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1.
Clin Genet ; 96(1): 72-84, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31001818

RESUMO

Variants in the chromodomain helicase DNA-binding protein 8 (CHD8) have been associated with intellectual disability (ID), autism spectrum disorders (ASDs) and overgrowth and CHD8 is one of the causative genes for OGID (overgrowth and ID). We investigated 25 individuals with CHD8 protein truncating variants (PTVs), including 10 previously unreported patients and found a male to female ratio of 2.7:1 (19:7) and a pattern of common features: macrocephaly (62.5%), tall stature (47%), developmental delay and/or intellectual disability (81%), ASDs (84%), sleep difficulties (50%), gastrointestinal problems (40%), and distinct facial features. Most of the individuals in this cohort had moderate-to-severe ID, some had regression of speech (37%), seizures (27%) and hypotonia (27%) and two individuals were non-ambulant. Our study shows that haploinsufficiency of CHD8 is associated with a distinctive OGID syndrome with pronounced autistic traits and supports a sex-dependent penetrance of CHD8 PTVs in humans.

2.
Nat Commun ; 10(1): 1178, 2019 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-30862802

RESUMO

Damage to alveoli, the gas-exchanging region of the lungs, is a component of many chronic and acute lung diseases. In addition, insufficient generation of alveoli results in bronchopulmonary dysplasia, a disease of prematurity. Therefore visualising the process of alveolar development (alveologenesis) is critical for our understanding of lung homeostasis and for the development of treatments to repair and regenerate lung tissue. Here we show live alveologenesis, using long-term, time-lapse imaging of precision-cut lung slices. We reveal that during this process, epithelial cells are highly mobile and we identify specific cell behaviours that contribute to alveologenesis: cell clustering, hollowing and cell extension. Using the cytoskeleton inhibitors blebbistatin and cytochalasin D, we show that cell migration is a key driver of alveologenesis. This study reveals important novel information about lung biology and provides a new system in which to manipulate alveologenesis genetically and pharmacologically.


Assuntos
Movimento Celular/fisiologia , Células Epiteliais/fisiologia , Organogênese/fisiologia , Alvéolos Pulmonares/embriologia , Actomiosina/antagonistas & inibidores , Actomiosina/fisiologia , Animais , Animais Recém-Nascidos , Movimento Celular/efeitos dos fármacos , Citocalasina D/farmacologia , Células Epiteliais/efeitos dos fármacos , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Microscopia Intravital , Camundongos , Camundongos Endogâmicos C57BL , Microscopia de Fluorescência , Modelos Animais , Organogênese/efeitos dos fármacos , Alvéolos Pulmonares/efeitos dos fármacos , Imagem com Lapso de Tempo
3.
Metab Eng ; 53: 59-68, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30772453

RESUMO

Recombineering-based redesign of bacterial genomes by adding, removing or editing large segments of genomic DNA is emerging as a powerful technique for expanding the range of functions that an organism can perform. Here, we describe a glyco-recoding strategy whereby major non-essential polysaccharide gene clusters in K-12 Escherichia coli are replaced with orthogonal glycosylation components for both biosynthesis of heterologous glycan structures and site-specific glycan conjugation to target proteins. Specifically, the native enterobacterial common antigen (ECA) and O-polysaccharide (O-PS) antigen loci were systematically replaced with ∼9-10 kbp of synthetic DNA encoding Campylobacter jejuni enzymes required for asparagine-linked (N-linked) protein glycosylation. Compared to E. coli cells carrying the same glycosylation machinery on extrachromosomal plasmids, glyco-recoded strains attached glycans to acceptor protein targets with equal or greater efficiency while exhibiting markedly better growth phenotypes and higher glycoprotein titers. Overall, our results define a convenient and reliable framework for bacterial glycome editing that provides a more stable route for chemical diversification of proteins in vivo and effectively expands the bacterial glycoengineering toolkit.


Assuntos
Proteínas de Bactérias , Campylobacter jejuni/genética , Escherichia coli , Edição de Genes , Família Multigênica , Polissacarídeos Bacterianos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Polissacarídeos Bacterianos/biossíntese , Polissacarídeos Bacterianos/genética
4.
Artigo em Inglês | MEDLINE | ID: mdl-30099598

RESUMO

Bacteria have garnered increased interest in recent years as a platform for the biosynthesis of a variety of glycomolecules such as soluble oligosaccharides, surface-exposed carbohydrates, and glycoproteins. The ability to engineer commonly used laboratory species such as Escherichia coli to efficiently synthesize non-native sugar structures by recombinant expression of enzymes from various carbohydrate biosynthesis pathways has allowed for the facile generation of important products such as conjugate vaccines, glycosylated outer membrane vesicles, and a variety of other research reagents for studying and understanding the role of glycans in living systems. This chapter highlights some of the key discoveries and technologies for equipping bacteria with the requisite biosynthetic machinery to generate such products. As the bacterial glyco-toolbox continues to grow, these technologies are expected to expand the range of glycomolecules produced recombinantly in bacterial systems, thereby opening up this platform to an even larger number of applications.

5.
Nat Commun ; 9(1): 3396, 2018 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-30127449

RESUMO

The original version of this Article contained an error in Figure 2, wherein the bottom right western blot panel in Figure 2a was blank. This has now been corrected in both the PDF and HTML versions of the Article.

6.
Nat Commun ; 9(1): 2686, 2018 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-30002445

RESUMO

The emerging discipline of bacterial glycoengineering has made it possible to produce designer glycans and glycoconjugates for use as vaccines and therapeutics. Unfortunately, cell-based production of homogeneous glycoproteins remains a significant challenge due to cell viability constraints and the inability to control glycosylation components at precise ratios in vivo. To address these challenges, we describe a novel cell-free glycoprotein synthesis (CFGpS) technology that seamlessly integrates protein biosynthesis with asparagine-linked protein glycosylation. This technology leverages a glyco-optimized Escherichia coli strain to source cell extracts that are selectively enriched with glycosylation components, including oligosaccharyltransferases (OSTs) and lipid-linked oligosaccharides (LLOs). The resulting extracts enable a one-pot reaction scheme for efficient and site-specific glycosylation of target proteins. The CFGpS platform is highly modular, allowing the use of multiple distinct OSTs and structurally diverse LLOs. As such, we anticipate CFGpS will facilitate fundamental understanding in glycoscience and make possible applications in on demand biomanufacturing of glycoproteins.

7.
Wellcome Open Res ; 3: 46, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29900417

RESUMO

Tatton-Brown-Rahman syndrome (TBRS; OMIM 615879), also known as the DNMT3A-overgrowth syndrome, is an overgrowth intellectual disability syndrome first described in 2014 with a report of 13 individuals with constitutive heterozygous DNMT3A variants. Here we have undertaken a detailed clinical study of 55 individuals with de novoDNMT3A variants, including the 13 previously reported individuals. An intellectual disability and overgrowth were reported in >80% of individuals with TBRS and were designated major clinical associations. Additional frequent clinical associations (reported in 20-80% individuals) included an evolving facial appearance with low-set, heavy, horizontal eyebrows and prominent upper central incisors; joint hypermobility (74%); obesity (weight ³2SD, 67%); hypotonia (54%); behavioural/psychiatric issues (most frequently autistic spectrum disorder, 51%); kyphoscoliosis (33%) and afebrile seizures (22%). One individual was diagnosed with acute myeloid leukaemia in teenage years. Based upon the results from this study, we present our current management for individuals with TBRS.

8.
Metab Eng ; 47: 488-495, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29702274

RESUMO

A synthetic pathway for production of the eukaryotic trimannosyl chitobiose glycan (mannose3-N-acetylglucosamine2, Man3GlcNAc2) and its transfer to specific asparagine residues in target proteins was previously engineered in Escherichia coli, providing this simple microbe with the ability to perform a complex post-translational protein modification. Here, we leveraged a flow cytometric fluorescence-based assay to improve Man3GlcNAc2 glycan biosynthesis in E. coli cells. Specifically, pathway improvements were identified, including reducing pathway enzyme expression levels and overexpressing nucleotide sugar biosynthesis genes, which enhanced production of lipid-linked Man3GlcNAc2 by nearly 50-fold to 13.9 µg/L. In turn, cells producing higher levels of the Man3GlcNAc2 substrate yielded up to 10 times more glycosylated acceptor protein (to ~ 14 mg/L) than their non-optimized counterparts. These results demonstrate the use of flow cytometry screening as a powerful tool for interrogating the surfaces of glyco-engineered bacteria and identifying meaningful improvements in glycan biosynthesis. We anticipate this approach will enable further optimization of bacterial glycan biosynthesis pathways using new strain engineering tools from metabolic engineering and synthetic biology.

9.
Dis Model Mech ; 11(1)2018 01 29.
Artigo em Inglês | MEDLINE | ID: mdl-29361513

RESUMO

We previously identified dipeptidylpeptidase 10 (DPP10) on chromosome 2 as a human asthma susceptibility gene, through positional cloning. Initial association results were confirmed in many subsequent association studies but the functional role of DPP10 in asthma remains unclear. Using the MRC Harwell N-ethyl-N-nitrosourea (ENU) DNA archive, we identified a point mutation in Dpp10 that caused an amino acid change from valine to aspartic acid in the ß-propeller region of the protein. Mice carrying this point mutation were recovered and a congenic line was established (Dpp10145D ). Macroscopic examination and lung histology revealed no significant differences between wild-type and Dpp10145D/145D mice. However, after house dust mite (HDM) treatment, Dpp10 mutant mice showed significantly increased airway resistance in response to 100 mg/ml methacholine. Total serum IgE levels and bronchoalveolar lavage (BAL) eosinophil counts were significantly higher in homozygotes than in control mice after HDM treatment. DPP10 protein is present in airway epithelial cells and altered expression is observed in both tissue from asthmatic patients and in mice following HDM challenge. Moreover, knockdown of DPP10 in human airway epithelial cells results in altered cytokine responses. These results show that a Dpp10 point mutation leads to increased airway responsiveness following allergen challenge and provide biological evidence to support previous findings from human genetic studies. This article has an associated First Person interview with the first author of the paper.


Assuntos
Asma/enzimologia , Asma/prevenção & controle , Dipeptidil Peptidases e Tripeptidil Peptidases/metabolismo , Sequência de Aminoácidos , Animais , Asma/complicações , Asma/patologia , Sequência de Bases , Dipeptidil Peptidases e Tripeptidil Peptidases/química , Dipeptidil Peptidases e Tripeptidil Peptidases/genética , Modelos Animais de Doenças , Células Epiteliais/metabolismo , Etilnitrosoureia , Genótipo , Homozigoto , Humanos , Hipersensibilidade/complicações , Hipersensibilidade/patologia , Inflamação/complicações , Inflamação/patologia , Mediadores da Inflamação/metabolismo , Pulmão/parasitologia , Pulmão/patologia , Camundongos , Camundongos Mutantes , Mutação/genética , Pyroglyphidae , Reprodutibilidade dos Testes
10.
Sci Rep ; 7(1): 1880, 2017 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-28500339

RESUMO

The question of how cell division orientation is determined is fundamentally important for understanding tissue and organ shape in both healthy or disease conditions. Here we provide evidence for cell contact-dependent orientation of planar cell division in the mammalian embryonic skin. We propose a model where the core planar polarity proteins Celsr1 and Frizzled-6 (Fz6) communicate the long axis orientation of interphase basal cells to neighbouring basal mitoses so that they align their horizontal division plane along the same axis. The underlying mechanism requires a direct, cell surface, planar polarised cue, which we posit depends upon variant post-translational forms of Celsr1 protein coupled to Fz6. Our hypothesis has parallels with contact-mediated division orientation in early C. elegans embryos suggesting functional conservation between the adhesion-GPCRs Celsr1 and Latrophilin-1. We propose that linking planar cell division plane with interphase neighbour long axis geometry reinforces axial bias in skin spreading around the mouse embryo body.

11.
J Psychopharmacol ; 31(5): 519-552, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28440103

RESUMO

Decisions about the use of psychotropic medication in pregnancy are an ongoing challenge for clinicians and women with mental health problems, owing to the uncertainties around risks of the illness itself to mother and fetus/infant, effectiveness of medications in pregnancy and risks to the fetus/infant from in utero exposure or via breast milk. These consensus guidelines aim to provide pragmatic advice regarding these issues. They are divided into sections on risks of untreated illness in pregnancy; general principles of using drugs in the perinatal period; benefits and harms associated with individual drugs; and recommendations for the management of specific disorders.


Assuntos
Transtornos Mentais/tratamento farmacológico , Período Pós-Parto/efeitos dos fármacos , Psicofarmacologia/normas , Psicotrópicos/uso terapêutico , Feminino , Humanos , Gravidez
12.
Dis Model Mech ; 10(4): 409-423, 2017 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-28237967

RESUMO

Lung diseases impose a huge economic and health burden worldwide. A key aspect of several adult lung diseases, such as idiopathic pulmonary fibrosis (IPF) and chronic obstructive pulmonary disease (COPD), including emphysema, is aberrant tissue repair, which leads to an accumulation of damage and impaired respiratory function. Currently, there are few effective treatments available for these diseases and their incidence is rising. The planar cell polarity (PCP) pathway is critical for the embryonic development of many organs, including kidney and lung. We have previously shown that perturbation of the PCP pathway impairs tissue morphogenesis, which disrupts the number and shape of epithelial tubes formed within these organs during embryogenesis. However, very little is known about the role of the PCP pathway beyond birth, partly because of the perinatal lethality of many PCP mouse mutant lines. Here, we investigate heterozygous Looptail (Lp) mice, in which a single copy of the core PCP gene, Vangl2, is disrupted. We show that these mice are viable but display severe airspace enlargement and impaired adult lung function. Underlying these defects, we find that Vangl2Lp/+ lungs exhibit altered distribution of actin microfilaments and abnormal regulation of the actin-modifying protein cofilin. In addition, we show that Vangl2Lp/+ lungs exhibit many of the hallmarks of tissue damage, including an altered macrophage population, abnormal elastin deposition and elevated levels of the elastin-modifying enzyme, Mmp12, all of which are observed in emphysema. In vitro, disruption of VANGL2 impairs directed cell migration and reduces the rate of repair following scratch wounding of human alveolar epithelial cells. Moreover, using population data from a birth cohort of young adults, all aged 31, we found evidence of an interactive effect between VANGL2 and smoking on lung function. Finally, we show that PCP genes VANGL2 and SCRIB are significantly downregulated in lung tissue from patients with emphysema. Our data reveal an important novel role for the PCP pathway in adult lung homeostasis and repair and shed new light on the genetic factors which may modify destructive lung diseases such as emphysema.


Assuntos
Envelhecimento/patologia , Polaridade Celular , Homeostase , Pulmão/patologia , Proteínas do Tecido Nervoso/genética , Cicatrização , Células A549 , Citoesqueleto de Actina/metabolismo , Animais , Movimento Celular , Regulação para Baixo/genética , Elastina/metabolismo , Embrião de Mamíferos/patologia , Técnicas de Silenciamento de Genes , Heterozigoto , Humanos , Pulmão/embriologia , Pulmão/fisiopatologia , Macrófagos/patologia , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Modelos Biológicos , Mutação/genética , Fenótipo , Polimorfismo Genético , Doença Pulmonar Obstrutiva Crônica/genética , Fumar/efeitos adversos , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo
13.
Health Technol Assess ; 20(74): 1-268, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27731292

RESUMO

BACKGROUND: Nausea and vomiting in pregnancy (NVP) affects up to 85% of all women during pregnancy, but for the majority self-management suffices. For the remainder, symptoms are more severe and the most severe form of NVP - hyperemesis gravidarum (HG) - affects 0.3-1.0% of pregnant women. There is no widely accepted point at which NVP becomes HG. OBJECTIVES: This study aimed to determine the relative clinical effectiveness and cost-effectiveness of treatments for NVP and HG. DATA SOURCES: MEDLINE, EMBASE, Cumulative Index to Nursing and Allied Health Literature, Cochrane Central Register of Controlled Trials, PsycINFO, Commonwealth Agricultural Bureaux (CAB) Abstracts, Latin American and Caribbean Health Sciences Literature, Allied and Complementary Medicine Database, British Nursing Index, Science Citation Index, Social Sciences Citation Index, Scopus, Conference Proceedings Index, NHS Economic Evaluation Database, Health Economic Evaluations Database, China National Knowledge Infrastructure, Cochrane Database of Systematic Reviews and Database of Abstracts of Reviews of Effects were searched from inception to September 2014. References from studies and literature reviews identified were also examined. Obstetric Medicine was hand-searched, as were websites of relevant organisations. Costs came from NHS sources. REVIEW METHODS: A systematic review of randomised and non-randomised controlled trials (RCTs) for effectiveness, and population-based case series for adverse events and fetal outcomes. Treatments: vitamins B6 and B12, ginger, acupressure/acupuncture, hypnotherapy, antiemetics, dopamine antagonists, 5-hydroxytryptamine receptor antagonists, intravenous (i.v.) fluids, corticosteroids, enteral and parenteral feeding or other novel treatment. Two reviewers extracted data and quality assessed studies. Results were narratively synthesised; planned meta-analysis was not possible due to heterogeneity and incomplete reporting. A simple economic evaluation considered the implied values of treatments. RESULTS: Seventy-three studies (75 reports) met the inclusion criteria. For RCTs, 33 and 11 studies had a low and high risk of bias respectively. For the remainder (n = 20) it was unclear. The non-randomised studies (n = 9) were low quality. There were 33 separate comparators. The most common were acupressure versus placebo (n = 12); steroid versus usual treatment (n = 7); ginger versus placebo (n = 6); ginger versus vitamin B6 (n = 6); and vitamin B6 versus placebo (n = 4). There was evidence that ginger, antihistamines, metoclopramide (mild disease) and vitamin B6 (mild to severe disease) are better than placebo. Diclectin® [Duchesnay Inc.; doxylamine succinate (10 mg) plus pyridoxine hydrochloride (10 mg) slow release tablet] is more effective than placebo and ondansetron is more effective at reducing nausea than pyridoxine plus doxylamine. Diclectin before symptoms of NVP begin for women at high risk of severe NVP recurrence reduces risk of moderate/severe NVP compared with taking Diclectin once symptoms begin. Promethazine is as, and ondansetron is more, effective than metoclopramide for severe NVP/HG. I.v. fluids help correct dehydration and improve symptoms. Dextrose saline may be more effective at reducing nausea than normal saline. Transdermal clonidine patches may be effective for severe HG. Enteral feeding is effective but extreme method treatment for very severe symptoms. Day case management for moderate/severe symptoms is feasible, acceptable and as effective as inpatient care. For all other interventions and comparisons, evidence is unclear. The economic analysis was limited by lack of effectiveness data, but comparison of costs between treatments highlights the implications of different choices. LIMITATIONS: The main limitations were the quantity and quality of the data available. CONCLUSION: There was evidence of some improvement in symptoms for some treatments, but these data may not be transferable across disease severities. Methodologically sound and larger trials of the main therapies considered within the UK NHS are needed. STUDY REGISTRATION: This study is registered as PROSPERO CRD42013006642. FUNDING: The National Institute for Health Research Health Technology Assessment programme.


Assuntos
Antieméticos/economia , Antieméticos/uso terapêutico , Hiperêmese Gravídica/tratamento farmacológico , Náusea/tratamento farmacológico , Antieméticos/administração & dosagem , Antieméticos/efeitos adversos , Ensaios Clínicos como Assunto , Terapias Complementares/economia , Terapias Complementares/métodos , Análise Custo-Benefício , Feminino , Hidratação/economia , Hidratação/métodos , Humanos , Hiperêmese Gravídica/terapia , Náusea/terapia , Gravidez
14.
JAMA ; 316(13): 1392-1401, 2016 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-27701665

RESUMO

Importance: Nausea and vomiting affects approximately 85% of pregnant women. The most severe form, hyperemesis gravidarum, affects up to 3% of women and can have significant adverse physical and psychological sequelae. Objective: To summarize current evidence on effective treatments for nausea and vomiting in pregnancy and hyperemesis gravidarum. Evidence Review: Databases were searched to June 8, 2016. Relevant websites and bibliographies were also searched. Titles and abstracts were assessed independently by 2 reviewers. Results were narratively synthesized; planned meta-analysis was not possible because of heterogeneity and incomplete reporting of findings. Findings: Seventy-eight studies (n = 8930 participants) were included: 67 randomized clinical trials (RCTs) and 11 nonrandomized studies. Evidence from 35 RCTs at low risk of bias indicated that ginger, vitamin B6, antihistamines, metoclopramide (for mild symptoms), pyridoxine-doxylamine, and ondansetron (for moderate symptoms) were associated with improved symptoms compared with placebo. One RCT (n = 86) reported greater improvements in moderate symptoms following psychotherapy (change in Rhodes score [range, 0 {no symptoms} to 40 {worst possible symptoms}], 18.76 [SD, 5.48] to 7.06 [SD, 5.79] for intervention vs 19.18 [SD, 5.63] to 12.81 [SD, 6.88] for comparator [P < .001]). For moderate-severe symptoms, 1 RCT (n = 60) suggested that pyridoxine-doxylamine combination taken preemptively reduced risk of recurrence of moderate-severe symptoms compared with treatment once symptoms begin (15.4% vs 39.1% [P < .04]). One RCT (n = 83) found that ondansetron was associated with lower nausea scores on day 4 than metoclopramide (mean visual analog scale [VAS] score, 4.1 [SD, 2.9] for ondansetron vs 5.7 [SD, 2.3] for metoclopramide [P = .023]) but not episodes of emesis (5.0 [SD, 3.1] vs 3.3 [SD, 3], respectively [P = .013]). Although there was no difference in trend in nausea scores over the 14-day study period, trend in vomiting scores was better in the ondansetron group (P = .042). One RCT (n = 159) found no difference between metoclopramide and promethazine after 24 hours (episodes of vomiting, 1 [IQR, 0-5] for metoclopramide vs 2 [IQR, 0-3] for promethazine [P = .81], VAS [0-10 scale] for nausea, 2 [IQR, 1-5] vs 2 [IQR, 1-4], respectively [P = .99]). Three RCTs compared corticosteroids with placebo or promethazine or metoclopramide in women with severe symptoms. Improvements were seen in all corticosteroid groups, but only a significant difference between corticosteroids vs metoclopramide was reported (emesis reduction, 40.9% vs 16.5% at day 2; 71.6% vs 51.2% at day 3; 95.8% vs 76.6% at day 7 [n = 40, P < .001]). For other interventions, evidence was limited. Conclusions and Relevance: For mild symptoms of nausea and emesis of pregnancy, ginger, pyridoxine, antihistamines, and metoclopramide were associated with greater benefit than placebo. For moderate symptoms, pyridoxine-doxylamine, promethazine, and metoclopramide were associated with greater benefit than placebo. Ondansetron was associated with improvement for a range of symptom severity. Corticosteroids may be associated with benefit in severe cases. Overall the quality of evidence was low.


Assuntos
Antieméticos/uso terapêutico , Hiperêmese Gravídica/terapia , Náusea/terapia , Complicações na Gravidez/terapia , Psicoterapia , Acupuntura , Corticosteroides/uso terapêutico , Doxilamina/uso terapêutico , Feminino , Gengibre , Antagonistas dos Receptores Histamínicos/uso terapêutico , Humanos , Ondansetron/uso terapêutico , Fitoterapia/métodos , Gravidez , Piridoxina/uso terapêutico , Ensaios Clínicos Controlados Aleatórios como Assunto , Complexo Vitamínico B/uso terapêutico , Vômito/terapia
15.
Neurology ; 86(24): 2251-7, 2016 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-27194385

RESUMO

OBJECTIVE: To investigate pregnancy outcomes following maternal use of pregabalin. METHODS: This multicenter, observational prospective cohort study compared pregnancy outcomes in women exposed to pregabalin with those of matched controls (not exposed to any medications known to be teratogenic or to any antiepileptic drugs). Teratology Information Services systematically collected data between 2004 and 2013. RESULTS: Data were collected from 164 exposed pregnancies and 656 controls. A significantly higher major birth defect rate in the pregabalin group was observed after exclusion of chromosomal aberration syndromes, and when cases with exposure during first trimester of pregnancy were analyzed separately (7/116 [6.0%] vs 12/580 [2.1%]; odds ratio 3.0, 95% confidence interval 1.2-7.9, p = 0.03). The rate of live births was lower in the pregabalin group (71.9% vs 85.2%, p < 0.001), primarily due to a higher rate of both elective (9.8% vs 5.0%, p = 0.02) and medically indicated (5.5% vs 1.8%, p = 0.008) pregnancy terminations. In the Cox proportional cause specific hazards model, pregabalin exposure was not associated with a significantly higher risk of spontaneous abortion. CONCLUSIONS: This study demonstrated a signal for increased risk of major birth defects after first trimester exposure to pregabalin. However, several limitations such as the small sample size, differences across groups in maternal conditions, and concomitant medication exposure exclude definitive conclusions, so these results call for confirmation through independent studies.


Assuntos
Anormalidades Induzidas por Medicamentos/epidemiologia , Fármacos do Sistema Nervoso Central/efeitos adversos , Pregabalina/efeitos adversos , Resultado da Gravidez/epidemiologia , Adulto , Fármacos do Sistema Nervoso Central/uso terapêutico , Europa (Continente) , Feminino , Humanos , Incidência , Farmacovigilância , Pregabalina/uso terapêutico , Gravidez , Primeiro Trimestre da Gravidez/efeitos dos fármacos , Modelos de Riscos Proporcionais , Estudos Prospectivos , Risco
16.
Open Biol ; 6(4): 150243, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-27110302

RESUMO

Currently, Streptococcus pneumoniae is responsible for over 14 million cases of pneumonia worldwide annually, and over 1 million deaths, the majority of them children. The major determinant for pathogenesis is a polysaccharide capsule that is variable and is used to distinguish strains based on their serotype. The capsule forms the basis of the pneumococcal polysaccharide vaccine (PPV23) that contains purified capsular polysaccharide from 23 serotypes, and the pneumococcal conjugate vaccine (PCV13), containing 13 common serotypes conjugated to CRM197 (mutant diphtheria toxin). Purified capsule from S. pneumoniae is required for pneumococcal conjugate vaccine production, and costs can be prohibitively high, limiting accessibility of the vaccine in low-income countries. In this study, we demonstrate the recombinant expression of the capsule-encoding locus from four different serotypes of S. pneumoniae within Escherichia coli. Furthermore, we attempt to identify the minimum set of genes necessary to reliably and efficiently express these capsules heterologously. These E. coli strains could be used to produce a supply of S. pneumoniae serotype-specific capsules without the need to culture pathogenic bacteria. Additionally, these strains could be applied to synthetic glycobiological applications: recombinant vaccine production using E. coli outer membrane vesicles or coupling to proteins using protein glycan coupling technology.


Assuntos
Cápsulas Bacterianas/metabolismo , Escherichia coli/metabolismo , Polissacarídeos Bacterianos/metabolismo , Recombinação Genética/genética , Streptococcus pneumoniae/metabolismo , Vias Biossintéticas/genética , Elementos de DNA Transponíveis/genética , Ensaio de Imunoadsorção Enzimática , Imunofluorescência , Genes Bacterianos , Loci Gênicos , Immunoblotting , Lipopolissacarídeos/metabolismo , Mutação/genética , Sorotipagem , Streptococcus pneumoniae/genética
17.
Glycobiology ; 26(4): 398-409, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26610891

RESUMO

Bacterial N-linking oligosaccharyl transferases (OTase enzymes) transfer lipid-linked glycans to selected proteins in the periplasm and were first described in the intestinal pathogen Campylobacter jejuni, a member of the ε-proteobacteria-subdivision of bacteria. More recently, orthologues from other ε-proteobacterial Campylobacter and Helicobacter species and a δ-proteobacterium, Desulfovibrio desulfuricans, have been described, suggesting that these two subdivisions of bacteria may be a source of further N-linked protein glycosylation systems. Whole-genome sequencing of both ε- and δ-proteobacteria from deep-sea vent habitats, a rich source of species from these subdivisions, revealed putative ORFs encoding OTase enzymes and associated adjacent glycosyltransferases similar to the C. jejuni N-linked glycosylation locus. We expressed putative OTase ORFs from the deep-sea vent species Nitratiruptor tergarcus, Sulfurovum lithotrophicum and Deferribacter desulfuricans in Escherichia coli and showed that they were able to functionally complement the C. jejuni OTase, CjPglB. The enzymes were shown to possess relaxed glycan specificity, transferring diverse glycan structures and demonstrated different glycosylation sequon specificities. Additionally, a permissive D. desulfuricans acceptor protein was identified, and we provide evidence that the N-linked glycan synthesized by N. tergarcus and S. lithotrophicum contains an acetylated sugar at the reducing end. This work demonstrates that deep-sea vent bacteria encode functional N-glycosylation machineries and are a potential source of biotechnologically important OTase enzymes.


Assuntos
Hexosiltransferases/genética , Proteínas de Membrana/genética , Polissacarídeos/metabolismo , Proteobactérias/genética , Escherichia coli/genética , Genoma Bacteriano , Glicosilação , Hexosiltransferases/biossíntese , Hexosiltransferases/metabolismo , Proteínas de Membrana/biossíntese , Proteínas de Membrana/metabolismo , Oceanos e Mares , Polissacarídeos/biossíntese , Proteobactérias/enzimologia , Especificidade por Substrato
18.
Am J Med Genet A ; 167A(8): 1916-20, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25900906

RESUMO

We describe two patients with microdeletion 1p35.2, intrauterine growth retardation, small stature, hypermetropia, hearing impairment and developmental delay. Both patients have long, myopathic facies, with fine eyebrows, small mouths and micrognathia. We postulate a role for the histone deacetylase HDAC1 in the facial phenotype and suggest that deletion of KPNA6 may prevent transmission of the 1p35.2 deletion from affected girls to any offspring through impaired zygotic genome activation.


Assuntos
Deleção Cromossômica , Cromossomos Humanos Par 1 , Deficiências do Desenvolvimento/terapia , Criança , Feminino , Humanos , Fenótipo
19.
PLoS Genet ; 10(5): e1004323, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24852022

RESUMO

Polarity coordinates cell movement, differentiation, proliferation and apoptosis to build and maintain complex epithelial tissues such as the mammary gland. Loss of polarity and the deregulation of these processes are critical events in malignant progression but precisely how and at which stage polarity loss impacts on mammary development and tumourigenesis is unclear. Scrib is a core polarity regulator and tumour suppressor gene however to date our understanding of Scrib function in the mammary gland has been limited to cell culture and transplantation studies of cell lines. Utilizing a conditional mouse model of Scrib loss we report for the first time that Scrib is essential for mammary duct morphogenesis, mammary progenitor cell fate and maintenance, and we demonstrate a critical and specific role for Scribble in the control of the early steps of breast cancer progression. In particular, Scrib-deficiency significantly induced Fra1 expression and basal progenitor clonogenicity, which resulted in fully penetrant ductal hyperplasia characterized by high cell turnover, MAPK hyperactivity, frank polarity loss with mixing of apical and basolateral membrane constituents and expansion of atypical luminal cells. We also show for the first time a role for Scribble in mammalian spindle orientation with the onset of mammary hyperplasia being associated with aberrant luminal cell spindle orientation and a failure to apoptose during the final stage of duct tubulogenesis. Restoring MAPK/Fra1 to baseline levels prevented Scrib-hyperplasia, whereas persistent Scrib deficiency induced alveolar hyperplasia and increased the incidence, onset and grade of mammary tumours. These findings, based on a definitive genetic mouse model provide fundamental insights into mammary duct maturation and homeostasis and reveal that Scrib loss activates a MAPK/Fra1 pathway that alters mammary progenitor activity to drive premalignancy and accelerate tumour progression.


Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Sistema de Sinalização das MAP Quinases , Neoplasias Mamárias Experimentais/metabolismo , Proteínas Proto-Oncogênicas c-fos/metabolismo , Animais , Polaridade Celular , Feminino , Homeostase , Hiperplasia , Glândulas Mamárias Animais/patologia , Neoplasias Mamárias Experimentais/patologia , Camundongos , Morfogênese
20.
Neurology ; 80(17): 1565-70, 2013 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-23553472

RESUMO

OBJECTIVES: Our objectives were to 1) determine whether first-trimester use of gabapentin is associated with an increased risk for major malformations; 2) examine rates of spontaneous abortions, therapeutic abortions, stillbirths, mean birth weight and gestational age at delivery; and 3) examine rates of poor neonatal adaptation syndrome following late pregnancy exposure. METHODS: The study design was prospective. Women were included who initially contacted the services between 5 and 8 weeks with a comparison group of women exposed to nonteratogens, collected in a similar manner. RESULTS: We have data on 223 pregnancy outcomes exposed to gabapentin and 223 unexposed pregnancies. The rates of major malformations were similar in both groups (p = 0.845). There was a higher rate of preterm births (p = 0.019) and low birth weight <2,500 g (p = 0.033) in the gabapentin group. Among infants who were exposed to gabapentin up until delivery, 23 of 61 (38%) were admitted to either the neonatal intensive care unit or special care nursery for observation and/or treatment, vs 6 of 201 (2.9%) live births in the comparison group (p < 0.001). There were 2 cases of possible poor neonatal adaptation syndrome in neonates exposed to gabapentin close to delivery, compared with none in the comparison group, although it must be noted that these infants were concomitantly exposed to other psychotropic drugs. Among the women who took gabapentin, the major indications were pain (n = 90; 43%) and epilepsy (n = 71; 34%); the remainder were for other indications, mostly psychiatric. CONCLUSION: Our results suggest that although this sample size is not large enough to make any definitive conclusions, and there was no comparator group treated with other antiepileptic drugs, gabapentin use in pregnancy does not appear to increase the risk for major malformations. This finding and the increased risk for low birth weight and preterm birth require further investigation.


Assuntos
Anormalidades Induzidas por Medicamentos/epidemiologia , Aminas/efeitos adversos , Anticonvulsivantes/efeitos adversos , Ácidos Cicloexanocarboxílicos/efeitos adversos , Resultado da Gravidez , Ácido gama-Aminobutírico/efeitos adversos , Anormalidades Induzidas por Medicamentos/etiologia , Adulto , Feminino , Gabapentina , Humanos , Gravidez , Estudos Prospectivos
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