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Background: Neurodegenerative disorders, including Alzheimer's disease (AD), have been linked to alterations in tryptophan (TRP) metabolism. However, no studies to date have systematically explored changes in the TRP pathway at both transcriptional and epigenetic levels. This study aimed to investigate transcriptomic, DNA methylomic (5mC) and hydroxymethylomic (5hmC) changes within genes involved in the TRP and nicotinamide adenine dinucleotide (NAD) pathways in AD, using three independent cohorts. Methods: DNA derived from post-mortem middle temporal gyrus (MTG) tissue from AD patients (n = 45) and age-matched controls (n = 35) was analyzed, along with DNA derived from blood samples from two independent cohorts: the German Study on Ageing, Cognition, and Dementia in Primary Care Patients (AgeCoDe) cohort (n = 96) and the Dutch BioBank Alzheimer Center Limburg (BBACL) cohort (n = 262). Molecular profiling, including assessing mRNA expression and DNA (hydroxy)methylation levels, was conducted using HumanHT-12 v4 Expression BeadChip and HM 450 K BeadChip arrays, respectively. Functional interactions between genes and identification of common phenotype-specific positive and negative elementary circuits were conducted using computational modeling, i.e. gene regulatory network (GRN) and network perturbational analysis. DNA methylation of IDO2 (cg11251498) was analyzed using pyrosequencing. Results: Twelve TRP- and twenty NAD-associated genes were found to be differentially expressed in the MTG of AD patients. Gene sets associated in the kynurenine pathway, the most common TRP pathway, and NAD pathway, showed enrichment at the mRNA expression level. Downstream analyses integrating data on gene expression, DNA (hydroxy)methylation, and AD pathology, as well as GRN and network perturbation analyses, identified IDO2, an immune regulatory gene, as a key candidate in AD. Notably, one CpG site in IDO2 (cg11251498) exhibited significant methylation differences between AD converters and non-converters in the AgeCoDe cohort. Conclusion: These findings reveal substantial transcriptional and epigenetic alterations in TRP- and NAD-pathway-associated genes in AD, highlighting IDO2 as a key candidate gene for further investigation. These genes and their encoded proteins hold potential as novel biomarkers and therapeutic targets for AD. [ABSTRACT FROM AUTHOR]

Alzheimer's disease is a neurodegenerative disorder clinically defined by gradual cognitive impairment and alteration in executive function. We conducted an epigenome-wide association study (EWAS) of a clinically and neuropathologically characterized cohort of 296 brains, including Alzheimer's disease (AD) and non-demented controls (ND), exploring the relationship with the RNA expression from matched donors. We detected 5246 CpGs and 832 regions differentially methylated, finding overlap with previous EWAS but also new associations. CpGs previously identified in ANK1, MYOC, and RHBDF2 were differentially methylated, and one of our top hits (GPR56) was not previously detected. ANK1 was differentially methylated at the region level, along with APOE and RHBDF2. Only a small number of genes showed a correlation between DNA methylation and RNA expression statistically significant. Multiblock partial least-squares discriminant analysis showed several CpG sites and RNAs discriminating AD and ND (AUC = 0.908) and strongly correlated with each other. Furthermore, the CpG site cg25038311 was negatively correlated with the expression of 22 genes. Finally, with the functional epigenetic module analysis, we identified a protein–protein network characterized by inverse RNA/DNA methylation correlation and enriched for "Regulation of insulin-like growth factor transport", with IGF1 as the hub gene. Our results confirm and extend the previous EWAS, providing new information about a brain region not previously explored in AD DNA methylation studies. The relationship between DNA methylation and gene expression is not significant for most of the genes in our sample, consistently with the complexities in the gene expression regulation. [ABSTRACT FROM AUTHOR]

The clinical diagnosis of neurodegenerative diseases is notoriously inaccurate and current methods are often expensive, time-consuming, or invasive. Simple inexpensive and noninvasive methods of diagnosis could provide valuable support for clinicians when combined with cognitive assessment scores. Biological processes leading to neuropathology progress silently for years and are reflected in both the central nervous system and vascular peripheral system. A blood-based screen to distinguish and classify neurodegenerative diseases is especially interesting having low cost, minimal invasiveness, and accessibility to almost any world clinic. In this study, we set out to discover a small set of blood transcripts that can be used to distinguish healthy individuals from those with Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, Friedreich's ataxia, or frontotemporal dementia. Using existing public datasets, we developed a machine learning algorithm for application on transcripts present in blood and discovered small sets of transcripts that distinguish a number of neurodegenerative diseases with high sensitivity and specificity. We validated the usefulness of blood RNA transcriptomics for the classification of neurodegenerative diseases. Information about features selected for the classification can direct the development of possible treatment strategies. [ABSTRACT FROM AUTHOR]

Understanding cell-to-cell variation at the molecular level provides relevant information about biological phenomena and is critical for clinical and biological research. Proteins carry important information not available from single-cell genomics and transcriptomics studies; however, due to the minute amount of proteins in single cells and the complexity of the proteome, quantitative protein analysis at the single-cell level remains challenging. Here, we report an integrated microfluidic platform in tandem with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF–MS) for the detection and quantification of targeted proteins from small cell ensembles (> 10 cells). All necessary steps for the assay are integrated on-chip including cell lysis, protein immunocapture, tryptic digestion, and co-crystallization with the matrix solution for MALDI-MS analysis. We demonstrate that our approach is suitable for protein quantification by assessing the apoptotic protein Bcl-2 released from MCF-7 breast cancer cells, ranging from 26 to 223 cells lysed on-chip (8.75 nL wells). A limit of detection (LOD) of 11.22 nM was determined, equivalent to 5.91 × 107 protein molecules per well. Additionally, the microfluidic platform design was further improved, establishing the successful quantification of Bcl-2 protein from MCF-7 cell ensembles ranging from 8 to 19 cells in 4 nL wells. The LOD in the smaller well designs for Bcl-2 resulted in 14.85 nM, equivalent to 3.57 × 107 protein molecules per well. This work shows the capability of our approach to quantitatively assess proteins from cell lysate on the MIMAS platform for the first time. These results demonstrate our approach constitutes a promising tool for quantitative targeted protein analysis from small cell ensembles down to single cells, with the capability for multiplexing through parallelization and automation. [ABSTRACT FROM AUTHOR]

Alzheimer's disease (AD) is the most common cause of dementia in older adults. However, the pathogenesis of AD remains to be fully understood and clinically effective treatments are lacking. Recent advances in single cell RNA sequencing offers an opportunity to characterize the heterogeneity of cell response and explore the molecular mechanism of complex diseases at a single cell level. Here, we present the application of the Ion AmpliSeq transcriptome approach to profile gene expression in single laser captured neurons as well as pooled 10 and 100 neurons from hippocampal CA1 of AD brains versus matching normal aged brains. Our results demonstrated the high sensitivity and high genome coverage of the AmpliSeq transcriptome in single cell sequencing. In addition to capturing the known changes related to AD, our data confirmed the diversity of neuronal profiles in AD brain, which allow the potential identification of single cell response that might be hidden in population analyses. Notably, we also revealed the extensive inhibition of olfactory signaling and confirmed the reduction of neurotransmitter receptors in AD hippocampus. We conclude that although single neuron data show more variance than data from 10 or 100 pooled neurons, single neuron data can be informative. These findings support the utility of the Ion AmpliSeq method for obtaining and analyzing gene expression data from single defined laser captured neurons. [ABSTRACT FROM AUTHOR]

We used Illumina Human HT-12 v4 arrays to compare RNA expression of middle temporal gyrus (MTG; BA21) in Alzheimer's disease (AD = 97) and non-demented controls (ND = 98). A total of 938 transcripts were highly differentially expressed (adj p < 0.01; log2 FC ≥ |0.500|, with 411 overexpressed and 527 underexpressed in AD. Our results correlated with expression profiling in neurons from AD and ND obtained by laser capture microscopy in MTG from an independent dataset (log2 FC correlation: r = 0.504; p = 2.2e-16). Additionally, selected effects were validated by qPCR. ANOVA analysis yielded no difference between genders in response to AD, but some gender specific genes were detected (e.g., IL8 and AGRN in males, and HSPH1 and GRM1 in females). Several transcripts were associated with Braak staging (e.g., AEBP1 and DNALI1), antemortem MMSE (e.g., AEBP1 and GFAP), and tangle density (e.g., RNU1G2, and DNALI1). At the pathway level, we detected enrichment of synaptic vesicle processes and GABAergic transmission genes. Finally, applying the Weighted Correlation Network Analysis, we identified four expression modules enriched for neuronal and synaptic genes, mitochondria-associated membrane, chemical stimulus and olfactory receptor and non-coding RNA metabolism genes. Our results represent an extensive description of MTG mRNA profiling in a large sample of AD and ND. These data provide a list of genes associated with AD, and correlated to neurofibrillary tangles density. In addition, these data emphasize the importance of mitochondrial membranes and transcripts related to olfactory receptors in AD. [ABSTRACT FROM AUTHOR]

For decades scholars have engaged in a lively debate about the distinctiveness of Canadian and American value systems. Lipset (1990) argues that divergent responses to the American Revolution led Canada to embrace Tory values of ‘peace, order, and good government’ while Americans pursued classical liberal values of ‘life, liberty, and the pursuit of happiness’. Other scholars question the extent, or even the existence, of the differences Lipset describes. Using surveys of students from Canadian and American universities close to the Canada-US border, we identify compelling support for Lipset’s hypothesised differences on the role of government in the least likely setting imaginable. [ABSTRACT FROM AUTHOR]

Recent epigenetic association studies have identified a new gene, ANK1, in the pathogenesis of Alzheimer’s disease (AD). Although strong associations were observed, brain homogenates were used to generate the data, introducing complications because of the range of cell types analyzed. In order to address the issue of cellular heterogeneity in homogenate samples we isolated microglial, astrocytes and neurons by laser capture microdissection from CA1 of hippocampus in the same individuals with a clinical and pathological diagnosis of AD and matched control cases. Using this unique RNAseq data set, we show that in the hippocampus, ANK1 is significantly (p<0.0001) up-regulated 4-fold in AD microglia, but not in neurons or astrocytes from the same individuals. These data provide evidence that microglia are the source of ANK1 differential expression previously identified in homogenate samples in AD. [ABSTRACT FROM AUTHOR]

Experimental evidence has demonstrated that several aspects of adult neural stem cells (NSCs), including their quiescence, proliferation, fate specification and differentiation, are regulated by epigenetic mechanisms. These control the expression of specific sets of genes, often including those encoding for small non-coding RNAs, indicating a complex interplay between various epigenetic factors and cellular functions. Previous studies had indicated that in addition to the neuropathology in Alzheimer's disease (AD), plasticity-related changes are observed in brain areas with ongoing neurogenesis, like the hippocampus and subventricular zone. Given the role of stem cells e.g. in hippocampal functions like cognition, and given their potential for brain repair, we here review the epigenetic mechanisms relevant for NSCs and AD etiology. Understanding the molecular mechanisms involved in the epigenetic regulation of adult NSCs will advance our knowledge on the role of adult neurogenesis in degeneration and possibly regeneration in the AD brain. [ABSTRACT FROM AUTHOR]

Transcription of DNA is essential for cell maintenance and survival; inappropriate localization of proteins that are involved in transcription would be catastrophic. In Alzheimer's disease brains, and in vitro studies, we have found qualitative and quantitative deficits in transport into the nucleus of DNA methyltransferase 1 (DNMT1) and RNA polymerase II (RNA pol II), accompanied by their abnormal sequestration in the cytoplasm. RAN (RAs-related Nuclear protein) knockdown, by siRNA and oligomeric Aβ42 treatment in neurons, replicate human data which indicate that transport disruption in AD may be mechanistically linked to reduced expression of RAN, a pivotal molecule in nucleocytoplasmic transport. In vitro studies also indicate a significant role for oligomeric Aβ42 in the observed phenomena. We propose a model in which reduced transcription regulators in the nucleus and their increased presence in the cytoplasm may lead to many of the cellular manifestations of Alzheimers disease. [ABSTRACT FROM AUTHOR]

Background: This study undertakes a systematic and comprehensive analysis of brain gene expression profiles of immune/inflammation-related genes in aging and Alzheimer's disease (AD). Methods: In a well-powered microarray study of young (20 to 59 years), aged (60 to 99 years), and AD (74 to 95 years) cases, gene responses were assessed in the hippocampus, entorhinal cortex, superior frontal gyrus, and post-central gyrus. Results: Several novel concepts emerge. First, immune/inflammation-related genes showed major changes in gene expression over the course of cognitively normal aging, with the extent of gene response far greater in aging than in AD. Of the 759 immune-related probesets interrogated on the microarray, approximately 40% were significantly altered in the SFG, PCG and HC with increasing age, with the majority upregulated (64 to 86%). In contrast, far fewer immune/inflammation genes were significantly changed in the transition to AD (approximately 6% of immune-related probesets), with gene responses primarily restricted to the SFG and HC. Second, relatively few significant changes in immune/inflammation genes were detected in the EC either in aging or AD, although many genes in the EC showed similar trends in responses as in the other brain regions. Third, immune/inflammation genes undergo gender-specific patterns of response in aging and AD, with the most pronounced differences emerging in aging. Finally, there was widespread upregulation of genes reflecting activation of microglia and perivascular macrophages in the aging brain, coupled with a downregulation of select factors (TOLLIP, fractalkine) that when present curtail microglial/macrophage activation. Notably, essentially all pathways of the innate immune system were upregulated in aging, including numerous complement components, genes involved in toll-like receptor signaling and inflammasome signaling, as well as genes coding for immunoglobulin (Fc) receptors and human leukocyte antigens I and II. Conclusions: Unexpectedly, the extent of innate immune gene upregulation in AD was modest relative to the robust response apparent in the aged brain, consistent with the emerging idea of a critical involvement of inflammation in the earliest stages, perhaps even in the preclinical stage, of AD. Ultimately, our data suggest that an important strategy to maintain cognitive health and resilience involves reducing chronic innate immune activation that should be initiated in late midlife. [ABSTRACT FROM AUTHOR]

DNA methylation [1,2] is capable of modulating coordinate expression of large numbers of genes across many different pathways, and may therefore warrant investigation for their potential role between genes and disease phenotype. In a rare set of monozygotic twins discordant for Alzheimer's disease (AD), significantly reduced levels of DNA methylation were observed in temporal neocortex neuronal nuclei of the AD twin. These findings are consistent with the hypothesis that epigenetic mechanisms may mediate at the molecular level the effects of life events on AD risk, and provide, for the first time, a potential explanation for AD discordance despite genetic similarities. [ABSTRACT FROM AUTHOR]

Gene expression profiles were assessed in the hippocampus, ento-rhinal cortex, superior-frontal gyrus, and postcentral gyrus across the lifespan of 55 cognitively intact individuals aged 20-99 years. Perspectives on global gene changes that are associated with brain aging emerged, revealing two overarching concepts. First, different regions of the forebrain exhibited substantially different gene profile changes with age. For example, comparing equally powered groups, 5,029 probe sets were significantly altered with age in the superior-frontal gyrus, compared with 1,110 in the entorhinal cortex. Prominent change occurred in the sixth to seventh decades across cortical regions, suggesting that this period is a critical transition point in brain aging, particularly in males. Second, clear gender differences in brain aging were evident, suggesting that the brain undergoes sexually dimorphic changes in gene expression not only in development but also in later life. Globally across all brain regions, males showed more gene change than females. Further, Gene Ontology analysis revealed that different categories of genes were predominantly affected in males vs. females. Notably, the male brain was characterized by global decreased catabolic and anabolic capacity with aging, with down-regulated genes heavily enriched in energy production and protein synthesis/transport categories. Increased immune activation was a prominent feature of aging in both sexes, with proportionally greater activation in the female brain. These data open opportunities to explore age-dependent changes in gene expression that set the balance between neurodegeneration and compensatory mechanisms in the brain and suggest that this balance is set differently in males and females, an intriguing idea. [ABSTRACT FROM AUTHOR]

Presents information on a study which demonstrated that the enzymatic reaction in the aRNA method results in linear amplification of RNAs. Feasibility of the use of dot blots to analyze levels of aRNA; Information on the human brains used in the study; RNA profiles of individual neurons of postmortem human brains; Discussion on the results of the study.

The ability to perform highly sensitive and multiplexed in-situ protein analysis is crucial to advance our understanding of normal physiology and disease pathogenesis. To achieve this goal, we here develop an approach using cleavable biotin-conjugated antibodies and cleavable fluorescent streptavidin (CFS). In this approach, protein targets are first recognized by the cleavable biotin-labeled antibodies. Subsequently, CFS is applied to stain the protein targets. Though layer-by-layer signal amplification using cleavable biotin-conjugated orthogonal antibodies and CSF, the protein detection sensitivity can be enhanced at least 10-fold, compared with the current in-situ proteomics methods. After imaging, the fluorophore and the biotin unbound to streptavidin are removed by chemical cleavage. The leftover streptavidin is blocked by biotin. Upon reiterative analysis cycles, a large number of different proteins with a wide range of expression levels can be profiled in individual cells at the optical resolution. Applying this approach, we have demonstrated that multiple proteins are unambiguously detected in the same set of cells, regardless of the protein analysis order. We have also shown that this method can be successfully applied to quantify proteins in formalin-fixed paraffin-embedded (FFPE) tissues. [ABSTRACT FROM AUTHOR]

An equivalent circuit obtained from microwave impedance and power data is proposed for a resonant tunneling diode. The four-element circuit consists of a series R and C in parallel with a nonlinear negative differential conductance -G, the combination in turn in series with a resistor r. The resistive cut-off frequency predicted by this model depends on both the RC and RG products. Detection at THz frequencies is also explained by the model. [ABSTRACT FROM AUTHOR]

Hypertension is a major health concern in the developed world, and its prevalence increases with advancing age. The impact of hypertension on the function of the renal and cardiovascular systems is well studied; however, its influence on the brain regions important for cognition has garnered less attention. We utilized the Cyp1a1‐Ren2 xenobiotic‐inducible transgenic rat model to mimic both the age of onset and rate of induction of hypertension observed in humans. Male, 15‐month‐old transgenic rats were fed 0.15% indole‐3‐carbinol (I3C) chow to slowly induce renin‐dependent hypertension over a 6‐week period. Systolic blood pressure significantly increased, eventually reaching 200 mmHg by the end of the study period. In contrast, transgenic rats fed a control diet without I3C did not show significant changes in blood pressure (145 mmHg at the end of study). Hypertension was associated with cardiac, aortic, and renal hypertrophy as well as increased collagen deposition in the left ventricle and kidney of the I3C‐treated rats. Additionally, rats with hypertension showed reduced savings from prior spatial memory training when tested on the hippocampus‐dependent Morris swim task. Motor and sensory functions were found to be unaffected by induction of hypertension. Taken together, these data indicate a profound effect of hypertension not only on the cardiovascular‐renal axis but also on brain systems critically important for learning and memory. Future use of this model and approach may empower a more accurate investigation of the influence of aging on the systems responsible for cardiovascular, renal, and neurological health. In this manuscript we demonstrate that an increase in blood pressure ‐ driven by a genetic upregulation of the renin‐angiotensin system ‐ in a fashion that closely mimics the timing and rate of onset of hypertension in the human alters spatial memory performance in the rat. [ABSTRACT FROM AUTHOR]