Vpliv levkocitnih glikozaminoglikanov na kemotaktično gibanje nevtrofilcev in limfocitovMihalič, Zala Nikita (Avtor) Bratkovič, Tomaž (Mentor) Kungl, Andreas (Komentor) glycosaminoglycans neutrophils T-cells chemotaxisChemokines and glycosaminoglycans play an important role in inflammation. Chemokines are small proteins, released by the tissue macrophages. They bind to their receptors and to the glycosaminoglycans, which results in the migration of leukocytes from blood to the site of inflammation. Glycosaminoglycans are polysaccharide molecules attached to core proteins, forming proteoglycans. Most of the studies are focused on proteoglycans on the surface of endothelial cells, but only few on leukocyte proteoglycans. In our study, we abolished the activity of glycosaminoglycans on neutrophils and T-cells by their digestion with heparinase C and competitive inhibition with the addition of unspecific glycosaminoglycan-binding protein or heparan sulphate. We observed differences in migration of treated and untreated cells with a Boyden chamber assay. Additionally, we compared the migration of cells upon different washing steps following digestion with heparinase C. To assess the level of proteoglycans on leukocytes we determined the expression of several anchoring proteins on neutrophils using quantitative polymerase chain reaction. For a further investigation of the glycosaminoglycan influence on neutrophil chemotaxis, we also performed a transendothelial migration assay. The digestion of glycosaminoglycans on neutrophils resulted in a decrease of chemotaxis. The addition of glycosaminoglycan-binding protein showed an increase in migration and the addition of heparan sulphate a slight increase, although it was significant only towards the middle concentration (0.15 μM) of chemokine. The digestion of T-cell glycosaminoglycans resulted in an increase in the migration towards the highest concentration (1.5 μM) of chemokine in all three conditions. A small difference in migration of neutrophils was observed depending on the different washing steps (i.e., whether or not the heparinase C was washed away from the cells). T-cells did not present any difference in migration. We showed that there is a big difference in expression of proteoglycans on the different cell types. The transendothelial assay revealed that endothelial glycosaminoglycans are more important for chemotaxis than neutrophil glycosaminoglycans. We proved that digestion of neutrophil glycosaminoglycans have an impact on migration but still the endothelial glycosaminoglycans are more important. However, T-cells presented no difference so further studies should be done to determine the role of glycosaminoglycans on leukocytes.[Z. N. Mihalič]20182019-11-23 14:43:46Magistrsko delo/naloga112945UDK: 577:615(043.3)COBISS_ID: 4647025sl