Abnormalities in the structure of fibrin fibers and blood platelets have been broadly studied and well correlated to functional abnormalities in the coagulation system due to disease, genetics or environmental factors. The importance of these and further ultrastructural investigations of fibrin fiber networks and platelets is therefore paramount in understanding iii the aetiology of haemorrhagic or thrombogenic tendencies and disorders. To study structural abnormalities and variations of fibrin fibers and platelets, electron microscopy is essential; unfortunately the use of electron microscopy necessitates several preparation steps to make a biological specimen stable enough to withstand the high vacuum environment of an electron microscope and also electron beam irradiation. The two most common procedures to accomplish this is chemical fixation and freeze fixation. Chemical fixation entails the chemical alteration of a specimen by means of the introduction of chemical bonds and cross-links that keep molecules and structures in place, followed by several rinsing and dehydration steps. Freeze fixation preserves biological specimens by the removal of thermal energy from the specimen at an extremely fast rate (> 105 In this study, methods to fixate fibrin fiber and platelet networks by freeze fixation was developed, optimised and subsequently compared to chemical fixation methods to ascertain the optimum preparation technique for transmission and scanning electron microscopy for ultrastructural studies of platelets and fibrin networks. K/s) allowing the water in a biological specimen to reach a super-cooled stabilized state (vitrification). The general consensus in the scientific community is that ultrastructural preservation by high pressure freeze fixation is superior to that of chemical fixation, although the facts are that different fixation methodologies have dissimilar chemical and physical interactions with different specimens and as a result different artefact introductions. Therefore the best possible specimen preparation method to ensure an accurate likeness of the fixated specimen to its in vivo condition needs to be ascertained and used. In this study, methods to fixate fibrin fiber and platelet networks by freeze fixation was developed, optimised and subsequently compared to chemical fixation methods to ascertain the optimum preparation technique for transmission and scanning electron microscopy for ultrastructural studies of platelets and fibrin networks. K/s) allowing the water in a biological specimen to reach a super-cooled stabilized state (vitrification). The general consensus in the scientific community is that ultrastructural preservation by high pressure freeze fixation is superior to that of chemical fixation, although the facts are that different fixation methodologies have dissimilar chemical and physical interactions with different specimens and as a result different artefact introductions. Therefore the best possible specimen preparation method to ensure an accurate likeness of the fixated specimen to its in vivo condition needs to be ascertained and used. Ultimately it was found that high pressure freezing coupled with freeze substitution is a superior method for fine structure preservation of fibrin fiber networks and platelets when utilizing transmission electron microscopy. Contrastingly for scanning electron microscopy ultrastructural studies it was found that chemical fixation is the more optimal method for the preparation of fibrin networks and platelets. / Dissertation (MSc)--University of Pretoria, 2013. / gm2014 / Anatomy / Unrestricted

Diabetes mellitus has emerged as a major public health problem with pandemic growth as the International Diabetes Federation estimates that there were 415 million diabetics in 2015 with that number reaching 642 million by 2040, affecting all regions of the world. Globally we are all interconnected when we deal with problems of climate change, water shortage, HIV or Ebola. The war against type 2 diabetes and other non-communicable diseases should be no different, as effective solutions will need expanded global engagement in science to win it. The risk of cardiovascular events in type 2 diabetes remains unchanged despite good control of diabetes and other cardiovascular risk factors. A better understanding of thrombogenicity in diabetes may help to identify novel therapeutic agents and a starting point would be to identify ultrastructural changes in diabetic erythrocytes, platelets and fibrin networks. In diabetes, thrombogenicity is enhanced and is characterised by: hyperactive platelets, higher levels of clotting factors and impaired fibrinolysis. Thus, in this research study, the technique of scanning electron microscopy (SEM) was used to identify ultrastructural abnormalities in erythrocytes, platelets and fibrin networks of diabetic subjects. Distinct abnormal morphological findings were observed in the erythrocytes, platelets and fibrin fibres of diabetic subjects in comparison to the controls. Physiological parameters such as platelet markers and tissue factor levels were also assessed. Flow cytometric analysis revealed hyperactive platelets in the diabetic subjects. The measurement of tissue factor in plasma was completed by using an ELISA. Tissue factor levels in the diabetic subjects were markedly elevated when compared to controls. Biomedical research has provided evidence that has led to the hypothesis that inflammation is the culprit behind almost most chronic illnesses. Hyperglycaemia, a key feature of diabetes, is known to promote a state of low-grade chronic inflammation. A natural method that can resolve acute and chronic inflammation is earthing. Earthing involves coupling your body to the Earth's surface energies by simply walking barefoot or being connected to a conductive device. When earthed, the electrons are conducted into the human body at the same electrical potential as the earth. It is also suggested that free electrons from the earth neutralize the positively charged free radicals that are the hallmark of chronic inflammation. In this study, earthing was accomplished with conductive adhesive patches placed on the sole of each foot and palm of each hand. An earthing cord was connected to the patches and led outdoors to be connected to a stainless-steel rod driven into the ground. Diabetic subjects were earthed for a session of two hours. Bloods were drawn before and just prior to the end of the two-hour session. Morphological SEM findings of the erythrocytes, platelets and fibrin networks at two-hours showed a remarkable difference when compared to findings at baseline. More importantly, the erythrocytes, platelets and fibrin findings revealed that they all almost reverted to looking like control erythrocytes, platelets and fibrin networks. It remains to be seen if earthing will reduce cardiovascular events in diabetics by improving morphology of cells involved in coagulation. / Thesis (PhD)--University of Pretoria, 2016. / Physiology / PhD / Unrestricted