Structural characterization and immunogenicity in wildtype and immune tolerant mice of degraded recombinant human interferon alpha2b

Purpose - To study the influence of protein structure on the immunogenicity in wildtype and immune tolerant mice of well-characterized degradation products of recombinant human interferon alpha2b (rhIFNα2b). Methods - RhIFNα2b was degraded by metal catalyzed oxidation (M), crosslinking with glutaraldehyde (G), oxidation with hydrogen peroxide (H) and incubation in a boiling water bath (B). The products were characterized with UV absorption, circular dichroism and fluorescence spectroscopy, gel permeation chromatography, reversed-phase HPLC, SDS-PAGE, Western blotting and mass spectrometry. The immunogenicity of the products was evaluated in wildtype mice and in transgenic mice immune tolerant for hIFNα2. Serum antibodies were detected by ELISA or surface plasmon resonance. Results - M-rhIFNα2b contained covalently aggregated rhIFNα2b with three methionines partly oxidized to methionine sulfoxides. G-rhIFNα2b contained covalent aggregates and did not show changes in secondary structure. H-rhIFNα2b was only chemically changed with four partly oxidized methionines. B-rhIFNα2b was largely unfolded and heavily aggregated. Native (N) rhIFNα2b was immunogenic in the wildtype mice but not in the transgenic mice, showing that the latter were immune tolerant for rhIFNα2b. The antirhIFNα2b antibody levels in the wildtype mice depended on the degradation product: M-rhIFNα2b > H-rhIFNα2b ~ N-rhIFNα2b >> B-rhIFNα2b; G-rhIFNα2b did not induce anti-rhIFNα2b antibodies. In the transgenic mice, only M-rhIFNα2b could break the immune tolerance. Conclusions - RhIFNα2b immunogenicity is related to its structural integrity. Moreover, the immunogenicity of aggregated rhIFNα2b depends on the structure and orientation of the constituent protein molecules and/or on the aggregate size.

Utilização de materiais anti-vibráteis e de isolamento acústico na via férrea

Trabalho Final de Mestrado para obtenção do grau de Mestre em Engenharia Civil na Área de Especialização de Vias de Comunicação e Transportes

Medidas anti-crise e a motivação : estudo no contexto de uma empresa multinacional de prestação de serviços

Mestrado em Gestão e Empreendedorismo

Anti-Corrosion performance of a new silane coating for corrosion protection of AZ31 Magnesium alloy in Hank’s solution

Mg alloys can be used as bioresorsable metallic implants. However, the high corrosion rate of magnesium alloys has limited their biomedical applications. Although Mg ions are essential to the human body, an excess may cause undesirable health effects. Therefore, surface treatments are required to enhance the corrosion resistance of magnesium parts, decreasing its rate to biocompatible levels and allowing its safe application as bioresorbable metallic implants. The application of biocompatible silane coatings is envisaged as a suitable strategy for retarding the corrosion process of magnesium alloys. In the current work, a new glycidoxypropyltrimethoxysilane (GPTMS) based coating was tested on AZ31 magnesium substrates subjected to different surface conditioning procedures before coating deposition. The surface conditioning included a short etching with hydrofluoric acid (HF) or a dc polarisation in alkaline electrolyte. The silane coated samples were immersed in Hank's solution and the protective performance of the coating was studied through electrochemical impedance spectroscopy (EIS). The EIS data was treated by new equivalent circuit models and the results revealed that the surface conditioning process plays a key role in the effectiveness of the silane coating. The HF treated samples led to the highest impedance values and delayed the coating degradation, compared to the mechanically polished samples or to those submitted to dc polarisation.

Da concorrência fiscal prejudicial ao uso e abuso das doutrinas judiciais anti-abuso : uma análise do caso português

Mestrado em Fiscalidade