Masters Degrees (Haematology and Cell Biology)
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Browsing Masters Degrees (Haematology and Cell Biology) by Author "Coetzee, M. J."
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Item Open Access Microparticles derived from stimulation of human umbilical endothelium(University of the Free State, 2013) Le Roux, Elzette; Meiring, S. M.; Coetzee, M. J.English: Endothelial microparticle reseach is currently a very novel and exciting topic in the field of haemosistasis and thrombosis. The role of microparticles in inflammatory and thrombotic disorders is however not fully understood. Dysfunction of endothelial cells is hypothesized to be a trigger of microparticle formation. In inflammatory disorders like sepsis and thrombotic disorders like atherosclerosis and thrombotic thrombocytopenic purpura, endothelial microparticle formation is altered and the numbers thereof may increase or decrease. It is not known if microparticles are the cause or the consequence of these disorders. To understand the role of endothelial microparticles in inflammation and thrombosis, the effect of inflammatory cytokines and coagulation stimuli was studied as well as combinations thereof on endothelial microparticle formation and on microparticle VWF and its regulating protease, ADAMTS-13 in HUVEC. In this study, the formation of microparticles in cultured human umbilical vein endothelial cells (HUVEC) was stimulated by different inflammatory agents: IL-6 (100 ng/ml), IL-8 (100 ng/ml) and TNF-α (100 ng/ml), coagulation stimuli: TF (2 U/ml) and thrombin (2 U/ml) and combinations thereof. The number of endothelial microparticles that formed was determined using flow cytometry. VWF and ADAMTS-13 levels of the microparticles were assessed by ELISAs and microparticle thrombin generation was measured by thrombin generation assays. VWF multimers were visualized by a Western Blot technique. IL-6 did not have any effect on HUVEC-derived microparticles due to the lack of the receptor for IL-6 on these cells. IL-8 only slightly increased effect on microparticle VWF and ADAMTS-13 levels. TNF-α had a significant effect on microparticle numbers and contributed to almost 80% of thrombin generated by the microparticles. It has however almost no effect on VWF levels. The coagulation stimulus TF, on the other hand, induced the highest increase in microparticle VWF levels and increased microparticle numbers impressively. Yet, it had no effect on the thrombin generation by the microparticles. TF in combination with TNF-α also induced an increase in microparticle VWF and a small decrease in ADAMTS-13 levels. So, TF may contribute to the increased VWF levels that are commonly found in TTP patients where inflammation and thrombosis occur. Interestingly, thrombin had a protective effect on the intact HUVEC by preventing microparticle formation. The combination stimuli of thrombin and inflammatory agents also had a protective effect on HUVEC. This highlighted the regulatory role of thrombin in intact endothelial cells and also the protection that it provides against thrombosis in extremely inflammatory environments. Endothelial microparticles can therefore be detrimental or beneficial, depending on the different stimuli and different environments. Inflammatory and coagulation stimuli may still pose a significant risk of clotting by altering microparticle quantity and content. This study contributes to understand the role that endothelial microparticles play in inflammation and thrombosis.