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It remains unclear whether activated inflammatory macrophages can adopt features of tissue-resident macrophages, or what mechanisms might mediate such a phenotypic conversion. Here we show that vitamin A is required for the phenotypic conversion of interleukin 4 (IL-4)-activated monocyte-derived F4/80intCD206+PD-L2+MHCII+ macrophages into macrophages with a tissue-resident F4/80hiCD206-PD-L2-MHCII-UCP1+ phenotype in the peritoneal cavity of mice and during the formation of liver granulomas in mice infected with Schistosoma mansoni. The phenotypic conversion of F4/80intCD206+ macrophages into F4/80hiCD206- macrophages was associated with almost complete remodeling of the chromatin landscape, as well as alteration of the transcriptional profiles. Vitamin A-deficient mice infected with S. mansoni had disrupted liver granuloma architecture and increased mortality, which indicates that failure to convert macrophages from the F4/80intCD206+ phenotype to F4/80hiCD206- may lead to dysregulated inflammation during helminth infection.
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Granuloma/imunologia , Fígado/imunologia , Macrófagos/imunologia , Esquistossomose mansoni/imunologia , Deficiência de Vitamina A/imunologia , Animais , Antígenos de Diferenciação/metabolismo , Citometria de Fluxo , Antígenos de Histocompatibilidade Classe II/metabolismo , Interleucina-4/imunologia , Lectinas Tipo C/metabolismo , Fígado/patologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos Alveolares/efeitos dos fármacos , Macrófagos Alveolares/imunologia , Macrófagos Alveolares/metabolismo , Receptor de Manose , Lectinas de Ligação a Manose/metabolismo , Camundongos , Cavidade Peritoneal/citologia , Proteína 2 Ligante de Morte Celular Programada 1/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Receptores de Superfície Celular/metabolismo , Schistosoma mansoni , Esquistossomose mansoni/patologia , Tretinoína/farmacologia , Proteína Desacopladora 1/metabolismo , Vitaminas/farmacologiaRESUMO
Additive manufacturing (AM) offers a variety of material manufacturing techniques for a wide range of applications across many industries. Most efforts at process optimization and exposure assessment for AM are centered around the manufacturing process. However, identifying the material allocation and potentially harmful exposures in end-of-life (EoL) management is equally crucial to mitigating environmental releases and occupational health impacts within the AM supply chain. This research tracks the allocation and potential releases of AM EoL materials within the US through a material flow analysis. Of the generated AM EoL materials, 58% are incinerated, 33% are landfilled, and 9% are recycled by weight. The generated data set was then used to examine the theoretical occupational hazards during AM EoL material management practices through generic exposure scenario assessment, highlighting the importance of ventilation and personal protective equipment at all stages of AM material management. This research identifies pollution sources, offering policymakers and stakeholders insights to shape pollution prevention and worker safety strategies within the US AM EoL management pathways.
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Exposição Ocupacional , Humanos , ReciclagemRESUMO
BACKGROUND: Patient access to body-powered and myoelectric upper limb prostheses in the United States is often restricted by a healthcare system that prioritizes prosthesis prescription based on cost and perceived value. Although this system operates on an underlying assumption that design differences between these prostheses leads to relative advantages and disadvantages of each device, there is limited empirical evidence to support this view. MAIN TEXT: This commentary article will review a series of studies conducted by our research team with the goal of differentiating how prosthesis design might impact user performance on a variety of interrelated domains. Our central hypothesis is that the design and actuation method of body-powered and myoelectric prostheses might affect users' ability to access sensory feedback and account for device properties when planning movements. Accordingly, other domains that depend on these abilities may also be affected. While our work demonstrated some differences in availability of sensory feedback based on prosthesis design, this did not result in consistent differences in prosthesis embodiment, movement accuracy, movement quality, and overall kinematic patterns. CONCLUSION: Collectively, our findings suggest that performance may not necessarily depend on prosthesis design, allowing users to be successful with either device type depending on the circumstances. Prescription practices should rely more on individual needs and preferences than cost or prosthesis design. However, we acknowledge that there remains a dearth of evidence to inform decision-making and that an expanded research focus in this area will be beneficial.
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Membros Artificiais , Desenho de Prótese , Extremidade Superior , Humanos , Extremidade Superior/fisiologia , Eletromiografia/instrumentação , Retroalimentação Sensorial/fisiologia , Fenômenos BiomecânicosRESUMO
Under the Toxic Substances Control Act (TSCA), the United States Environmental Protection Agency (USEPA) is required to determine whether a new chemical substance poses an unreasonable risk to human health or the environment before the chemical is manufactured in or imported into the United States. This manuscript provides a review of the process used to evaluate the risk associated with a chemical based on the scenarios and models used in the evaluation. Specifically, the Generic Scenarios and Emission Scenario Documents developed by the USEPA were reviewed, along with background documentation prepared by USEPA to identify the core elements of the environmental release and occupational exposure scenarios used to assess the risk of the chemical being evaluated. Additionally, this contribution provides an overview of methods used to model occupational exposures and environmental releases as part of the chemical evaluation process used in other jurisdictions, along with work being performed to improve these models. Finally, the alternative methods to evaluate occupational exposures and environmental releases that may be used as part of the decision-making process regarding a chemical are identified. The contribution provides a path forward for reducing the time required and improving the chemical evaluation of the unreasonable risk determination regarding the manufacture or import of a chemical.
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Exposição Ocupacional , Estados Unidos , Humanos , Exposição Ocupacional/prevenção & controle , Medição de Risco/métodos , Fatores de Risco , Exposição AmbientalRESUMO
Type 2 immunity plays an essential role in the maintenance of metabolic homeostasis and its disruption during obesity promotes meta-inflammation and insulin resistance. Infection with the helminth parasite Schistosoma mansoni and treatment with its soluble egg antigens (SEA) induce a type 2 immune response in metabolic organs and improve insulin sensitivity and glucose tolerance in obese mice, yet, a causal relationship remains unproven. Here, we investigated the effects and underlying mechanisms of the T2 ribonuclease omega-1 (ω1), one of the major S mansoni immunomodulatory glycoproteins, on metabolic homeostasis. We show that treatment of obese mice with plant-produced recombinant ω1, harboring similar glycan motifs as present on the native molecule, decreased body fat mass, and improved systemic insulin sensitivity and glucose tolerance in a time- and dose-dependent manner. This effect was associated with an increase in white adipose tissue (WAT) type 2 T helper cells, eosinophils, and alternatively activated macrophages, without affecting type 2 innate lymphoid cells. In contrast to SEA, the metabolic effects of ω1 were still observed in obese STAT6-deficient mice with impaired type 2 immunity, indicating that its metabolic effects are independent of the type 2 immune response. Instead, we found that ω1 inhibited food intake, without affecting locomotor activity, WAT thermogenic capacity or whole-body energy expenditure, an effect also occurring in leptin receptor-deficient obese and hyperphagic db/db mice. Altogether, we demonstrate that while the helminth glycoprotein ω1 can induce type 2 immunity, it improves whole-body metabolic homeostasis in obese mice by inhibiting food intake via a STAT6-independent mechanism.
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Ingestão de Alimentos , Endorribonucleases/uso terapêutico , Glicoproteínas/uso terapêutico , Proteínas de Helminto/uso terapêutico , Obesidade/tratamento farmacológico , Tecido Adiposo/efeitos dos fármacos , Tecido Adiposo/metabolismo , Animais , Células Cultivadas , Endorribonucleases/farmacologia , Glicoproteínas/farmacologia , Proteínas de Helminto/farmacologia , Locomoção , Macrófagos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Recombinantes/farmacologia , Proteínas Recombinantes/uso terapêutico , Schistosoma mansoni/enzimologia , Linfócitos T Auxiliares-Indutores/efeitos dos fármacos , Termogênese , Nicotiana/genética , Nicotiana/metabolismoRESUMO
Understanding the fundamental characteristics of prosthetic movement control is imperative in improving prosthesis design and training. This study quantified how using an upper limb prosthesis affected performance during goal-directed reaching tasks. Nine prosthesis users with unilateral transradial limb absence and nine healthy controls completed a series of goal-directed reaching movements with different goals: one spatial and three temporal with different goal frequencies. We quantified end-point accuracy, smoothness, and peak speed for the spatial task and temporal accuracy, horizontal distance, and speed for the temporal task. For the temporal task, we also used a goal-equivalent manifold (GEM) approach to decompose variability in movement distance and speed into those perpendicular and tangential to the GEM. Detrended fluctuation analysis (DFA) quantified the temporal persistence of each time series. For the spatial task, movements made with prostheses were less smooth, had larger end-point errors, and had slower peak speed compared to those with control limbs (p < 0.041). For the temporal task, movements made with prostheses and intact limbs of prosthesis users and control limbs were similar in distance and speed and had similar timing errors (p > 0.138). Timing errors, distance, speed, and GEM deviations were corrected similarly between prosthetic limbs and control limbs (p > 0.091). The mean and variability of distance, speed, and perpendicular deviations decreased with increased goal frequency (p < 0.001). Our results suggest that prosthesis users have a sufficient internal model to successfully complete ballistic movements but are unable to accurately complete movements requiring substantial feedback.
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Membros Artificiais , Humanos , Objetivos , Extremidade Superior , Movimento , Fatores de TempoRESUMO
This study proposes a set of key decision-making features of the contaminated site remediation process to assist in selecting the most appropriate decision support method(s). Using a case study consistent with the requirements of the U.S. regulation for contaminated sites management, this article shows that suitable Multiple Criteria Decision Analysis methods can be selected based on a dynamic and evolving problem structuring. The selected methods belong to the family of PROMETHEE methods and can provide ranking recommendations of the considered alternatives using variable structures of the criteria, evaluation of the alternatives and exploitation of the preference model. It was found that in order to support a quick and up-to-date application of powerful decision support techniques in the process of remediation of contaminated sites, decision analysts and stakeholders should interact and co-develop the process. This research also displays how such interactions can guarantee a transparent and traceable decision recommendation so that stakeholders can better understand why some alternatives perform comprehensively better than others when a multitude of inputs is used in the decision-making process.
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OBJECTIVES: Hereditary spastic paraplegias (HSPs) are a heterogenous group of rare neurodegenerative disorders that present with lower limb spasticity. It is known as complicated HSP if spasticity is accompanied by additional features such as cognitive impairment, cerebellar syndrome, thin corpus callosum, or neuropathy. Most HSP families show autosomal dominant (AD) inheritance. On the other hand, autosomal recessive (AR) cases are also common because of the high frequency of consanguineous marriages in our country. This study aimed to investigate the clinical and genetic aetiology in a group of HSP patients. PATIENTS AND METHODS: We studied 21 patients from 17 families. Six of them presented with recessive inheritance. All index patients were screened for ATL1 and SPAST gene mutations to determine the prevalence of the most frequent types of HSP in our cohort. Whole exome sequencing was performed for an AD-HSP family, in combination with homozygosity mapping for five selected AR-HSP families. RESULTS: Two novel causative variants were identified in PLP1 and SPG11 genes, respectively. Distribution of HSP mutations in our AD patients was found to be similar to European populations. CONCLUSION: Our genetic studies confirmed that clinical analysis can be misleading when defining HSP subtypes. Genetic testing is an important tool for diagnosis and genetic counselling. However, in the majority of AR HSP cases, a genetic diagnosis is not possible.
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Paraplegia Espástica Hereditária , Estudos de Coortes , Proteínas de Ligação ao GTP , Testes Genéticos , Humanos , Proteínas de Membrana , Mutação , Proteínas , Espastina , TurquiaRESUMO
BACKGROUND: Etoposide (E) at 100 mg/m2 combined with Cisplatin (P) at 20 mg/m2 represents an induction 2-day regimen embedded in our clinical practice for patients with advanced GCT or TN at high risk of early death. We evaluated 24/7 Em-EP administration to a combined GCT-TN cohort at our Emergency Cancer Treatment Centre (ECTC) to determine its efficacy within the acute setting. METHODS: Patients who received Em-EP during a five-year interval were identified from electronic databases at Imperial College Healthcare NHS Trust. Data collected included demographics, treatment details and clinical outcome. RESULTS: Em-EP was administered in the emergency setting to 104 patients, predominantly young adults (median age 35, range 17-71). Half the cases were GCT (n = 52): 22 male (6 seminomas, 13 non-seminomas); 30 female (2 dysgerminomas, 28 non-dysgerminomas). The other 50% were treated for TN (n = 52): 45 gestational (GTN) and 7 non-gestational. Most patients received Em-EP for a new cancer diagnosis (n = 100, 96%), within 24 h (n = 93, 89%) and out-of-hours (n = 74, 70%). Indications for Em-EP included symptomatic disease (n = 66, 63%), high-burden disease, (n = 51, 49%) and organ failure requiring Intensive Care Unit support (n = 9, 9%). Neutropenic sepsis was observed in 5%. Four-week overall survival after Em-EP administration was 98%. CONCLUSIONS: Despite the potentially fatal complications encountered in the acute setting, early mortality with Em-EP is low at our ECTC. Specialist units that treat unwell patients with advanced GCT or TN should consider making Em-EP available 24/7 for emergency administration. Its efficacy within a prospective cohort and in other platinum-sensitive malignancies requires evaluation.
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Antineoplásicos Fitogênicos/uso terapêutico , Cisplatino/uso terapêutico , Serviços Médicos de Emergência , Etoposídeo/uso terapêutico , Doença Trofoblástica Gestacional/tratamento farmacológico , Doença Aguda , Adolescente , Adulto , Idoso , Antineoplásicos Fitogênicos/administração & dosagem , Antineoplásicos Fitogênicos/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Cisplatino/administração & dosagem , Cisplatino/efeitos adversos , Atenção à Saúde , Etoposídeo/administração & dosagem , Etoposídeo/efeitos adversos , Feminino , Febre/etiologia , Seguimentos , Doença Trofoblástica Gestacional/mortalidade , Humanos , Masculino , Pessoa de Meia-Idade , Neutropenia/etiologia , Gravidez , Estudos Prospectivos , Estudos Retrospectivos , Sepse/etiologia , Taxa de Sobrevida , Resultado do Tratamento , Adulto JovemRESUMO
Despite extensive efforts, half of patients with rare movement disorders such as hereditary spastic paraplegias and cerebellar ataxias remain genetically unexplained, implicating novel genes and unrecognized mutations in known genes. Non-coding DNA variants are suspected to account for a substantial part of undiscovered causes of rare diseases. Here we identified mutations located deep in introns of POLR3A to be a frequent cause of hereditary spastic paraplegia and cerebellar ataxia. First, whole-exome sequencing findings in a recessive spastic ataxia family turned our attention to intronic variants in POLR3A, a gene previously associated with hypomyelinating leukodystrophy type 7. Next, we screened a cohort of hereditary spastic paraplegia and cerebellar ataxia cases (n = 618) for mutations in POLR3A and identified compound heterozygous POLR3A mutations in â¼3.1% of index cases. Interestingly, >80% of POLR3A mutation carriers presented the same deep-intronic mutation (c.1909+22G>A), which activates a cryptic splice site in a tissue and stage of development-specific manner and leads to a novel distinct and uniform phenotype. The phenotype is characterized by adolescent-onset progressive spastic ataxia with frequent occurrence of tremor, involvement of the central sensory tracts and dental problems (hypodontia, early onset of severe and aggressive periodontal disease). Instead of the typical hypomyelination magnetic resonance imaging pattern associated with classical POLR3A mutations, cases carrying c.1909+22G>A demonstrated hyperintensities along the superior cerebellar peduncles. These hyperintensities may represent the structural correlate to the cerebellar symptoms observed in these patients. The associated c.1909+22G>A variant was significantly enriched in 1139 cases with spastic ataxia-related phenotypes as compared to unrelated neurological and non-neurological phenotypes and healthy controls (P = 1.3 × 10-4). In this study we demonstrate that (i) autosomal-recessive mutations in POLR3A are a frequent cause of hereditary spastic ataxias, accounting for about 3% of hitherto genetically unclassified autosomal recessive and sporadic cases; and (ii) hypomyelination is frequently absent in POLR3A-related syndromes, especially when intronic mutations are present, and thus can no longer be considered as the unifying feature of POLR3A disease. Furthermore, our results demonstrate that substantial progress in revealing the causes of Mendelian diseases can be made by exploring the non-coding sequences of the human genome.
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Deficiência Intelectual/genética , Espasticidade Muscular/genética , Atrofia Óptica/genética , RNA Polimerase III/genética , Paraplegia Espástica Hereditária/genética , Ataxias Espinocerebelares/genética , Idoso , Técnicas de Cultura de Células , Éxons/genética , Feminino , Estudos de Associação Genética , Humanos , Células-Tronco Pluripotentes Induzidas , Deficiência Intelectual/diagnóstico por imagem , Deficiência Intelectual/fisiopatologia , Íntrons/genética , Masculino , Pessoa de Meia-Idade , Espasticidade Muscular/diagnóstico por imagem , Espasticidade Muscular/fisiopatologia , Mutação , Atrofia Óptica/diagnóstico por imagem , Atrofia Óptica/fisiopatologia , Linhagem , Fenótipo , Paraplegia Espástica Hereditária/diagnóstico por imagem , Paraplegia Espástica Hereditária/fisiopatologia , Ataxias Espinocerebelares/diagnóstico por imagem , Ataxias Espinocerebelares/fisiopatologiaRESUMO
Diabetes mellitus and neurodegeneration are common diseases for which shared genetic factors are still only partly known. Here, we show that loss of the BiP (immunoglobulin heavy-chain binding protein) co-chaperone DNAJC3 leads to diabetes mellitus and widespread neurodegeneration. We investigated three siblings with juvenile-onset diabetes and central and peripheral neurodegeneration, including ataxia, upper-motor-neuron damage, peripheral neuropathy, hearing loss, and cerebral atrophy. Exome sequencing identified a homozygous stop mutation in DNAJC3. Screening of a diabetes database with 226,194 individuals yielded eight phenotypically similar individuals and one family carrying a homozygous DNAJC3 deletion. DNAJC3 was absent in fibroblasts from all affected subjects in both families. To delineate the phenotypic and mutational spectrum and the genetic variability of DNAJC3, we analyzed 8,603 exomes, including 506 from families affected by diabetes, ataxia, upper-motor-neuron damage, peripheral neuropathy, or hearing loss. This analysis revealed only one further loss-of-function allele in DNAJC3 and no further associations in subjects with only a subset of the features of the main phenotype. Our findings demonstrate that loss-of-function DNAJC3 mutations lead to a monogenic, recessive form of diabetes mellitus in humans. Moreover, they present a common denominator for diabetes and widespread neurodegeneration. This complements findings from mice in which knockout of Dnajc3 leads to diabetes and modifies disease in a neurodegenerative model of Marinesco-Sjögren syndrome.
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Diabetes Mellitus Tipo 1/genética , Regulação da Expressão Gênica , Proteínas de Choque Térmico HSP40/genética , Proteínas de Choque Térmico/genética , Atrofia de Múltiplos Sistemas/genética , Adolescente , Adulto , Ataxia/genética , Diabetes Mellitus Tipo 1/diagnóstico por imagem , Chaperona BiP do Retículo Endoplasmático , Exoma/genética , Feminino , Fibroblastos , Proteínas de Choque Térmico HSP40/metabolismo , Homozigoto , Humanos , Masculino , Modelos Moleculares , Atrofia de Múltiplos Sistemas/diagnóstico por imagem , Mutação , Linhagem , Fenótipo , Radiografia , Análise de Sequência de DNA , Adulto JovemRESUMO
Hereditary spastic paraplegias (HSPs) are clinically and genetically heterogeneous neurological conditions. Their main pathogenic mechanisms are thought to involve alterations in endomembrane trafficking, mitochondrial function, and lipid metabolism. With a combination of whole-genome mapping and exome sequencing, we identified three mutations in REEP2 in two families with HSP: a missense variant (c.107T>A [p.Val36Glu]) that segregated in the heterozygous state in a family with autosomal-dominant inheritance and a missense change (c.215T>A [p.Phe72Tyr]) that segregated in trans with a splice site mutation (c.105+3G>T) in a family with autosomal-recessive transmission. REEP2 belongs to a family of proteins that shape the endoplasmic reticulum, an organelle that was altered in fibroblasts from an affected subject. In vitro, the p.Val36Glu variant in the autosomal-dominant family had a dominant-negative effect; it inhibited the normal binding of wild-type REEP2 to membranes. The missense substitution p.Phe72Tyr, in the recessive family, decreased the affinity of the mutant protein for membranes that, together with the splice site mutation, is expected to cause complete loss of REEP2 function. Our findings illustrate how dominant and recessive inheritance can be explained by the effects and nature of mutations in the same gene. They have also important implications for genetic diagnosis and counseling in clinical practice because of the association of various modes of inheritance to this new clinico-genetic entity.
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Proteínas de Membrana/genética , Paraplegia Espástica Hereditária/genética , Sequência de Aminoácidos , Animais , Células COS , Chlorocebus aethiops , Mapeamento Cromossômico , Exoma , Feminino , Heterozigoto , Humanos , Masculino , Proteínas de Membrana/metabolismo , Dados de Sequência Molecular , Mutação de Sentido Incorreto , Linhagem , Fenótipo , Paraplegia Espástica Hereditária/patologiaRESUMO
OBJECTIVES: To analyse and describe the clinical and genetic spectrum of Charcot-Marie-Tooth disease (CMT) caused by mutations in the neurofilament light polypeptide gene (NEFL). METHODS: Combined analysis of newly identified patients with NEFL-related CMT and all previously reported cases from the literature. RESULTS: Five new unrelated patients with CMT carrying the NEFL mutations P8R and N98S and the novel variant L311P were identified. Combined data from these cases and 62 kindreds from the literature revealed four common mutations (P8R, P22S, N98S and E396K) and three mutational hotspots accounting for 37 (55%) and 50 (75%) kindreds, respectively. Eight patients had de novo mutations. Loss of large-myelinated fibres was a uniform feature in a total of 21 sural nerve biopsies and 'onion bulb' formations and/or thin myelin sheaths were observed in 14 (67%) of them. The neurophysiological phenotype was broad but most patients with E90K and N98S had upper limb motor conduction velocities <38 m/s. Age of onset was ≤3 years in 25 cases. Pyramidal tract signs were described in 13 patients and 7 patients were initially diagnosed with or tested for inherited ataxia. Patients with E90K and N98S frequently presented before age 3 years and developed hearing loss or other neurological features including ataxia and/or cerebellar atrophy on brain MRI. CONCLUSIONS: NEFL-related CMT is clinically and genetically heterogeneous. Based on this study, however, we propose mutational hotspots and relevant clinical-genetic associations that may be helpful in the evaluation of NEFL sequence variants and the differential diagnosis with other forms of CMT.
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Doença de Charcot-Marie-Tooth/genética , Doença de Charcot-Marie-Tooth/patologia , Mutação/genética , Proteínas de Neurofilamentos/genética , Axônios/patologia , Ataxia Cerebelar/genética , Ataxia Cerebelar/patologia , Genótipo , Humanos , Linhagem , Fenótipo , Nervo Sural/patologiaRESUMO
We performed whole exome sequencing on a patient with Charcot-Marie-Tooth disease type 1 and identified a de novo mutation in PMP2, the gene that encodes the myelin P2 protein. This mutation (p.Ile52Thr) was passed from the proband to his one affected son, and segregates with clinical and electrophysiological evidence of demyelinating neuropathy. We then screened a cohort of 136 European probands with uncharacterized genetic cause of Charcot-Marie-Tooth disease and identified another family with Charcot-Marie-Tooth disease type 1 that has a mutation affecting an adjacent amino acid (p.Thr51Pro), which segregates with disease. Our genetic and clinical findings in these kindred demonstrate that dominant PMP2 mutations cause Charcot-Marie-Tooth disease type 1.
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Doença de Charcot-Marie-Tooth/genética , Proteína P2 de Mielina/genética , Adolescente , Exoma/genética , Feminino , Predisposição Genética para Doença/genética , Haplótipos , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Condução Nervosa/genética , Linhagem , Adulto JovemRESUMO
Axons damaged by acute injury, toxic insults, or neurodegenerative diseases execute a poorly defined autodestruction signaling pathway leading to widespread fragmentation and functional loss. Here, we describe an approach to study Wallerian degeneration in the Drosophila L1 wing vein that allows for analysis of axon degenerative phenotypes with single-axon resolution in vivo. This method allows for the axotomy of specific subsets of axons followed by examination of progressive axonal degeneration and debris clearance alongside uninjured control axons. We developed new Flippase (FLP) reagents using proneural gene promoters to drive FLP expression very early in neural lineages. These tools allow for the production of mosaic clone populations with high efficiency in sensory neurons in the wing. We describe a collection of lines optimized for forward genetic mosaic screens using MARCM (mosaic analysis with a repressible cell marker; i.e., GFP-labeled, homozygous mutant) on all major autosomal arms (â¼95% of the fly genome). Finally, as a proof of principle we screened the X chromosome and identified a collection eight recessive and two dominant alleles of highwire, a ubiquitin E3 ligase required for axon degeneration. Similar unbiased forward genetic screens should help rapidly delineate axon death genes, thereby providing novel potential drug targets for therapeutic intervention to prevent axonal and synaptic loss.
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Axônios , Drosophila/genética , Alelos , Animais , Genes Dominantes , Genes de Insetos , Genes Recessivos , Mosaicismo , Asas de Animais/irrigação sanguíneaRESUMO
Spastic paraplegia 46 refers to a locus mapped to chromosome 9 that accounts for a complicated autosomal-recessive form of hereditary spastic paraplegia (HSP). With next-generation sequencing in three independent families, we identified four different mutations in GBA2 (three truncating variants and one missense variant), which were found to cosegregate with the disease and were absent in controls. GBA2 encodes a microsomal nonlysosomal glucosylceramidase that catalyzes the conversion of glucosylceramide to free glucose and ceramide and the hydrolysis of bile acid 3-O-glucosides. The missense variant was also found at the homozygous state in a simplex subject in whom no residual glucocerebrosidase activity of GBA2 could be evidenced in blood cells, opening the way to a possible measurement of this enzyme activity in clinical practice. The overall phenotype was a complex HSP with mental impairment, cataract, and hypogonadism in males associated with various degrees of corpus callosum and cerebellar atrophy on brain imaging. Antisense morpholino oligonucleotides targeting the zebrafish GBA2 orthologous gene led to abnormal motor behavior and axonal shortening/branching of motoneurons that were rescued by the human wild-type mRNA but not by applying the same mRNA containing the missense mutation. This study highlights the role of ceramide metabolism in HSP pathology.
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Neurônios Motores/patologia , Paraplegia Espástica Hereditária/enzimologia , Paraplegia Espástica Hereditária/genética , Proteínas de Peixe-Zebra/genética , beta-Glucosidase/genética , Adolescente , Adulto , Idoso , Animais , Encéfalo/patologia , Criança , Pré-Escolar , Família , Feminino , Glucosilceramidase , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Mutação/genética , Neuroimagem , Linhagem , Adulto Jovem , Peixe-ZebraRESUMO
Hereditary spastic paraplegias (HSPs) form a heterogeneous group of neurological disorders. A whole-genome linkage mapping effort was made with three HSP-affected families from Spain, Portugal, and Tunisia and it allowed us to reduce the SPG26 locus interval from 34 to 9 Mb. Subsequently, a targeted capture was made to sequence the entire exome of affected individuals from these three families, as well as from two additional autosomal-recessive HSP-affected families of German and Brazilian origins. Five homozygous truncating (n = 3) and missense (n = 2) mutations were identified in B4GALNT1. After this finding, we analyzed the entire coding region of this gene in 65 additional cases, and three mutations were identified in two subjects. All mutated cases presented an early-onset spastic paraplegia, with frequent intellectual disability, cerebellar ataxia, and peripheral neuropathy as well as cortical atrophy and white matter hyperintensities on brain imaging. B4GALNT1 encodes ß-1,4-N-acetyl-galactosaminyl transferase 1 (B4GALNT1), involved in ganglioside biosynthesis. These findings confirm the increasing interest of lipid metabolism in HSPs. Interestingly, although the catabolism of gangliosides is implicated in a variety of neurological diseases, SPG26 is only the second human disease involving defects of their biosynthesis.
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Disfunção Cognitiva/genética , Gangliosídeos/biossíntese , Paraplegia Espástica Hereditária/genética , Adolescente , Adulto , Idade de Início , Brasil , Ataxia Cerebelar/genética , Criança , Pré-Escolar , Mapeamento Cromossômico/métodos , Exoma , Feminino , Galactosiltransferases/genética , Galactosiltransferases/metabolismo , Gangliosídeos/genética , Predisposição Genética para Doença , Alemanha , Homozigoto , Humanos , Lactente , Metabolismo dos Lipídeos , Masculino , Mutação de Sentido Incorreto , Linhagem , Portugal , Espanha , Paraplegia Espástica Hereditária/metabolismo , Tunísia , Adulto JovemRESUMO
Demands for quick and accurate life cycle assessments create a need for methods to rapidly generate reliable life cycle inventories (LCI). Data mining is a suitable tool for this purpose, especially given the large amount of available governmental data. These data are typically applied to LCIs on a case-by-case basis. As linked open data becomes more prevalent, it may be possible to automate LCI using data mining by establishing a reproducible approach for identifying, extracting, and processing the data. This work proposes a method for standardizing and eventually automating the discovery and use of publicly available data at the United States Environmental Protection Agency for chemical-manufacturing LCI. The method is developed using a case study of acetic acid. The data quality and gap analyses for the generated inventory found that the selected data sources can provide information with equal or better reliability and representativeness on air, water, hazardous waste, on-site energy usage, and production volumes but with key data gaps including material inputs, water usage, purchased electricity, and transportation requirements. A comparison of the generated LCI with existing data revealed that the data mining inventory is in reasonable agreement with existing data and may provide a more-comprehensive inventory of air emissions and water discharges. The case study highlighted challenges for current data management practices that must be overcome to successfully automate the method using semantic technology. Benefits of the method are that the openly available data can be compiled in a standardized and transparent approach that supports potential automation with flexibility to incorporate new data sources as needed.
Assuntos
Monitoramento Ambiental , United States Environmental Protection Agency , Reprodutibilidade dos Testes , Estados UnidosRESUMO
Spinal muscular atrophy is a disorder of lower motor neurons, most commonly caused by recessive mutations in SMN1 on chromosome 5q. Cases without SMN1 mutations are subclassified according to phenotype. Spinal muscular atrophy, lower extremity-predominant, is characterized by lower limb muscle weakness and wasting, associated with reduced numbers of lumbar motor neurons and is caused by mutations in DYNC1H1, which encodes a microtubule motor protein in the dynein-dynactin complex and one of its cargo adaptors, BICD2. We have now identified 32 patients with BICD2 mutations from nine different families, providing detailed insights into the clinical phenotype and natural history of BICD2 disease. BICD2 spinal muscular atrophy, lower extremity predominant most commonly presents with delayed motor milestones and ankle contractures. Additional features at presentation include arthrogryposis and congenital dislocation of the hips. In all affected individuals, weakness and wasting is lower-limb predominant, and typically involves both proximal and distal muscle groups. There is no evidence of sensory nerve involvement. Upper motor neuron signs are a prominent feature in a subset of individuals, including one family with exclusively adult-onset upper motor neuron features, consistent with a diagnosis of hereditary spastic paraplegia. In all cohort members, lower motor neuron features were static or only slowly progressive, and the majority remained ambulant throughout life. Muscle MRI in six individuals showed a common pattern of muscle involvement with fat deposition in most thigh muscles, but sparing of the adductors and semitendinosus. Muscle pathology findings were highly variable and included pseudomyopathic features, neuropathic features, and minimal change. The six causative mutations, including one not previously reported, result in amino acid changes within all three coiled-coil domains of the BICD2 protein, and include a possible 'hot spot' mutation, p.Ser107Leu present in four families. We used the recently solved crystal structure of a highly conserved region of the Drosophila orthologue of BICD2 to further-explore how the p.Glu774Gly substitution inhibits the binding of BICD2 to Rab6. Overall, the features of BICD2 spinal muscular atrophy, lower extremity predominant are consistent with a pathological process that preferentially affects lumbar lower motor neurons, with or without additional upper motor neuron involvement. Defining the phenotypic features in this, the largest BICD2 disease cohort reported to date, will facilitate focused genetic testing and filtering of next generation sequencing-derived variants in cases with similar features.
Assuntos
Proteínas Associadas aos Microtúbulos/genética , Atrofia Muscular Espinal/genética , Atrofia Muscular Espinal/patologia , Adolescente , Adulto , Idoso , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Músculo Esquelético/patologia , Mutação/genética , Linhagem , Fenótipo , Ligação Proteica , Coluna Vertebral/patologia , Adulto JovemRESUMO
Mutations in VCP have been reported to account for a spectrum of phenotypes that include inclusion body myopathy with Paget's disease of the bone and frontotemporal dementia, hereditary spastic paraplegia, and 1-2% of familial amyotrophic lateral sclerosis. We identified a novel VCP mutation (p.Glu185Lys) segregating in an autosomal dominant Charcot-Marie-Tooth disease type 2 family. Functional studies showed that the Glu185Lys variant impaired autophagic function leading to the accumulation of immature autophagosomes. VCP mutations should thus be considered for genetically undefined Charcot-Marie-Tooth disease type 2.