title: Dietary fat modulation of extracellular vesicles
Creators: name: Alanzeei, Amal
Publisher: name: University of Reading
Year: 2022

description: Cardiovascular diseases (CVDs) are considered as the major cause of death worldwide. Dietary fats have been reported to be involved in the causation and prevention of CVD. It is advised to cut down the consumption of SFA (saturated fatty acids) and TFA (trans fatty acids) due to their negative impact on CVD risk, particularly with regard to levels of blood cholesterol. On the other hand, n-3 PUFAs (n-3 polyunsaturated fatty acids) are reported to protect against CVD due to their role in reducing blood TAG (triacylglycerol). Extracellular vesicles (EVs) are small membrane-bound vesicles released from all cell types and their levels are reported to be increased in CVD, suggesting their potential use as a disease biomarker. Studying EVs as potential biomarkers involves their purification from contaminating particles; this is of particular importance in postprandial studies after a high fat meal, where the number of lipoproteins particles (whose size overlaps with that of EVs) increases substantially. The first study in this thesis examined whether there was co-isolation and/or interference of lipoproteins during EV isolation and analysis using size-dependent isolation methods. Following consumption of a high fat meal by healthy volunteers, EVs were isolated from platelet-free plasma by size exclusion chromatography (SEC) and the EV fractions were shown to have very little contamination with the apolipoproteins, apoB48 or apoB100. Lipoprotein fractions prepared from platelet-free plasma by density gradient centrifugation and analysed by flow cytometry (FCM) to visualise annexin V-positive, platelet-derived and endothelial�derived EVs were virtually devoid of EVs. Furthermore, when purified lipoproteins were applied to size exclusion columns, they eluted later than EVs, with little cross-contamination. These findings together suggested that even after consuming high fat meal, EVs could be isolated and analysed without significant contamination from lipoproteins 7 The second study described in this thesis explored the effects of high fat meals containing different commercially available fats on the number and thrombogenic activity of EVs during the postprandial period. Numbers and thrombogenic activity of EVs increased during the postprandial period, reaching a peak at 4h, but there was no difference in response to the type of fat in the meal, and in particular, to interesterified (IE) fats compared to traditional commercial fats. The final study was based on published data showing that n-3 PUFA supplements decrease numbers of circulating EVs, advancing on this by comparing the effects of fish oil supplements with two oily fish meals per week on EV number, composition and procoagulant activity. Supplementation with fish oil significantly decreased both number and thrombogenic capacity of EVs, while consumption of oily fish at a level achievable in the diet had no effect on either EV number or thrombogenic capacity. In summary, numbers of EVs are increased during the postprandial period after a high fat meal and the EVs are more thrombogenic, but the type of fat in the meal does not appear to influence either the number or thrombogenicity, and the analysis of EVs during the postprandial period is not hampered by the presence of lipoproteins. In a chronic intervention study, n-3 PUFA from fish oil supplements reduced EV number and procoagulant activity, but consumption of two oily fish meals per week had no effect.