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Parkinson’s disease is a neurodegenerative disorder characterized by the progressive dysfunction and loss of dopamine (DA) neurons of the substantia nigra pars compacta (SNc). Several pathways of programmed cell death are likely to play a role in DA neuron death, such as apoptosis, necrosis, pyroptosis, ferroptosis as well as cell death associated with proteasomal and mitochondrial dysfunction. A better understanding of the molecular mechanisms underlying DA neuron death could inform the design of drugs that promote neuron survival. Necroptosis is a recently characterized regulated cell death mechanism that exhibits morphological features common to both apoptosis and necrosis. It requires activation of an intracellular pathway involving receptor-interacting protein 1 (RIP1) and its kinase (RIP1 kinase, RIPK1), receptor-interacting protein 3 (RIP3) and its kinase (RIP3 kinase, RIPK3), and mixed lineage kinase domain like pseudokinase (MLKL). The potential involvement of this programmed cell death pathway in the pathogenesis of PD has been studied by analyzing the biomarkers for necroptosis, such as the levels and oligomerization of pRIPK3 and pMLKL, in several PD preclinical models and in PD human tissue. While there is evidence that other types of cell death also have a role in DA neuron death, most studies support the hypothesis that this cell death mechanism is activated in PD tissues. Thus drugs that prevent or reduce necroptosis may provide neuroprotection for PD. In this review, we summarize the findings from these studies. We also discuss how manipulating necroptosis might open a novel therapeutic approach to reduce neuronal degeneration in PD.

ObjectivesAlexander disease (AD) is a rare disorder of the CNS. Diagnosis is based on clinical symptoms, typical MRI findings, and mutations in the glial fibrillary acid protein (GFAP) gene. In this case study, we describe a new mutation (p.L58P) in GFAP that caused a phenotype of adult-onset AD (AOAD).MethodsIn our outpatient clinic, a patient presented with cerebellar and bulbar symptoms after brain concussion. We used MRI and performed next-generation exome sequencing (NGS) to find mutations in GFAP to diagnose AD. The mutation was then transfected into HeLa cell lines to prove its pathogenicity.ResultsThe brain MRI finding showed typical AD alterations. The NGS found a heterozygous variant of unknown significance in GFAP (c.173T>C; p.L58P). After transfecting HeLa cell lines with this mutation, we showed that GFAP-L58P formed pathogenic clusters of cytoplasmic aggregates.DiscussionWe have found a new mutation that causes AOAD. We recommend that AOAD is included in the diagnostic workup in adult patients with gait ataxia and cerebellar and bulbar symptoms in association with a traumatic head injury.

The Substantia Nigra pars compacta (SNc) is a midbrain dopaminergic nucleus that plays a key role in modulating motor and cognitive functions. It is crucially involved in several disorders, particularly Parkinson's disease, which is characterized by a progressive loss of SNc dopaminergic cells. Electrophysiological studies on SNc neurons are of paramount importance to understand the role of dopaminergic transmission in health and disease. Here, we provide an extensive protocol to prepare SNc-containing mouse brain slices and record the electrical activity of dopaminergic cells. We describe all the necessary steps, including mouse transcardiac perfusion, brain extraction, slice cutting, and patch-clamp recordings.

Introduction: The study aims at investigating psychometric properties of the Edinburgh cognitive and behavioural ALS screen (ECAS) in Parkinson’s (PD) and Huntington’s (HD) diseases. The sensitivity and specificity of the ECAS in highlighting HD and PD cognitive-behavioural features and in differentiating between these two populations and from healthy controls (HC) were evaluated. Moreover, correlations between the ECAS and traditional cognitive measures, together with core clinical features, were analysed. Methods: Seventy-three PD patients, 38 HD patients, and 49 education-matched healthy participants were enrolled. Participants were administered the ECAS, together with other cognitive screening tools and psychological questionnaires. Patients’ behavioural assessment was also carried out with carers. Results: The ECAS distinguished between HD patients and HC and between the two clinical syndromes with high sensitivity and specificity. Even if the diagnostic accuracy of the ECAS in distinguishing between PD and HC was low, the PD cognitive phenotype was very well described by the ECAS performances. Convergent validity of the ECAS against other traditional cognitive screening was observed, as well as correlations with psychological aspects and typical clinical features, especially for the HD group. Conclusions: The ECAS represents a rapid and feasible tool, useful also in other neurodegenerative disorders affecting verbal-motor abilities than the amyotrophic lateral sclerosis such as PD and HD. Clinical applications in these neurodegenerative conditions require further investigations and, probably, some adaptations of the original test.

Huntington's disease (HD) is a neurodegenerative disorder characterized by motor disturbances, cognitive decline, and behaviour changes. A well-recognized feature of advanced HD is dysphagia, which leads to malnutrition and aspiration pneumonia, the latter being the primary cause of death in HD. Previous studies have underscored the importance of dysphagia in HD patients with moderate-to-advanced stage disease, but it is unclear whether dysphagia affects patients already at an early stage of disease and whether genetic or clinical factors can predict its severity. We performed fiberoptic endoscopic evaluation of swallowing (FEES) in 61 patients with various stages of HD. Dysphagia was found in 35% of early-stage, 94% of moderate-stage, and 100% of advanced-stage HD. Silent aspiration was found in 7.7% of early-stage, 11.8% of moderate-stage, and 27.8% of advanced-stage HD. A strong correlation was observed between disease progression and dysphagia severity: worse dysphagia was associated with worsening of motor symptoms. Dysphagia severity as assessed by FEES correlated with Huntington’s Disease Dysphagia Scale scores (a self-report questionnaire specific for evaluating swallowing in HD). The present findings add to our understanding of dysphagia onset and progression in HD. A better understanding of dysphagia onset and progression in HD may inform guidelines for standard clinical care in dysphagia, its recognition, and management.

BackgroundMutations in the PARK2 gene encoding the protein parkin cause autosomal recessive juvenile parkinsonism (ARJP), a neurodegenerative disease characterized by dysfunction and death of dopamine (DA) neurons in the substantia nigra pars compacta. Since a neuroprotective therapy for ARJP does not exist, research efforts aimed at discovering potential targets for neuroprotection are critically needed.A previous study demonstrated that loss of parkin function or expression of parkin mutants associated with ARJP causesan accumulation of glutamate kainate receptors (KARs) in human brain tissues and an increase of KAR-mediated currents in neuronsin vitro. MethodsBased on the hypothesisthat such KAR hyper-activation may contribute to the death of nigralDA neurons, we investigated the effect of KAR antagonism on the DA neuron dysfunction and death that occur in the parkinQ311X mouse, a model of human parkin-induced toxicity. ResultsWe found that early accumulation of KARs occurs in the DA neurons of the parkinQ311X mouse model and that chronic administration of the KAR antagonist UBP310 prevents DA neuron loss. This neuroprotective effect was associated with rescue of the abnormal firing rate of nigral DA neurons and downregulation of GluK2, the key KAR subunit. ConclusionsThis study provides novel evidence ofa causal role of glutamate KARs in the DA neuron dysfunction and loss occurring in a mouse model of human parkin-induced toxicity. Our results support KAR as a potential target in the development of a neuroprotective therapy for ARJP.

The vacuolar protein sorting 35 (VPS35) gene located on chromosome 16 has recently emerged as a cause of late-onset familial Parkinson’s disease (PD) (PARK17). The gene encodes a 796-residue protein nearly ubiquitously expressed in human tissues. The protein localizes on endosomes where it assembles with other peripheral membrane proteins to form the retromer complex. How VPS35 mutations induce dopaminergic neuron degeneration in humans is still unclear. Because the retromer complex recycles the receptors that mediate the transport of hydrolase to lysosome, it has been suggested that VPS35 mutations lead to impaired lysosomal and autophagy function. Recent studies also demonstrated that VPS35 and the retromer complex influence mitochondrial homeostasis, suggesting that VPS35 mutations elicit mitochondrial dysfunction. More recent studies have identified a key role of VPS35 in neurotransmission, whilst others reported a functional interaction between VPS35 and other genes associated with familial PD, including α-SYNUCLEIN-PARKIN–LRRK2. Here, we review the biological role of VPS35 protein, the VPS35 mutations identified in human PD patients, and the potential molecular mechanism by which VPS35 mutations can induce progressive neurodegeneration in PD.

Neurodevelopmental disorders (NDDs) are a group of diseases whose symptoms arise during childhood or adolescence and that impact several higher cognitive functions such as learning, sociability and mood. Accruing evidence suggests that a shared pathogenic mechanism underlying these diseases is the dysfunction of glutamatergic synapses. We summarize present knowledge on autism spectrum disorders (ASD), intellectual disability (ID), Down syndrome (DS), Rett syndrome (RS) and attention-deficit hyperactivity disorder (ADHD), highlighting the involvement of glutamatergic synapses and receptors in these disorders. The most commonly shared defects involve α-amino-3-hydroxy-5-methyl- 4-isoxazole propionic acid receptors (AMPARs), N-methyl-. d-aspartate receptors (NMDARs) and metabotropic glutamate receptors (mGluRs), whose functions are strongly linked to synaptic plasticity, affecting both cell-autonomous features as well as circuit formation. Moreover, the major scaffolding proteins and, thus, the general structure of the synapse are often deregulated in neurodevelopmental disorders, which is not surprising considering their crucial role in the regulation of glutamate receptor positioning and functioning. This convergence of defects supports the definition of neurodevelopmental disorders as a continuum of pathological manifestations, suggesting that glutamatergic synapses could be a therapeutic target to ameliorate patient symptomatology.

Mutations in the PARKIN gene cause early-onset Parkinson’s disease (PD). Despite the high proportion of still missing phenotyping data in the literature devoted to early-onset PD, studies suggest that, as compared with late-onset PD, PARKIN patients show dystonia at onset and extremely dose-sensitive levodopa-induced dyskinesia (LID). What pathophysiological mechanisms underpin such early and atypical dyskinesia in patients with PARKIN mutations? Though the precise mechanisms underlying dystonia and LID are still unclear, evidence suggests that hyperkinetic disorders in PD are a behavioural expression of maladaptive functional and morphological changes at corticostriatal synapses induced by long-term dopamine (DA) depletion. However, since the dyskinesia in PARKIN patients can also be present at onset, other mechanisms beside the well-established DA depletion may play a role in the development of dyskinesia in these patients. Because cortical and striatal neurons express parkin protein, and parkin modulates the function of ionotropic glutamatergic receptors (iGluRs), an intriguing explanation may rest on the potential role of parkin in directly controlling the glutamatergic corticostriatal synapse transmission. We discuss the novel theory that loss of parkin function can dysregulate transmission at the corticostriatal synapses where they cause early maladaptive changes that co-occur with the changes stemming from DA loss. This hypothesis suggests an early striatal synaptopathy; it could lay the groundwork for pharmacological treatment of dyskinesias and LID in patients with PARKIN mutations.

Growing evidence suggests that amyotrophic lateral sclerosis (ALS) is a multisystem neurodegenerative disease that primarily affects motor neurons and, though less evidently, other neuronal systems. About 75% of sporadic and familial ALS patients show a subclinical degeneration of small-diameter fibers, as measured by loss of intraepidermal nerve fibers (IENFs), but the underlying biological causes are unknown. Small-diameter fibers are derived from small-diameter sensory neurons, located in dorsal root ganglia (DRG), whose biochemical hallmark is the expression of type III intermediate filament peripherin. We tested here the hypothesis that small-diameter DRG neurons of ALS mouse model SOD1(G93A)suffer from axonal stress and investigated the underlying molecular mechanism. We found that SOD1(G93A)mice display small fiber pathology, as measured by IENF loss, which precedes the onset of the disease. In vitro small-diameter DRG neurons of SOD1(G93A)mice show axonal stress features and accumulation of a peripherin splice variant, named peripherin56, which causes axonal stress through disassembling light and medium neurofilament subunits (NFL and NFM, respectively). Our findings first demonstrate that small-diameter DRG neurons of the ALS mouse model SOD1(G93A)display axonal stress in vitro and in vivo, thus sustaining the hypothesis that the effects of ALS disease spread beyond motor neurons. These results suggest a molecular mechanism for the small fiber pathology found in ALS patients. Finally, our data agree with previous findings, suggesting a key role of peripherin in the ALS pathogenesis, thus highlighting that DRG neurons mirror some dysfunctions found in motor neurons.

Huntington's Disease (HD) is a neurodegenerative disorder caused by a CAG expansion in the exon-1 of the IT15 gene encoding the protein Huntingtin. Expression of mutated Huntingtin in humans leads to dysfunction and ultimately degeneration of selected neuronal populations of the striatum and cerebral cortex. Current available HD therapy relies on drugs to treat chorea and control psychiatric symptoms, however, no therapy has been proven to slow down disease progression or prevent disease onset. Thus, although 24 years have passed since HD gene identification, HD remains a relentless progressive disease characterized by cognitive dysfunction and motor disability that leads to death of the majority of patients, on average 10-20 years after its onset. Up to now several molecular pathways have been implicated in the process of neurodegeneration involved in HD and have provided potential therapeutic targets. Based on these data, approaches currently under investigation for HD therapy aim on the one hand at getting insight into the mechanisms of disease progression in a human-based context and on the other hand at silencing mHTT expression by using antisense oligonucleotides. An innovative and still poorly investigated approach is to identify new factors that increase neurogenesis and/or induce reprogramming of endogenous neuroblasts and parenchymal astrocytes to generate new healthy neurons to replace lost ones and/or enforce neuroprotection of pre-existent striatal and cortical neurons. Here, we review studies that use human disease-in-a-dish models to recapitulate HD pathogenesis or are focused on promoting in vivo neurogenesis of endogenous striatal neuroblasts and direct neuronal reprogramming of parenchymal astrocytes, which combined with neuroprotective protocols bear the potential to re-establish brain homeostasis lost in HD.

Loss-of-function caused by mutations in the parkin gene (PARK2) lead to early-onset familial Parkinson's disease. Recently, mechanistic studies proved the ability of parkin in regulating mitochondria homeostasis and microtubule (MT) stability. Looking at these systems during aging of PARK2 knockout mice, we found that loss of parkin induced an accelerated (over)acetylation of MT system both in dopaminergic neuron cell bodies and fibers, localized in the substantia nigra and corpus striatum, respectively. Interestingly, in PARK2 knockout mice, changes of MT stability preceded the alteration of mitochondria transport. Moreover, in-cell experiments confirmed that loss of parkin affects mitochondria mobility and showed that this defect depends on MT system as it is rescued by paclitaxel, a well-known MT-targeted agent. Furthermore, both in PC12 neuronal cells and in patients' induced pluripotent stem cell-derived midbrain neurons, we observed that parkin deficiencies cause the fragmentation of stable MTs. Therefore, we suggest that parkin acts as a regulator of MT system during neuronal aging, and we endorse the hypothesis that MT dysfunction may be crucial in the pathogenesis of Parkinson's disease. (C) 2017 Elsevier Inc. All rights reserved.

Huntington's disease (HD) is a neurodegenerative disorder caused by the expansion of a CAG repeat in the IT15 gene that encodes the protein huntingtin (htt). Evidence shows that mutant htt causes mitochondrial depolarization and fragmentation, but the underlying molecular mechanism has yet to be clarified. Bax/Bak and BNip3 are pro-apoptotic members of the Bcl-2 family protein whose activation triggers mitochondrial depolarization and fragmentation inducing cell death. Evidence suggests that Bax/Bak and BNip3 undergo activation upon mutant htt expression but whether these proteins are required for mitochondrial depolarization and fragmentation induced by mutant htt is unclear. Our results show that BNip3 knock-out cells are protected from mitochondrial damage and cell death induced by mutant htt whereas Bax/Bak knock-out cells are not. Moreover, deletion of BNip3 C-terminal transmembrane domain, required for mitochondrial targeting, suppresses mitochondrial depolarization and fragmentation in a cell culture model of HD. Hence, our results suggest that changes in mitochondrial morphology and transmembrane potential, induced by mutant htt protein, are dependent and linked to BNip3 and not to Bax/Bak activation. These results provide new compelling evidence that underlies the molecular mechanisms by which mutant htt causes mitochondrial dysfunction and cell death, suggesting BNip3 as a potential target for HD therapy.

Loss of function mutations in the gene PARK2, which encodes the protein parkin, cause autosomal recessive juvenile parkinsonism, a neurodegenerative disease characterized by degeneration of the dopaminergic neurons localized in the substantia nigra pars compacta. No therapy is effective in slowing disease progression mostly because the pathogenesis of the disease is yet to be understood. From accruing evidence suggesting that the protein parkin directly regulates synapses it can be hypothesized that PARK2 gene mutations lead to early synaptic damage that results in dopaminergic neuron loss over time. We review evidence that supports the role of parkin in modulating excitatory and dopaminergic synapse functions. We also discuss how these findings underpin the concept that autosomal recessive juvenile parkinsonism can be primarily a synaptopathy. Investigation into the molecular interactions between parkin and synaptic proteins may yield novel targets for pharmacologic interventions.

The complex architecture of adult brain derives from tightly regulated migration and differentiation of precursor cells generated during embryonic neurogenesis. Changes at transcriptional level of genes that regulate migration and differentiation may lead to neurodevelopmental disorders. Androgen receptor (AR) is a transcription factor that is already expressed during early embryonic days. However, AR role in the regulation of gene expression at early embryonic stage is yet to be determinate. Long non-coding RNA (lncRNA) Sox2 overlapping transcript (Sox2OT) plays a crucial role in gene expression control during development but its transcriptional regulation is still to be clearly defined. Here, using Bicalutamide in order to pharmacologically inactivated AR, we investigated whether AR participates in the regulation of the transcription of the lncRNASox2OTat early embryonic stage. We identified a new DNA binding region upstream of Sox2 locus containing three androgen response elements (ARE), and found that AR binds such a sequence in embryonic neural stem cells and in mouse embryonic brain. Our data suggest that through this binding, AR can promote the RNA polymerase II dependent transcription of Sox2OT. Our findings also suggest that AR participates in embryonic neurogenesis through transcriptional control of the long non-coding RNA Sox2OT.

Itch is thought to represent the peculiar response to stimuli conveyed by somatosensory pathways shared with pain through the activation of specific neurons and receptors. It can occur in association with dermatological, systemic and neurological diseases, or be the side effect of certain drugs. However, some patients suffer from chronic idiopathic itch that is frequently ascribed to psychological distress and for which no biomarker is available to date. We investigated three multigenerational families, one of which diagnosed with joint hypermobility syndrome/Ehlers-Danlos syndrome hypermobility type (JHS/EDS-HT), characterized by idiopathic chronic itch with predominantly proximal distribution. Skin biopsy was performed in all eight affected members and revealed in six of them reduced intraepidermal nerve fibre density consistent with small fibre neuropathy. Whole exome sequencing identified two COL6A5 rare variants co-segregating with chronic itch in eight affected members and absent in non-affected members, and in one unrelated sporadic patient with type 1 painless diabetic neuropathy and chronic itch. Two families and the diabetic patient carried the nonsense c.6814G>T (p.Glu2272*) variant and another family carried the missense c.6486G>C (p.Arg2162Ser) variant. Both variants were predicted as likely pathogenic by in silico analyses. The two variants were rare (minor allele frequency < 0.1%) in 6271 healthy controls and absent in 77 small fibre neuropathy and 167 JHS/EDS-HT patients without itch. Null-allele test on cDNA from patients' fibroblasts of both families carrying the nonsense variant demonstrated functional haploinsufficiency due to activation of nonsense mediated RNA decay. Immunofluorescence microscopy and western blotting revealed marked disorganization and reduced COL6A5 synthesis, respectively. Indirect immunofluorescence showed reduced COL6A5 expression in the skin of patients carrying the nonsense variant. Treatment with gabapentinoids provided satisfactory itch relief in the patients carrying the mutations. Our findings first revealed an association between COL6A5 gene and familiar chronic itch, suggesting a new contributor to the pathogenesis of neuropathic itch and identifying a new candidate therapeutic target.

Background RAB39B pathogenic variants cause X-linked Parkinsonism associated with Intellectual Disability, known as Waisman syndrome, a very rare disorder that has been mainly identified through exome sequencing in large Parkinson's disease cohorts. In this study we searched for pathogenic variants in RAB39B in two Italian families affected by X-linked early-onset Parkinsonism and Intellectual Disability. Methods Three patients received neurological evaluation and underwent RAB39B sequencing. Results Two novel RAB39B frameshift variants were found to result in the absence of RAB39B protein (family 1: c.137dupT; family 2: c.371delA). Patients showed unilateral rest tremor and bradykinesia; one of them also displayed an early-onset postural tremor. Paramagnetic substance deposition in the substantia nigra, globus pallidi, red nucleus, putamen and pulvinar was assessed by brain imaging. Two patients also showed moderate calcification of globus pallidi. Conclusion In this study we highlight the evidence that X-linked early-onset Parkinsonism associated with Intellectual Disability occurs as a pattern of clinical and neuroimaging features attributable to RAB39B pathogenic variants.

Mutations in Cu/Zn-superoxide dismutase (SOD1) cause familial amyotrophic lateral sclerosis (ALS). Previous papers showed that mutant SOD1 accumulates and undergoes misfolding in motor neurons and that the specific interaction of mutant SOD1 with derlin-1 leads to endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR). Because evidence shows that mutant SOD1 expression also damages sensory neurons, we hypothesized that, similarly to motor neurons, the sensory neurons of ALS mouse model SOD1(G93A) accumulate mutant/misfolded SOD1 and suffer from ER stress and UPR activation. Our results reveal that SOD1(G93A) sensory neurons accumulate mutant/misfolded SOD1 but, surprisingly, do not suffer from ER stress and UPR activation. Moreover, the sensory neurons do not express detectable levels of the SOD1 interactor derlin-1. These results suggest a potential molecular mechanism underlying the differential vulnerability of motor and sensory neurons to mutant SOD1 toxicity.

Mitochondrial defects that affect cellular energy metabolism have long been implicated in the etiology of Huntington's disease (HD). Indeed, several studies have found defects in the mitochondrial functions of the central nervous system and peripheral tissues of HD patients. In this study, we investigated the in vivo oxidative metabolism of exercising muscle in HD patients. Ventilatory and cardiometabolic parameters and plasma lactate concentrations were monitored during incremental cardiopulmonary exercise in twenty-five HD subjects and twenty-five healthy subjects. The total exercise capacity was normal in HD subjects but notably the HD patients and presymptomatic mutation carriers had a lower anaerobic threshold than the control subjects. The low anaerobic threshold of HD patients was associated with an increase in the concentration of plasma lactate. We also analyzed in vitro muscular cell cultures and found that HD cells produce more lactate than the cells of healthy subjects. Finally, we analyzed skeletal muscle samples by electron microscopy and we observed striking mitochondrial structural abnormalities in two out of seven HD subjects. Our findings confirm mitochondrial abnormalities in HD patients' skeletal muscle and suggest that the mitochondrial dysfunction is reflected functionally in a low anaerobic threshold and an increased lactate synthesis during intense physical exercise. © 2010 Movement Disorder Society.

Huntington's disease (HD) is a genetically dominant condition caused by expanded CAG repeats. These repeats code for a glutamine tract in the HD gene product huntingtin (htt), which is a protein expressed in almost all tissues. Although most HD symptoms reflect preferential neuronal death in specific brain regions, even before the HD gene was identified numerous reports had described additional abnormalities in the peripheral tissues of HD patients, including weight loss, altered glucose homeostasis, and sub-cellular abnormalities in fibroblasts, lymphocytes and erythrocytes. Several years have elapsed since the HD mutation was discovered, and analyses of peripheral tissues from HD patients have helped to understand the molecular pathogenesis of the disease and revealed that the molecular mechanisms through which mutated htt leads to cell dysfunction are widely shared between central nervous system (CNS) and peripheral tissues. These studies show that in peripheral tissues, mutated htt causes accumulation of intracellular protein aggregates, impairment of energetic metabolism, transcriptional deregulation and hyperactivation of programmed cell-death mechanisms. Here, we review the current knowledge of peripheral tissue alterations in HD patients and in animal models of HD and focus on how this information can be used to identify potential therapeutic possibilities and biomarkers for disease progression.

We investigated the role of PPAR gamma coactivator 1alpha (PGC-1alpha) in muscle dysfunction in Huntington's disease (HD). We observed reduced PGC-1alpha and target genes expression in muscle of HD transgenic mice. We produced chronic energy deprivation in HD mice by administering the catabolic stressor beta-guanidinopropionic acid (GPA), a creatine analogue that reduces ATP levels, activates AMP-activated protein kinase (AMPK), which in turn activates PGC-1alpha. Treatment with GPA resulted in increased expression of AMPK, PGC-1alpha target genes, genes for oxidative phosphorylation, electron transport chain and mitochondrial biogenesis, increased oxidative muscle fibers, numbers of mitochondria and motor performance in wild-type, but not in HD mice. In muscle biopsies from HD patients, there was decreased PGC-1alpha, PGC-1beta and oxidative fibers. Oxygen consumption, PGC-1alpha, NRF1 and response to GPA were significantly reduced in myoblasts from HD patients. Knockdown of mutant huntingtin resulted in increased PGC-1alpha expression in HD myoblast. Lastly, adenoviral-mediated delivery of PGC-1alpha resulted increased expression of PGC-1alpha and markers for oxidative muscle fibers and reversal of blunted response for GPA in HD mice. These findings show that impaired function of PGC-1alpha plays a critical role in muscle dysfunction in HD, and that treatment with agents to enhance PGC-1alpha function could exert therapeutic benefits. Furthermore, muscle may provide a readily accessible tissue in which to monitor therapeutic interventions.

Background and Objective: Counterfactual thinking (CFT) refers to the generation of mental simulations of alternatives to past events, actions and outcomes. CFT is a pervasive cognitive feature in every-day life and is closely related to decision-making, planning and problem-solving - all of which are cognitive processes linked to unimpaired frontal lobe functioning. Huntington's Disease (HD) is a neurodegenerative disorder characterised by motor, behavioral and cognitive dysfunctions. Because an impairment in frontal and executive functions has been described in HD, we hypothesised that HD patients may have a CFT impairment. Methods: Tests of spontaneous counterfactual thoughts and counterfactual-derived inferences were administered to 24 symptomatic HD patients and 24 age- and sex-matched healthy subjects. Results: Our results show a significant impairment in the spontaneous generation of CFT and low performance on the Counterfactual Inference Test (CIT) in HD patients. Low performance on the spontaneous CFT test significantly correlates with impaired attention abilities, verbal fluency and frontal lobe efficiency, as measured by Trail Making Test - Part A, Phonemic Verbal Fluency Test and FAB. Conclusions: Spontaneous CFT and the use of this type of reasoning are impaired in HD patients. This deficit may be related to frontal lobe dysfunction, which is a hallmark of HD. Because CFT has a pervasive role in patients' daily lives regarding their planning, decision making and problem solving skills, cognitive rehabilitation may improve HD patients' ability to analyse current behaviors and future actions.

Mutated huntingtin (htt) is ubiquitously expressed in tissues of Huntington's disease (HD) patients. In the brain, the mutated protein leads to neuronal cell dysfunction and death, associated with formation of htt-positive inclusions. Given increasing evidence of abnormalities in HD skeletal muscle, we extensively analyzed primary muscle cell cultures from seven HD subjects (including two unaffected mutation carriers). Myoblasts from presymptomatic and symptomatic HD subjects showed cellular abnormalities in vitro, namely mitochondrial depolarization, cytochrome c release, increased caspase-3, -8, and -9 activities, and defective cell differentiation. Another notable feature was the formation of htt inclusions in differentiated myotubes. This study helps to advance current knowledge about the downstream effects of the htt mutation in human tissues. Further applications may include drug screening using this human cellular model.

Although loss-of-function mutations in the PARK2 gene, the gene that encodes the protein parkin, cause autosomal recessive juvenile parkinsonism, the responsible molecular mechanisms remain unclear. Evidence suggests that a loss of parkin dysregulates excitatory synapses. Here we show that parkin interacts with the kainate receptor (KAR) GluK2 subunit and regulates KAR function. Loss of parkin function in primary cultured neurons causes GluK2 protein to accumulate in the plasma membrane, potentiates KAR currents and increases KAR-dependent excitotoxicity. Expression in the mouse brain of a parkin mutant causing autosomal recessive juvenile parkinsonism results in GluK2 protein accumulation and excitotoxicity. These findings show that parkin regulates KAR function in vitro and in vivo, and suggest that KAR upregulation may have a pathogenetic role in parkin-related autosomal recessive juvenile parkinsonism., Loss-of-function mutations in the PARK2 gene are implicated in autosomal recessive juvenile parkinsonism, but the mechanisms are unclear. Here, the authors show that these mutations cause accumulation of the kainate receptor subunit GluK2 in the plasma membrane of neurons, which facilitates neuronal death.

Heterogeneous nuclear ribonucleoproteinK(hnRNP K) is an RNA-binding protein implicated inRNAmetabolism. Here, we investigated the role of hnRNP K in synapse function. We demonstrated that hnRNP K regulates dendritic spine density and long-term potentiation (LTP) in cultured hippocampal neurons from embryonic rats. LTP requires the extracellular signal-regulated kinase (ERK)1/2-mediated phosphorylation and cytoplasmic accumulation of hnRNP K. Moreover, hnRNP K knockdown prevents ERK cascade activation and GluA1-S845 phosphorylation and surface delivery, which are essential steps for LTP. These findings establish hnRNP K as a new critical regulator of synaptic transmission and plasticity in hippocampal neurons. © 2014 the authors.

The pathogenetic role of vascular endothelial growth factor (VEGF) in long-term retinal and kidney complications of diabetes has been demonstrated. Conversely, little is known in diabetic neuropathy. We examined the modulation of VEGF pathway at mRNA and protein level on dorsal root ganglion (DRG) neurons and Schwann cells (SC) induced by hyperglycaemia. Moreover, we studied the effects of VEGF neutralization on hyperglycemic DRG neurons and streptozotocin-induced diabetic neuropathy. Our findings demonstrated that DRG neurons were not affected by the direct exposition to hyperglycaemia, whereas showed an impairment of neurite outgrowth ability when exposed to the medium of SC cultured in hyperglycaemia. This was mediated by an altered regulation of VEGF and FLT-1 receptors. Hyperglycaemia increased VEGF and FLT-1 mRNA without changing their intracellular protein levels in DRG neurons, decreased intracellular and secreted protein levels without changing mRNA level in SC, while reduced the expression of the soluble receptor sFLT-1 both in DRG neurons and SC. Bevacizumab, a molecule that inhibits VEGF activity preventing the interaction with its receptors, restored neurite outgrowth and normalized FLT-1 mRNA and protein levels in co-cultures. In diabetic rats, it both prevented and restored nerve conduction velocity and nociceptive thresholds. We demonstrated that hyperglycaemia early affected neurite outgrowth through the impairment of SC-derived VEGF/FLT-1 signaling and that the neutralization of SC-secreted VEGF was protective both in vitro and in vivo models of diabetic neuropathy. Copyright

Amyotrophic lateral sclerosis (ALS) is a fatal neuromuscular disorder characterized by motor neuron (MN) degeneration that leads to progressive skeletal muscle atrophy and paralysis. Most ALS cases are sporadic, but a small percentage (5–10%) are familial. Many hypotheses have been formulated to explain the pathogenesis of sporadic ALS (sALS), including autoimmune reactions to calcium channels on MNs, glutamate-induced excitotoxic injury, exposure to toxins or latent infections, disorganization of intermediate filaments, and loss of neurotrophic support to MNs.1 The latter hypothesis is of particular interest, because adult muscle fibers produce molecules that influence MN survival, axonal growth, and maintenance of synaptic connections. Among these trophic factors, insulin-like growth factor-1 (IGF-1) has a key role; it is involved in muscle and nerve tissue anabolism and thus induces muscle hypertrophy and promotes neuronal survival.2–6 The neurotrophic effect of IGF-1 was the starting point for three clinical trials based on subcutaneous injections of human recombinant IGF-1 in ALS patients. Disappointingly, these studies did not show a significant positive effect of IGF-1 therapy on disease progression or survival in ALS patients.7–9 However, the therapeutic role of IGF-1 in ALS is still debated. Indeed, Kaspar and colleagues demonstrated that treatment with adeno-associated virus/IGF-1 retrogradely transported from muscle to MNs of the spinal cord led to therapeutic benefits in the G93A transgenic mouse model.10 This effect was further increased with associated physical exercise.11 More recently, Dobrowolny et al. reported that muscle-restricted expression of IGF-1 isoforms maintained muscle integrity, stabilized neuromuscular junctions, reduced inflammation in the spinal cord, enhanced motor neuronal survival, and delayed the onset and slowed disease progression in sodium dismutase 1 (SOD1) G93A mice.12, 13 Both these studies reappraised the potential role of the skeletal muscle and IGF-1 signaling as a target for treatment in ALS patients. Moreover, a recent study also showed that overexpression of IGF-1 in muscle attenuates disease in a mouse model of spinal and bulbar muscular atrophy.14 The IGF-1 signaling system is complex and regulated by many factors.15, 16 The components of the IGF-1 system include a growth factor, IGF-1, which is a single-chain polypeptide with a molecular weight of approximately 7.5 kDa. The IGF-1 receptor (IGF-1R) is a membrane glycoprotein of 300–350 kDa. It consists of two α subunits (135 kDa each) containing the ligand-binding site, two β subunits (90 kDa each) containing the hydrophobic transmembrane domain, a short extracellular region, and a tyrosine kinase domain in its cytoplasmic portion.17 Besides IGF-1R, six IGF-binding proteins (IGF-BPs) have been identified. This is a family of secreted proteins that specifically bind IGF-1 with affinities that are equal to or greater than those of the IGF-1R. In terms of IGF-1 function, it is now well established that some of the IGF-BPs, such as -BP2, -BP4, and -BP6, inhibit IGF-1. BP5 potentiates IGF-1 actions, whereas -BP1 and -BP3 can inhibit or potentiate IGF-1.18–22 In addition, numerous data also demonstrate evidence for IGF-1–independent actions of IGF-BPs, which are unexpected.23, 24 The lack of information available on the expression levels of the IGF-1 system in skeletal muscle of ALS patients led us to investigate the expression of IGF-1; IGF-BP3, -BP4, -BP5; and IGF-1R β subunit (IGF-1Rβ) in skeletal muscle specimens and primary muscle cell cultures obtained from sALS patients.

Huntington's disease (HD) is a rare hereditary neurodegenerative disorder characterized in over 90 percent of cases by chorea as the presenting motor symptom. We report a 54-year-old male who presented with Parkinsonism as the initial symptom of the disease. Genetic analysis revealed expansion of 40 CAG repeats, and brain MRI showed both severe caudate nuclei and cortical atrophy. Single-photon emission computed tomography (SPECT) imaging of the dopamine transporter showed nigrostriatal pathway degeneration. Here, we also describe his 2 years of clinical followup after ensuing dopaminergic stimulation.

Reduced Brain-Derived Neurotrophic Factor (BDNF) levels have been described in a number of patho-physiological conditions, most notably, in Huntington's disease (HD), a progressive neurodegenerative disorder. Since BDNF is also produced in blood, we have undertaken the measurement of its peripheral levels in the attempt to identify a possible link with HD prognosis and/or its progression. Here we evaluated BDNF level in 398 blood samples including 138 controls, 56 preHD, and 204 HD subjects. We found that BDNF protein levels were not reliably different between groups, whether measured in plasma (52 controls, 26 preHD, 105 HD) or serum (39 controls, 5 preHD, 29 HD). Our experience, and a re-analysis of the literature highlighted that intra-group variability and methodological aspects affect this measurement, especially in serum. We also assessed BDNF mRNA levels in blood samples from 47 controls, 25 preHD, and 70 HD subjects, and found no differences among the groups. We concluded that levels of BDNF in human blood were not informative (mRNA levels or plasma protein level) nor reliable (serum protein levels) as HD biomarkers. We also wish to warn the scientific community in interpreting the significance of changes measured in BDNF protein levels in serum from patients suffering from different conditions.

A defective expression or activity of neurotrophic factors, such as brain- and glial-derived neurotrophic factors, contributes to neuronal damage in Huntington's disease (HD). Here, we focused on transforming growth factor-β (TGF-β(1) ), a pleiotropic cytokine with an established role in mechanisms of neuroprotection. Asymptomatic HD patients showed a reduction in TGF-β(1) levels in the peripheral blood, which was related to trinucleotide mutation length and glucose hypometabolism in the caudate nucleus. Immunohistochemical analysis in post-mortem brain tissues showed that TGF-β(1) was reduced in cortical neurons of asymptomatic and symptomatic HD patients. Both YAC128 and R6/2 HD mutant mice showed a reduced expression of TGF-β(1) in the cerebral cortex, localized in neurons, but not in astrocytes. We examined the pharmacological regulation of TGF-β(1) formation in asymptomatic R6/2 mice, where blood TGF-β(1) levels were also reduced. In these R6/2 mice, both the mGlu2/3 metabotropic glutamate receptor agonist, LY379268, and riluzole failed to increase TGF-β(1) formation in the cerebral cortex and corpus striatum, suggesting that a defect in the regulation of TGF-β(1) production is associated with HD. Accordingly, reduced TGF-β(1) mRNA and protein levels were found in cultured astrocytes transfected with mutated exon 1 of the human huntingtin gene, and in striatal knock-in cell lines expressing full-length huntingtin with an expanded glutamine repeat. Taken together, our data suggest that serum TGF-β(1) levels are potential biomarkers of HD development during the asymptomatic phase of the disease, and raise the possibility that strategies aimed at rescuing TGF-β(1) levels in the brain may influence the progression of HD.

Searching brain and peripheral biomarkers is a requisite to cure Huntington's disease (HD). To search for markers indicating the rate of brain neurodegenerative changes in the various disease stages, we quantified changes in brain atrophy in subjects with HD. We analyzed the cross-sectional and longitudinal rate of brain atrophy, quantitatively measured by fully-automated multiparametric magnetic resonance imaging, as fractional gray matter (GM, determining brain cortex volume), white matter (WM, measuring the volume of axonal fibers), and corresponding cerebral spinal fluid (CSF, a measure of global brain atrophy), in 94 gene-positive subjects with presymptomatic to advanced HD, and age-matched healthy controls. Each of the three brain compartments we studied (WM, GM, and CSF) had a diverse role and their time courses differed in the development of HD. GM volume decreased early in life. Its decrease was associated with decreased serum brain-derived-neurotrophic-factor and started even many years before onset symptoms, then decreased slowly in a nonlinear manner during the various symptomatic HD stages. WM volume loss also began in the presymptomatic stage of HD a few years before manifest symptoms appear, rapidly decreasing near to the zone-of-onset. Finally, the CSF volume increase began many years before age at onset. Its volume measured in presymptomatic subjects contributed to improve the CAG-based model of age at onset prediction. The progressive CSF increase depended on CAG mutation size and continued linearly until the last stages of HD, perhaps representing the best marker of progression rate and severity in HD (R(2)= 0.25, P < 0.0001).

Huntington's disease (HD) is a neurodegenerative disorder characterized by motor, cognitive, and psychiatric symptoms and by a progressive degeneration of neurons in basal ganglia and in brain cortex. Brain-derived neurotrophic factor (BDNF) is a pro-survival factor for striatal neurons. Some evidence implicates a brain BDNF deficiency, related to mutated huntingtin expression, in the selective vulnerability of striatal neurons in HD. We compared BDNF serum levels in 42 patients with HD (range 28–72 years, mean age 51.9 ± 11.5), and 42 age-matched healthy subjects (range 25–68 years, mean age 48.2 ± 12.5). We evaluated the potential relationship between BDNF serum levels, CAG repeat number (range 40–54, mean 44.8 ± 3.4) and duration of illness (range 6–228 months, mean 103.6 ± 62.1). Serum BDNF levels were significantly lower in patients than in age-matched healthy subjects. Lower BDNF levels were associated with a longer CAG repeat length and a longer duration of illness. Severity of the illness, as assessed by the Unified Huntington's Disease Rating Scale (UHDRS) motor and cognitive scores, was negatively related to serum BDNF levels. These results in vivo confirm that the huntingtin mutation causes BDNF production to decline and show that the BDNF deficiency is detectable in HD patients' sera. Further studies on a larger sample size should confirm whether BDNF concentrations in patients' serum could be a useful clinical marker related to the patients' disease phenotype. © 2007 Wiley-Liss, Inc.

An alteration of the retinoid pathway can influence the development of uterine leiomyomas in animal models, and retinoids have shown efficacy in inhibiting the growth of this benign tumor both in vitro and in vivo. However, the underlying mechanisms and biological implications are unclear. The present study was based on the demonstration of an accumulation of full-length retinoid X receptor α (RXRα) in leiomyomas that was not associated with a modification of its gene expression. This accumulation was shown to increase the transcription of the RXR-responsive gene cellular retinoic acid binding protein II (CRABP-II) and to be linked to the cellular redistribution of the receptor and to its retarded degradation via the ubiquitin/proteasome pathway. Accordingly, treatment with a specific proteasome inhibitor but not with protease inhibitors strongly inhibited the degradation of full-length RXRα in cells deriving from both myometrium and leiomyoma, but the formation of RXRα/ubiquitin conjugates was differentially regulated between the two cell types. Moreover, full-length RXRα accumulated in leiomyomas was abnormally phosphorylated at serine/threonine residues relative to myometrial tissue. The ligand to RXRα, 9-cis-retinoic acid, induced the receptor breakdown in smooth muscle cells deriving from both normal and tumor tissue, whereas a MAPK-specific inhibitor was able to reduce RXRα levels only in leiomyoma cells. These results suggest that switching of the ubiquitin/proteasome-dependent degradation of RXRα by phosphorylation in leiomyomas may be responsible for the accumulation of the receptor and the consequent dysregulation of retinoic acid target genes. The ability of retinoids to modify this molecular alteration may be the rationale for their use in the treatment of leiomyomas.

Until fairly recently, interest in stem cells was restricted to neurobiology studies on the principles of embryonic development. This situation has changed rapidly in the last few years when neuronal stems and precursors were isolated in vitro, thus allowing expansion and controlled differentiation of selective populations of neuronal cells. This theoretically unlimited reserve would then supply specific cells for transplantation in diseases characterized by widespread degeneration of selective cell populations as motor neurons in Amyotrophic Lateral Sclerosis (ALS). The recent evidence of cell transdifferentiation has further amplified the potential therapeutic use of stem cells. Stem cell technology is at an early stage but the desperate need for a therapy in ALS patients may legitimize clinical trials in absence of conclusive scientific evidence. This paper discusses the premises for stem cell therapy in ALS.