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Genes belonging to neuroplasticity, monoamine, circadian rhythm, and transcription factor pathways were investigated as modulators of antidepressant efficacy. The present study aimed (1) to replicate previous findings in an independent sample with treatment-resistant depression (TRD), and (2) to perform a pathway analysis to investigate the possible molecular mechanisms involved. 220 patients with major depressive disorder who were non-responders to a previous antidepressant were treated with venlafaxine for 4-6 weeks and in case of non-response with escitalopram for 4-6 weeks. Symptoms were assessed using the Montgomery Asberg Depression Rating Scale. The phenotypes were response and remission to venlafaxine, non-response (TRDA) and non-remission (TRDB) to neither venlafaxine nor escitalopram. 50 tag SNPs in 14 genes belonging to the pathways of interest were tested for association with phenotypes. Molecular pathways (KEGG database) that included one or more of the genes associated with the phenotypes were investigated also in the STAR*D sample. The associations between ZNF804A rs7603001 and response, CREB1 rs2254137 and remission were replicated, as well as CHL1 rs2133402 and lower risk of TRD. Other CHL1 SNPs were potential predictors of TRD (rs1516340, rs2272522, rs1516338, rs2133402). The MAPK1 rs6928 SNP was consistently associated with all the phenotypes. The protein processing in endoplasmic reticulum pathway (hsa04141) was the best pathway that may explain the mechanisms of MAPK1 involvement in antidepressant response. Signals in genes previously associated with antidepressant efficacy were confirmed for CREB1, ZNF804A and CHL1. These genes play pivotal roles in synaptic plasticity, neural activity and connectivity.

Neurodegenerative diseases, as Parkinson's disease (PD), involve irreversible neural cell damage and impairment. In PD, there is a selective degeneration of the dopaminergic neurons leading to motor symptoms. A common finding in PD neurodegeneration is the increase of reactive oxygen species (ROS), leading to oxidative stress. To date there are only interventions to relieve PD symptoms, however progress has been made in the development of therapies that target the immune system or use its components as therapeutic agents; among these, mesenchymal stem cells (MSCs), which are able to express neuroprotective factors as cytokines, chemokines and angiogenic molecules, collectively named secretome, that accumulate in MSC culture medium. However, lasting cell-free administration of secretome in vitro or in vivo is challenging. We used the conditioned media from rat adipose tissue-derived MSCs (RAA-MSCs) to check for neuroprotective activity towards pro-oxidizing agents such as hydrogen peroxide (H 2 O 2 ) or the dopaminergic selective toxin 6-hydroxydopamine (6-OHDA) that is commonly used to model PD neurodegeneration. When neuroblastoma SH-SY5Y cells were pre-conditioned with 100% RAA-MSC media, then treated with H 2 O 2 and 6-OHDA, mortality and ROS generation were reduced. We implemented the controlled release of RAA-MSC secretome from injectable biodegradable hydrogels that offer a possible in situ implant with mini-invasive techniques. The hydrogels were composed of type I bovine collagen (COLL) and low-molecular-weight hyaluronic acid (LMWHA) or COLL and polyethylene glycol (PEG). Hydrogels were suitable for RAA-MSC embedding up to 48h and secretome from these RAA-MSCs was active and counteracted 6-OHDA toxicity, with upregulation of the antioxidant enzyme sirtuin 3 (SIRT3). These results support a biomaterials-based approach for controlled delivery of MSC-produced neuroprotective factors in a PD-relevant experimental context., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)

In this study, we evaluated the anti-amyloid effect of functionalized nanoliposomes (mApoE-PA-LIP) in a mouse model of Alzheimer's disease with use of positron emission tomography and β-amyloid (Aβ)-targeted tracer [ 11 C]Pittsburgh compound B ([ 11 C]PIB). APP23 mice were injected with mApoE-PA-LIP or saline (3 times per week for 3 weeks) and [ 11 C]PIB imaging was performed at baseline, after the treatment and after 3 months follow-up period, accompanied by Aβ immunohistochemistry and ELISA. After the treatment, [ 11 C]PIB binding ratios between mApoE-PA-LIP and saline groups were equivalent in all analyzed brain regions; however, in the saline group, binding ratios increased from the baseline, whereas no increase was detected in the mApoE-PA-LIP group. During the additional follow-up, [ 11 C]PIB binding increased significantly from baseline in both groups, and binding ratios correlated with the immunohistochemically defined Aβ load. This study further supports the use of [ 11 C]PIB positron emission tomography imaging as a biomarker of Aβ deposition in APP23 mice and highlights the benefits of noninvasive follow-up, that is, using baseline data for animal stratification and normalization of treatment effects to baseline values, for future anti-amyloid treatment studies., (Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.)

Spinal cord injury (SCI) is a complex pathology that evolves after primary acute mechanical injury, causing further damage to the spinal cord tissue that exacerbates clinical outcomes. Based on encouraging results from preclinical experiments, some cell treatments being translated into clinical practice demonstrate promising and effective improvement in sensory/motor function. Combinatorial treatments of cell and drug/biological factors have been demonstrated to be more effective than cell treatments alone. Recent advances have led to the development of biomaterials aiming to promote in situ cell delivery for SCI, together with combinatorial strategies using drugs/biomolecules to achieve a maximized multitarget approach. This review provides an overview of single and combinatorial regenerative cell treatments as well as potential delivery options to treat SCI., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The failure of clinical trials largely focused on mild to moderate stages of Alzheimer disease has suggested to the scientific community that the effectiveness of Amyloid-β (Aβ)-centered treatments should be evaluated starting as early as possible, well before irreversible brain damage has occurred. Accordingly, also the preclinical development of new therapies should be carried out taking into account this suggestion. In the present investigation we evaluated the efficacy of a treatment with liposomes multifunctionalized for crossing the blood-brain barrier and targeting Aβ, carried out on young APP/PS1 Tg mice, taken as a model of pre-symptomatic disease stage. Liposomes were administered once a week to Tg mice for 7months, starting at the age of 5months and up to the age of 12 when they display AD-like cognitive and brain biochemical/anatomical features. The treatment prevented the onset of the long-term memory impairment and slowed down the deposition of brain Aβ; at anatomical level, prevented both ventricle enlargement and entorhinal cortex thickness reduction, otherwise occurring in untreated mice. Strikingly, these effects were maintained 3months after treatment discontinuation. An increase of Aβ levels in the liver was detected at the end of the treatment, then followed also by reduction of brain Amyloid Precursor Protein and increase of Aβ-degrading enzymes. These results suggest that the treatment promotes brain Aβ clearance by a peripheral 'sink' effect and ultimately affects Aβ turnover in the brain. Worth of note, the treatment was apparently not toxic for all the organs analyzed, in particular for brain, as suggested by the lower brain TNF-α and MDA levels, and by higher level of SOD activity in treated mice. Together, these findings promote a very early treatment with multi-functional liposomes as a well-tolerated nanomedicine-based approach, potentially suitable for a disease-modifying therapy of AD, able to delay or prevent relevant features of the disease., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Introduction: Schizophrenia (SCZ) is one of the most disabling psychiatric disorders. Genetic factors play an important role in both SCZ liability and its treatment outcome. In the present paper, we investigated the effects of several single nucleotide polymorphisms (SNPs) within ten strong candidate genes involved with antipsychotics (APs) mechanisms of action., Methods: Two independent samples were investigated in the present study. Totals of 176 SCZ subjects and 326 controls of Korean ancestry, and 83 SCZ subjects and 194 controls of Italian ancestry were recruited and genotyped. SCZ risk and other parameters were also investigated., Results: Concerning APs response, only a nominal association with HOMER1 rs3822568 in the Korean sample was found. In the haplotype analysis, rs9801117 C-rs12668837 C-rs4621754 A haplotype within ESYT2 and NCAPG2 genes was associated with APs response in the same sample. As for secondary outcomes, rs7439 within PKDCC and rs12668837 within NCAPG2 were associated with SCZ risk in the Italian sample. In the haplotype analysis, rs2788478 G-rs2657375 T-rs1039621 A within the region between WDR60 and ESYT genes and rs2013 C (ESYT2)-rs6459896 A (NCAPG2) haplotypes were associated with SCZ in the same sample. No association was found in the Korean sample. Finally, our exploratory data suggest a possible modulation of HOMER1, ARC, BDNF, TXNRD2, WDR60, and ESYT2 genes in the APs response to specific symptom clusters., Conclusion: Our results did not support a primary role for the genes investigated in the APs response. On the other hand, our secondary results suggest a possible involvement of NACPG2 and PKDCC in SCZ liability. Finally, our exploratory findings may deserve further investigations in specific studies.

Amyotrophic lateral sclerosis (ALS) is a multifactorial disease, possibly with contributions from genetics and lifestyle. We examined variants in genes relevant to energy metabolism and physical activity in a case-control association study, with the aim of assessing genetics and physical activity as contributors to ALS risk. A well-characterized sample of Italian ALS patients (101) and controls (101) from the EURALS Consortium underwent a questionnaire interview on demographic, physical and other lifestyle habits, and venipuncture for DNA extraction. The genes selected were sirtuin 3 (SIRT3), peroxisome proliferator-activated receptor-γ coactivator-1α (PPARGC1A) and apolipoprotein E (APOE). Genetic studies suggested, for the first time, a protective role of the SIRT3 single-nucleotide polymorphism rs4980329 in ALS risk, and a contribution of the APOE-ε2 allele, which was more frequent in ALS patients than in controls. A joint analysis coupling genetic data and sporting activity revealed opposite roles of APOE-ε2 and SIRT3 rs3825075, the former being more frequent in physically active ALS patients and the latter more frequent in physically inactive patients. These findings suggest a contribution to ALS risk of genetic and environmental factors involved in energy metabolism, and stress the importance of a multifactorial analysis for evaluating this risk.

Few studies have examined the relationship between CSF and structural biomarkers, and cognitive function in MCI. We examined the relationship between cognitive function, hippocampal volume and cerebrospinal fluid (CSF) Aβ 42 and tau in 145 patients with MCI. Patients were assessed on cognitive tasks from the Cambridge Neuropsychological Test Automated Battery (CANTAB), the Geriatric Depression Scale and the Functional Activities Questionnaire. Hippocampal volume was measured using magnetic resonance imaging (MRI), and CSF markers of Aβ 42 , tau and p-tau 181 were also measured. Worse performance on a wide range of memory and sustained attention tasks were associated with reduced hippocampal volume, higher CSF tau and p-tau 181 and increased tau/Aβ 42 ratio. Memory tasks were also associated with lower ability to conduct functional activities of daily living, providing a link between AD biomarkers, memory performance and functional outcome. These results suggest that biomarkers of Aβ and tau are strongly related to cognitive performance as assessed by the CANTAB, and have implications for the early detection and characterization of incipient AD., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Several antipsychotics and antidepressants have been associated with QTc prolongation or other electrocardiogram (ECG) alterations, but their impact is still debated and other risk factors are known to affect QTc. We investigated the effect of antidepressants and antipsychotics on QTc and other ECG intervals/waves in three samples. Two discovery samples (cross-sectional sample n = 145 and prospective sample n = 68, naturalistic treatment) and a replication prospective sample (Clinical Antipsychotic Trials of Intervention Effectiveness, n = 515, randomized treatment) were analysed. In both prospective samples, baseline/follow-up changes in ECG parameters were analysed in relation to the number of psychotropic drugs stratified according to their known cardiovascular risk. In the cross-sectional sample, ECG parameters were compared among drugs with different risk profile. The possible effect of single nucleotide polymorphisms (SNPs) in the CACNA1C gene on QTc was also investigated. There was no evidence of mean QTc prolongation or increased risk of clinically relevant QTc prolongation (≥20 msec.) in association with psychotropic drugs stratified according to their known cardiovascular risk. The prescription of drugs with cardiovascular risk was less common in older individuals or individuals with cardiovascular comorbidities. Other factors (gender, baseline QTc, renal function) affected QTc. rs1006737 and SNPs in linkage disequilibrium with it modulated QTc duration/changes in all samples. An association between risk drugs and shorter RR interval or higher heart rate was found in all samples. A relevant effect of psychotropic drugs with cardiovascular risk on QTc duration was not observed. A number of factors other than psychotropic drugs may influence QTc. CACNA1C rs1006737 may modulate QTc in patients treated with psychotropic drugs., (© 2016 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).)

Fatal Familial Insomnia (FFI) is a genetic prion disease caused by a point mutation in the prion protein gene (PRNP) characterized by prominent thalamic atrophy, diffuse astrogliosis and moderate deposition of PrP Sc in the brain. Here, for the first time, we demonstrate that the olfactory mucosa (OM) of patients with FFI contains trace amount of PrP Sc detectable by PMCA and RT-QuIC. Quantitative PMCA analysis estimated a PrP Sc concentration of about 1 × 10 -14  g/ml. In contrast, PrP Sc was not detected in OM samples from healthy controls and patients affected by other neurodegenerative disorders, including Alzheimer's disease, Parkinson's disease and frontotemporal dementia. These results indicate that the detection limit of these assays is in the order of a single PrP Sc oligomer/molecule with a specificity of 100%.

Background: Amyloid-β oligomers (AβO) are species mainly involved in the synaptic and cognitive dysfunction in Alzheimer's disease. Although their action has been described mainly at neuronal level, it is now clear that glial cells govern synaptic activity in their resting state, contributing to new learning and memory establishment. In contrast, when activated, they may lead to synaptic and cognitive dysfunction. Using a reliable acute AβO-mediated mouse model of AD, we explored whether the memory alteration AβOs induce relies on the activation of glial cells, and if Toll-like receptor 4 (TLR4), pivotal in the initiation of an immune response, is involved., Methods: C57 naïve mice were given a single intracerebroventricular injection of synthetic AβO-containing solution (1μM), which induces substantial impairment in the establishment of recognition memory. Then, first we assessed glial cell activation at different times post-injection by western blot, immunohistochemistry and ELISA in the hippocampus. After that we explored the efficacy of pre-treatment with anti-inflammatory drugs (indomethacin and an IL-1β receptor antagonist) to prevent impairment in the novel object recognition task, and compared AβO's effects in TLR4 knockout mice., Results: A single AβO injection rapidly activated glial cells and increased pro-inflammatory cytokine expression. Both anti-inflammatory drugs prevented the AβO-mediated impairment in memory establishment. A selective TLR4 receptor antagonist abolished AβO's action on memory, and in TLR4 knockout mice it had no effect on either memory or glial activation., Conclusions: These data provide new information on AβO's mechanism of action, indicating that besides direct action at the synapses, they also act through the immune system, with TLR4 playing a major role. This suggests that in a potential therapeutic setting inflammation must be considered as well., (Copyright © 2016. Published by Elsevier Inc.)

Aggregation of amyloid-β peptide (Aβ) is a key event in the pathogenesis of Alzheimer's disease (AD). We investigated the effects of nanoliposomes decorated with the retro-inverso peptide RI-OR2-TAT (Ac-rGffvlkGrrrrqrrkkrGy-NH 2 ) on the aggregation and toxicity of Aβ. Remarkably low concentrations of these peptide inhibitor nanoparticles (PINPs) were required to inhibit the formation of Aβ oligomers and fibrils in vitro, with 50% inhibition occurring at a molar ratio of ~1:2000 of liposome-bound RI-OR2-TAT to Aβ. PINPs also bound to Aβ with high affinity (K d =13.2-50 nM), rescued SHSY-5Y cells from the toxic effect of pre-aggregated Aβ, crossed an in vitro blood-brain barrier model (hCMEC/D3 cell monolayer), entered the brains of C57 BL/6 mice, and protected against memory loss in APP SWE transgenic mice in a novel object recognition test. As the most potent aggregation inhibitor that we have tested so far, we propose to develop PINPs as a potential disease-modifying treatment for AD., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Resting state electroencephalographic (EEG) rhythms reflect the fluctuation of cortical arousal and vigilance in a typical clinical setting, namely the EEG recording for few minutes with eyes closed (i.e., passive condition) and eyes open (i.e., active condition). Can this procedure be back-translated to C57 (wild type) mice for aging studies? On-going EEG rhythms were recorded from a frontoparietal bipolar channel in 85 (19 females) C57 mice. Male mice were subdivided into 3 groups: 25 young (4.5-6 months), 18 middle-aged (12-15 months), and 23 old (20-24 months) mice to test the effect of aging. EEG power density was compared between short periods (about 5 minutes) of awake quiet behavior (passive) and dynamic exploration of the cage (active). Compared with the passive condition, the active condition induced decreased EEG power at 1-2 Hz and increased EEG power at 6-10 Hz in the group of 85 mice. Concerning the aging effects, the passive condition showed higher EEG power at 1-2 Hz in the old group than that in the others. Furthermore, the active condition exhibited a maximum EEG power at 6-8 Hz in the former group and 8-10 Hz in the latter. In the present conditions, delta and theta EEG rhythms reflected changes in cortical arousal and vigilance in freely behaving C57 mice across aging. These changes resemble the so-called slowing of resting state EEG rhythms observed in humans across physiological and pathological aging. The present EEG procedures may be used to enhance preclinical phases of drug discovery in mice for understanding the neurophysiological effects of new compounds against brain aging., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Introduction: With the increase of the population's average age, Alzheimer's disease (AD) is becoming one of the most disabling diseases worldwide. Recently, neurodevelopment processes have been involved in the AD etiopathogenesis. Genetic studies in this field could contribute to our knowledge and suggest new molecular targets for possible treatments., Methods: Our primary aim was to investigate the associations among single nucleotide polymorphisms (SNPs) within neurodevelopment related genes (BDNF, ST8SIA2, C15orf32, NCAPG2, ESYT2, WDR60, LOC154822, VIPR2, GSK3B, NR1I2, ZNF804A, SP4) and AD. A number of exploratory analyses was also performed to evaluate the associations with the presence of behavioral and psychiatric symptoms of dementia (BPSD), as well as with variations in hematological parameters. Two independent samples were investigated, one of 228 Greek subjects and one sample of 229 Italian subjects, including 156Alzheimer's Disease patients CE patients and 301 healthy controls. All patients were affected by late onset AD (LOAD)., Results: None of the analyzed SNPs was associated with AD in our samples. In the exploratory analyses, several genetic variants were associated with inflammation parameters in the Greek sample and in the merged one, suggesting a relationship among these genes and the modulation of inflammation and the immune response. Other exploratory analyses showed associations among several SNPs and psychiatric symptomatology in the Greek sample, suggesting a possible modulation of these variants on psychiatric comorbidities in AD., Conclusions: Although we failed to find a direct relationship between AD and the genetic variants investigated, possible connections with inflammation and psychiatric symptoms were suggested., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Alzheimer's disease (AD) is histopathologically characterized by neurodegeneration, the formation of intracellular neurofibrillary tangles and extracellular Aβ deposits that derive from proteolytic processing of the amyloid precursor protein (APP). As rodents do not normally develop Aβ pathology, various transgenic animal models of AD were designed to overexpress human APP with mutations favouring its amyloidogenic processing. However, these mouse models display tremendous differences in the spatial and temporal appearance of Aβ deposits, synaptic dysfunction, neurodegeneration and the manifestation of learning deficits which may be caused by age-related and brain region-specific differences in APP transgene levels. Consequentially, a comparative temporal and regional analysis of the pathological effects of Aβ in mouse brains is difficult complicating the validation of therapeutic AD treatment strategies in different mouse models. To date, no antibodies are available that properly discriminate endogenous rodent and transgenic human APP in brains of APP-transgenic animals. Here, we developed and characterized rat monoclonal antibodies by immunohistochemistry and Western blot that detect human but not murine APP in brains of three APP-transgenic mouse and one APP-transgenic rat model. We observed remarkable differences in expression levels and brain region-specific expression of human APP among the investigated transgenic mouse lines. This may explain the differences between APP-transgenic models mentioned above. Furthermore, we provide compelling evidence that our new antibodies specifically detect endogenous human APP in immunocytochemistry, FACS and immunoprecipitation. Hence, we propose these antibodies as standard tool for monitoring expression of endogenous or transfected APP in human cells and APP expression in transgenic animals., (© 2016 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.)

15B3 is a monoclonal IgM antibody that selectively detects pathological aggregates of the prion protein (PrP). We report the unexpected finding that 15B3 also recognizes oligomeric but not monomeric forms of amyloid-β (Aβ)42, an aggregating peptide implicated in the pathogenesis of Alzheimer's disease (AD). The 15B3 antibody: i) inhibits the binding of synthetic Aβ42 oligomers to recombinant PrP and neuronal membranes; ii) prevents oligomer-induced membrane depolarization; iii) antagonizes the inhibitory effects of oligomers on the physiological pharyngeal contractions of the nematode Caenorhabditis elegans; and iv) counteracts the memory deficits induced by intracerebroventricular injection of Aβ42 oligomers in mice. Thus this antibody binds to pathologically relevant forms of Aβ, and offers a potential research, diagnostic, and therapeutic tool for AD.

The neuropathological hallmarks of Alzheimer's disease (AD) are extracellular plaques built up by the accumulation of the amyloid-β protein precursor (AβPP)-derived peptide β (Aβ), and intracellular tangles of hyperphosphorylated tau protein. Sirtuin 2 (SIRT2) is a member of the sirtuin family, featuring conserved enzymes with deacetylase activity and involved in several cell molecular pathways. We investigated the importance of SIRT2 inhibition in AD. We inhibited SIRT2 by small molecules (AGK-2, AK-7) and examined AβPP metabolism in H4-SW neuroglioma cells overexpressing AβPP and two AD transgenic mouse models (3xTg-AD and APP23). The in vitro studies suggested that the inhibition of SIRT2 reduced Aβ production; in vivo data showed an improvement of cognitive performance in the novel object recognition test, and an effect on AβPP proteolytic processing leading to a reduction of soluble β-AβPP and an increase of soluble α-AβPP protein. In 3xTg-AD mice, we noticed that total tau protein level rose. Overall, our pre-clinical data support a role for SIRT2 inhibition in the improvement of cognitive performance and the modulation of molecular mechanisms relevant for AD, thus deserving attention as possible therapeutic strategy.

The term oligomeropathies defines the neurodegenerative disorders associated with protein misfolding, where small soluble aggregates (oligomers 4-200 KDa) are the cause of neuronal dysfunction and are responsible for spreading the pathology. The ability of these soluble β-sheet conformers to induce neuronal damage has been investigated in direct challenge with the monomeric and fibrillary structures, showing that only the oligomeric species affected the neurons. β amyloid oligomers were initially purified from Alzheimer brains and obtained using synthetic peptides. Together with the neuronal death, synaptic dysfunction, loss of spines, and LTP impairment were seen with the direct application of β amyloid oligomers. Similar results have been described with proteins associated with other neurodegenerative disorders. The biological activities of oligomeric forms of α synuclein have been described in Parkinson's disease and Lewy body dementia. Detrimental effects have been associated with the oligomeric forms of prion, tau, and huntingtin, the key proteins in prion diseases, frontotemporal dementia, and Huntington's disease, respectively. The molecular mechanisms of the oligomer-related toxic effects can be summarized under three headings: nonspecific perturbance of cellular and intracellular membranes, specific interaction with various cellular entities, and amyloid pore channel formation. To characterize and distinguish oligomer actions better, we compared the ability of β amyloid and α synuclein oligomers to induce cognitive impairment when applied directly into the brain in the same acute mouse model. We also investigated the role of inflammatory components. © 2016 International Parkinson and Movement Disorder Society., (© 2016 International Parkinson and Movement Disorder Society.)

Background: In the field of Alzheimer's disease (AD), the validation of biomarkers for early AD diagnosis and for use as a surrogate outcome in AD clinical trials is of considerable research interest., Objective: To characterize the clinical profile and genetic, neuroimaging and neurophysiological biomarkers of prodromal AD in amnestic mild cognitive impairment (aMCI) patients enrolled in the IMI WP5 PharmaCog (also referred to as the European ADNI study)., Methods: A total of 147 aMCI patients were enrolled in 13 European memory clinics. Patients underwent clinical and neuropsychological evaluation, magnetic resonance imaging (MRI), electroencephalography (EEG) and lumbar puncture to assess the levels of amyloid β peptide 1-42 (Aβ42), tau and p-tau, and blood samples were collected. Genetic (APOE), neuroimaging (3T morphometry and diffusion MRI) and EEG (with resting-state and auditory oddball event-related potential (AO-ERP) paradigm) biomarkers were evaluated., Results: Prodromal AD was found in 55 aMCI patients defined by low Aβ42 in the cerebrospinal fluid (Aβ positive). Compared to the aMCI group with high Aβ42 levels (Aβ negative), Aβ positive patients showed poorer visual (P = 0.001), spatial recognition (P < 0.0005) and working (P = 0.024) memory, as well as a higher frequency of APOE4 (P < 0.0005), lower hippocampal volume (P = 0.04), reduced thickness of the parietal cortex (P < 0.009) and structural connectivity of the corpus callosum (P < 0.05), higher amplitude of delta rhythms at rest (P = 0.03) and lower amplitude of posterior cingulate sources of AO-ERP (P = 0.03)., Conclusion: These results suggest that, in aMCI patients, prodromal AD is characterized by a distinctive cognitive profile and genetic, neuroimaging and neurophysiological biomarkers. Longitudinal assessment will help to identify the role of these biomarkers in AD progression., (© 2016 The Association for the Publication of the Journal of Internal Medicine.)

Previous studies suggest that genetic variants in CHRNA7, which encodes for the major subunit of the acetylcholine receptor (α7-nAChR), are associated with the clinical response to cholinesterase inhibitors (ChEI) in Alzheimer's disease (AD) patients. We sought to replicate the association of two SNPs in the CHRNA7 gene, rs6494223 and rs8024987, with response to ChEI treatment in an Italian cohort of 169 AD patients, further extending the study to gene-level analysis. None of the tested variants was associated with clinical response. However, rs6494223 showed a consistent effect direction (OR = 1.4; p = 0.17), which after meta-analysis with previous study yielded a significant result (OR = 1.57, p = 0.02, I2 = 0%).

Many cells of the nervous system have been shown to release exosomes, a subclass of secreted vesicles of endosomal origin capable of transferring biomolecules among cells: this transfer modality represents a novel physiological form of intercellular communication between neural cells. Herein, we demonstrated that progranulin (PGRN), a protein targeted to the classical secretory pathway, is also secreted in association with exosomes by human primary fibroblasts. Moreover, we demonstrated that null mutations in the progranulin gene (GRN), a major cause of frontotemporal dementia, strongly reduce the number of released exosomes and alter their composition. In vitro GRN silencing in SHSY-5Y cells confirmed a role of PGRN in the control of exosome release. It is believed that depletion of PGRN in the brain might cause neurodegeneration in GRN-associated frontotemporal dementia. We demonstrated that, along with shortage of the circulating PGRN, GRN null mutations alter intercellular communication. Thus, a better understanding of the role played by exosomes in GRN-associated neurodegeneration is crucial for the development of novel therapies for these diseases., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Frontotemporal lobar degeneration (FTLD) includes a spectrum of disorders characterized by changes of personality and social behavior and, often, a gradual and progressive language dysfunction. In the last years, several efforts have been fulfilled in identifying both genetic mutations and pathological proteins associated with FTLD. The molecular bases undergoing the onset and progression of the disease remain still unknown. Recent literature prompts an involvement of RNA metabolism in FTLD, particularly microRNAs (miRNAs). Dysregulation of miRNAs in several disorders, including neurodegenerative diseases, and increasing importance of circulating miRNAs in different pathologies has suggested to implement the study of their possible application as biological markers and new therapeutic targets; moreover, miRNA-based therapy is becoming a powerful tool to deepen the function of a gene, the mechanism of a disease, and validate therapeutic targets. Regarding FTLD, different studies showed that miRNAs are playing an important role. For example, several reports have evaluated miRNA regulation of the progranulin gene suggesting that it is under their control, as described for miR-29b, miR-107, and miR-659. More recently, it has been demonstrated that TMEM106B gene, which protein is elevated in FTLD-TDP brains, is repressed by miR-132/212 cluster; this post-transcriptional mechanism increases intracellular levels of progranulin, affecting its pathways. These findings if confirmed could suggest that these microRNAs have a role as potential targets for some related-FTLD genes. In this review, we focus on the emerging roles of the miRNAs in the pathogenesis of FTLD.

Background/aims: Suicidal behavior (SB) in bipolar disorder (BD) is a complex multifactorial event resulting from an interaction of genetic, neurobiological and psychosocial factors. Recent studies identified new possible mechanisms, suggesting a role for sirtuins (SIRTs 1-7), a family of nicotinamide adenine dinucleotide-dependent enzymes with a multifaceted role in the central nervous system. The aims of the present study were: (1) to investigate the effects of the rs10997870 SIRT1 gene variant on SB in BD; (2) to explore the effects of the same gene variant on specific depressive symptoms at the severest episode., Methods: One hundred and eighty DSM-IV bipolar outpatients were enrolled in a naturalistic cohort study. The rs10997870 polymorphism within the SIRT1 gene was analyzed., Results: An association between the GG genotype and SB was detected (lifetime: p = 0.015). Compared to other genotypes, GG carriers presented more frequently psychomotor agitation (p = 0.009) and a higher Hamilton Depression Rating Scale total score (p = 0.014) at the severest depressive episode. SB and psychomotor agitation were found to be associated with GG carriers and G allele in a multivariate analysis as well., Conclusion: Our findings suggest a role of the rs10997870 SIRT1 gene variant in SB amongst BD patients and its association with specific depressive symptoms. Despite a number of limitations of this exploratory study, our results may provide new insight into the mechanisms underlying SB in BD., (© 2016 S. Karger AG, Basel.)

Many efforts have been performed in order to understand the role of recruited macrophages in the progression of spinal cord injury (SCI). Different studies revealed a pleiotropic effect played by these cells associated to distinct phenotypes (M1 and M2), showing a predictable spatial and temporal distribution in the injured site after SCI. Differently, the role of activated microglia in injury progression has been poorly investigated, mainly because of the challenges to target and selectively modulate them in situ. A delivery nanovector tool (poly-ε-caprolactone-based nanoparticles) able to selectively treat/target microglia has been developed and used here to clarify the temporal and spatial involvement of the pro-inflammatory response associated to microglial cells in SCI. We show that a treatment with nanoparticles loaded with minocycline, the latter a well-known anti-inflammatory drug, when administered acutely in a SCI mouse model is able to efficiently modulate the resident microglial cells reducing the pro-inflammatory response, maintaining a pro-regenerative milieu and ameliorating the behavioral outcome up to 63 days post injury. Furthermore, by using this selective delivery tool we demonstrate a mechanistic link between early microglia activation and M1 macrophages recruitment to the injured site via CCL2 chemokine, revealing a detrimental contribution of pro-inflammatory macrophages to injury progression after SCI., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Stem cell therapy with human mesenchymal stem cells (hMSCs) represents a promising strategy in spinal cord injury (SCI). However, both systemic and parenchymal hMSCs administrations show significant drawbacks as a limited number and viability of stem cells in situ. Biomaterials able to encapsulate and sustain hMSCs represent a viable approach to overcome these limitations potentially improving the stem cell therapy. In this study, we evaluate a new agarose/carbomer based hydrogel which combines different strategies to optimize hMSCs viability, density and delivery of paracrine factors. Specifically, we evaluate a new loading procedure on a lyophilized scaffold (soaked up effect) that reduces mechanical stress in encapsulating hMSCs into the hydrogel. In addition, we combine arginine-glycine-aspartic acid (RGD) tripeptide and 3D extracellular matrix deposition to increase the capacity to attach and maintain healthy hMSCs within the hydrogel over time. Furthermore, the fluidic diffusion from the hydrogel toward the injury site is improved by using a cling film that oriented efficaciously the delivery of paracrine factors in vivo. Finally, we demonstrate that an improved combination as here proposed of hMSCs and biomimetic hydrogel is able to immunomodulate significantly the pro-inflammatory environment in a SCI mouse model, increasing M2 macrophagic population and promoting a pro-regenerative environment in situ., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Drug-effect phenotypes in human lymphoblastoid cell lines recently allowed to identify CHL1 (cell adhesion molecule with homology to L1CAM), GAP43 (growth-associated protein 43) and ITGB3 (integrin beta 3) as new candidates for involvement in the antidepressant effect. CHL1 and ITGB3 code for adhesion molecules, while GAP43 codes for a neuron-specific cytosolic protein expressed in neuronal growth cones; all the three gene products are involved in synaptic plasticity. Sixteen polymorphisms in these genes were genotyped in two samples (n=369 and 90) with diagnosis of major depressive episode who were treated with antidepressants in a naturalistic setting. Phenotypes were response, remission and treatment-resistant depression. Logistic regression including appropriate covariates was performed. Genes associated with outcomes were investigated in the Sequenced Treatment Alternatives to Relieve Depression (STAR*D) genome-wide study (n=1861) as both individual genes and through a pathway analysis (Reactome and String databases). Gene-based analysis suggested CHL1 rs4003413, GAP43 rs283393 and rs9860828, ITGB3 rs3809865 as the top candidates due to their replication across the largest original sample and the STAR*D cohort. GAP43 molecular pathway was associated with both response and remission in the STAR*D, with ELAVL4 representing the gene with the highest percentage of single nucleotide polymorphisms (SNPs) associated with outcomes. Other promising genes emerging from the pathway analysis were ITGB1 and NRP1. The present study was the first to analyze cell adhesion genes and their molecular pathways in antidepressant response. Genes and biomarkers involved in neuronal adhesion should be considered by further studies aimed to identify predictors of antidepressant response.

The gray mouse lemur (Microcebus murinus) is considered a useful primate model for translational research. In the framework of IMI PharmaCog project (Grant Agreement n°115009, www.pharmacog.org), we tested the hypothesis that spectral electroencephalographic (EEG) markers of motor and locomotor activity in gray mouse lemurs reflect typical movement-related desynchronization of alpha rhythms (about 8-12 Hz) in humans. To this aim, EEG (bipolar electrodes in frontal cortex) and electromyographic (EMG; bipolar electrodes sutured in neck muscles) data were recorded in 13 male adult (about 3 years) lemurs. Artifact-free EEG segments during active state (gross movements, exploratory movements or locomotor activity) and awake passive state (no sleep) were selected on the basis of instrumental measures of animal behavior, and were used as an input for EEG power density analysis. Results showed a clear peak of EEG power density at alpha range (7-9 Hz) during passive state. During active state, there was a reduction in alpha power density (8-12 Hz) and an increase of power density at slow frequencies (1-4 Hz). Relative EMG activity was related to EEG power density at 2-4 Hz (positive correlation) and at 8-12 Hz (negative correlation). These results suggest for the first time that the primate gray mouse lemurs and humans may share basic neurophysiologic mechanisms of synchronization of frontal alpha rhythms in awake passive state and their desynchronization during motor and locomotor activity. These EEG markers may be an ideal experimental model for translational basic (motor science) and applied (pharmacological and non-pharmacological interventions) research in Neurophysiology.

Studies on sirtuins (SIRT), a family of proteins with deacetylase activity, have provided convergent evidence of the key role of these enzymes in aging-linked physiological functions. The link between SIRT1 and longevity has emerged in model organism but few data are available in humans, in particular relying on longitudinal studies. Here, we assessed whether a genetic variant within SIRT1 gene promoter (rs12778366) was associated to human longevity. We analyzed 586 genomic DNA (gDNA) collected in the study "Treviso Longeva" (TRELONG), including elderly over 70 years of age from the municipality of Treviso, a town in the Northeast of Italy, with a 11-year follow-up. We genotyped SIRT1 rs12778366 by real-time polymerase chain reaction (RT-PCR) allelic discrimination assay. A cross-sectional analysis performed by comparing people over and under 85 years of age did not evidence association between rs12778366 and longevity. When we performed a longitudinal analysis considering mortality as dependent variable, we did not observe an association of rs12778366 with longevity in the whole population (corrected P-value = 0.33). However, when we stratified the TRELONG subjects according to circulating level of interleukin-6 (IL-6), a predictor of disability and mortality, we found that rs12778366 (TC+CC) carriers were at increased risk of mortality in comparison to the TT reference group (corrected P-value = 0.03, HR 1.47). Our data do not support a major role of rs12778366 in human longevity, but the stratified analysis on IL-6 suggests that this variant may be involved in the detrimental effect of high circulating IL-6 in the elderly.

Amyotrophic Lateral Sclerosis (ALS) is a progressive and fatal disease due to motoneuron degeneration. Magnetic resonance imaging (MRI) is becoming a promising non-invasive approach to monitor the disease course but a direct correlation with neuropathology is not feasible in human. Therefore in this study we aimed to examine MRI changes in relation to histopathology in two mouse models of ALS (C57BL6/J and 129S2/SvHsd SOD1G93A mice) with different disease onset and progression. A longitudinal in vivo analysis of T2 maps, compared to ex vivo histological changes, was performed on cranial motor nuclei. An increased T2 value was associated with a significant tissue vacuolization that occurred prior to motoneuron loss in the cranial nuclei of C57 SOD1G93A mice. Conversely, in 129Sv SOD1G93A mice, which exhibit a more severe phenotype, MRI detected a milder increase of T2 value, associated with a milder vacuolization. This suggests that alteration within brainstem nuclei is not predictive of a more severe phenotype in the SOD1G93A mouse model. Using an ex vivo paradigm, Diffusion Tensor Imaging was also applied to study white matter spinal cord degeneration. In contrast to degeneration of cranial nuclei, alterations in white matter and axons loss reflected the different disease phenotype of SOD1G93A mice. The correspondence between MRI and histology further highlights the potential of MRI to monitor progressive motoneuron and axonal degeneration non-invasively in vivo. The identification of prognostic markers of the disease nevertheless requires validation in multiple models of ALS to ensure that these are not merely model-specific. Eventually this approach has the potential to lead to the development of robust and validated non-invasive imaging biomarkers in ALS patients, which may help to monitor the efficacy of therapies.

Fatal familial insomnia (FFI) and a genetic form of Creutzfeldt-Jakob disease (CJD178) are clinically different prion disorders linked to the D178N prion protein (PrP) mutation. The disease phenotype is determined by the 129 M/V polymorphism on the mutant allele, which is thought to influence D178N PrP misfolding, leading to the formation of distinctive prion strains with specific neurotoxic properties. However, the mechanism by which misfolded variants of mutant PrP cause different diseases is not known. We generated transgenic (Tg) mice expressing the mouse PrP homolog of the FFI mutation. These mice synthesize a misfolded form of mutant PrP in their brains and develop a neurological illness with severe sleep disruption, highly reminiscent of FFI and different from that of analogously generated Tg(CJD) mice modeling CJD178. No prion infectivity was detectable in Tg(FFI) and Tg(CJD) brains by bioassay or protein misfolding cyclic amplification, indicating that mutant PrP has disease-encoding properties that do not depend on its ability to propagate its misfolded conformation. Tg(FFI) and Tg(CJD) neurons have different patterns of intracellular PrP accumulation associated with distinct morphological abnormalities of the endoplasmic reticulum and Golgi, suggesting that mutation-specific alterations of secretory transport may contribute to the disease phenotype.

The age-specific prevalence rates of dementia vary widely. Studies focusing on specific age groups are needed to provide reliable estimates for healthcare providers and policy makers. We estimated the prevalence of dementia, dementia subtypes and cognitive impairment in "InveCe.Ab" (ClinicalTrials.gov, NCT01345110), a single-step multidimensional population-based study of 70-74-year olds living in Abbiategrasso (Milan, Italy). We also looked for associations with socio-demographic factors and the presence of the apolipoprotein E-ɛ4 allele. The overall dementia prevalence was 3% (95%CI: 2.1-4.1%) [Alzheimer's disease (AD): 1.2% (95%CI 0.6-1.9%); vascular dementia (VD): 1.4% (95%CI: 0.8-2.2%)]. Being single was found to be a risk factor for vascular dementia; subjects born in southern Italy were shown to be at greater risk both of overall dementia and of vascular dementia. The prevalence of cognitive impairment, with or without subjective cognitive complaints (cognitive impairment, no dementia, CIND) was 7.8% (95%CI: 6.4-9.4%). As regards the CIND subgroups, the prevalence of subjects with subjective cognitive complaints (mild cognitive impairment, MCI) was 5.0% (95%CI 3.9-6.3%), while the prevalence of those without MCI (CIND-other) was 2.8% (95%CI: 1.9-3.8). The males had a higher risk of MCI and CIND-other; the older subjects were more likely to have MCI, and those born in north-eastern Italy to have CIND-other. The prevalence of AD was higher among the apolipoprotein E-ɛ4 carriers. Our data highlight the importance of dementia and cognitive impairment in the transitional period from adulthood to old age, and reveal the presence of different associations with socio-demographic and genetic factors., (Copyright © 2014 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.)

Alzheimer's disease is experimentally modeled in transgenic (Tg) mice overexpressing mutated forms of the human amyloid precursor protein either alone or combined with mutated presenilins and tau. In the present study, we developed a systematic approach to compare double (TASTPM) and triple (APP/PS2/Tau) Tg mice by serial magnetic resonance imaging and spectroscopy analysis from 4 to 26 months of age to define homologous biomarkers between mice and humans. Hippocampal atrophy was found in Tg mice compared with WT. In APP/PS2/Tau the effect was age-dependent, whereas in TASTPM it was detectable from the first investigated time point. Importantly, both mice displayed an age-related entorhinal cortex thinning and robust striatal atrophy, the latter associated with a significant loss of synaptophysin. Hippocampal magnetic resonance spectroscopy revealed lower glutamate levels in both Tg mice and a selective myo-inositol increase in TASTPM. This noninvasive magnetic resonance imaging analysis, revealed common biomarkers between humans and mice, and could, thus, be promoted as a fully translational tool to be adopted in the preclinical investigation of therapeutic approaches., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Alzheimer's disease (AD) is the most common form of dementia characterized by synaptic dysfunction, memory loss, neuroinflammation, and neuronal cell death. Amyloid-β (Aβ), recognized as the main culprit of AD, aggregates and accumulates in the extracellular compartment as neuritic plaques, after deregulation of its production or clearance. Apolipoprotein E (ApoE) plays a major role in Aβ clearance and its expression is transcriptionally regulated by the liver X receptor and retinoid X receptors (RXRs) system. Bexarotene (BEXA), an RXR agonist that increases ApoE expression and microglia phagocytosis has been proposed as a promising therapy for AD, resolving both the amyloid pathology and memory loss. Despite the first compelling report, however, multiple failures have been documented, raising concern about whether BEXA could in fact become a novel disease-modifying strategy for AD. To help clarify this, we investigated the effect of BEXA in vivo at multiple levels in TASTPM transgenic mice. Seven-day oral administration of BEXA to these mice did not achieve any significant memory improvement, plaque reduction, or enhancement of microglial cell activation. No differences were found when specifically investigating the microglial phagocytic state in vivo. In addition, a brain structural analysis with magnetic resonance did not detect any BEXA-mediated change in the volume reduction of the main affected brain areas in our mice. These results suggest that BEXA has no beneficial effects on the multi-factorial pathologic phenotype of AD mice.

Background: Dopaminergic degeneration is a major finding in brains of patients with Parkinson's disease (PD), together with Lewy bodies, intraneuronal inclusions mainly composed of the fibrillogenic protein α-synuclein (α-syn). The familial-PD-related protein DJ-1 was reported to reduce dopaminergic degeneration triggered by α-syn or by the dopaminergic-selective neurotoxin 6-hydroxydopamine (6-OHDA)., Objective: The aim was to further investigate the role of DJ-1 in dopaminergic degeneration and to see whether a cell-permeable recombinant form of DJ-1 (TAT-DJ-1) could restore dopamine depletion in vivo, thus representing an innovative therapeutic approach., Methods: We developed in vitro (PC12/TetOn cells and mouse primary mesencephalic neurons) and in vivo models [including DJ-1 knockout (-/-) mice] to investigate DJ-1 in dopaminergic degeneration., Results: We found that in PC12/TetOn cells overexpressing α-syn with the familial-PD linked mutation A30P, DJ-1 silencing increased α-syn (A30P) toxicity. Primary mesencephalic neurons from DJ-1 (-/-) mice were more vulnerable to a cell-permeable form of α-syn (TAT-α-syn) and to 6-OHDA. Intrastriatally administered TAT-DJ-1 reduced 6-OHDA toxicity in vivo in C57BL/6 mice. Finally, when we injected TAT-α-syn (A30P) in the striatum of DJ-1 (-/-) animals, dopamine was depleted more than in the control strain., Conclusion: DJ-1 appears to have a protective role against dopaminergic degeneration triggered by α-syn or 6-OHDA, reinforcing the possible therapeutic importance of this protein in PD., (© 2014 S. Karger AG, Basel.)

The treatment of bipolar disorder (BD) usually requires combination therapies, with the critical issue of the emergence of adverse drug reactions (ADRs) and the possibility of low treatment adherence. Genetic polymorphisms are hypothesized to modulate the pharmacodynamics of psychotropic drugs, representing potential biological markers of ADRs. This study investigated genes involved in the regulation of neuroplasticity (BDNF, ST8SIA2), second messenger cascades (GSK3B, MAPK1, and CREB1), circadian rhythms (RORA), transcription (SP4, ZNF804A), and monoaminergic system (HTR2A and COMT) in the risk of neurological, psychic, autonomic, and other ADRs. Two independent samples of BD patients naturalistically treated were included (COPE-BD n = 147; STEP-BD n = 659). In the COPE-BD 34 SNPs were genotyped, while in the STEP-BD polymorphisms in the selected genes were extracted from the genome-wide dataset. Each ADRs group was categorized as absent-mild or moderate-severe and logistic regression with appropriate covariates was applied to identify possible risk genotypes/alleles. 58.5 and 93.5 % of patients were treated with mood stabilizers, 44.2 and 50.7 % were treated with antipsychotics, and 69.4 and 46.1 % were treated with antidepressants in the COPE-BD and STEP-BD, respectively. Our findings suggested that ST8SIA2 may be associated with psychic ADRs, as shown in the COPE-BD (rs4777989 p = 0.0017) and STEP-BD (rs56027313, rs13379489 and rs10852173). A cluster of RORA SNPs around rs2083074 showed an effect on psychic ADRs in the STEP-BD. Trends supporting the association between HTR2A and autonomic ADRs were found in both samples. Confirmations are needed particularly for ST8SIA2 and RORA since the few available data regarding their role in relation to psychotropic ADRs.

The text describes a preventive clinical trial with drug treatment in a very rare neurodegenerative disease (Fatal familial Insomnia, FFI) designed with the help of individuals at genetic risk of developing the disease, asymptomatic carriers, who have agreed to be exposed over a 10-year period to doxycycline, an antibiotic with anti-prion activity. At least 10 carriers of the FFI mutation over 42 y old will be treated with doxycycline (100 mg/die) and the incidence of the disease will be compared to that of an historical dataset. For ethical reasons a randomized, double-blind, placebo-controlled trial was not feasible, however the study design and the statistical analysis ensure the scientific value of the results. This approach might represent an important breakthrough in terms of potential therapy and knowledge of rare diseases that could give some hopes to these neglected patients.

Several studies have established the sortilin-related receptor gene (SORL1) as a susceptibility locus for Alzheimer's disease (AD). Single nucleotide polymorphisms of SORL1 reported in literature as being associated with AD were investigated in an Italian case-control data set, and their role as a risk factor of conversion to AD was studied in an independent sample of subjects diagnosed with mild cognitive impairment (MCI) at baseline. rs641120, rs2070045, and rs1010159 were genotyped in 734 subjects diagnosed with AD (n = 338) and MCI (n = 181) and in healthy controls (n = 215). Our results confirmed the association between rs641120 and AD (p = 0.01). In the MCI cohort, rs1010159 was associated with conversion to AD (HR = 1.56, p = 0.002). Taken together, these findings confirm that SORL1 is associated with AD and might be a potential tool for identifying MCI subjects at high risk of conversion to AD.

Alzheimer's disease is characterized by the accumulation and deposition of plaques of β-amyloid (Aβ) peptide in the brain. Given its pivotal role, new therapies targeting Aβ are in demand. We rationally designed liposomes targeting the brain and promoting the disaggregation of Aβ assemblies and evaluated their efficiency in reducing the Aβ burden in Alzheimer's disease mouse models. Liposomes were bifunctionalized with a peptide derived from the apolipoprotein-E receptor-binding domain for blood-brain barrier targeting and with phosphatidic acid for Aβ binding. Bifunctionalized liposomes display the unique ability to hinder the formation of, and disaggregate, Aβ assemblies in vitro (EM experiments). Administration of bifunctionalized liposomes to APP/presenilin 1 transgenic mice (aged 10 months) for 3 weeks (three injections per week) decreased total brain-insoluble Aβ1-42 (-33%), assessed by ELISA, and the number and total area of plaques (-34%) detected histologically. Also, brain Aβ oligomers were reduced (-70.5%), as assessed by SDS-PAGE. Plaque reduction was confirmed in APP23 transgenic mice (aged 15 months) either histologically or by PET imaging with [(11)C]Pittsburgh compound B (PIB). The reduction of brain Aβ was associated with its increase in liver (+18%) and spleen (+20%). Notably, the novel-object recognition test showed that the treatment ameliorated mouse impaired memory. Finally, liposomes reached the brain in an intact form, as determined by confocal microscopy experiments with fluorescently labeled liposomes. These data suggest that bifunctionalized liposomes destabilize brain Aβ aggregates and promote peptide removal across the blood-brain barrier and its peripheral clearance. This all-in-one multitask therapeutic device can be considered as a candidate for the treatment of Alzheimer's disease., (Copyright © 2014 the authors 0270-6474/14/3414013-09$15.00/0.)

Introduction: It has been reported that elderly subjects have a compromised ability to produce melatonin nightly, and that reduced melatonin levels may be a risk factor for cancer. The purpose of this study was to evaluate the relationship between melatonin levels and chronic diseases in a cohort of elderly subjects using the Charlson comorbidity index (CCI)., Design: We performed a secondary data analysis of a longitudinal study of a representative, age-stratified, sample population., Setting: The Treviso Longeva (Trelong) study, in Treviso, Italy., Participants: A total of 114 men and 146 women, aged 77 years and older, still alive after 7 years of follow-up., Measurements: As an estimation of serum melatonin secretion levels, urinary 6-sulfatoxymelatonin (aMT6s) was assayed in the urine of 260 elderly subjects using an enzyme-linked immunosorbent assay (ELISA) kit (product 01-EK-M6S, ALPCO Immunoassays, Windham, NH). All aMT6s levels were creatinine standardized ([aMT6s]/[creatinine]), and the CCI was calculated., Results: The melatonin levels decreased with aging despite not reaching statistical significance, and the decrease was more evident in males than in females (40.5 ng vs 47.0 ng aMT6s/mg creatinine, ns). Melatonin levels were significantly lower in patients reporting insomnia (p=0.05). The CCI score was inversely correlated with the levels of melatonin (p=0.03). Melatonin levels of subjects affected by CCI pathologies were significantly lower than those of healthy subjects (p=0.03) and of subjects suffering from diseases not included in the CCI and, therefore, less severe (p=0.03)., Conclusion: Melatonin appears to be a marker of disease state and severity, as well as of sleep disorders, in the elderly. These early findings would confirm the protective role of melatonin against several chronic diseases. The benefits of this agent as a possible medication should be more thoroughly clinically tested.

Spinal cord injury (SCI) is the result of a traumatic primary event followed by a so-called secondary injury, which is characterized by a large spectrum of biochemical cellular pathways able to spread the lesion, worsening neurologic recovery. A growing number of potential therapeutic interventions to counteract different neurodegenerative mechanisms of SCI have been proposed, but they did not show relevant efficacy when translated as clinical treatments. Different reasons could explain these disappointing results: on one side the multifactorial evolution of SCI after the primary injury that limits the beneficial effect of just one targeted treatment and, on the other, the restricted access of pharmacological therapies to the spinal cord. For these reasons, recently, a growing interest has been shown in the development of alternative delivery strategies to administer drugs and/or biological/cellular therapies into the spine (hydrogel and nanoparticles)., (© 2014 Wiley Periodicals, Inc.)

Background: The purpose of the study was to evaluate the applicability of (18) F-labelled amyloid imaging positron emission tomography (PET) agent [ (18) F]flutemetamol to detect changes in brain beta-amyloid (Aβ) deposition in vivo in APP23, Tg2576 and APPswe-PS1dE9 mouse models of Alzheimer's disease. We expected that the high specific activity of [ (18) F]flutemetamol would make it an attractive small animal Aβ imaging agent., Methods: [ (18) F]flutemetamol uptake in the mouse brain was evaluated in vivo at 9 to 22 months of age with an Inveon Multimodality PET/CT camera (Siemens Medical Solutions USA, Knoxville, TN, USA). Retention in the frontal cortex (FC) was evaluated by Logan distribution volume ratios (DVR) and FC/cerebellum (CB) ratios during the late washout phase (50 to 60 min). [ (18) F]flutemetamol binding to Aβ was also evaluated in brain slices by in vitro and ex vivo autoradiography. The amount of Aβ in the brain slices was determined with Thioflavin S and anti-Aβ1-40 immunohistochemistry., Results: In APP23 mice, [ (18) F]flutemetamol retention in the FC increased from 9 to 18 months. In younger mice, DVR and FC/CB50-60 were 0.88 (0.81) and 0.88 (0.89) at 9 months (N = 2), and 0.98 (0.93) at 12 months (N = 1), respectively. In older mice, DVR and FC/CB50-60 were 1.16 (1.15) at 15 months (N = 1), 1.13 (1.16) and 1.35 (1.35) at 18 months (N = 2), and 1.05 (1.31) at 21 months (N = 1). In Tg2576 mice, DVR and FC/CB50-60 showed modest increasing trends but also high variability. In APPswe-PS1dE9 mice, DVR and FC/CB50-60 did not increase with age. Thioflavin S and anti-Aβ1-40 positive Aβ deposits were present in all transgenic mice at 19 to 22 months, and they co-localized with [ (18) F]flutemetamol binding in the brain slices examined with in vitro and ex vivo autoradiography., Conclusions: Increased [ (18) F]flutemetamol retention in the brain was detected in old APP23 mice in vivo. However, the high specific activity of [ (18) F]flutemetamol did not provide a notable advantage in Tg2576 and APPswe-PS1dE9 mice compared to the previously evaluated structural analogue [(11)C]PIB. For its practical benefits, [ (18) F]flutemetamol imaging with a suitable mouse model like APP23 is an attractive alternative.

Background: Frontotemporal dementia (FTD) is a complex disorder characterised by a broad range of clinical manifestations, differential pathological signatures, and genetic variability. Mutations in three genes-MAPT, GRN, and C9orf72--have been associated with FTD. We sought to identify novel genetic risk loci associated with the disorder., Methods: We did a two-stage genome-wide association study on clinical FTD, analysing samples from 3526 patients with FTD and 9402 healthy controls. To reduce genetic heterogeneity, all participants were of European ancestry. In the discovery phase (samples from 2154 patients with FTD and 4308 controls), we did separate association analyses for each FTD subtype (behavioural variant FTD, semantic dementia, progressive non-fluent aphasia, and FTD overlapping with motor neuron disease [FTD-MND]), followed by a meta-analysis of the entire dataset. We carried forward replication of the novel suggestive loci in an independent sample series (samples from 1372 patients and 5094 controls) and then did joint phase and brain expression and methylation quantitative trait loci analyses for the associated (p<5 × 10(-8)) single-nucleotide polymorphisms., Findings: We identified novel associations exceeding the genome-wide significance threshold (p<5 × 10(-8)). Combined (joint) analyses of discovery and replication phases showed genome-wide significant association at 6p21.3, HLA locus (immune system), for rs9268877 (p=1·05 × 10(-8); odds ratio=1·204 [95% CI 1·11-1·30]), rs9268856 (p=5·51 × 10(-9); 0·809 [0·76-0·86]) and rs1980493 (p value=1·57 × 10(-8), 0·775 [0·69-0·86]) in the entire cohort. We also identified a potential novel locus at 11q14, encompassing RAB38/CTSC (the transcripts of which are related to lysosomal biology), for the behavioural FTD subtype for which joint analyses showed suggestive association for rs302668 (p=2·44 × 10(-7); 0·814 [0·71-0·92]). Analysis of expression and methylation quantitative trait loci data suggested that these loci might affect expression and methylation in cis., Interpretation: Our findings suggest that immune system processes (link to 6p21.3) and possibly lysosomal and autophagy pathways (link to 11q14) are potentially involved in FTD. Our findings need to be replicated to better define the association of the newly identified loci with disease and to shed light on the pathomechanisms contributing to FTD., Funding: The National Institute of Neurological Disorders and Stroke and National Institute on Aging, the Wellcome/MRC Centre on Parkinson's disease, Alzheimer's Research UK, and Texas Tech University Health Sciences Center., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Introduction: Curcumin is a neuroprotective compound that inhibits the formation of amyloid oligomers and fibrils and binds to β-amyloid plaques in Alzheimer's disease (AD). We aimed to synthesize an (18)F-labeled curcumin derivate ([(18)F]4) and to characterize its positron emission tomography (PET) tracer-binding properties to β-amyloid plaques in a transgenic APP23 mouse model of AD., Methods: We utilized facile one-pot synthesis of [(18)F]4 using nucleophilic (18)F-fluorination and click chemistry. Binding of [(18)F]4 to β-amyloid plaques in the transgenic APP23 mouse brain cryosections was studied in vitro using heterologous competitive binding against PIB. [(18)F]4 uptake was studied ex vivo in rodents and in vivo using PET/computed tomography of transgenic APP23 and wild-type control mice., Results: The radiochemical yield of [(18)F]4 was 21 ± 11%, the specific activity exceeded 1TBq/μmol, and the radiochemical purity exceeded 99.3% at the end of synthesis. In vitro studies of [(18)F]4 with the transgenic APP23 mouse revealed high β-amyloid plaque binding. In vivo and ex vivo studies demonstrated that [(18)F]4 has fast clearance from the blood, moderate metabolism but low blood-brain barrier (BBB) penetration., Conclusions: [(18)F]4 was synthesized in high yield and excellent quality. In vitro studies, metabolite profile, and fast clearance from the blood indicated a promising tracer for Aβ imaging. However, [(18)F]4 has low in vivo BBB penetration and thus further studies are needed to reveal the reason for this and to possibly overcome this issue., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Background: Creutzfeldt-Jakob disease (CJD) is a fatal, untreatable prion encephalopathy. Previous studies showed that doxycycline is effective in in-vitro and in-vivo models of disease, and patients with CJD who received compassionate treatment with doxycycline showed increased survival time compared with historical series. We therefore did a randomised, double-blind study of doxycycline versus placebo in CJD., Methods: We recruited patients older than 18 years old who had a diagnosis of definite or probable sporadic CJD or genetic forms of the disease via Italian reference centres and the French national referral system. Patients were randomly assigned (ratio 1:1) to receive oral doxycycline (100 mg daily) or placebo under double-blind conditions from the day of randomisation to death. Centralised randomisation was done independently of enrolment or evaluation of patients using a minimisation method in Italy and a simple randomisation in France. Participants, caregivers, and clinicians were masked to group assignment. The primary efficacy variable was the survival time from randomisation. Interim analyses were planned to detect a significant effect of treatment as early as possible. This trial is registered with EudraCT, 2006-001858-27 for the Italian study and 2007-005553-34 for the French study., Findings: From April 12, 2007, to Aug 19, 2010, in Italy, and from Jan 30, 2009, to Jan 10, 2012, in France, 121 patients with CJD were enrolled in the study, 62 of whom were randomly assigned to the treatment group and 59 to the placebo group. The first interim analysis showed absence of superiority of doxycycline compared with placebo, and the trial was stopped for futility. Efficacy analyses did not show significant differences between patients treated with doxycycline and placebo with regard to survival times (HR 1.1, 95% CI 0.8-1.7, p=0.50). Serious adverse events were judged not to be related to treatment, whereas a relation was deemed probable or possible for five non-serious adverse events that occurred in each treatment group., Interpretation: Doxycycline at a dose of 100 mg per day was well tolerated but did not significantly affect the course of CJD, at variance with the results of previous observational studies. Our experience could be useful in the design of large multinational controlled trials of potential anti-prion molecules in this rare disease., Funding: Agenzia Italiana Farmaco, Italian Ministry of Health, AIEnP, and French Ministry of Health., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Human sirtuins are seven proteins with deacetylase activity that are emerging as key modulators of basic physiological functions. Some evidence links SIRT3 to longevity in mammals. This study aimed to investigate whether variants within SIRT3 gene were associated to human longevity. We analyzed 549 genomic DNA collected during the prospective study "Treviso Longeva," including elderly over 70 years of age from the municipality of Treviso, a small city in the northeast of Italy. We genotyped SIRT3 rs3825075, rs4980329, and rs11555236 single nucleotide polymorphisms (SNPs) by real-time polymerase chain reaction allelic discrimination assay. A cross-sectional analysis performed by comparing people over and under 85 years of age did not evidence association among the SIRT3 SNPs and longevity. However, when we performed a longitudinal analysis considering mortality as a dependent variable, we observed an association of SIRT3 rs11555236 and rs4980329 with longevity in the whole population (p values corrected for potential confounders = 0.04 and 0.03, respectively). After stratification according to gender, the same SNPs were associated to female longevity only (p values corrected for potential confounders = 0.03 and 0.02, respectively). Finally, as rs11555236 was reported to be in linkage disequilibrium with a putative functional enhancer within the SIRT3 gene, we assessed whether rs11555236 genotypes correlated with a different level of SIRT3 protein in peripheral blood mononuclear cells. We found an increased level of SIRT3 in subjects homozygous for the (T) allele. We suggest that SIRT3 genetic variability might be relevant for the modulation of human longevity in the Italian population.

The possibility to control the fate of the cells responsible for secondary mechanisms following spinal cord injury (SCI) is one of the most relevant challenges to reduce the post traumatic degeneration of the spinal cord. In particular, microglia/macrophages associated inflammation appears to be a self-propelling mechanism which leads to progressive neurodegeneration and development of persisting pain state. In this study we analyzed the interactions between poly(methyl methacrylate) nanoparticles (PMMA-NPs) and microglia/macrophages in vitro and in vivo, characterizing the features that influence their internalization and ability to deliver drugs. The uptake mechanisms of PMMA-NPs were in-depth investigated, together with their possible toxic effects on microglia/macrophages. In addition, the possibility to deliver a mimetic drug within microglia/macrophages was characterized in vitro and in vivo. Drug-loaded polymeric NPs resulted to be a promising tool for the selective administration of pharmacological compounds in activated microglia/macrophages and thus potentially able to counteract relevant secondary inflammatory events in SCI., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Environmental enrichment (EE) is a non-pharmacological intervention reported to counteract pathological signs in models of Alzheimer's disease (AD). We developed EE protocols in APP23 mice and evaluated how they influenced cognitive decline and brain amyloid-β (Aβ) burden. We also investigated the involvement of sirtuins (SIRTs) as a possible molecular mediator of EE, by assessing hippocampal and cortical mRNA and protein levels of the SIRT family members (SIRT1 to SIRT7). APP23 transgenic mice were moved to EE cages (TG-EEs) starting from 3 months of age. TG-EEs were compared to transgenic mice housed in standard cages (TG-SHs) and to wild-type littermates in the two housing conditions (WT-EEs and WT-SHs). At 7 months of age, all mice were tested for behavioral performance with Morris Water Maze (MWM) and visual novel Object Recognition Test (vORT). After a month, a group underwent biochemical analyses, while another group continued in the EE environment till 18 months of age, when Aβ plaque load was assessed. At 7 months, TG-SHs had impaired behavioral performance in MWM and vORT. In contrast, TG-EE mice had restored behavioral performance. At 8 months, EE did not affect AβPP expression or processing, Aβ40/42, pGlu-Aβ3-40/3-42, or Aβ oligomer level. The expression of two Aβ degrading enzymes (insulin degrading enzyme and neprilysin) was not modulated by EE. Brain sirtuin mRNA and protein levels were unchanged, while brain-derived neurotrophic factor expression increased after EE. Aβ deposition was attenuated in 18-month-old TG-EE mice, without apparent reduction of neuroinflammatory signs. We suggest that EE had a beneficial effect on cognitive performance and lessened long-term Aβ accumulation, but brain sirtuin expression was not modulated when cognitive impairment was restored.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by cerebral accumulation of extracellular amyloid β (Aβ) and neurofibrillary tangles made of hyperphosphorylated tau protein, two main lesions which appear sequentially during the disease progression. In the last decade numerous studies have proposed small soluble aggregates of Aβ, known as oligomers, as the species responsible for synaptic dysfunction, memory loss and neurodegeneration typical of AD. In vitro and in vivo experiments have identified Aβ oligomers as the elements that can alter synaptic function by a reversible mechanism, which gradually becomes permanent when exposure is continuous. Here we show that intracerebroventricular (ICV) injection in mice of a solution containing specifically Aβ1-42 oligomers substantially affects their memory when tested in the novel object recognition task. This acute mouse model enabled us to distinguish whether oligomers were affecting specific phases of the memory processing. A single injection of Aβ1-42 oligomers before memory consolidation abolished information processing, leading to memory impairment, whereas no such effects were observed when the injection was done once the information had been processed, indicating that the oligomers affect memory consolidation rather than retrieval. Beside Aβ1-42, Aβ1-40 oligomers also impaired memory, and both isoforms were antagonized by the anti-Aβ4G8 monoclonal antibody. This simple and reliable paradigm is useful to investigate the mechanisms through which Aβ oligomers interfere with neuronal processes and to test the efficacy of new therapeutic approaches specifically against these species. We tested several molecules by direct coincubation with Aβ oligomers, ICV injections preceding Aβ oligomers, and the systemic treatment with drugs that cross the blood brain barrier. We also examined the proposed involvement of cellular prion protein as a mediator of the oligomer-induced memory impairment.