Virulence factors of piglets septicemia causing Aeromonas hydrophila

This work aims to contribute to determine the resistance profile to different antibiotics (ampicillin, gentamicin, penicillin G, oxytetracycline, lincomycin, neomycin, streptomycin, enrofloxacin, colistin sulfate, trimethoprim, sulfamide, tulathromycin, ceftiofur, amoxicillin/clavulanic acid), to assess genetic determinants associated to aminoglycoside antibiotics resistance, namely the presence of genes encoding acetyltransferases (AAC), phosphotransferases (APH) and nucletildiltranferases (ANT), determined by PCR studies, and to search for potentially pathogenic features as the production of extracellular lipases and proteases and the presence of genes encoding for putative virulence factors as aerolysin and related toxins, lipase proteins and type III secretion system component

Mental Retardation: Case Series of Dona Estefânia Hospital Child Development Centre

Segundo a DSM IV a Deficiência Mental (DM) define-se como o funcionamento intelectual global inferior à média (QI < 70) associado a perturbações do comportamento adaptativo com início antes dos 18 anos. Procurou-se caracterizar retrospectivamente a população de crianças com DM observadas no Centro de Desenvolvimento do Hospital de Dona Estefânia (CDHDE), entre Janeiro 2005 e Junho 2007. Foram avaliados os dados epidemiológicos, gravidade, etiologia, co-morbilidade e intervenção proposta. Do total de 232 processos clínicos observados, 185 apresentavam DM. Classificaram-se em DM ligeira 112 (61%), DM moderada 54 (29%), DM grave 17 (9%) e profunda 2 (1%). Foram definidas etiologias em 86 crianças (46%) sendo a taxa de diagnóstico mais elevada na DM de maior gravidade. Observou-se uma elevada variabilidade de etiologias: as mais frequentemente encontradas foram as doenças genéticas, prematuridade e patologia associada. Foi detectada co-morbilidade em 123 crianças (66%), sendo a mais frequente as do foro oftalmológico (57 crianças, 46%). Foram propostas e sinalizadas para apoio a totalidade das crianças com DM, 47% em intervenção precoce e 58% em educação especial, das quais 5% usufruiram, por curto período, do apoio simultaneo de educadora de Intervenção Precoce e de docente do Ensino Especial, durante o período inicial de integração em jardim de infância. Observou-se um predomínio do sexo masculino. Foi efectuada caracterização clínica e funcional das crianças seguidas no CDHDE com o diagnóstico de DM e encontraram-se semelhanças entre os dados presentes e os descritos na literatura. Contudo alguns dados diferem de outras casuísticas decorrente, muito provavelmente decorrente da heterogeneidade da população estudada, quer do ponto de vista etiológico, quer no referente aos grupos etários, provavelmente condicionada, pela política assistencial

Three-dimensional culture of single embryonic stem-derived neural/stem progenitor cells in fibrin hydrogels: neuronal network formation and matrix remodelling

In an attempt to improve the efficacy of neural stem/progenitor cell (NSPC) based therapies, fibrin hydrogels are being explored to provide a favourable microenvironment for cell survival and differentiation following transplantation. In the present work, the ability of fibrin to support the survival, proliferation, and neuronal differentiation of NSPCs derived from embryonic stem (ES) cells under monolayer culture was explored. Single mouse ES-NSPCs were cultured within fibrin (fibrinogen concentration: 6 mg/ml) under neuronal differentiation conditions up to 14 days. The ES-NSPCs retained high cell viability and proliferated within small-sized spheroids. Neuronal differentiation was confirmed by an increase in the levels of ßIII-tubulin and NF200 over time. At day 14, cell-matrix constructs mainly comprised NSPCs and neurons (46.5% ßIII-tubulin + cells). Gamma-aminobutyric acid (GABA)ergic and dopaminergic/noradrenergic neurons were also observed, along with a network of synaptic proteins. The ES-NSPCs expressed matriptase and secreted MMP-2/9, with MMP-2 activity increasing along time. Fibronectin, laminin and collagen type IV deposition was also detected. Fibrin gels prepared with higher fibrinogen concentrations (8/10 mg/ml) were less permissive to neurite extension and neuronal differentiation, possibly owing to their smaller pore area and higher rigidity. Overall, it is shown that ES-NSPCs within fibrin are able to establish neuronal networks and to remodel fibrin through MMP secretion and extracellular matrix (ECM) deposition. This three-dimensional (3D) culture system was also shown to support cell viability, neuronal differentiation and ECM deposition of human ES-NSPCs. The settled 3D platform is expected to constitute a valuable tool to develop fibrin-based hydrogels for ES-NSPC delivery into the injured central nervous system.The authors would like to acknowledge Prof. Domingos Henrique (Instituto de Medicina Molecular, Lisbon) for providing the ES 46C cell line. This work was supported by FEDER funds through the Programa Operacional Factores de Competitividade – COMPETE (FCOMP‐01–0124‐FEDER‐021125) and by National funds FCT – Fundação para a Ciência e a Tecnologia (PTDC/SAU‐BMA/118869/2010). A.R.B. and M.J. Oliveira are supported by FCT (SFRH/BD/86200/2012; Investigator FCT)

Rotary orbital suspension culture of embryonic stem cell-derived neural stem/progenitor cells: impact of hydrodynamic culture on aggregate yield, morphology and cell phenotype

Embryonic stem (ES)-derived neural stem/progenitor cells (ES-NSPCs) constitute a promising cell source for application in cell therapies for the treatment of central nervous system disorders. In this study, a rotary orbital hydrodynamic culture system was applied to single-cell suspensions of ES-NSPCs, to obtain homogeneously-sized ES-NSPC cellular aggregates (neurospheres). Hydrodynamic culture allowed the formation of ES-NSPC neurospheres with a narrower size distribution than statically cultured neurospheres, increasing orbital speeds leading to smaller-sized neurospheres and higher neurosphere yield. Neurospheres formed under hydrodynamic conditions (72 h at 55 rpm) showed higher cell compaction and comparable percentages of viable, dead, apoptotic and proliferative cells. Further characterization of cellular aggregates provided new insights into the effect of hydrodynamic shear on ES-NSPC behaviour. Rotary neurospheres exhibited reduced protein levels of N-cadherin and ß-catenin, and higher deposition of laminin (without impacting fibronectin deposition), matrix metalloproteinase-2 (MMP-2) activity and percentage of neuronal cells. In line with the increased MMP-2 activity levels found, hydrodynamically-cultured neurospheres showed higher outward migration on laminin. Moreover, when cultured in a 3D fibrin hydrogel, rotary neurospheres generated an increased percentage of neuronal cells. In conclusion, the application of a constant orbital speed to single-cell suspensions of ES-NSPCs, besides allowing the formation of homogeneously-sized neurospheres, promoted ES-NSPC differentiation and outward migration, possibly by influencing the expression of cell–cell adhesion molecules and the secretion of proteases/extracellular matrix proteins. These findings are important when establishing the culture conditions needed to obtain uniformly-sized ES-NSPC aggregates, either for use in regenerative therapies or in in vitro platforms for biomaterial development or pharmacological screening.The authors would like to acknowledge Professor Domingos Henrique (Instituto de Medicina Molecular, Lisbon) for providing the ES 46C cell line. This study was supported by FEDER funds through the Programa Operacional Factores de Competitividade – COMPETE (Grant No. FCOMP‐01‐0124‐FEDER‐021125) and by National Funds through FCT – Fundação para a Ciência e a Tecnologia (Grant No. PTDC/SAU‐BMA/118869/2010). I. F. Amaral is supported by QREN through programme ON.2 (Grant No. NORTE‐07‐0124‐FEDER‐000005) and M. J. Oliveira is an Investigator FCT Fellow

Usefulness of ACTH stimulation test in the differential diagnosis of precocious pubarche

Introdução Nos doentes com pubarca precoce, o gold‐standard para o diagnóstico diferencial entre pubarca precoce idiopática (PPI) e a forma não clássica da hiperplasia congénita da suprarrenal (HCSR‐NC) é a prova de Synacthen. Esta permite também estimar a reserva adrenal de cortisol nos doentes com HCSR‐NC. Objetivos Comparar as características clínicas e perfil hormonal basal dos doentes com pubarca precoce; avaliar a importância da prova de Synacthen no diagnóstico diferencial entre PPI e HCSR‐NC e na determinação da reserva adrenal de cortisol. Material e métodos Estudo transversal de doentes com pubarca precoce que realizaram prova de Synacthen. ResultadosForam incluídos 43 doentes, com idade mediana de 7,5 anos (3,5‐9,4), sendo 37 (86,0%) do sexo feminino. Na prova de Synacthen, 37 (86,0%) foram classificados como PPI e 6 (14,0%) como HCSR‐NC. Não houve diferenças significativas entre os 2 grupos quanto às características clínicas e doseamentos basais de ACTH, cortisol e androgénios da suprarrenal. A 17‐OHP basal e estimulada foi mais elevada nos doentes com HCSR‐NC (p = 0,001 e p < 0,001, respetivamente) (basal: 4,62 ± 3,70 ng/ml [0,80‐10,50]; estimulada: 35,41 ± 24,87 ng/ml [12,0‐80,2]) do que nos doentes com PPI (basal: 1,04 ± 0,77 ng/ml [0,22‐3,80]; estimulada: 4,18 ± 1,71 ng/ml [1,0‐8,96]). O cut‐off basal habitualmente proposto (< 2,0 ng/ml) para a distinção entre estes grupos não o permitiu em 2 doentes, que apenas foram diagnosticados após realização da prova de Synacthen. Dois doentes com HCSR‐NC (33,3%) tiveram cortisol após estimulação < 18 μg/dl, revelando necessidade de tratamento com glucocorticoide em stress. Os doentes com HCSR‐NC com valores mais elevados de 17‐OHP basal tiveram valores de cortisol mais baixos após estimulação (p = 0,004; r = ‐0,43). Conclusão A realização desta prova foi útil para distinguir os doentes com HCSR‐NC e PPI, pois nenhum valor de 17‐OHP basal permitia fazer o diagnóstico diferencial definitivo. Em alguns doentes com HCSR‐NC a prova revelou secreção inapropriada de cortisol em stress, contribuindo para a decisão terapêutica

Inhibition of fucosylation in human invasive ductal carcinoma reduces E-selectin ligand expression, cell proliferation, and ERK1/2 and p38 MAPK activation

Breast cancer tissue overexpresses fucosylated glycans, such as sialyl-Lewis X/A (sLeX/A), and a-1,3/4-fucosyltransferases (FUTs) in relation to increased disease progression and metastasis. These glycans in tumor circulating cells mediate binding to vascular E-selectin, initiating tumor extravasation. However, their role in breast carcinogenesis is still unknown. Here, we aimed to define the contribution of the fucosylated structures, including sLeX/A, to cell adhesion, cell signaling, and cell proliferation in invasive ductal carcinomas (IDC), the most frequent type of breast cancer. We first analyzed expression of E-selectin ligands in IDC tissue and established primary cell cultures from the tissue. We observed strong reactivity with E-selectin and anti-sLeX/A antibodies in both IDC tissue and cell lines, and expression of a-1,3/4 FUTs FUT4, FUT5, FUT6, FUT10, and FUT11. To further assess the role of fucosylation in IDC biology, we immortalized a primary IDC cell line with human telomerase reverse transcriptase to create the ‘CF1_T cell line’. Treatment with 2-fluorofucose (2-FF), a fucosylation inhibitor, completely abrogated its sLeX/A expression and dramatically reduced adherence of CF1_T cells to E-selectin under hemodynamic flow conditions. In addition, 2-FF-treated CF1_T cells showed a reduced migratory ability, as well as decreased cell proliferation rate. Notably, 2-FF treatment lowered the growth factor expression of CF1_T cells, prominently for FGF2, vascular endothelial growth factor, and transforming growth factor beta, and negatively affected activation of signal-regulating protein kinases 1 and 2 and p38 mitogen-activated protein kinase signaling pathways. These data indicate that fucosylation licenses several malignant features of IDC, such as cell adhesion, migration, proliferation, and growth factor expression, contributing to tumor progression.The authors acknowledge the financial support from the LPCC/Pfizer 2011 and Portuguese Foundation for Science and Technology (FCT)—SFRH/BD/100970/ 2014 (MAC), SFRH/BD/81860/2011 (MS), and the United States National Institutes of Health National Heart Blood Institute (NHLBI Grant HL107146, RS). We also thank Dr Nicole Okeley from Seattle Genetics for the valuable help and opinions

A Common Variant in the CDK8 Gene Is Associated with Sporadic Pituitary Adenomas in the Portuguese Population: a Case-Control Study

The majority of pituitary adenomas occur in a sporadic context, and in the absence of known genetic predisposition. Three common variants at the NEBL (rs2359536), PCDH15 (rs10763170) and CDK8 (rs17083838) loci were previously associated with sporadic pituitary adenomas in the Han Chinese population, but these findings have not yet been replicated in any other population. The aim of this case-control study was to assess if these variants are associated with susceptibility to sporadic pituitary adenomas in the Portuguese population. Genotype and allele frequencies were determined in 570 cases and in 546 controls. The CDK8 rs17083838 minor allele (A allele) was significantly associated with sporadic pituitary adenomas, under an additive (odds ratio (OR) 1.73, 95% confidence interval (CI) 1.19-2.50, p = 0.004) and dominant (OR 1.82, 95% CI 1.24-2.68, p = 0.002) inheritance model. The NEBL rs2359536 and PCDH15 rs10763170 variants were not associated with the overall risk for the disease, although a borderline significant association was observed between the PCDH15 rs10763170 minor allele (T allele) and somatotrophinomas (dominant model, OR 1.55, 95% CI 1.02-2.35, p = 0.035). These findings suggest that the CDK8 rs17083838 variant, and possibly the PCDH15 rs10763170 variant, may increase susceptibility to sporadic pituitary adenomas in the Portuguese population.info:eu-repo/semantics/publishedVersio

Sarcoidose multissistémica: quando somos nós os primeiros a suspeitar!

A Sarcoidose é uma doença inflamatória crónica, granulomatosa, de envolvimento multissistémico e de etiologia desconhecida, que apresenta um espectro alargado de manifestações clínicas. Entre as mais frequentes, destaca-se o envolvimento pulmonar, cutâneo e oftalmológico. Através de um relato de um caso clínico pretende-se discutir as principais manifestações oftalmológicas, o pleomorfismo desta doença e a importância de um diagnóstico precoce. MATERIAL E MÉTODOS: Descrição e discussão de um caso clínico. Doente do sexo feminino, 58 anos, observada no S.U. de Oftalmologia por diminuição de acuidade visual (AV) no olho direito (OD) com cerca de uma semana de evolução. A AV era de 5/10 no OD e de 6/10 no olho esquerdo, apresentando bilateralmente reacção de câmara anterior com precipitados queráticos endoteliais, vitrite e, na fundoscopia, tortuosidade e engurgitamento venosos com periflebite e lesões em “pingos de cera”. Apresentava ainda edema da papila no OD. Concomitantemente referia no último mês aparecimento de nódulos cutâneos nos membros inferiores (MI), queixas álgicas articulares, tosse seca e episódios de febre intermitente acompanhados por suores nocturnos. Encaminhou-se a doente para observação pela Medicina Interna. Foi pedida avaliação laboratorial e imagiológica, bem como biopsia de lesão nodular dos MI. Foram ainda realizados Retinografia, Angiografia Fluouresceínica e Tomografia de Coerência Óptica. RESULTADOS: A avaliação clínica, laboratorial e imagiológica desta doente revelou aspectos sugestivos de Sarcoidose com afecção multissistémica, incluíndo uma panuveíte bilateral com compromisso visual. CONCLUSÃO: A Sarcoidose é uma patologia multiorgânica com frequente envolvimento ocular, sendo que as manifestações oftalmológicas podem constituir a forma de apresentação da doença ou fornecer pistas úteis para o diagnóstico. O diagnóstico precoce é essencial para a correcta orientação do doente e para um tratamento adequado e atempado, minimizando potenciais complicações graves, oculares e sistémicas, que podem advir desta patologia. REFERÊNCIAS BIBLIOGRÁFICAS: Carl P. Herbort et al, “International Criteria for the Diagnosis of Ocular Sarcoidosis: Results of the First International Workshop on Ocular sarcoidosis”, Ocular Immunology and Inflammation, 17, 160-169, 2009 Hilario Nunes et al, Sarcoidosis- Review , Orphanet Journal of Rare diseases 2007, 2:46 Costa F., Arrobas A., Sarcoidose Extratorácica, Artigo de Revisão, Revista Portuguesa de Pneumologia, Vol XIV nº1, Jan/Fev 200