Studying T Cells N-Glycosylation by Imaging Flow Cytometry

Imaging flow cytometry is an emerging imaging technology that combines features of both conventional flow cytometry and fluorescence microscopy allowing quantification of the imaging parameters. The analysis of protein posttranslational modifications by glycosylation using imaging flow cytometry constitutes an important bioimaging tool in the glycobiology field. This technique allows quantification of the glycan fluorescence intensity, co-localization with proteins, and evaluation of the membrane/cytoplasmic expression. In this chapter we provide the guidelines to analyze glycan expression, particularly the ß1,6 GlcNAc branched N-glycans, on the membrane of intestinal T cells from inflammatory bowel disease patients.This work was supported by grants from the Portuguese Foundation for Science and Technology (FCT), project grants (PTDC/DTPPIC/0560/2014; PTDC/BBB-EBI/0786/2012; EXPL/BIMMEC/0149/2012), “financiados no âmbito do Programa Operacional Temático Factores de Competitividade (COMPETE) e comparticipado pelo fundo Comunitário Europeu FEDER,” e do Quadro de Referência Estratégia Nacional QREN. This work was further supported by a Portuguese grant from “Grupo de Estudo da Doença Infl amatória Intestinal” (GEDII). This work had also the fi nantial support of FCT/MEC through National Funds and, when applicable, co-fi nanced by the FEDER via the PT2020 Partnership Agreement under the 4293 Unit I&D. S.S.P. (SFRH/BPD/63094/2009) also acknowledges FCT. A.M.D. PD/BD/105982/2014 also acknowledges FCT and BiotechHealth Doctoral Programme. The Institute of Molecular Pathology and Immunology of the University of Porto (IPATIMUP) integrates the Institute for Research and Innovation in Health (I3S), which is partially supported by the Portuguese Foundation for Science and Technology (FCT). Data was acquired at the Bioimaging Center for Biomaterials and Regenerative Therapies (b.IMAGE, INEB, Porto, Portugal)

COVID-19 in a Pediatric Cohort—Retrospective Review of Chest Computer Tomography Findings

Background: Radiological features of the novel 2019 coronavirus disease (COVID-19) have been mainly described in adults. Available literature states that imaging findings in children are similar but less pronounced. The aim of this study is to describe and illustrate the chest computer tomography (CT) features of pediatric COVID-19. Results: This retrospective study was based on the review of all the chest CTs performed in pediatric patients with confirmed COVID-19 disease between March 8th and May 26th 2020 (n = 24). The presence of comorbidities and coinfection was assessed, as well as timing of CT examination in relation to the onset of symptoms. CT findings were categorized as typical, indeterminate, atypical, and negative for COVID-19 according to International Expert Consensus Statement on Chest Imaging in Pediatric COVID-19 Patient Management. This study found that CT findings were abnormal in 17 (71%) patients, with 5 (21%), 9 (38%), and 3 (13%) patients considered to have typical, indeterminate, and atypical findings, respectively. The most common CT patterns were multiple ground-glass opacities (58%), followed by consolidations (50%). Six patients showed predominantly peripheral distribution of parenchymal abnormalities. A halo sign was identified in 3 patients and a perilobular pattern was identified in one of the cases with typical findings. Conclusions: Chest CT findings in children infected with SARS-CoV-2 can be subtle or absent. Besides recognizing typical findings, radiologists should be able to identify features that favor different or concomitant diagnosis.info:eu-repo/semantics/publishedVersio

Levels of Angiopoietin 2 Are Predictive for Mortality in Patients Infected With Yellow Fever Virus

In 2018 there was a large yellow fever outbreak in São Paulo, Brazil, with a high fatality rate. Yellow fever virus can cause, among other symptoms, hemorrhage and disseminated intravascular coagulation, indicating a role for endothelial cells in disease pathogenesis. Here, we conducted a case-control study and measured markers related to endothelial damage in plasma and its association with mortality. We found that angiopoietin 2 is strongly associated with a fatal outcome and could serve as a predictive marker for mortality. This could be used to monitor severe cases and provide care to improve disease outcome

Dysregulation of T cell receptor N-glycosylation: A molecular mechanism involved in ulcerative colitis

The incidence of inflammatory bowel disease is increasing worldwide and the underlying molecular mechanisms are far from being fully elucidated. Herein, we evaluated the role of N-glycosylation dysregulation in T cells as a key mechanism in the ulcerative colitis (UC) pathogenesis. The evaluation of the branched N-glycosylation levelsandprofile of intestinalTcell receptor (TCR)wereassessedin colonic biopsies fromUCpatientsand healthy controls. Expression alterations of the glycosyltransferase gene MGAT5 were also evaluated. We demonstrated thatUCpatients exhibit a dysregulation ofTCRbranchedN-glycosylationonlamina propriaTlymphocytes. Patients with severe UC showed the most pronounced defect on N-glycan branching in T cells. Moreover, UC patients showed a significant reduction of MGAT5 gene transcription in T lymphocytes. In this study, we disclose for the first time that a deficiency in branched N-glycosylation on TCR due to a reduced MGAT5 gene expression is a new molecular mechanism underlying UC pathogenesis, being a potential novel biomarker with promising clinical and therapeutic applications.This work was supported by grants from the Portuguese Foundation for Science and Technology (FCT), project grants (PTDC/ CVT/111358/2009; PTDC/BBB-EBI/0786/2012; EXPL/ BIM-MEC/0149/2012), ‘financiados no âmbito do Programa Operacional Temático Factores de Competitividade (COMPETE) e comparticipado pelo fundo Comunitário Europeu FEDER’, e do Quadro de Referência Estratégia Nacio-nal QREN. This work was further supported by a portuguese grant from ‘Grupo de Estudo da Doenc¸a Inflamatória Intestinal’ (GEDII). S.S.P. (SFRH/BPD/63094/2009); S.C. (SFRH/BD/ 77386/2011) also acknowledge FCT. IPATIMUP is an Associate Laboratory of the Portuguese Ministry of Science, Technology and Higher Education, and is partially supported by FCT

Integração de métodos multicritério na busca da sustentabilidade agrícola para a produção de tomates no município de São José de Ubá-RJ.

Este estudo discute alternativas para os sistemas produtivos da terra no plantio de tomates em São José de Ubá-RJ e as relações de trabalho passíveis de serem colocadas em pratica de forma a promover uma agricultura sustentavel na região, priorizando os aspectos economicos, ambientais e culturais do problema. A introdução da questão cultural aumenta a complexidade da questão, sendo necessário o uso de uma metodologia capaz de lidar com toda a subjetividade envolvida nesse processo de tomada de decisão. O estudo visa contribuir para a conquista de um processo sustentavel na região, estabelecendo um processo de tomada de decisão pautado nas opinioes do agricultor, respeitando principalmente as questões culturais do problema. As características do ambiente de tomada de decisão indicaram o uso de uma combinação de métodos de apoio a decisão multicritério - MACBETH e VIP Analysis - para a seleção da melhor alternativa capaz de possibilitar o alcance dos objetivos propostos

DNA methylation regulates expression of VEGF-R2 (KDR) and VEGF-R3 (FLT4)

Abstract Background Vascular Endothelial Growth Factors (VEGFs) and their receptors (VEGF-Rs) are important regulators for angiogenesis and lymphangiogenesis. VEGFs and VEGF-Rs are not only expressed on endothelial cells but also on various subtypes of solid tumors and leukemias contributing to the growth of the malignant cells. This study was performed to examine whether VEGF-R2 (KDR) and VEGF-R3 (FLT4) are regulated by DNA methylation. Methods Real-time (RT) PCR analysis was performed to quantify KDR and FLT4 expression in some ninety leukemia/lymphoma cell lines, human umbilical vein endothelial cells (HUVECs) and dermal microvascular endothelial cells (HDMECs). Western blot analyses and flow cytometric analyses confirmed results at the protein level. After bisulfite conversion of DNA we determined the methylation status of KDR and FLT4 by DNA sequencing and by methylation specific PCR (MSP). Western blot analyses were performed to examine the effect of VEGF-C on p42/44 MAPK activation. Results Expression of KDR and FLT4 was observed in cell lines from various leukemic entities, but not in lymphoma cell lines: 16% (10/62) of the leukemia cell lines expressed KDR, 42% (27/65) were FLT4 positive. None of thirty cell lines representing six lymphoma subtypes showed more than marginal expression of KDR or FLT4. Western blot analyses confirmed KDR and FLT4 protein expression in HDMECs, HUVECs and in cell lines with high VEGF-R mRNA levels. Mature VEGF-C induced p42/44 MAPK activation in the KDR- /FLT4+ cell line OCI-AML1 verifying the model character of this cell line for VEGF-C signal transduction studies. Bisulfite sequencing and MSP revealed that GpG islands in the promoter regions of KDR and FLT4 were unmethylated in HUVECs, HDMECs and KDR + and FLT4 + cell lines, whereas methylated cell lines did not express these genes. In hypermethylated cell lines, KDR and FLT4 were re-inducible by treatment with the DNA demethylating agent 5-Aza-2'deoxycytidine, confirming epigenetic regulation of both genes. Conclusions Our data show that VEGF-Rs KDR and FLT4 are silenced by DNA methylation. However, if the promoters are unmethylated, other factors (e.g. transactivation factors) determine the extent of KDR and FLT4 expression

Optimal functional outcome measures for assessing treatment for Dupuytren's disease: A systematic review and recommendations for future practice

This article is available through the Brunel Open Access Publishing Fund. Copyright © 2013 Ball et al.; licensee BioMed Central Ltd.Background: Dupuytren's disease of the hand is a common condition affecting the palmar fascia, resulting in progressive flexion deformities of the digits and hence limitation of hand function. The optimal treatment remains unclear as outcomes studies have used a variety of measures for assessment. Methods: A literature search was performed for all publications describing surgical treatment, percutaneous needle aponeurotomy or collagenase injection for primary or recurrent Dupuytren’s disease where outcomes had been monitored using functional measures. Results: Ninety-one studies met the inclusion criteria. Twenty-two studies reported outcomes using patient reported outcome measures (PROMs) ranging from validated questionnaires to self-reported measures for return to work and self-rated disability. The Disability of Arm, Shoulder and Hand (DASH) score was the most utilised patient-reported function measure (n=11). Patient satisfaction was reported by eighteen studies but no single method was used consistently. Range of movement was the most frequent physical measure and was reported in all 91 studies. However, the methods of measurement and reporting varied, with seventeen different techniques being used. Other physical measures included grip and pinch strength and sensibility, again with variations in measurement protocols. The mean follow-up time ranged from 2 weeks to 17 years. Conclusions: There is little consistency in the reporting of outcomes for interventions in patients with Dupuytren’s disease, making it impossible to compare the efficacy of different treatment modalities. Although there are limitations to the existing generic patient reported outcomes measures, a combination of these together with a disease-specific questionnaire, and physical measures of active and passive individual joint Range of movement (ROM), grip and sensibility using standardised protocols should be used for future outcomes studies. As Dupuytren’s disease tends to recur following treatment as well as extend to involve other areas of the hand, follow-up times should be standardised and designed to capture both short and long term outcomes

Monovarietal extra-virgin olive oil classification: a fusion of human sensory attributes and an electronic tongue

Olive oil quality grading is traditionally assessed by human sensory evaluation of positive and negative attributes (olfactory, gustatory, and final olfactorygustatory sensations). However, it is not guaranteed that trained panelist can correctly classify monovarietal extra-virgin olive oils according to olive cultivar. In this work, the potential application of human (sensory panelists) and artificial (electronic tongue) sensory evaluation of olive oils was studied aiming to discriminate eight single-cultivar extra-virgin olive oils. Linear discriminant, partial least square discriminant, and sparse partial least square discriminant analyses were evaluated. The best predictive classification was obtained using linear discriminant analysis with simulated annealing selection algorithm. A low-level data fusion approach (18 electronic tongue signals and nine sensory attributes) enabled 100 % leave-one-out cross-validation correct classification, improving the discrimination capability of the individual use of sensor profiles or sensory attributes (70 and 57 % leave-one-out correct classifications, respectively). So, human sensory evaluation and electronic tongue analysis may be used as complementary tools allowing successful monovarietal olive oil discrimination.This work was co-financed by FCT/MEC and FEDER under Program PT2020 (Project UID/EQU/50020/2013); by Fundacao para a Ciencia e Tecnologia under the strategic funding of UID/BIO/04469/2013 unit; and by Project POCTEP through Project RED/AGROTEC-Experimentation network and transfer for development of agricultural and agro industrial sectors between Spain and Portugal

Ecological research in the Large-scale Biosphere-Atmosphere Experiment in Amazonia: Early results

Copyright by the Ecological Society of America ©2004 Michael Keller, Ane Alencar, Gregory P. Asner, Bobby Braswell, Mercedes Bustamante, Eric Davidson, Ted Feldpausch, Erick Fernandes, Michael Goulden, Pavel Kabat, Bart Kruijt, Flavio Luizão, Scott Miller, Daniel Markewitz, Antonio D. Nobre, Carlos A. Nobre, Nicolau Priante Filho, Humberto da Rocha, Pedro Silva Dias, Celso von Randow, and George L. Vourlitis 2004. ECOLOGICAL RESEARCH IN THE LARGE-SCALE BIOSPHERE– ATMOSPHERE EXPERIMENT IN AMAZONIA: EARLY RESULTS. Ecological Applications 14:3–16. http://dx.doi.org/10.1890/03-6003The Large-scale Biosphere–Atmosphere Experiment in Amazonia (LBA) is a multinational, interdisciplinary research program led by Brazil. Ecological studies in LBA focus on how tropical forest conversion, regrowth, and selective logging influence carbon storage, nutrient dynamics, trace gas fluxes, and the prospect for sustainable land use in the Amazon region. Early results from ecological studies within LBA emphasize the variability within the vast Amazon region and the profound effects that land-use and land-cover changes are having on that landscape. The predominant land cover of the Amazon region is evergreen forest; nonetheless, LBA studies have observed strong seasonal patterns in gross primary production, ecosystem respiration, and net ecosystem exchange, as well as phenology and tree growth. The seasonal patterns vary spatially and interannually and evidence suggests that these patterns are driven not only by variations in weather but also by innate biological rhythms of the forest species. Rapid rates of deforestation have marked the forests of the Amazon region over the past three decades. Evidence from ground-based surveys and remote sensing show that substantial areas of forest are being degraded by logging activities and through the collapse of forest edges. Because forest edges and logged forests are susceptible to fire, positive feedback cycles of forest degradation may be initiated by land-use-change events. LBA studies indicate that cleared lands in the Amazon, once released from cultivation or pasture usage, regenerate biomass rapidly. However, the pace of biomass accumulation is dependent upon past land use and the depletion of nutrients by unsustainable land-management practices. The challenge for ongoing research within LBA is to integrate the recognition of diverse patterns and processes into general models for prediction of regional ecosystem function