The inhibitory effect of an RGD-human chitin-binding domain fusion protein on the adhesion of fibroblasts to reacetylated chitosan films

Biomaterials used for tissue engineering applications must provide a structural support for the tissue development and also actively interact with cells, promoting adhesion, proliferation, and differentiation. To achieve this goal, adhesion molecules may be used, such as the tripeptide Arg-Gly-Asp (RGD). A method based on the use of a carbohydrate-binding module, with affinity for chitin, was tested as an alternative approach to the chemical grafting of bioactive peptides. This approach would simultaneously allow the production of recombinant peptides (alternatively to peptide synthesis) and provide a simple way for the specific and strong adsorption of the peptides to the biomaterial. A fusion recombinant protein, containing the RGD sequence fused to a human chitin-binding module (ChBM), was expressed in E. coli. The adhesion of fibroblasts to reacetylated chitosan (RC) films was the model system selected to analyze the properties of the obtained proteins. Thus, the evaluation of cell attachment and proliferation on polystyrene surfaces and reacetylated chitosan films, coated with the recombinant proteins, was performed using mouse embryo fibroblasts 3T3. The results show that the recombinant proteins affect negatively fibroblasts anchorage to the materials surface, inhibiting its adhesion and proliferation. We also conclude that this negative effect is fundamentally due to the human chitin-binding domain.Fundação para a Ciência e a Tecnologia (FCT) - SFRH/BD/27359/2006, POCTI/BIO/45356/200

Grazing, disturbance and plant soil interactions in northern grasslands

Abstract Plants and soil organisms are closely linked. Plants are the sole source of carbon in the soil and soil organisms are responsible for recycling of nutrients, making them available for plant growth. To understand the function of a system, it is important to understand the interactions between the soil and plants. These interactions have mainly been studied in temperate areas, with few studies in the arctic and subarctic. The aim of this thesis was to investigate the effect of ecological disturbances in sub- and low-arctic grasslands on soil organisms and plant-soil feedback relationships. The effect of removal of vegetation, replanting of a local plant species, and different components of grazing (trampling, defoliation and return of nutrients) on soil decomposer organisms were studied. Whether short term effects of defoliation depended on plant species community was also studied, as well as whether defoliation in the field could create changes in the soil system systems that affect the growth of seedlings. Experiments were conducted under both controlled greenhouse conditions and in field sites. The results showed that physical disturbance (removal of vegetation and trampling) reduced the abundance and diversity of soil biota. Defoliation increased soil decomposer abundance in the short term. Plant species composition did not affect soil biota and only in a few cases did it changes their responses to defoliation. In the long-term, effects of fertilization and defoliation on the soil biota were context-dependent. However, defoliation did create changes in the soil that reduced the growth of seedlings planted into the soil. Furthermore, plant species community and spatial heterogeneity (revealed by blocking) had important effects on the soil communities