Show simple item record

dc.contributor.advisorMeiring, S. M.
dc.contributor.authorLe Roux, Elzette
dc.contributor.otherCoetzee, M. J.
dc.date.accessioned2015-09-08T09:38:46Z
dc.date.available2015-09-08T09:38:46Z
dc.date.issued2013
dc.identifier.urihttp://hdl.handle.net/11660/1193
dc.description.abstractEnglish: 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.en_ZA
dc.description.abstractAfrikaans: Endoteel-mikropartikelnavorsing is tans ’n baie nuwe en belowende onderwerp in hemostase en trombose. Die rol van mikropartikels in inflammatoriese en trombotiese toestande is nog onbekend. Wanfunksie van endoteelselle word as een van die oorsake van mikropartikelvorming bestempel. In inflammatoriese en trombotiese siektetoestande, soos sepsis, arterosklerose en trombotiese trombositopeniese purpura, vind daar verskeie veranderings in die vorming van mikropartikels plaas. Dit is nog onbekend of mikropartikels die oorsaak of die gevolg van hierdie siektetoestande is. Om ’n beter begrip van die rol van endoteelmikropartikels te verkry, het ons die uitwerking van inflammatoriese sitokine en stollingsfaktore asook kombinasies van die twee groepe op endoteel-mikropartikelvorming in HUVEC ondersoek. Ons het ook die effek daarvan op die uitdrukking van von Willebrand faktor (VWF) en sy regulerende protease, ADAMTS-13, bestudeer. Ons het in hierdie studie die vorming van mikropartikels gestimuleer deur menslike naelstringendoteelselle (HUVEC) met inflammatoriese agente, IL-6 (100 ng/ml), IL-8 (100 ng/ml) en TNF-α (100 ng/ml), asook stollingsfaktore, TF (2 U/ml) en trombien (2 U/ml) en kombinasies daarvan, te behandel. Die doel van die studie was nie alleen om die hoeveelheid HUVEC-mikropartikels wat vorm te bepaal nie, maar ook om die ADAMTS-13 en VWF-vlakke, VWF-multimeerpatroon en die trombiengenerasie van die mikropartikels te bepaal. Die hoeveelheid mikropartikels is bepaal deur vloeisitometrie te gebruik. VWF- en ADAMTS-13-vlakke is bepaal deur ELISA tegnieke en die trombiengenerasie is gemeet deur kinetiese reaksies. ’n “Western” kladtegniek is gebruik om die VWF-multimere te besigtig. Interleukien-6 het geen uitwerking op die vorming van mikropartikels in HUVEC gehad nie. Die rede hiervoor is dat hierdie tipe endoteelselle geen reseptor vir IL-6 besit nie. Interleukien-8 het die mikropartikel VWF- en ADAMTS-13-vlakke slegs in ’n geringe mate verhoog. Tumor-nekrose-faktor-alfa het die hoeveelheid van endoteel-mikropartikels betekenisvol verhoog. Dit het sowat 80% bygedra tot die trombiengenerasie van die mikropartikels, maar het geen effek op die VWF-inhoud van die mikropartikels gehad nie. Tumor-nekrose-faktor-alfa is dus ’n kragtige stimulant van endoteelselle en is moontlik verantwoordelik vir die trombotiese neigings in trombotiese en inflammatoriese siektetoestande. Die stollingsfaktor TF het mikropartikelvlakke die meeste verhoog, maar het geen uitwerking op die trombiengenerasie van mikropartikels getoon nie. Stimulasie met weefselfaktor in kombinasie met TNF-α het ook ‘n verhoging in mikropartikel-VWF- en ’n klein verlaging in ADAMTS-13 vlakke veroorsaak. Dit is dus moontlik dat TF betrokke is by die verhoogde VWF-vlakke wat in TTP-pasiente voorkom. Inflammasie en trombose speel ‘n hoogs waarskynlike rol in hierdie siektetoestand. Trombien het ‘n beskermende uitwerking op ongeskonde HUVEC, aangesien dit die vorming van mikropartikels bekamp. Die kombinasiestimuli van trombien en sitokine het ook ‘n beskermende uitwerking op HUVEC-selle getoon. Dit beklemtoon dus die regulerende rol van trombien in hemostase, veral op ongeskonde endoteelselle, asook die beskermingseffek daarvan teen trombose in oormatige stollings- en inflammatoriese toestande. Endoteel-mikropartikels kan dus nadelig of voordelig vir die liggaam wees na aanleiding van verskillende behandelings en in verskillende omgewings. Inflammatoriese en stollingsfaktore is wel risikofaktore vir stolling, aangesien dit die aantal en inhoud van mikropartikes beïnvloed. Hierdie studie dra by tot die kennis van mikropartikels afkomstig van endoteelselle asook die rol daarvan in inflammasie en trombose.af
dc.language.isoenen_ZA
dc.publisherUniversity of the Free Stateen_ZA
dc.subjectThrombosis -- Treatmenten_US
dc.subjectBlood -- Coagulationen_ZA
dc.subjectBlood coagulation disordersen_ZA
dc.subjectInflammation -- Treatmenten_ZA
dc.subjectCytokinesen_ZA
dc.subjectDissertation (M.Med.Sc. (Haematology and Cell Biology))--University of the Free State, 2013en_ZA
dc.titleMicroparticles derived from stimulation of human umbilical endotheliumen_ZA
dc.typeDissertationen_ZA
dc.rights.holderUniversity of the Free Stateen_ZA


Files in this item

Thumbnail

This item appears in the following Collection(s)

Show simple item record