Protection of cattle against a natural infection of Fasciola hepatica by vaccination with recombinant cathepsin L1 (rFhCL1)

The liver fluke, Fasciola hepatica causes liver fluke disease, or fasciolosis, in ruminants such as cattle and sheep. An effective vaccine against the helminth parasite is essential to reduce our reliance on anthelmintics, particularly in light of frequent reports of resistance to some frontline drugs. In our study, Friesian cattle (13 per group) were vaccinated with recombinant F. hepatica cathepsin L1 protease (rFhCL1) formulated in mineral-oil based adjuvants, Montanide (TM) ISA 70VG and ISA 206VG. Following vaccination the animals were exposed to fluke-contaminated pastures for 13 weeks. At slaughter, there was a significant reduction in fluke burden of 48.2% in the cattle in both vaccinated groups, relative to the control non-vaccinated group, at p

Fluorobenzoyl dipeptidyl derivatives as inhibitors of the Fasciola hepatica cysteine protease cathepsin L1

Cathepsins are known to have many important physiological roles and provide a viable target for inhibition. Fluorobenzoyl dipeptide derivatives were synthesized and tested for biological activity in an effort to find an efficient inhibitor of the cysteine protease cathepsin L. Thirty-six novel inhibitors (1-36) were synthesized from protected amino acids via the standard DCC/HOBt coupling protocol, containing a benzyl ester or a nitrile as an electrophilic warhead. The activity of the inhibitors was evaluated against cathepsin L and IC50 values calculated. Modification of both amino acids and terminal groups afforded compounds with single digit micromolar inhibition. Results utilizing the benzoyl-L-leucine-glycine nitrile backbone are comparable to that for the commercially available inhibitor 39.

The importance of pH in regulating the function of the Fasciola hepatica cathepsin L1 cysteine protease

The helminth parasite Fasciola hepatica secretes cathepsin L cysteine proteases to invade its host, migrate through tissues and digest haemoglobin, its main source of amino acids. Here we investigated the importance of pH in regulating the activity and functions of the major cathepsin L protease FheCL1. The slightly acidic pH of the parasite gut facilitates the auto-catalytic activation of FheCL1 from its inactive proFheCL1 zymogen; this process was approximately 40-fold faster at pH 4.5 than at pH 7.0. Active mature FheCL1 is very stable at acidic and neutral conditions (the enzyme retained approximately 45% activity when incubated at 37 degrees C and pH 4.5 for 10 days) and displayed a broad pH range for activity peptide substrates and the protein ovalbumin, peaking between pH 5.5 and pH 7.0. This pH profile likely reflects the need for FheCL1 to function both in the parasite gut and in the host tissues. FheCL1, however, could not cleave its natural substrate Hb in the pH range pH 5.5 and pH 7.0; digestion occurred only at pH

Micro-environmental conditions modulate protein secretion and infectivity of the Trichinella spiralis L1 larva

After digestion of infected meat the free L1 of Trichinella spp. penetrate the intestinal mucosa where they moult to the mature adult stage. We have used proteomics to identify changes in protein secretion during in vitro culture of free T. spiralis muscle larvae under different environmental conditions, and to correlate these changes with their infectivity in mice. Muscle larvae were cultured in different media (RPMI-1640, C-199 and HBSS) under conditions of anaerobiosis, microaerobiosis and in 5% CO(2) at 37 degrees C. Following incubation the larval excretory/secretory proteins were analysed by two-dimensional gel electrophoresis and the larvae were used to orally infect naïve CD1 mice. For all culture media tested, infectivity of the L1 was preserved following incubation in anaerobic conditions. In contrast, the infectivity of worms cultured in nutrient-rich media was almost completely abolished in both microaerobiosis and in the presence of 5% CO(2). Some infectivity was retained in poor or reduced culture media. Comparative analysis of larval infectivity and protein secretion showed that loss of infectivity correlated with the appearance of non-tyvelosylated proteins that in turn may be related to the onset of moulting.

Proteomic analysis of the excretory-secretory proteins of the Trichinella spiralis L1 larva, a nematode parasite of skeletal muscle

Trichinella spiralis is an intracellular nematode parasite of mammalian skeletal muscle. Infection of the muscle cell leads to the formation of a host-parasite complex that results in profound alterations to the host cell and a re-alignment of muscle-specific gene expression. The role of parasite excretory-secretory (ES) proteins in mediating these effects is currently unknown, largely due to the difficulty in identifying and assigning function to individual proteins. In this study, a global proteomics approach was used to analyse the ES proteins from T. spiralis muscle larvae. Following 2-DE of ES proteins,MALDI-TOF-MS and LC-MS/MS were used to identify the peptide spots. Specific Trichinella EST databases were assembled and used to analyse the data. Despite the current absence of a Trichinella genome-sequencing project, 43 out of 52 protein spots analysed were identified and included the major secreted glycoproteins. Other novel proteins were identified from matches with sequences in the T. spiralis database. Our results demonstrate the value of proteomics as a tool for the identification of Trichinella ES proteins and in the study of the molecular mechanism underpinning the formation of the host-parasite complex during Trichinella infections.

The prognostic significance of PD-L1 in bladder cancer

Immunotherapy is a promising strategy for the treatment of various types of cancer. An antibody that targets programmed death ligand-1 (PD-L1) pathway has been shown to be active towards various types of cancer, including melanoma and lung cancer. MPDL3280A, an anti‑PD-L1 antibody, has shown clear clinical activity in PD-L1-overexpressing bladder cancer with an objective response rate of 40-50%, resulting in a breakthrough therapy designation granted by FDA. These events pronounce the importance of targeting the PD-L1 pathway in the treatment of bladder cancer. In the present study, we investigated the prognostic significance of the expression of three genes in the PD-L1 pathway, including PD-L1, B7.1 and PD-1, in three independent bladder cancer datasets in the Gene Expression Omnibus database. PD-L1, B7.1 and PD-1 were significantly associated with clinicopathological parameters indicative of a more aggressive phenotype of bladder cancer, such as a more advanced stage and a higher tumor grade. In addition, a high level expression of PD-L1 was associated with reduced patient survival. Of note, the combination of PD-L1 and B7.1 expression, but not other combinations of the three genes, were also able to predict patient survival. Our findings support the development of anti-PD-L1, which blocks PD-L1-PD-1 and B7.1-PD-L1 interactions, in treatment of bladder cancer. The observations were consistent in the three independent bladder cancer datasets consisting of a total of 695 human bladder specimens. The datasets were then assessed and it was found that the expression levels of the chemokine CC-motif ligand (CCL), CCL3, CCL8 and CCL18, were correlated with the PD-L1 expression level, while ADAMTS13 was differentially expressed in patients with a different survival status (alive or deceased). Additional investigations are required to elucidate the role of these genes in the PD-L1-mediated immune system suppression and bladder cancer progression. In conclusion, findings of this study suggested that PD-L1 is an important prognostic marker and a therapeutic target for bladder cancer.

Immune-derived PD-L1 gene expression defines a subgroup of stage II/III colorectal cancer patients with favorable prognosis that may be harmed by adjuvant chemotherapy

A recent phase 2 study of metastatic colorectal carcinoma (CRC) patients showed that mismatch repair gene status was predictive of clinical response to PD-1-targeting immune checkpoint blockade. Further examination revealed strong correlation between PD-L1 protein expression and microsatellite instability (MSI) in stage IV CRC, suggesting that the amount of PD-L1 protein expression could identify late stage patients who may benefit from immunotherapy. To assess whether the clinical associations between PD-L1 gene expression and MSI identified in metastatic CRC are also present in stage II/III CRC, we used in silico analysis to elucidate the cell types expressing the PD-L1 gene. We found a significant association of PD-L1 gene expression with MSI in early stage CRC (P < 0.001) and show that unlike in non-CRC tumors, PD-L1 is derived predominantly from the immune infiltrate. We demonstrate that PD-L1 gene expression has positive prognostic value in the adjuvant disease setting (PD-L1low v PD-L1high HR = 9.09; CI, 2.11-39.10). PD-L1 gene expression had predictive value, as patients with high PD-L1 expression appear to be harmed by standard-of-care treatment (HR = 4.95; CI,1.10-22.35). Building on the promising results from the metastatic CRC PD-1-targeting trial, we provide compelling evidence that PD-L1high/MSI/immunehigh stage II/III CRC patients should not receive standard chemotherapy. This conclusion supports the rationale to clinically evaluate this patient subgroup for PD-1 blockade treatment.

Competências das entidades inspectoras ou organismos de inspecção de acordo com a norma NP EN 45 004

O regulamento técnico relativo ao projecto, construção, exploração e manutenção de redes de gás combustível canalizado tem sofrido algumas alterações ao longo dos últimos treze anos. Este facto está directamente relacionado com o projecto de introdução do gás natural (GN) no território nacional, considerado como uma das prioridades na política energética dos últimos governos. A área geográfica de concessão de serviço público de importação de gás natural e do seu transporte e fornecimento através da rede de alta pressão encontra-se actualmente alargada a todos os municípios do continente, por força da Portaria n.º 2/2001, de 2 de Janeiro.

La percezione della norma linguistica attraverso il periodico La Crusca per voi

Riassunto: Dedicato «alle scuole e agli amatori della lingua», ai lettori interessati e preoccupati delle sorti della lingua nazionale, il periodico semestrale La Crusca per voi esce per la prima volta nell’ottobre 1990 con una Giustificazione e Un po’ di storia, in cui il fondatore Giovanni Nencioni spiega le ragioni dell’istituzione di un vero e proprio «consultorio linguistico», destinato ad accogliere le richieste, i dubbi, le perplessità sul corretto impiego della lingua italiana proveniente da persone di ogni livello sociale e culturale. Scopo di questo lavoro è studiare la tipologia dei quesiti posti agli accademici della Crusca, con particolare attenzione a fenomeni di carattere lessicale, negli ultimi diciotto numeri del periodico (dal 2005 al 2013, con qualche prelievo da uscite precedenti). Conclude il lavoro una breve indagine a campione sulla diffusione e la traducibilità di alcuni termini stranieri in italiano.

Información de moluscos y mariscos : Norma 61-25: Establecimientos de ventas de alimentos

This Spanish language sheet explains SC regulation 61-25 dealing with retail food establishments and shellfish. It also has a daily shellstock receiving record that can be used by restaurants for record keeping.

A implementação da norma NP EN ISO 50001:2012 em instituições da administração pública portuguesa: caso de estudo

Tese de mestrado integrado em Engenharia da Energia e do Ambiente, apresentada à Universidade de Lisboa, através da Faculdade de Ciências, 2015

Contributos para a revisão da Norma ISO 9001:2008 na perspetiva de uma aproximação à realidade organizacional

Este trabalho de investigação pretende dar o seu contributo para a renovação da norma ISO 9001:2008, na perspetiva de uma maior aproximação à realidade organizacional, através da análise da perceção dos especialistas/profissionais da qualidade. Numa primeira fase foi realizada uma abordagem ao estado da arte, relativamente ao conceito qualidade, à organização International Organization for Standardization (ISO), ao processo de alteração da norma, ao referencial normativo ISO 9001:2008, ao comportamento ao nível mundial da certificação segundo a norma ISO 9001:2008 e finalmente ao processo de preparação da revisão da norma ISO 9001:2008 a ser divulgada em 2015, cuja publicação na versão final, está prevista para novembro de 2015. Com a finalidade de apurar as opiniões/sugestões dos especialistas da área realizou-se um questionário que foi colocado online nas principais redes sociais e também enviado para alguns auditores, consultores, gestores/técnicos da qualidade e gestores de topo. Participaram 60 profissionais. Após da recolha dos dados, estes foram tratados através da utilização do software statistical package for the social sciences (SPSS). Os resultados obtidos demonstraram que a perceção dos profissionais vai de encontro às alterações que já estão propostas para a nova norma ISO 9001:2015. Das alterações sugeridas é de realçar, uma maior adequação da norma às diversas áreas de negócio, maior exigência nos aspetos ligados às organizações, adequação de conceitos e objetivos, compatibilidade com outros referenciais, complementar áreas como segurança, finanças e/ou responsabilidade social e maior envolvimento das organizações no processo de renovação da norma. No que concerne às expetativas, os profissionais esperam que a nova norma ISO 9001:2015 venha facilitar a integração e compreensão dos vários sistemas, maior adequação e credibilização junto das organizações e uma melhoria generalizada. Desta forma com o presente trabalho, espera-se ter contribuído para a clarificação das dificuldades sentidas na adequação do atual referencial à realidade organizacional. Assim como dar um contributo para futura apresentação aos responsáveis pelo acompanhamento da atividade normativa em Portugal (comité técnico CT80).

Norma Contabilística e de Relato financeiro 5 divulgações de partes relacionadas

Dissertação de Mestrado apresentada ao Instituto Superior de Contabilidade e Administração do Porto para a obtenção do Grau de Mestre em Auditoria.

O Impacto da implementação da norma contabilística relato financeiro 19: contratos de construção

Dissertação de Mestrado submetida ao Instituto Superior de Contabilidade e Administração do Porto para a obtenção do grau de Mestre em Auditoria. Trabalho efectuado sob a orientação do Professor Doutor Eduardo Sá e Silva