| dc.description.abstract | Glycan-based interactions have been the subject of an increasing number of studies in recent years, as their importance in biological systems is becoming more understood. As glycans are present on the majority of cell surfaces, they are involved in a wide range of biological interactions from cell-cell to hostpathogen interactions. Developments in sensitive force-probes have led to an increased interest in elucidating the mechanisms involved in these weak molecular interactions and their roles in biological systems. This thesis utilizes the atomic force-microscope to study glycan-based interactions in two different biological systems.
In this thesis, Atlantic salmon (Salmo salar) yolk sac fry are used as a model system to study mucosal surfaces. AFM experiments are used to study the adhesive and mechanical properties of the skin mucosa of germ-free and conventionally reared salmon fry. Bacterial adhesion of the pathogens Aeromonas salmonicida and Yersinia ruckeri to mucins and the mucosal surface were studied using live AFM-probes, while nanoindentation was used to elucidate the mechanical properties of the skin mucosa. The main glycan type present on mucins is sialic acids. Sialic acids (Sias) are often the terminal unit on glycans, and as a result they are often involved in interactions. In this thesis, BJAB lymphoma cells are used as a model system to study the effect of sialic acid expression on cell surface properties. The cells are modified with the Sias N-acetylneuraminic acid (Neu5Ac) or N-glycolylneuraminic acid (Neu5Gc) and are probed with a hydrophobically modified AFM-tip to study how the Sia-expression influences cell surface properties.
A method for studying intact salmon fry under the AFM was established. Adhesion studies revealed adhesion between pathogens and mucins isolated from salmon, with a preference for mucins isolated from the gut. Removal of sialic acid increased adhesion of A. salmonicida, particularly to gut mucins. Both A. salmonicida and Y. ruckeri showed no adhesion to the intact salmon mucosa. Due to low phase angle measurements, we hypothesize that mucins have low internal adhesion and are released to wash away the pathogens. Nanoindentation experiments reveal increasing stiffness with age and that the germ-free salmon have a softer surface in early development, indicating that the microbiota contribute to the barrier. Results from the studies on serum protein and BJAB cells show a higher frequency of hydrophobic interactions on proteins and cells expressing Neu5Ac, consistent with the hypothesis based on the sialic acid structure. The result from both systems highlights the importance of sialic acids in influencing the cell properties and demonstrates the usefulness of AFM for studying diverse biological systems and elucidating the roles of glycans in biological interactions. | en_US |
