Evaluation of a novel Escherichia coli fusion system for overproduction of recombinant immunogenic proteins

Recombinant protein production has been widely applied for therapeutic and diagnostic applications, namely for polyclonal antibody production. Antibodies are usually raised against a specific protein by immunization of animals with the purified protein. The bacterium Escherichia coli is one of the most used host cells for the bio-production of proteins, but it still presents some drawbacks: many proteins of biomedical interest are difficult to express properly in this host system, resulting in insoluble protein aggregates. Gene fusion technology has been employed to optimize recombinant protein production in E. coli. Fusion partners have also been used to potentially increase protein immunogenicity. In this work, the overproduction and immunopotentiating properties of a novel fusion system were studied. Novel fusion tags, Fh8 and H, were fused to five target proteins with diagnostic interests: CP12, a 12 kDa surface protein from Cryptosporidium parvum oocysts; CWP, a cyst wall protein from Giardia lamblia; ENT, a surface protein from Entamoeba histolytica cysts; TgOWP, a Toxoplasma gondii oocyst wall protein; and Frutalin, a recombinant lectin from Artocarpus incisa seeds. Production yields of all Fh8-fused proteins, H-fused proteins and non-fused recombinant proteins were compared and polyclonal antibodies were raised against CP12, CWP and ENT nonfused and H-fused antigens. Overall, the results showed that the fusion of both Fh8 and H tags to all target proteins improved their production in comparison with the respective non-fused target proteins. Moreover, the H tag efficiently increased CP12, CWP and ENT specific immunogenicity without being removed from the fusion antigens and without coadministration of adjuvants, resulting in a more effective and earlier immune response. The overproduction and immunopotentiating effects observed for this novel fusion system make it a unique alternative for recombinant protein production in E. coli and for immunodiagnostic and immunoprophylactic purposes

Atributos químicos dos solos sob diferentes usos e manejos em uma sub-bacia no noroeste do Estado de São Paulo

Changes resulting from the soils use and cropping affect the structure and biological activity, then interfeing on the chemical attributes. This research had the objective of evaluating the soil chemical attributes of sub-river basin Jardim Novo Horizonte, in order to verify the effect of different soil uses and management. Soils in the sub-river basin are Red Oxisol and Red-Yellow Alfisol. Six and four spots were evaluated in the first and second soils, respectively, totaling 10 spots along the sub-river basin with five replications each. The following systems were sampled from the Oxisol: area cropped with mango, four areas under pasture and area with annual crop. Two areas under pasture, one with annual crop and another with forest fragment area were sampled in the Alfisol. The Alfisol showed better chemical attributes compared with Oxisol, higher content of organic matter, providing better support capacity and, consequently, greater the soil vegetal cover. Annual crops area had better conditions soils fertility for both, and the pastures worst, due to inadequate management. The chemical degradation of the Oxisol by the use and management, mainly in the areas cultivated with pasture, causes more susceptibility of soil erosion for the sub-river basin studied.As alterações decorrentes do uso e ocupação dos solos afetam a sua estrutura e atividade biológica, afetando assim, seus atributos químicos. A presente pesquisa teve por objetivo avaliar os atributos químicos dos solos da sub-bacia Jardim Novo Horizonte, com o intuito de verificar o efeito dos diferentes usos e manejos do solo. Os solos da sub-bacia são o Latossolo Vermelho e o Argissolo Vermelho-Amarelo. As coletas de solos foram realizadas em 10 locais ao longo da sub-bacia, sendo seis localizados no Latossolo e quatro no Argissolo, e em cada local cinco repetições. Os usos foram: cultura da manga, quatro locais com pastagem e cultura anual, no Latossolo, e dois locais com pastagem, cultura anual e área com fragmento de mata, no Argissolo. O Argissolo apresentou melhores atributos químicos em relação ao Latossolo, maiores conteúdos de matéria orgânica, propiciando melhor capacidade de suporte e, consequentemente, maior cobertura vegetal ao solo. As áreas com culturas anuais apresentaram melhores condições de fertilidade para os dois solos, e as pastagens as piores, devido ao manejo inadequado das mesmas. A degradação química do Latossolo pelo uso e manejo, sobretudo nas áreas cultivadas com pastagens, causa maior suscetibilidade à erosão dos solos da sub-bacia estudada

Effect of hypertonic saline treatment on the inflammatory response after hydrochloric acid-induced lung injury in pigs

OBJECTIVES: Hypertonic saline has been proposed to modulate the inflammatory cascade in certain experimental conditions, including pulmonary inflammation caused by inhaled gastric contents. The present study aimed to assess the potential anti-inflammatory effects of administering a single intravenous dose of 7.5% hypertonic saline in an experimental model of acute lung injury induced by hydrochloric acid. METHODS: Thirty-two pigs were anesthetized and randomly allocated into the following four groups: Sham, which received anesthesia and were observed; HS, which received intravenous 7.5% hypertonic saline solution (4 ml/kg); acute lung injury, which were subjected to acute lung injury with intratracheal hydrochloric acid; and acute lung injury + hypertonic saline, which were subjected to acute lung injury with hydrochloric acid and treated with hypertonic saline. Hemodynamic and ventilatory parameters were recorded over four hours. Subsequently, bronchoalveolar lavage samples were collected at the end of the observation period to measure cytokine levels using an oxidative burst analysis, and lung tissue was collected for a histological analysis. RESULTS: Hydrochloric acid instillation caused marked changes in respiratory mechanics as well as blood gas and lung parenchyma parameters. Despite the absence of a significant difference between the acute lung injury and acute lung injury + hypertonic saline groups, the acute lung injury animals presented higher neutrophil and tumor necrosis factor alpha (TNF-α), interleukin (IL)-6 and IL-8 levels in the bronchoalveolar lavage analysis. The histopathological analysis revealed pulmonary edema, congestion and alveolar collapse in both groups; however, the differences between groups were not significant. Despite the lower cytokine and neutrophil levels observed in the acute lung injury + hypertonic saline group, significant differences were not observed among the treated and non-treated groups. CONCLUSIONS: Hypertonic saline infusion after intratracheal hydrochloric acid instillation does not have an effect on inflammatory biomarkers or respiratory gas exchange

Production and characterization of recombinant frutalin expressed in the yeast Pichia Pastoris

CNPq, CAPES, FUNCAP, FCT, ALBAN, UMINHO, UFC, UNIFO