Evolution of the Aging Brain Transcriptome and Synaptic Regulation

Alzheimer's disease and other neurodegenerative disorders of aging are characterized by clinical and pathological features that are relatively specific to humans. To obtain greater insight into how brain aging has evolved, we compared age-related gene expression changes in the cortex of humans, rhesus macaques, and mice on a genome-wide scale. A small subset of gene expression changes are conserved in all three species, including robust age-dependent upregulation of the neuroprotective gene apolipoprotein D (APOD) and downregulation of the synaptic cAMP signaling gene calcium/calmodulin-dependent protein kinase IV (CAMK4). However, analysis of gene ontology and cell type localization shows that humans and rhesus macaques have diverged from mice due to a dramatic increase in age-dependent repression of neuronal genes. Many of these age-regulated neuronal genes are associated with synaptic function. Notably, genes associated with GABA-ergic inhibitory function are robustly age-downregulated in humans but not in mice at the level of both mRNA and protein. Gene downregulation was not associated with overall neuronal or synaptic loss. Thus, repression of neuronal gene expression is a prominent and recently evolved feature of brain aging in humans and rhesus macaques that may alter neural networks and contribute to age-related cognitive changes

Chronic Apocynin Treatment Attenuates Beta Amyloid Plaque Size and Microglial Number in hAPP(751)SL Mice

Background: NADPH oxidase is implicated in neurotoxic microglial activation and the progressive nature of Alzheimer’s Disease (AD). Here, we test the ability of two NADPH oxidase inhibitors, apocynin and dextromethorphan (DM), to reduce learning deficits and neuropathology in transgenic mice overexpressing human amyloid precursor protein with the Swedish and London mutations (hAPP(751)SL). Methods: Four month old hAPP(751)SL mice were treated daily with saline, 15 mg/kg DM, 7.5 mg/kg DM, or 10 mg/kg apocynin by gavage for four months. Results: Only hAPP(751)SL mice treated with apocynin showed reduced plaque size and a reduction in the number of cortical microglia, when compared to the saline treated group. Analysis of whole brain homogenates from all treatments tested (saline, DM, and apocynin) demonstrated low levels of TNFa, protein nitration, lipid peroxidation, and NADPH oxidase activation, indicating a low level of neuroinflammation and oxidative stress in hAPP(751)SL mice at 8 months of age that was not significantly affected by any drug treatment. Despite in vitro analyses demonstrating that apocynin and DM ameliorate Ab-induced extracellular superoxide production and neurotoxicity, both DM and apocynin failed to significantly affect learning and memory tasks or synaptic density in hAPP(751)SL mice. To discern how apocynin was affecting plaque levels (plaque load) and microglial number in vivo, in vitro analysis of microglia was performed, revealing no apocynin effects on beta-amyloid (Ab) phagocytosis, microglial proliferation, or microglial survival. Conclusions: Together, this study suggests that while hAPP(751)SL mice show increases in microglial number and plaque load, they fail to exhibit elevated markers of neuroinflammation consistent with AD at 8 months of age, which may be a limitation of this animal model. Despite absence of clear neuroinflammation, apocynin was still able to reduce both plaque size and microglial number, suggesting that apocynin may have additional therapeutic effects independent of anti-inflammatory characteristics

Expression of Nestin by Neural Cells in the Adult Rat and Human Brain

Neurons and glial cells in the developing brain arise from neural progenitor cells (NPCs). Nestin, an intermediate filament protein, is thought to be expressed exclusively by NPCs in the normal brain, and is replaced by the expression of proteins specific for neurons or glia in differentiated cells. Nestin expressing NPCs are found in the adult brain in the subventricular zone (SVZ) of the lateral ventricle and the subgranular zone (SGZ) of the dentate gyrus. While significant attention has been paid to studying NPCs in the SVZ and SGZ in the adult brain, relatively little attention has been paid to determining whether nestin-expressing neural cells (NECs) exist outside of the SVZ and SGZ. We therefore stained sections immunocytochemically from the adult rat and human brain for NECs, observed four distinct classes of these cells, and present here the first comprehensive report on these cells. Class I cells are among the smallest neural cells in the brain and are widely distributed. Class II cells are located in the walls of the aqueduct and third ventricle. Class IV cells are found throughout the forebrain and typically reside immediately adjacent to a neuron. Class III cells are observed only in the basal forebrain and closely related areas such as the hippocampus and corpus striatum. Class III cells resemble neurons structurally and co-express markers associated exclusively with neurons. Cell proliferation experiments demonstrate that Class III cells are not recently born. Instead, these cells appear to be mature neurons in the adult brain that express nestin. Neurons that express nestin are not supposed to exist in the brain at any stage of development. That these unique neurons are found only in brain regions involved in higher order cognitive function suggests that they may be remodeling their cytoskeleton in supporting the neural plasticity required for these functions

Real-Time Imaging and Quantification of Amyloid-β Peptide Aggregates by Novel Quantum-Dot Nanoprobes

Background: Protein aggregation plays a major role in the pathogenesis of neurodegenerative disorders, such as Alzheimer’s disease. However, direct real-time imaging of protein aggregation, including oligomerization and fibrillization, has never been achieved. Here we demonstrate the preparation of fluorescent semiconductor nanocrystal (quantum dot; QD)-labeled amyloid-b peptide (QDAb) and its advanced applications. Methodology/Principal Findings: The QDAb construct retained Ab oligomer-forming ability, and the sizes of these oligomers could be estimated from the relative fluorescence intensities of the imaged spots. Both QDAb coaggregation with intact Ab42 and insertion into fibrils were detected by fluorescence microscopy. The coaggregation process was observed by real-time 3D imaging using slit-scanning confocal microscopy, which showed a typical sigmoid curve with 1.5 h in the lag-time and 12 h until saturation. Inhibition of coaggregation using an anti-Ab antibody can be observed as 3D images on a microscopic scale. Microglia ingested monomeric QDAb more significantly than oligomeric QDAb, and the ingested QDAb was mainly accumulated in the lysosome. Conclusions/Significance: These data demonstrate that QDAb is a novel nanoprobe for studying Ab oligomerization an

Kihi-to, a herbal traditional medicine, improves Abeta(25–35)-induced memory impairment and losses of neurites and synapses

<p>Abstract</p> <p>Background</p> <p>We previously hypothesized that achievement of recovery of brain function after the injury requires the reconstruction of neuronal networks, including neurite regeneration and synapse reformation. Kihi-to is composed of twelve crude drugs, some of which have already been shown to possess neurite extension properties in our previous studies. The effect of Kihi-to on memory deficit has not been examined. Thus, the goal of the present study is to determine the <it>in vivo </it>and <it>in vitro </it>effects of Kihi-to on memory, neurite growth and synapse reconstruction.</p> <p>Methods</p> <p>Effects of Kihi-to, a traditional Japanese-Chinese traditional medicine, on memory deficits and losses of neurites and synapses were examined using Alzheimer's disease model mice. Improvements of Aβ(25–35)-induced neuritic atrophy by Kihi-to and the mechanism were investigated in cultured cortical neurons.</p> <p>Results</p> <p>Administration of Kihi-to for consecutive 3 days resulted in marked improvements of Aβ(25–35)-induced impairments in memory acquisition, memory retention, and object recognition memory in mice. Immunohistochemical comparisons suggested that Kihi-to attenuated neuritic, synaptic and myelin losses in the cerebral cortex, hippocampus and striatum. Kihi-to also attenuated the calpain increase in the cerebral cortex and hippocampus. When Kihi-to was added to cells 4 days after Aβ(25–35) treatment, axonal and dendritic outgrowths in cultured cortical neurons were restored as demonstrated by extended lengths of phosphorylated neurofilament-H (P-NF-H) and microtubule-associated protein (MAP)2-positive neurites. Aβ(25–35)-induced cell death in cortical culture was also markedly inhibited by Kihi-to. Since NF-H, MAP2 and myelin basic protein (MBP) are substrates of calpain, and calpain is known to be involved in Aβ-induced axonal atrophy, expression levels of calpain and calpastatin were measured. Treatment with Kihi-to inhibited the Aβ(25–35)-evoked increase in the calpain level and decrease in the calpastatin level. In addition, Kihi-to inhibited Aβ(25–35)-induced calcium entry.</p> <p>Conclusion</p> <p>In conclusion Kihi-to clearly improved the memory impairment and losses of neurites and synapses.</p

Molecular Insights into the Pathogenesis of Alzheimer's Disease and Its Relationship to Normal Aging

Alzheimer's disease (AD) is a complex neurodegenerative disorder that diverges from the process of normal brain aging by unknown mechanisms. We analyzed the global structure of age- and disease-dependent gene expression patterns in three regions from more than 600 brains. Gene expression variation could be almost completely explained by four transcriptional biomarkers that we named BioAge (biological age), Alz (Alzheimer), Inflame (inflammation), and NdStress (neurodegenerative stress). BioAge captures the first principal component of variation and includes genes statistically associated with neuronal loss, glial activation, and lipid metabolism. Normally BioAge increases with chronological age, but in AD it is prematurely expressed as if some of the subjects were 140 years old. A component of BioAge, Lipa, contains the AD risk factor APOE and reflects an apparent early disturbance in lipid metabolism. The rate of biological aging in AD patients, which cannot be explained by BioAge, is associated instead with NdStress, which includes genes related to protein folding and metabolism. Inflame, comprised of inflammatory cytokines and microglial genes, is broadly activated and appears early in the disease process. In contrast, the disease-specific biomarker Alz was selectively present only in the affected areas of the AD brain, appears later in pathogenesis, and is enriched in genes associated with the signaling and cell adhesion changes during the epithelial to mesenchymal (EMT) transition. Together these biomarkers provide detailed description of the aging process and its contribution to Alzheimer's disease progression

Influence of the Stability of a Fused Protein and Its Distance to the Amyloidogenic Segment on Fibril Formation

Conversion of native proteins into amyloid fibrils is irreversible and therefore it is difficult to study the interdependence of conformational stability and fibrillation by thermodynamic analyses. Here we approached this problem by fusing amyloidogenic poly-alanine segments derived from the N-terminal domain of the nuclear poly (A) binding protein PABPN1 with a well studied, reversibly unfolding protein, CspB from Bacillus subtilis. Earlier studies had indicated that CspB could maintain its folded structure in fibrils, when it was separated from the amyloidogenic segment by a long linker. When CspB is directly fused with the amyloidogenic segment, it unfolds because its N-terminal chain region becomes integrated into the fibrillar core, as shown by protease mapping experiments. Spacers of either 3 or 16 residues between CspB and the amyloidogenic segment were not sufficient to prevent this loss of CspB structure. Since the low thermodynamic stability of CspB (ΔGD = 12.4 kJ/mol) might be responsible for unfolding and integration of CspB into fibrils, fusions with a CspB mutant with enhanced thermodynamic stability (ΔGD = 26.9 kJ/mol) were studied. This strongly stabilized CspB remained folded and prevented fibril formation in all fusions. Our data show that the conformational stability of a linked, independently structured protein domain can control fibril formation

Genome-wide association reveals genetic effects on human Aβ₄₂and τ protein levels in cerebrospinal fluids: a case control study

<p>Abstract</p> <p>Background</p> <p>Alzheimer's disease (AD) is common and highly heritable with many genes and gene variants associated with AD in one or more studies, including APOE ε2/ε3/ε4. However, the genetic backgrounds for normal cognition, mild cognitive impairment (MCI) and AD in terms of changes in cerebrospinal fluid (CSF) levels of Aβ_1-42, T-tau, and P-tau_181P, have not been clearly delineated. We carried out a genome-wide association study (GWAS) in order to better define the genetic backgrounds to these three states in relation to CSF levels.</p> <p>Methods</p> <p>Subjects were participants in the Alzheimer's Disease Neuroimaging Initiative (ADNI). The GWAS dataset consisted of 818 participants (mainly Caucasian) genotyped using the Illumina Human Genome 610 Quad BeadChips. This sample included 410 subjects (119 Normal, 115 MCI and 176 AD) with measurements of CSF Aβ_1-42, T-tau, and P-tau_181PLevels. We used PLINK to find genetic associations with the three CSF biomarker levels. Association of each of the 498,205 SNPs was tested using additive, dominant, and general association models while considering APOE genotype and age. Finally, an effort was made to better identify relevant biochemical pathways for associated genes using the ALIGATOR software.</p> <p>Results</p> <p>We found that there were some associations with APOE genotype although CSF levels were about the same for each subject group; CSF Aβ_1-42levels decreased with APOE gene dose for each subject group. T-tau levels tended to be higher among AD cases than among normal subjects. From adjusted result using APOE genotype and age as covariates, no SNP was associated with CSF levels among AD subjects. <it>CYP19A1 </it>'aromatase' (rs2899472), <it>NCAM2</it>, and multiple SNPs located on chromosome 10 near the <it>ARL5B </it>gene demonstrated the strongest associations with Aβ_1-42in normal subjects. Two genes found to be near the top SNPs, <it>CYP19A1 </it>(rs2899472, p = 1.90 × 10^-7) and <it>NCAM2 </it>(rs1022442, p = 2.75 × 10^-7) have been reported as genetic factors related to the progression of AD from previous studies. In AD subjects, APOE ε2/ε3 and ε2/ε4 genotypes were associated with elevated T-tau levels and ε4/ε4 genotype was associated with elevated T-tau and P-tau_181Plevels. Pathway analysis detected several biological pathways implicated in Normal with CSF β-amyloid peptide (Aβ_1-42).</p> <p>Conclusions</p> <p>Our genome-wide association analysis identified several SNPs as important factors for CSF biomarker. We also provide new evidence for additional candidate genetic risk factors from pathway analysis that can be tested in further studies.</p

An Epigenetic Blockade of Cognitive Functions in the Neurodegenerating Brain

Cognitive decline is a debilitating feature of most neurodegenerative diseases of the central nervous system, including Alzheimer’s disease. The causes leading to such impairment are only poorly understood and effective treatments are slow to emerge. Here we show that cognitive capacities in the neurodegenerating brain are constrained by an epigenetic blockade of gene transcription that is potentially reversible. This blockade is mediated by histone deacetylase 2, which is increased by Alzheimer’s-disease-related neurotoxic insults in vitro, in two mouse models of neurodegeneration and in patients with Alzheimer’s disease. Histone deacetylase 2 associates with and reduces the histone acetylation of genes important for learning and memory, which show a concomitant decrease in expression. Importantly, reversing the build-up of histone deacetylase 2 by short-hairpin-RNA-mediated knockdown unlocks the repression of these genes, reinstates structural and synaptic plasticity, and abolishes neurodegeneration-associated memory impairments. These findings advocate for the development of selective inhibitors of histone deacetylase 2 and suggest that cognitive capacities following neurodegeneration are not entirely lost, but merely impaired by this epigenetic blockade

Association of Plasma Aß Peptides with Blood Pressure in the Elderly

Background Aß peptides are often considered as catabolic by-products of the amyloid ß protein precursor (APP), with unknown physiological functions. However, several biological properties have been tentatively attributed to these peptides, including a role in vasomotion. We assess whether plasma Aß peptide levels might be associated with systolic and diastolic blood pressure values (SBP and DBP, respectively). Methodology/Principal Findings Plasma Aß1-40 and Aß1-42 levels were measured using an xMAP-based assay in 1,972 individuals (none of whom were taking antihypertensive drugs) from 3 independent studies: the French population-based 3C and MONA-LISA (Lille) studies (n = 627 and n = 769, respectively) and the Australian, longitudinal AIBL study (n = 576). In the combined sample, the Aß1-42/ Aß1-40 ratio was significantly and inversely associated with SBP (p = 0.03) and a similar trend was observed for DBP (p = 0.06). Using the median age (69) as a cut-off, the Aß1-42/Aß1-40 ratio was strongly associated with both SBP and DBP in elderly individuals (p = 0.002 and p = 0.03, respectively). Consistently, a high Aß1-42/ Aß1-40 ratio was associated with a lower risk of hypertension in both the combined whole sample (odds ratio [OR], 0.71; 95% confidence interval [CI], 0.56-0.90) and (to an even greater extent) in the elderly subjects (OR, 0.53; 95% CI, 0.37–0.75). Lastly, all these associations appeared to be primarily driven by the level of plasma Aß1-40. Conclusion The plasma Aß1-42/Aß1-40 ratio is inversely associated with SBP, DBP and the risk of hypertension in elderly subjects, suggesting that Aß peptides affect blood pressure in vivo. These results may be particularly relevant in Alzheimer\u27s disease, in which a high Aß1-42/Aß1-40 plasma ratio is reportedly associated with a decreased risk of incident disease