OC dissolution profile. OC-PF and OC-SD improved memory deficits of 5xFAD mice in behavioral studies. OC-PF and OC-SD exhibited significant attenuation of the accumulation of Aβ plaques and tau phosphorylation in the brain of 5xFAD female mice. Both formulations markedly suppressed C3AR1 (complement component 3a receptor 1) activity by targeting the downstream marker STAT3. Collectively, these results demonstrate the potential for the application of OC-PF as a prospective nutraceutical or dietary supplement to control the progression of amyloid pathogenesis associated with AD.

Effect of exercise training on amyloid-like protein aggregates among patients with heart failure

Abstract Funding Acknowledgements Type of funding sources: Other. Main funding source(s): MG and CS were supported by a PhD FCT grant (SFRH/BD/128893/2017) and by an individual grant from CAPES [BEX 0554/14-6], respectively. This work was financially supported by the project POCI-01-0145-FEDER-030011, funded by FEDER, through COMPETE2020-POCI, and by national funds, through FCT/MCTES (PTDC/MEC-CAR/30011/2017). iBiMED is a research unit supported by the Portuguese Foundation for Science and Technology (REF: UID/BIM/04501/2020) and FEDER/Compete2020 funds). Introduction Amyloid-like protein aggregates play a decisive role in the pathology of heart failure. Alterations in protein homeostasis, in particular, the clearance of toxic amyloid-like aggregates are emerging therapeutic targets in cardiovascular medicine. The clinical benefits of cardiac rehabilitation and exercise training are widely accepted in heart failure; however, little is known about the potential benefit of exercise training in amyloid-like protein aggregates. Purpose To assess the effects of a moderate-intensity exercise training program on amyloid-like protein aggregates levels among patients with heart failure with reduced ejection fraction. Methods Eighteen subjects participated in the study; eight patients (age: 66.6 ± 5.9 years; FEVE: 38.4 ± 8.9%) with heart failure with reduced ejection fraction participated in a 3-month exercise training program (2 x 60 min sessions per week of moderate-intensity aerobic and resistance exercise). Ten healthy subjects (age: 68. 4 ± 3.1 years) were recruited to an age-matched reference group. Amyloid-like protein aggregates were assessed before and after 3 months of exercise training. Clinical data, medication, anthropometrics, and cardiorespiratory fitness were also assessed. Thioflavin T (ThT) dye fluorescence was used to quantify the plasma levels of amyloid-like aggregates and the Fourier transform infrared spectroscopy (FTIR) was applied to evaluate the conformation of cross-β-sheet structures characteristic of amyloid protein aggregates. Results Exercise program improved cardiorespiratory fitness by 14.0 ± 17.1% (17.4 ± 3.2 to 19.7 ± 2.9 ml/kg/min) and reduced NT-proBNP levels by 16.5% (34.2) (median concentration of 632 pg/mL (720.8) to 517.5 pg/mL (707.0)) in the heart failure patients. A slight decrease of amyloid-like aggregates levels was observed in post-exercise training samples (a reduction of 3.1%); interestingly, after the exercise training program, the heart failure patients showed levels of amyloid-like aggregates similar to the reference group (1132.0 ± 114.2 vs. 1094.8 ± 132.9 a.u.). Additionally, the PLS-R multivariate analysis of the amide I region of the FTIR spectra revealed enrichment of antiparallel β-sheets (1693 cm-1) assigned to amyloid-like oligomers in the samples of heart failure patients before, but not after, the exercise program. Of note, oligomeric species, as intermediates of amyloid assembly, can contribute to the increase of amyloid burden, but also, some have been reported to be highly reactive and toxic to cells, being key elements of amyloid pathogenesis. Conclusions Our preliminary results indicate that 3 months of exercise training may have significant effects on amyloid-like oligomers, and start hindering the formation of the larger ThT-positive aggregates among patients with heart failure. Abstract Figure.

The inhibition of cellular toxicity of amyloid-β by dissociated transthyretin

The protective effect of transthyretin (TTR) on cellular toxicity of β-amyloid (Aβ) has been previously reported. TTR is a tetrameric carrier of thyroxine in blood and cerebrospinal fluid, the pathogenic aggregation of which causes systemic amyloidosis. However, studies have documented a protective effect of TTR against cellular toxicity of pathogenic Aβ, a protein associated with Alzheimer's disease. TTR binds Aβ, alters its aggregation, and inhibits its toxicity both in vitro and in vivo In this study, we investigate whether the amyloidogenic ability of TTR and its antiamyloid inhibitory effect are associated. Using protein aggregation and cytotoxicity assays, we found that the dissociation of the TTR tetramer, required for its amyloid pathogenesis, is also necessary to prevent cellular toxicity from Aβ oligomers. These findings suggest that the Aβ-binding site of TTR may be hidden in its tetrameric form. Aided by computational docking and peptide screening, we identified a TTR segment that is capable of altering Aβ aggregation and toxicity, mimicking TTR cellular protection. EM, immune detection analysis, and assessment of aggregation and cytotoxicity revealed that the TTR segment inhibits Aβ oligomer formation and also promotes the formation of nontoxic, nonamyloid amorphous aggregates, which are more sensitive to protease digestion. Finally, this segment also inhibits seeding of Aβ catalyzed by Aβ fibrils extracted from the brain of an Alzheimer's patient. Together, these findings suggest that mimicking the inhibitory effect of TTR with peptide-based therapeutics represents an additional avenue to explore for the treatment of Alzheimer's disease.

A Preliminary Study of Cu Exposure Effects upon Alzheimer's Amyloid Pathology.

A large body of evidence indicates that dysregulation of cerebral biometals (Fe, Cu, Zn) and their interactions with amyloid precursor protein (APP) and Aβ amyloid may contribute to the Alzheimer’s disease (AD) Aβ amyloid pathology. However, the molecular underpinnings associated with the interactions are still not fully understood. Herein we have further validated the exacerbation of Aβ oligomerization by Cu and H2O2 in vitro. We have also reported that Cu enhanced APP translations via its 5′ untranslated region (5′UTR) of mRNA in SH-SY5Y cells, and increased Aβ amyloidosis and expression of associated pro-inflammatory cytokines such as MCP-5 in Alzheimer’s APP/PS1 doubly transgenic mice. This preliminary study may further unravel the pathogenic role of Cu in Alzheimer’s Aβ amyloid pathogenesis, warranting further investigation.

Effect of amyloid toxicity or chronic cerebral hypoperfusion on brain insulin resistance in a rat model with intracerebroventricular streptozotocin

Sporadic Alzheimer’s disease (AD) is a multifactorial neurodegenerative disorder affected by amyloid and vascular pathogenesis. Brain insulin resistance (BIR) has been suggested as one of the pathomechanisms of sporadic AD. We investigated how the amyloid and vascular pathogenesis of AD interacts with BIR. We examined experimental groups mimicking amyloid pathogenesis following intracerebroventriculr (icv) injection of amyloid β or vascular pathogenesis following permanent ligation of the bilateral common carotid arteries in Wistar rats that had undergone icv injection of streptozotocin. Behavioral tests and pathologic studies were performed. Cognitive impairments were induced by BIR superimposed by amyloid or vascular pathogenesis. Neuroinflammation in the white matter and hippocampus was aggravated by an interaction between BIR and vascular pathogenesis. Amyloid-associated pathology in the white matter was enhanced by BIR and vascular pathogenesis. Tau-associated pathology in the hippocampus was altered by BIR in a relation with amyloid or vascular pathogenesis. Our study may provide useful experimental insights based on an integrated approach to the influence of amyloid and vascular pathogenesis on BIR, permitting better understanding of the heterogeneous pathogenesis of sporadic AD. Pathologic responses in sporadic AD may differ depending on amyloid and vascular pathogenesis and may sometimes be synergistically aggravated when combined with BIR.

Associative Interactions among Zinc, Apolipoprotein E, and Amyloid-β in the Amyloid Pathology.

Zinc and apolipoprotein E (apoE) are reportedly involved in the pathology of Alzheimer’s disease. To investigate the associative interaction among zinc, apoE, and amyloid-β (Aβ) and its role in amyloid pathogenesis, we performed various biochemical and immunoreactive analyses using brain tissues of Tg2576 mice and synthetic Aβ and apoE peptides. On amyloid plaques or in brain lysates of Tg2576 mice, apoE and Aβ immunoreactivities increased after zinc chelation and were restored by its subsequent replacement. Zinc depletion dissociated apoE/Aβ complexes or larger-molecular sizes of Aβ oligomers/aggregates into smaller-molecular sizes of apoE and/or Aβ monomers/complexes. In the presence of zinc, synthetic apoE and/or Aβ peptides aggregated into larger-molecular sizes of oligomers or complexes. Endogenous proteases or plasmin in brain lysates degraded apoE and/or Aβ complexes, and their proteolytic activity increased with zinc depletion. These biochemical findings suggest that zinc associates with apoE and Aβ to encourage the formation of apoE/Aβ complexes or large aggregates, raising the deposition of zinc-rich amyloid plaques. In turn, the presence of abundant zinc around and within apoE/Aβ complexes may block the access or activity of Aβ-degrading antibodies or proteases. These results support the plausibility of chelation strategy aiming at reducing amyloid pathology in Alzheimer’s disease.

IL-1\u03b2-driven amyloid plaque clearance is associated with an expansion of transcriptionally reprogrammed microglia

BackgroundNeuroinflammation is thought to contribute to the pathogenesis of Alzheimer’s disease (AD), yet numerous studies have demonstrated a beneficial role for neuroinflammation in amyloid plaque clearance. We have previously shown that sustained expression of IL-1β in the hippocampus of APP/PS1 mice decreases amyloid plaque burden independent of recruited CCR2+ myeloid cells, suggesting resident microglia as the main phagocytic effectors of IL-1β-induced plaque clearance. To date, however, the mechanisms of IL-1β-induced plaque clearance remain poorly understood.MethodsTo determine whether microglia are involved in IL-1β-induced plaque clearance, APP/PS1 mice induced to express mature human IL-1β in the hippocampus via adenoviral transduction were treated with the Aβ fluorescent probe methoxy-X04 (MX04) and microglial internalization of fibrillar Aβ (fAβ) was analyzed by flow cytometry and immunohistochemistry. To assess microglial proliferation, APP/PS1 mice transduced with IL-1β or control were injected intraperitoneally with BrdU and hippocampal tissue was analyzed by flow cytometry. RNAseq analysis was conducted on microglia FACS sorted from the hippocampus of control or IL-1β-treated APP/PS1 mice. These microglia were also sorted based on MX04 labeling (MX04+ and MX04− microglia).ResultsResident microglia (CD45loCD11b+) constituted > 70% of the MX04+ cells in both Phe- and IL-1β-treated conditions, and < 15% of MX04+ cells were recruited myeloid cells (CD45hiCD11b+). However, IL-1β treatment did not augment the percentage of MX04+ microglia nor the quantity of fAβ internalized by individual microglia. Instead, IL-1β increased the total number of MX04+ microglia in the hippocampus due to IL-1β-induced proliferation. In addition, transcriptomic analyses revealed that IL-1β treatment was associated with large-scale changes in the expression of genes related to immune responses, proliferation, and cytokine signaling.ConclusionsThese studies show that IL-1β overexpression early in amyloid pathogenesis induces a change in the microglial gene expression profile and an expansion of microglial cells that facilitates Aβ plaque clearance.

Mild cognitive impairment has similar alterations as Alzheimer's disease in gut microbiota

ObjectiveGut microbiota changes before the onset of Alzheimer's disease (AD) and the alterations could be detected in the stage of mild cognitive impairment (MCI). The findings might offer diagnostic biomarkers before the onset of dementia. BackgroundAD is the most common cause of dementia, and MCI is the predementia state. Recent studies suggest the alterations in the gut microbial communities associated with AD, whereas the microbiota in MCI before the onset of dementia has not been discovered and characterized in humans. New/Updated HypothesisWe hypothesize that the dysbiosis happens in the MCI stage. Patients with AD and MCI have decreased microbial diversity, and changes in gut microbiota could be detected for early detection of AD. In our preliminary study, we identified differences between AD and normal controls in 11 genera from the feces and 11 genera from the blood. No difference in genera between AD and MCI was detected. Using the diagnostic model from fecal samples with all different genera input, 93% (28 in 30) of patients with MCI could be identified correctly. Major Challenges for the HypothesisThe diagnosis of MCI and AD in the study was based on symptoms and neuroimaging, and AD biomarkers should be included for precise diagnosis in further validating studies. Besides, as the microbiota changes longitudinally, their relationship with the progress of dementia needs to be studied in the prospective studies. Linkage to Other Major TheoriesEscherichia was observed increased at genus level in both fecal and blood samples from AD and MCI. For AD biomarker, postmortem brain tissue from patients with AD showed lipopolysaccharides and gram-negative Escherichia coli fragments colocalize with amyloid plaque. In this way, the amyloid pathogenesis for AD would be triggered during MCI by gut microbiota shifting. Besides, systemic inflammatory reactions caused by compounds secreted by bacteria may impair the blood-brain barrier and promote neuroinflammation and/or neurodegeneration. Furthermore, abnormal metabolites caused by microbial gene functions have an impact on neurodegeneration.

Subchronic Pathobiological Response Following Chronic Repetitive Mild Traumatic Brain Injury in an Aged Preclinical Model of Amyloid Pathogenesis.

Repetitive mild traumatic brain injury (r-mTBI) is a risk factor for Alzheimer disease (AD). The precise nature of how r-mTBI leads to, or precipitates, AD pathogenesis remains unclear. In this study, we explore subchronic effects of chronic r-mTBI (12-impacts) administered over 1-month in aged-PS1/APP mice and littermate controls. We investigate specific mechanisms that may elucidate the molecular link between AD and r-mTBI, focusing primarily on amyloid and tau pathology, amyloid processing, glial activation states, and associated clearance mechanisms. Herein, we demonstrate r-mTBI in aged PS1/APP mice does not augment, glial activation, amyloid burden, or tau pathology (with exception of pS202-positive Tau) 1 month after exposure to the last-injury. However, we observed a decrease in brain soluble Aβ42 levels without any appreciable change in peripheral soluble Aβ42 levels. This was accompanied by an increase in brain insoluble to soluble Aβ42 ratio in injured PS1/APP mice compared with sham injury. A parallel reduction in phagocytic receptor, triggering receptor expressed on myeloid cells 2, was also observed. This study demonstrates very subtle subchronic effects of r-mTBI on a preexisting amyloid pathology background, which may be on a continuum toward a slow and worsening neurodegenerative outcome compared with sham injury, and therefore, have many implications, especially in the elderly population exposed to TBI.