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Alzheimer’s disease (AD) is a tragic neurodegenerative disease affecting more than 5 million Americans. Circadian disruptions impact nearly all AD patients, with reversal of sleep/wake cycles and agitation in the evening being common disturbances that manifest early in disease. These alterations support a role for circadian dysfunction as a driver of AD, emphasizing a critical need to investigate the therapeutic potential of circadian-modulating interventions. One of the most powerful regulators of the circadian system is the daily feed/fast cycle. Here we show that time-restricted feeding (TRF) without caloric restriction, improved key disease components including behavior, disease pathology and transcription in the APP23 mouse model of Alzheimer’s disease. We found that TRF had the remarkable capability of simultaneously reducing amyloid deposition, increasing Aβ42 clearance, improving sleep and hyperactivity, and normalizing transcription of circadian, AD and neuroinflammation-associated genes in APP23 mice. Thus, our study unveils for the first time that circadian modulation through timed feeding has far-reaching effects beyond metabolism and affects the brain as the substrate for neurodegeneration. Since the pleiotropic effects of TRF can substantially modify disease trajectory, this intervention has immediate translational value, addressing the crucial need for accessible approaches to reduce or halt AD progression.

Multiple System Atrophy is a rare neurodegenerative disease with alpha-synuclein aggregation in glial cytoplasmic inclusions and either predominant olivopontocerebellar atrophy or striatonigral degeneration, leading to dysautonomia, parkinsonism, and cerebellar ataxia. One prior genome-wide association study in mainly clinically diagnosed patients with Multiple System Atrophy failed to identify genetic variants predisposing for the disease. Since the clinical diagnosis of Multiple System Atrophy yields a high rate of misdiagnosis when compared to the neuropathological gold standard, we studied common genetic variation in only autopsy-confirmed cases (N = 731) and controls (N = 2,898).The most strongly disease-associated markers were rs16859966 on chromosome 3 (P = 8.6 × 10−7, odds ratio (OR) = 1.58, [95% confidence interval (CI) = 1.32-1.89]), rs7013955 on chromosome 8 (P = 3.7 × 10−6, OR = 1.8 [1.40-2.31]), and rs116607983 on chromosome 4 (P = 4.0 × 10−6, OR = 2.93 [1.86-4.63]), all of which were supported by at least one additional genotyped and several imputed single nucleotide polymorphisms with P-values below 5 × 10−5. The genes closest to the chromosome 3 locus are ZIC1 and ZIC4 encoding the zinc finger proteins of cerebellum 1 and 4 (ZIC1 and ZIC4).Since mutations of ZIC1 and ZIC4 and paraneoplastic autoantibodies directed against ZIC4 are associated with severe cerebellar dysfunction, we conducted immunohistochemical analyses in brain tissue of the frontal cortex and the cerebellum from 24 Multiple System Atrophy patients. Strong immunohistochemical expression of ZIC4 was detected in a subset of neurons of the dentate nucleus in all healthy controls and in patients with striatonigral degeneration, whereas ZIC4 positive neurons were significantly reduced in patients with olivopontocerebellar atrophy.These findings point to a potential ZIC4-mediated vulnerability of neurons in Multiple System Atrophy.

Multiple systems atrophy (MSA) is a rare neurodegenerative disorder characterized by the accumulation of α-synuclein in glial cells and neurodegeneration in the striatum, substantia nigra, and cerebellum. Aberrant miRNA regulation has been associated with neurodegeneration, including alterations of specific miRNAs in brain tissue, serum, and cerebrospinal fluid from MSA patients. Still, a causal link between deregulation of miRNA networks and pathological changes in the transcriptome remains elusive. We profiled ~ 800 miRNAs in the striatum of MSA patients in comparison to healthy individuals to identify specific miRNAs altered in MSA. In addition, we performed a parallel screening of 700 transcripts associated with neurodegeneration to determine the impact of miRNA deregulation on the transcriptome. We identified 60 miRNAs with abnormal levels in MSA brains that are involved in extracellular matrix receptor interactions, prion disease, inflammation, ubiquitin-mediated proteolysis, and addiction pathways. Using the correlation between miRNA expression and the abundance of their known targets, miR-124-3p, miR-19a-3p, miR-27b-3p, and miR-29c-3p were identified as key regulators altered in MSA, mainly contributing to neuroinflammation. Finally, our study also uncovered a potential link between Alzheimer's disease (AD) and MSA pathologies that involves miRNAs and deregulation of BACE1. Our results provide a comprehensive appraisal of miRNA alterations in MSA and their effect on the striatal transcriptome, supporting that aberrant miRNA expression is highly correlated with changes in gene transcription associated with MSA neuropathology, in particular those driving inflammation, disrupting myelination, and potentially impacting α-synuclein accumulation via deregulation of autophagy and prion mechanisms.

Introduction Dementia with Lewy bodies (DLB) and Parkinson's disease dementia (PDD) are characterized by cognitive alterations, visual hallucinations, and motor impairment. Diagnosis is based on type and timing of clinical manifestations; however, determination of clinical subtypes is challenging. The utility of blood DNA methylation as a biomarker for Lewy body disorders (LBD) is mostly unexplored. Methods We performed a cross‐sectional analysis of blood methylation in 42 DLB and 50 PDD cases applying linear models to compare groups and logistic least absolute shrinkage and selection operator regression to explore the discriminant power of methylation signals. Results DLB blood shows differential methylation compared to PDD. Some methylation changes associate with core features of LBD. Sets of probes show high predictive value to discriminate between variants. Discussion Our study is the first to explore LBD blood methylation. Despite overlapping clinical presentation, we detected differential epigenetic signatures that, if confirmed in independent cohorts, could be developed into useful biomarkers.

Recent discoveries demonstrate a critical role for circadian rhythms and sleep in immune system homeostasis. Both innate and adaptive immune responses - ranging from leukocyte mobilization, trafficking, and chemotaxis to cytokine release and T cell differentiation -are mediated in a time of day-dependent manner. The National Institutes of Health (NIH) recently sponsored an interdisciplinary workshop, "Sleep Insufficiency, Circadian Misalignment, and the Immune Response," to highlight new research linking sleep and circadian biology to immune function and to identify areas of high translational potential. This Review summarizes topics discussed and highlights immediate opportunities for delineating clinically relevant connections among biological rhythms, sleep, and immune regulation.

Prenatal stress (PS) impacts early postnatal behavioural and cognitive development. This process of 'fetal programming' is mediated by the effects of the prenatal experience on the developing hypothalamic-pituitary-adrenal (HPA) axis and autonomic nervous system (ANS). The HPA axis is a dynamic system regulating homeostasis, especially the stress response, and is highly sensitive to adverse early life experiences. We review the evidence for the effects of PS on fetal programming of the HPA axis and the ANS. We derive a multi-scale multi-species approach to devising preclinical and clinical studies to identify early non-invasively available pre- and postnatal biomarkers of these programming effects. Such approach would identify adverse postnatal brain developmental trajectories, a prerequisite for designing therapeutic interventions. The multiple scales include the biomarkers reflecting changes in the brain epigenome, metabolome, microbiome and the ANS activity gauged via an array of advanced non-invasively obtainable properties of fetal heart rate fluctuations. The proposed framework has the potential to reveal mechanistic links between maternal stress during pregnancy and changes across these physiological scales. Such biomarkers may hence be useful as early and non-invasive predictors of neurodevelopmental trajectories influenced by the PS. We conclude that studies into PS effects must be conducted on multiple scales derived from concerted observations in multiple animal models and human cohorts performed in an interactive and iterative manner and deploying machine learning for data synthesis, identification and validation of the best non-invasive biomarkers., Comment: Focused review, 13 pages, 5 figures

Introduction Circadian alterations are prevalent in Alzheimer's disease (AD) and may contribute to cognitive impairment, behavioral symptoms, and neurodegeneration. Epigenetic mechanisms regulate the circadian clock, and changes in DNA methylation have been reported in AD brains, but the pathways that mediate circadian deregulation in AD are incompletely understood. We hypothesized that aberrant DNA methylation may affect circadian rhythms in AD. Methods We investigated DNA methylation, transcription, and expression of BMAL1 , a positive regulator of the circadian clock, in cultured fibroblasts and brain samples from two independent cohorts of aging and AD. Results DNA methylation modulated rhythmic expression of clock genes in cultured fibroblasts. Moreover, rhythmic methylation of BMAL1 was altered in AD brains and fibroblasts and correlated with transcription cycles. Discussion Our results indicate that cycles of DNA methylation contribute to the regulation of BMAL1 rhythms in the brain. Hence, aberrant epigenetic patterns may be linked to circadian alterations in AD.

Huntington’s disease (HD) is an autosomal-dominant neurodegenerative movement disorder that presents with prominent cognitive and psychiatric dysfunction. Brain-derived neurotrophic factor (BDNF) plays an important role in the pathophysiology of HD, as well as other neurodegenerative and psychiatric disorders, and epigenetic alterations in the complex BDNF promoter have been associated with its deregulation in pathological conditions. BDNF has gained increased attention as a potential biomarker of disease; but currently, the conflicting results from measurements of BDNF in different biofluids difficult the assessment of its utility as a biomarker for HD. Here, we measured BDNF protein levels in plasma (n = 85) and saliva (n = 81) samples from premanifest and manifest HD patients and normal controls using ELISA assays. We further examined DNA methylation levels of BDNF promoter IV using DNA derived from whole blood of HD patients and healthy controls (n = 40) using pyrosequencing. BDNF protein levels were not significantly different in plasma samples across diagnostic groups. Plasma BDNF was significantly correlated with age in control subjects but not in HD patients, nor were significant gender effects observed. Similar to plasma, salivary BDNF was correlated with age only in control subjects, with no gender effects observed. Importantly, we detected significantly lower levels of salivary BDNF in premanifest and manifest HD patients compared to control subjects, with lower BDNF levels being observed in premanifest patients within a predicted 10 years to disease onset. Salivary and plasma BDNF levels were not significantly correlated with one another, suggesting different origins. DNA methylation at four out of the 12 CpG sites studied in promoter IV were significantly altered in HD patients in comparison to controls. Interestingly, methylation at three of these CpG sites was inversely correlated to the Hospital Anxiety and Depression Scale (HADS) scores. BDNF promoter methylation was not correlated with motor or cognitive scores in HD patients, and was not associated with sex or age in neither disease nor control groups. Conclusion: Our studies show that BDNF protein levels are decreased in saliva; and BDNF promoter methylation increased in blood in HD subjects when compared to controls. These findings suggest that salivary BDNF measures may represent an early marker of disease onset and DNA methylation at the BDNF promoter IV, could represent a biomarker of psychiatric symptoms in HD patients.

Parkinson’s Disease (PD) is a common neurodegenerative disorder currently diagnosed based on the presentation of characteristic movement symptoms. Unfortunately, patients exhibiting these symptoms have already undergone significant dopaminergic neuronal loss. Earlier diagnosis, aided by molecular biomarkers specific to PD, would improve overall patient care. Epigenetic mechanisms, which are modified by both environment and disease pathophysiology, are emerging as important components of neurodegeneration. Alterations to the PD methylome have been reported in epigenome-wide association studies. However, the extent to which methylation changes correlate with disease progression has not yet been reported; nor the degree to which methylation is affected by PD medication. We performed a longitudinal genome-wide methylation study surveying ~850,000 CpG sites in whole blood from 189 well-characterized PD patients and 191 control individuals obtained at baseline and at a follow-up visit ~2 y later. We identified distinct patterns of methylation in PD cases versus controls. Importantly, we identified genomic sites where methylation changes longitudinally as the disease progresses. Moreover, we identified methylation changes associated with PD pathology through the analysis of PD cases that were not exposed to anti-parkinsonian therapy. In addition, we identified methylation sites modulated by exposure to dopamine replacement drugs. These results indicate that DNA methylation is dynamic in PD and changes over time during disease progression. To the best of our knowledge, this is the first longitudinal epigenome-wide methylation analysis for Parkinson’s disease and reveals changes associated with disease progression and in response to dopaminergic medications in the blood methylome.

We review evidence supporting the role of early life programming in the susceptibility for adult neurodegenerative diseases while highlighting questions and proposing avenues for future research to advance our understanding of this fundamental process. The key elements of this phenomenon are chronic stress, neuroinflammation triggering microglial polarization, microglial memory and their connection to neurodegeneration. We review the mediating mechanisms which may function as early biomarkers of increased susceptibility for neurodegeneration. Can we devise novel early life modifying interventions to steer developmental trajectories to their optimum?. Fil: Desplats, Paula. University of California; Estados Unidos Fil: Gutierrez, Ashley M.. University of California; Estados Unidos Fil: Antonelli, Marta Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Biología Celular y Neurociencia "Prof. Eduardo de Robertis". Universidad de Buenos Aires. Facultad de Medicina. Instituto de Biología Celular y Neurociencia; Argentina Fil: Frasch, Martin G.. University of Washington; Estados Unidos

Neuroinflammation is a common pathological correlate of HIV-associated neurocognitive disorders (HAND) in individuals on antiretroviral therapy (ART). Triggering receptor expressed on myeloid cells 2 (TREM2) regulates neuroinflammation, clears extracellular Amyloid (A)-β, surveys for damaged neurons, and orchestrates microglial differentiation. TREM2 has not been studied in HIV+ brain tissues. In this retrospective study, we investigated TREM2 expression levels and localization to microglia, Aβ protein levels, and tumor necrosis factor (TNF)-α transcript levels in the frontal cortices of 52 HIV+ decedents. All donors had been on ART; 14 were cognitively normal (CN), 17 had an asymptomatic neurocognitive impairment (ANI), and 21 had a minor neurocognitive disorder (MND). Total TREM2 protein levels were increased in the soluble and decreased in the membrane-enriched fractions of MND brain tissues compared to CN; however, brains from MND Hispanics showed the most robust alterations in TREM2 as well as significantly increased TNF-α mRNA and Aβ levels when compared to CN Hispanics. Significant alterations in the expression of total TREM2 protein and transcripts for TNF-α were not observed in non-Hispanics, despite higher levels of Aβ in the non-Hispanic CN group compared to the non-Hispanic MND groups. These findings show that decreased and increased TREM2 in membrane-bound fractions and in soluble-enriched fractions, respectively, is associated with increased Aβ and neuroinflammation in this cohort of HIV+ brains, particularly those identifying as Hispanics. These findings suggest a role for TREM2 in the brain of HIV+ individuals may deserve more investigation as a biomarker for HAND and as a possible therapeutic target. OPEN SCIENCE BADGES: This article has received a badge for *Open Materials* because it provided all relevant information to reproduce the study in the manuscript. The complete Open Science Disclosure form for this article can be found at the end of the article. More information about the Open Practices badges can be found at https://cos.io/our-services/open-science-badges/.

Insulin and its receptor are widely expressed in a variety of tissues throughout the body including liver, adipose tissue, liver and brain. The insulin receptor is expressed as two functionally distinct isoforms, differentiated by a single 12 amino acid exon. The two receptor isoforms, designated IR/A and IR/B, are expressed in a highly tissue and cell specific manner and relative proportions of the different isoforms vary during development, aging and disease states. The high degree of similarity between the two isoforms has prevented detailed studies as differentiation of the two isoforms by traditional immunological methods cannot be achieved. We describe here a new in situ RT-PCR/ FISH assay that allows for the visualization of IR/A and IR/B in tissue along with tissue specific markers. We used this new method to show for the first time that IR/A and IR/B are both expressed in neurons in the adult human brain. Thus, we present a method that enables the investigation of IR/A and IR/B insulin receptor isoform expression in situ in various tissues.

Neural stem cells (NSCs) have been considered as potential therapy in Alzheimer's disease (AD) but their use is hampered by the poor survival of grafted cells. Supply of neurotrophic factors to the grafted cells has been proposed as a way to augment survival of the stem cells. In this context, we investigated the utility of Cerebrolysin (CBL), a peptidergic mixture with neurotrophic-like properties, as an adjunct to stem cell therapy in an APP transgenic (tg) model of AD. We grafted murine NSCs into the hippocampus of non-tg and APP tg that were treated systemically with CBL and analyzed after 1, 3, 6 and 9months post grafting. Compared to vehicle-treated non-tg mice, in the vehicle-treated APP tg mice there was considerable reduction in the survival of the grafted NSCs. Whereas, CBL treatment enhanced the survival of NSCs in both non-tg and APP tg with the majority of the surviving NSCs remaining as neuroblasts. The NSCs of the CBL treated mice displayed reduced numbers of caspase-3 and TUNEL positive cells and increased brain derived neurotrophic factor (BDNF) and furin immunoreactivity. These results suggest that CBL might protect grafted NSCs and as such be a potential adjuvant therapy when combined with grafting.

Multiple system atrophy (MSA) is a fatal neurodegenerative disorder characterized by the pathological accumulation of alpha-synuclein (α-syn) in oligodendrocytes. Therapeutic efforts to stop or delay the progression of MSA have yielded suboptimal results in clinical trials, and there are no efficient treatments currently available for MSA patients. We hypothesize that combining therapies targeting different aspects of the disease may lead to better clinical outcomes. To test this hypothesis, we combined the use of a single-chain antibody targeting α-syn modified for improved central nervous system penetration (CD5-D5) with an unconventional anti-inflammatory treatment (lenalidomide) in the myelin basic protein (MBP)-α-syn transgenic mouse model of MSA. While the use of either CD5-D5 or lenalidomide alone had positive effects on neuroinflammation and/or α-syn accumulation in this mouse model of MSA, the combination of both approaches yielded better results than each single treatment. The combined treatment reduced astrogliosis, microgliosis, soluble and aggregated α-syn levels, and partially improved behavioral deficits in MBP-α-syn transgenic mice. These effects were associated with an activation of the Akt signaling pathway, which may mediate cytoprotective effects downstream tumor necrosis factor alpha (TNFα). These results suggest that a strategic combination of treatments may improve the therapeutic outcome in trials for MSA and related neurodegenerative disorders. Electronic supplementary material The online version of this article (doi:10.1186/s40478-016-0409-1) contains supplementary material, which is available to authorized users.

Parkinson's disease and dementia with Lewy bodies are neurodegenerative disorders characterized by accumulation of α-synuclein (α-syn). Recently, single-chain fragment variables (scFVs) have been developed against individual conformational species of α-syn. Unlike more traditional monoclonal antibodies, these scFVs will not activate or be endocytosed by Fc receptors. For this study, we investigated an scFV directed against oligomeric α-syn fused to the LDL receptor-binding domain from apolipoprotein B (apoB). The modified scFV showed enhanced brain penetration and was imported into neuronal cells through the endosomal sorting complex required for transport (ESCRT) pathway, leading to lysosomal degradation of α-syn aggregates. Further analysis showed that the scFV was effective at ameliorating neurodegenerative pathology and behavioral deficits observed in the mouse model of dementia with Lewy bodies/Parkinson's disease. Thus, the apoB modification had the effect of both increasing accumulation of the scFV in the brain and directing scFV/α-syn complexes for degradation through the ESCRT pathway, leading to improved therapeutic potential of immunotherapy.

Abnormal accumulation and propagation of the neuronal protein α-synuclein has been hypothesized to underlie the pathogenesis of Parkinson's disease, dementia with Lewy bodies and multiple system atrophy. Here we report a de novo-developed compound (NPT100-18A) that reduces α-synuclein toxicity through a novel mechanism that involves displacing α-synuclein from the membrane. This compound interacts with a domain in the C-terminus of α-synuclein. The E83R mutation reduces the compound interaction with the 80-90 amino acid region of α-synuclein and prevents the effects of NPT100-18A. In vitro studies showed that NPT100-18A reduced the formation of wild-type α-synuclein oligomers in membranes, reduced the neuronal accumulation of α-synuclein, and decreased markers of cell toxicity. In vivo studies were conducted in three different α-synuclein transgenic rodent models. Treatment with NPT100-18A ameliorated motor deficits in mThy1 wild-type α-synuclein transgenic mice in a dose-dependent manner at two independent institutions. Neuropathological examination showed that NPT100-18A decreased the accumulation of proteinase K-resistant α-synuclein aggregates in the CNS and was accompanied by the normalization of neuronal and inflammatory markers. These results were confirmed in a mutant line of α-synuclein transgenic mice that is prone to generate oligomers. In vivo imaging studies of α-synuclein-GFP transgenic mice using two-photon microscopy showed that NPT100-18A reduced the cortical synaptic accumulation of α-synuclein within 1 h post-administration. Taken together, these studies support the notion that altering the interaction of α-synuclein with the membrane might be a feasible therapeutic approach for developing new disease-modifying treatments of Parkinson's disease and other synucleinopathies.

Objective:We aimed to investigate whether HIV latency in the CNS might have adverse molecular, pathologic, and clinical consequences.Methods:This was a case-control comparison of HIV-1 seropositive (HIV+) patients with clinical and neuropathologic examination. Based on the levels of HIV-1 DNA, RNA, and p24 in the brain, cases were classified as controls, latent HIV CNS infection, and HIV encephalitis (HIVE). Analysis of epigenetic markers including BCL11B, neurodegeneration, and neuroinflammation was performed utilizing immunoblot, confocal microscopy, immunochemistry/image analysis, and qPCR. Detailed antemortem neurocognitive data were available for 23 out of the 32 cases.Results:HIV+ controls (n = 12) had no detectable HIV-1 DNA, RNA, or p24 in the CNS; latent HIV+ cases (n = 10) showed high levels of HIV-1 DNA but no HIV RNA or p24; and HIVE cases (n = 10) had high levels of HIV-1 DNA, RNA, and p24. Compared to HIV+ controls, the HIV+ latent cases displayed moderate cognitive impairment with neurodegenerative and neuroinflammatory alterations, although to a lesser extent than HIVE cases. Remarkably, HIV+ latent cases showed higher levels of BCL11B and other chromatin modifiers involved in silencing. Increased BCL11B was associated with deregulation of proinflammatory genes like interleukin-6, tumor necrosis factor–α, and CD74.Conclusion:Persistence of latent HIV-1 infection in the CNS was associated with increased levels of chromatin modifiers, including BCL11B. Alteration of these epigenetic factors might result in abnormal transcriptomes, leading to inflammation, neurodegeneration, and neurocognitive impairment. BCL11B and other epigenetic factors involved in silencing might represent potential targets for HIV-1 involvement of the CNS.

UnlabelledAlzheimer's disease (AD) is characterized by the progressive accumulation of amyloid β (Aβ) and microtubule associate protein tau, leading to the selective degeneration of neurons in the neocortex, limbic system, and nucleus basalis, among others. Recent studies have shown that α-synuclein (α-syn) also accumulates in the brains of patients with AD and interacts with Aβ and tau, forming toxic hetero-oligomers. Although the involvement of α-syn has been investigated extensively in Lewy body disease, less is known about the role of this synaptic protein in AD. Here, we found that reducing endogenous α-syn in an APP transgenic mouse model of AD prevented the degeneration of cholinergic neurons, ameliorated corresponding deficits, and recovered the levels of Rab3a and Rab5 proteins involved in intracellular transport and sorting of nerve growth factor and brain-derived neurotrophic factor. Together, these results suggest that α-syn might participate in mechanisms of vulnerability of selected neuronal populations in AD and that reducing α-syn might be a potential approach to protecting these populations from the toxic effects of Aβ.Significance statementReducing endogenous α-synuclein (α-syn) in an APP transgenic mouse model of Alzheimer's disease (AD) prevented the degeneration of cholinergic neurons, ameliorated corresponding deficits, and recovered the levels of Rab3a and Rab5 proteins involved in intracellular transport and sorting of nerve growth factor and brain-derived neurotrophic factor. These results suggest that α-syn might participate in mechanisms of vulnerability of selected neuronal populations in AD and that reducing α-syn might be a potential approach to protecting these populations from the toxic effects of amyloid β.

Background HIV-associated neurocognitive disorders (HAND) continue to be a common morbidity associated with chronic HIV infection. It has been shown that HIV proteins (e.g., gp120) released from infected microglial/macrophage cells can cause neuronal damage by triggering inflammation and oxidative stress, activating aberrant kinase pathways, and by disrupting mitochondrial function and biogenesis. Previous studies have shown that FK506, an immunophilin ligand that modulates inflammation and mitochondrial function and inhibits calcineurin, is capable of rescuing the neurodegenerative pathology in models of Parkinson’s disease, Alzheimer’s disease, and Huntington’s disease. In this context, the main objective of this study was to evaluate if FK506 could rescue the neuronal degeneration and mitochondrial alterations in a transgenic (tg) animal model of HIV1-gp120 neurotoxicity. Methods GFAP-gp120 tg mice were treated with FK506 and analyzed for neuropathology, behavior, mitochondrial markers, and calcium flux by two-photon microscopy. Results We found that FK506 reduced the neuronal cell loss and neuro-inflammation in the gp120 tg mice. Moreover, while vehicle-treated gp120 tg mice displayed damaged mitochondria and increased neuro-inflammatory markers, FK506 rescued the morphological mitochondrial alterations and neuro-inflammation while increasing levels of optic atrophy 1 and mitofusin 1. By two-photon microscopy, calcium levels were not affected in the gp120 tg mice and no effects of FK506 were detected. However, at a functional level, FK506 ameliorated the gp120 tg mice hyperactivity in the open field. Conclusions Together, these results suggest that FK506 might be potentially neuroprotective in patients with HAND by mitigating inflammation and mitochondrial alterations.

α-Synuclein (α-syn) has been implicated in neurological disorders with parkinsonism, including Parkinson's disease and Dementia with Lewy body. Recent studies have shown α-syn oligomers released from neurons can propagate from cell-to-cell in a prion-like fashion exacerbating neurodegeneration. In this study, we examined the role of the endosomal sorting complex required for transport (ESCRT) pathway on the propagation of α-syn. α-syn, which is transported via the ESCRT pathway through multivesicular bodies for degradation, can also target the degradation of the ESCRT protein-charged multivesicular body protein (CHMP2B), thus generating a roadblock of endocytosed α-syn. Disruption of the ESCRT transport system also resulted in increased exocytosis of α-syn thus potentially increasing cell-to-cell propagation of synuclein. Conversely, delivery of a lentiviral vector overexpressing CHMP2B rescued the neurodegeneration in α-syn transgenic mice. Better understanding of the mechanisms of intracellular trafficking of α-syn might be important for understanding the pathogenesis and developing new treatments for synucleinopathies.

Forkhead box protein p1 (Foxp1), a transcription factor showing highly enriched expression in the striatum, has been implicated in central nervous system (CNS) development, but its role in the mature brain is unknown. In order to ascertain functional roles for Foxp1 in the CNS, we have identified gene targets for Foxp1 both in vitro and in vivo using genome-wide expression microarrays and chromatin-immunoprecipitation followed by high-throughput sequencing (ChIP-seq) assays. We found that mouse Foxp1 overexpression in striatal cells elicited expression changes of genes related to immune signaling, transcriptional regulation and a manually curated Huntington's disease (HD)-signaling pathway. Similar results were found when the gene expression data set was integrated with Foxp1-binding data determined from ChIP-seq analysis. In vivo lentiviral-mediated overexpression of human FOXP1 in the context of mutant huntingtin (Htt) protein resulted in a robust downregulation of glial cell-associated, immune genes, including those encoding a variety of cytokines and chemokines. Furthermore, Foxp1-induced expression changes were significantly negatively correlated with those changes elicited by mutant Htt protein in several different HD mouse models, and most significantly in post-mortem caudate from human HD subjects. We finally show that Foxp1 interacts with mutant Htt protein in mouse brain and is present in nuclear Htt aggregates in the striatum of R6/1 transgenic mice. These findings implicate Foxp1 as a key repressor of immune signaling in the CNS and suggest that the loss of Foxp1-mediated gene regulation in HD contributes to the immune dysfunction in this disease. We further suggest that Foxp1-regulated pathways might be important mediators of neuronal-glial cell communication.

Parkinson's disease (PD) and dementia with Lewy bodies are common disorders of the aging population and characterized by the progressive accumulation of α-synuclein (α-syn) in the central nervous system. Aggregation of α-syn into oligomers with a ring-like appearance has been proposed to play a role in toxicity. However, the molecular mechanisms and the potential sequence of events involved in the formation of pore-like structures are unclear. We utilized computer modeling and cell-based studies to investigate the process of oligomerization of wild-type and A53T mutant α-syn in membranes. The studies suggest that α-syn penetrates the membrane rapidly, changing its conformation from α-helical towards a coiled structure. This penetration facilitates the incorporation of additional α-syn monomers in the complex, and the subsequent displacement of phospholipids and the formation of oligomers in the membrane. This process occurred more rapidly, and with a more favorable energy of interaction, for mutant A53T compared with wild-type α-syn. After 4 ns of simulation of the protein-membrane model, α-syn had penetrated through two-thirds of the membrane. By 9 ns, the penetration of the annular α-syn oligomers can result in the formation of pore-like structures that fully perforate the lipid bilayer. Experimental incubation of recombinant α-syn in synthetic membranes resulted in the formation of similar pore-like complexes. Moreover, mutant (A53T) α-syn had a greater tendency to accumulate in neuronal membrane fractions in cell cultures, resulting in greater neuronal permeability, as demonstrated with the calcein efflux assay. These studies provide a sequential molecular explanation for the process of α-syn oligomerization in the membrane, and support the role of formation of pore-like structures in the pathogenesis of the neurodegenerative process in PD.

DNA methylation is a major epigenetic modification that regulates gene expression. Dnmt1, the maintenance DNA methylation enzyme, is abundantly expressed in the adult brain and is mainly located in the nuclear compartment, where it has access to chromatin. Hypomethylation of CpG islands at intron 1 of the SNCA gene has recently been reported to result in overexpression of α-synuclein in Parkinson disease (PD) and related disorders. We therefore investigated the mechanisms underlying altered DNA methylation in PD and dementia with Lewy bodies (DLB). We present evidence of reduction of nuclear Dnmt1 levels in human postmortem brain samples from PD and DLB patients as well as in the brains of α-synuclein transgenic mice models. Furthermore, sequestration of Dnmt1 in the cytoplasm results in global DNA hypomethylation in human and mouse brains, involving CpG islands upstream of SNCA, SEPW1, and PRKAR2A genes. We report that association of Dnmt1 and α-synuclein might mediate aberrant subcellular localization of Dnmt1. Nuclear Dnmt1 levels were partially rescued by overexpression of Dnmt1 in neuronal cell cultures and in α-synuclein transgenic mice brains. Our results underscore a novel mechanism for epigenetic dysregulation in Lewy body diseases, which might underlie the decrease in DNA methylation reported for PD and DLB.

Neurodegenerative disorders such as Alzheimer disease, Parkinson disease, frontotemporal dementia, Huntington disease and Creutzfeldt-Jakob disease (CJD) are characterized by progressive accumulation of protein aggregates in selected brain regions. Protein misfolding and templated assembly into aggregates might result from an imbalance between protein synthesis, aggregation and clearance. Although protein misfolding and aggregation occur in most neurodegenerative disorders, the concept of spreading and infectivity of aggregates in the CNS has, until now, been confined to prion diseases such as CJD and bovine spongiform encephalopathy. Emerging evidence, however, suggests that prion-like spreading, involving secreted proteins such as amyloid-β and cytosolic proteins such as tau, huntingtin and α-synuclein, can occur in other neurodegenerative disorders. The underlying molecular mechanisms and the therapeutic implications of the new data are discussed in this article.

Huntington's disease (HD) is a progressive neurodegenerative disorder characterized by motor, psychiatric, and cognitive abnormalities. In this present study, we tested whether abnormal motor behavior in a mouse model of HD, the R6/1 transgenic (Tg) mice, was associated with changes in cerebellar lipid composition and gene expression. We report altered motor behavior, which was associated with abnormal expression of glycosyltransferase genes in the cerebellum of R6/1 Tg mice. Cerebellar wet weight and total ganglioside concentration was significantly lower in R6/1 Tg mice than in wild-type (Wt) mice. Furthermore, the Purkinje cell-enriched ganglioside LD1 and the granule cell-enriched ganglioside GD1a were significantly lower in R6/1 Tg mice than in Wt mice. The myelin-enriched lipid sulfatides was also reduced in the cerebellum of R6/1 Tg mice. In contrast to the R6/1 Tg mice, total cerebellar ganglioside concentration did not differ between HD and control subjects. However, expression of several cerebellar glycosyltransferases genes was significantly less in HD subjects than in control subjects. Our findings indicate that the R6/1 Tg mice have severe cerebellar glycosphingolipid (GSL) abnormalities that may account, in part, for their abnormal motor behavior. Although the cerebellar lipid abnormalities found in the R6/1 Tg mice were not found in these HD subjects, the R6/1 Tg mice may be useful for evaluating the role of GSLs in cerebellar development.

Cell proliferation of neural progenitors in the subventricular zone (SVZ) of Parkinson disease (PD) patients and animal models is decreased. It was previously demonstrated that the neurotransmitter dopamine modulates cell proliferation in the embryonic brain. The aim of the present study was to analyze whether oral treatment with the dopamine receptor agonist pramipexole (PPX) modulates adult neurogenesis in the SVZ/olfactory bulb system in a dopaminergic lesion model. 6-Hydroxydopamine (6-OHDA) lesioned adult rats received either PPX (1.0 mg/kg) or PBS orally twice daily and bromodeoxyuridine (BrdU, a cell proliferation marker) for 10 days and were perfused immediately after treatment or 4 weeks after PPX withdrawal. Stereological analysis revealed a significant augmentation in SVZ proliferation by PPX. Consecutively, enhanced neuronal differentiation and more new neurons were present in the olfactory bulb 4 weeks after PPX withdrawal. In addition, dopaminergic neurogenesis was increased in the olfactory bulb after PPX treatment. Motor activity as assessed by using an open field paradigm was permanently increased even after long term PPX withdrawal. In addition, we demonstrate that D2 and D3 receptors are present on adult rat SVZ-derived neural progenitors in vitro, and PPX specifically increased mRNA levels of epidermal growth factor receptor (EGF-R) and paired box gene 6 (Pax6). Oral PPX treatment selectively increases adult neurogenesis in the SVZ-olfactory bulb system by increasing proliferation and cell survival of newly generated neurons. Analyzing the neurogenic fate decisions mediated by D2/D3 signaling pathways may lead to new avenues to induce neural repair in the adult brain.

We have explored genome-wide expression of genes related to glycobiology in exon 1 transgenic Huntington's disease (HD) mice using a custom-designed GLYCOv2 chip and Affymetrix microarray analyses. We validated, using quantitative real-time PCR, abnormal expression levels of genes encoding glycosyltransferases in the striatum of R6/1 transgenic mice, as well as in postmortem caudate from human HD subjects. Many of these genes show differential regional expression within the CNS, as indicated by in situ hybridization analysis, suggesting region-specific regulation of this system in the brain. We further show disrupted patterns of glycolipids (acidic and neutral lipids) and/or ganglioside levels in both the forebrain of the R6/1 transgenic mice and caudate samples from human HD subjects. These findings reveal novel disruptions in glycolipid/ganglioside metabolic pathways in the pathology of HD and suggest that the development of new targets to restore glycosphingolipid balance may act to ameliorate some symptoms of HD.

We have identified and cataloged 54 genes that exhibit predominant expression in the striatum. Our hypothesis is that such mRNA molecules are likely to encode proteins that are preferentially associated with particular physiological processes intrinsic to striatal neurons, and therefore might contribute to the regional specificity of neurodegeneration observed in striatal disorders such as Huntington's disease (HD). Expression of these genes was measured simultaneously in the striatum of HD R6/1 transgenic mice using Affymetrix oligonucleotide arrays. We found a decrease in expression of 81% of striatum-enriched genes in HD transgenic mice. Changes in expression of genes associated with G-protein signaling and calcium homeostasis were highlighted. The most striking decrement was observed for a newly identified subunit of the sodium channel, beta 4, with dramatic decreases in expression beginning at 8 weeks of age. A subset of striatal genes was tested by real-time PCR in caudate samples from human HD patients. Similar alterations in expression were observed in human HD and the R6/1 model for the striatal genes tested. Expression of 15 of the striatum-enriched genes was measured in 6-hydroxydopamine-lesioned rats to determine their dependence on dopamine innervation. No changes in expression were observed for any of these genes. These findings demonstrate that mutant huntingtin protein causes selective deficits in the expression of mRNAs responsible for striatum-specific physiology and these may contribute to the regional specificity of degeneration observed in HD.

The role of sucrose in cyanobacteria is still not fully understood. It is generally considered a salt-response molecule, and particularly, in Synechocystis sp. strain PCC 6803, it is referred as a secondary osmolyte. We showed that sucrose accumulates transiently in Synechocystis cells at early stages of a salt shock, which could be ascribed to salt activation of sucrose-phosphate synthase (SPS, UDP-glucose: d -fructose-6-phosphate 2-α- d -glucosyltransferase; EC 2.4.1.14), the key enzyme in sucrose synthesis pathway, and to an increase of the expression of the SPS encoding gene. Experiments with a mutant strain impaired in sucrose biosynthesis showed that sucrose is essential in stationary phase cells to overcome a later salt stress. Taken together, these results led us to suggest a more intricate function for sucrose than to be an osmoprotectant compound.

The α-proteobacteria phylogenetically related to the Roseobacter clade are predominantly responsible for the degradation of organosulfur compounds, including the algal osmolyte dimethylsulfoniopropionate (DMSP). Silicibacter sp. strain TM1040, isolated from a DMSP-producing Pfiesteria piscicida dinoflagellate culture, degrades DMSP, producing 3-methylmercaptopropionate. TM1040 possesses three lophotrichous flagella and is highly motile, leading to a hypothesis that TM1040 interacts with P. piscicida through a chemotactic response to compounds produced by its dinoflagellate host. A combination of a rapid chemotaxis screening assay and a quantitative capillary assay were used to measure chemotaxis of TM1040. These bacteria are highly attracted to dinoflagellate homogenates; however, the response decreases when homogenates are preheated to 80°C. To help identify the essential attractant molecules within the homogenates, a series of pure compounds were tested for their ability to serve as attractants. The results show that TM1040 is strongly attracted to amino acids and DMSP metabolites, while being only mildly responsive to sugars and the tricarboxylic acid cycle intermediates. Adding pure DMSP, methionine, or valine to the chemotaxis buffer resulted in a decreased response to the homogenates, indicating that exogenous addition of these chemicals blocks chemotaxis and suggesting that DMSP and amino acids are essential attractant molecules in the dinoflagellate homogenates. The implication of Silicibacter sp. strain TM1040 chemotaxis in establishing and maintaining its interaction with P. piscicida is discussed.

The recent years have witnessed an exponential growth in the knowledge of epigenetic mechanisms, and piling evidence now links DNA methylation and histone modifications with a wide range of physiological processes from embryonic development to memory formation and behavior. Not surprisingly, deregulation of epigenetic modifications is associated with human diseases as well.

Alzheimer disease (AD) is characterized by widespread neurodegeneration throughout the association cortex and limbic system, deposition of amyloid-β peptide (Aβ) in the neuropil and around the blood vessels, and formation of neurofibrillary tangles. The endopeptidase neprilysin has been successfully used to reduce the accumulation of Aβ following intracranial viral vector delivery or ex vivo manipulated intracranial delivery. These therapies have relied on direct injections into the brain, whereas a clinically desirable therapy would involve i.v. infusion of a recombinant enzyme. We previously characterized a recombinant neprilysin that contained a 38-amino acid brain-targeting domain. Recombinant cell lines have been generated expressing this brain-targeted enzyme (ASN12). In this report, we characterize the ASN12 recombinant protein for pharmacology in a mouse as well as efficacy in two APPtg mouse models of AD. The recombinant ASN12 transited to the brain with a t½ of 24 h and accumulated to 1.7% of injected dose at 24 h following i.v. delivery. We examined pharmacodynamics in the tg2576 APPtg mouse with the prion promoter APP695 SWE mutation and in the Line41 mThy1 APP751 mutation mouse. Treatment of either APPtg mouse resulted in reduced Aβ, increased neuronal synapses, and improved learning and memory. In addition, the Line41 APPtg mice showed increased levels of C-terminal neuropeptide Y fragments and increased neurogenesis. These results suggest that the recombinant brain-targeted neprilysin, ASN12, may be an effective treatment for AD and warrant further investigation in clinical trials.

Background In dementia with Lewy bodies (DLB) abnormal interactions between α-synuclein (α-syn) and beta amyloid (Aβ) result in selective degeneration of neurons in the neocortex, limbic system and striatum. However, factors rendering these neurons selectively vulnerable have not been fully investigated. The metabotropic glutamate receptor 5 (mGluR5) has been shown to be up regulated in DLB and might play a role as a mediator of the neurotoxic effects of Aβ and α-syn in vulnerable neuronal populations. In this context, the main objective of the present study was to investigate the role of mGluR5 as a mediator of the neurotoxic effects of α-syn and Aβ in the hippocampus. Results We generated double transgenic mice over-expressing amyloid precursor protein (APP) and α-syn under the mThy1 cassette and investigated the relationship between α-syn cleavage, Aβ, mGluR5 and neurodegeneration in the hippocampus. We found that compared to the single tg mice, the α-syn/APP tg mice displayed greater accumulation of α-syn and mGluR5 in the CA3 region of the hippocampus compared to the CA1 and other regions. This was accompanied by loss of CA3 (but not CA1) neurons in the single and α-syn/APP tg mice and greater loss of MAP 2 and synaptophysin in the CA3 in the α-syn/APP tg. mGluR5 gene transfer using a lentiviral vector into the hippocampus CA1 region resulted in greater α-syn accumulation and neurodegeneration in the single and α-syn/APP tg mice. In contrast, silencing mGluR5 with a lenti-shRNA protected neurons in the CA3 region of tg mice. In vitro, greater toxicity was observed in primary hippocampal neuronal cultures treated with Aβ oligomers and over-expressing α-syn; this effect was attenuated by down-regulating mGluR5 with an shRNA lentiviral vector. In α-syn-expressing neuronal cells lines, Aβ oligomers promoted increased intracellular calcium levels, calpain activation and α-syn cleavage resulting in caspase-3-dependent cell death. Treatment with pharmacological mGluR5 inhibitors such as 2-Methyl-6-(phenylethynyl)pyridine (MPEP) and 3-((2-Methyl-4-thiazolyl)ethynyl)pyridine (MTEP) attenuated the toxic effects of Aβ in α-syn-expressing neuronal cells. Conclusions Together, these results support the possibility that vulnerability of hippocampal neurons to α-syn and Aβ might be mediated via mGluR5. Moreover, therapeutical interventions targeting mGluR5 might have a role in DLB.

HIV involvement of the CNS continues to be a significant problem despite successful use of combination antiretroviral therapy (cART). Drugs of abuse can act in concert with HIV proteins to damage glia and neurons, worsening the neurotoxicity caused by HIV alone. Methamphetamine (METH) is a highly addictive psychostimulant drug, abuse of which has reached epidemic proportions and is associated with high-risk sexual behavior, increased HIV transmission, and development of drug resistance. HIV infection and METH dependence can have synergistic pathological effects, with preferential involvement of frontostriatal circuits. At the molecular level, epigenetic alterations have been reported for both HIV-1 infection and drug abuse, but the neuropathological pathways triggered by their combined effects are less known. We investigated epigenetic changes in the brain associated with HIV and METH. We analyzed postmortem frontal cortex tissue from 27 HIV seropositive individuals, 13 of which had a history of METH dependence, in comparison to 14 cases who never used METH. We detected changes in the expression of DNMT1, at mRNA and protein levels, that resulted in the increase of global DNA methylation. Genome-wide profiling of DNA methylation in a subset of cases, showed differential methylation on genes related to neurodegeneration; dopamine metabolism and transport; and oxidative phosphorylation. We provide evidence for the synergy of HIV and METH dependence on the patterns of DNA methylation on the host brain, which results in a distinctive landscape for the comorbid condition. Importantly, we identified new epigenetic targets that might aid in understanding the aggravated neurodegenerative, cognitive, motor and behavioral symptoms observed in persons living with HIV and addictions.

Alzheimer's disease (AD) is associated with the formation of toxic amyloid-β (Aβ)42 oligomers, and recent evidence supports a role for Aβ dimers as building blocks for oligomers. Molecular dynamics simulation studies have identified clans for the dominant conformations of Aβ42 forming dimers; however, it is unclear if a larger spectrum of dimers is involved and which set(s) of dimers might evolve to oligomers verse fibrils. Therefore, for this study we generated multiple structural conformations of Aβ42, using explicit all-atom molecular dynamics, and then clustering the different structures based on key conformational similarities. Those matching a selection threshold were then used to model a process of oligomerization. Remarkably, we showed a greater diversity in Aβ dimers than previously described. Depending on the clan family, different types of Aβ dimers were obtained. While some had the tendency to evolve into oligomeric rings, others formed fibrils of diverse characteristics. Then we selected the dimers that would evolve to membranephilic annular oligomers. Nearly one third of the 28 evaluated annular oligomers had the dimer interfaces between the neighboring Aβ42 monomers with possible salt bridges between the residue K28 from one side and either residue E22 or D23 on the other. Based on these results, key amino acids were identified for point mutations that either enhanced or suppressed the formation and toxicity of oligomer rings. Our studies suggest a greater diversity of Aβ dimers. Understanding the structure of Aβ dimers might be important for the rationale design of small molecules that block formation of toxic oligomers.