With great appreciation, I would like to accentuate the substantial contributions of the following people who made this project possible: To Dr. CMT Fourie (my supervisor), Prof. JM van Rooyen (my co–supervisor) and Prof. AE Schutte (my co–supervisor) whose gracious advise, patient guidance, commitment and support have enabled me to plan, analyse, interpret and write this project in a scientific manner. It has been an educational experience for me, thank you. To Mr. LS Wyldbore for the language editing of this dissertation. I thank all the participants, researchers, field workers and supporting staff of the PURE study. The financial assistance of the National Research Foundation (DAAD–NRF) towards this research is hereby acknowledged. A special thanks to my parents, sister, Albert, family and friends, thank you for the never–ending love, support, patience and understanding that you gave me throughout this project. Last, but not the least, a special thank to God for giving me the opportunity, talent, determination and endurance to complete this project. / Thesis (M.Sc. (Physiology))--North-West University, Potchefstroom Campus, 2012.

With great appreciation, I would like to accentuate the substantial contributions of the following people who made this project possible: To Dr. CMT Fourie (my supervisor), Prof. JM van Rooyen (my co–supervisor) and Prof. AE Schutte (my co–supervisor) whose gracious advise, patient guidance, commitment and support have enabled me to plan, analyse, interpret and write this project in a scientific manner. It has been an educational experience for me, thank you. To Mr. LS Wyldbore for the language editing of this dissertation. I thank all the participants, researchers, field workers and supporting staff of the PURE study. The financial assistance of the National Research Foundation (DAAD–NRF) towards this research is hereby acknowledged. A special thanks to my parents, sister, Albert, family and friends, thank you for the never–ending love, support, patience and understanding that you gave me throughout this project. Last, but not the least, a special thank to God for giving me the opportunity, talent, determination and endurance to complete this project. / Thesis (M.Sc. (Physiology))--North-West University, Potchefstroom Campus, 2012.

Motivation - The burden of cardiovascular diseases (CVD) is increasing in developing countries worldwide, but even more so in sub-Saharan Africa. Due to rapid urbanisation, black populations experience lifestyle changes (e.g. unhealthy diet, increased access to alcohol and tobacco) that predispose them to increased obesity and cardiovascular risk. In this study, attention will be given to cardiovascular alterations, specifically arterial calcification, in lean and overweight/obese women nearing or already experiencing menopause. These include elevated blood pressure, large artery stiffness (indicated by increased central pulse pressure (cPP)) and carotid intima-media thickness (CIMT). Other factors linked to arterial calcification include the level of obesity as well as low bone mineral density. Ectopic calcification plays a significant role in cardiovascular morbidity and mortality, especially in renal failure patients, osteoporotic and elderly women. Factors contributing to the development and progression of arterial calcification include calciotropic hormones and altered bone metabolism, particularly in older postmenopausal women. This is due to the lack of protective effects of oestrogen against vascular alterations and bone loss after menopause. Previous studies have shown that increased bone resorption indicated by elevated levels of c-telopeptide of type I collagen (CTX), parathyroid hormone (PTH), low 25- hydroxycholecalciferol (25(OH)D3) and parathyroid hormone to 25-hydroxycholecalciferol ratio (PTH:25(OH)D3) are independently linked to arterial stiffening, CIMT and vascular calcification. Knowledge on the contribution of altered bone metabolism and associated calciotropic hormones on cardiovascular health in Africans is limited. Previous studies on ectopic calcification in South Africans focused on men and renal failure patients. This study will explore the possible role of altered calcium regulation and bone metabolism in the development of arterial calcification and CVD in older African women. Aim - The aim of this study was to investigate the associations of brachial and central pressures and CIMT with PTH, PTH:25(OH)D3 and CTX, a marker of bone resorption, in lean and overweight/obese African women older than 46 years. Methodology - This sub-study forms part of the Prospective Urban Rural Epidemiology (PURE) study. A total of 434 urban and rural women older than 46 years were included in the study. Women infected with the human immunodeficiency virus (HIV) were excluded from the study. The study was reviewed and approved by the Ethics Committee of the North-West University (Potchefstroom campus) and all participants signed an informed consent form prior to enrolment into the project. Field workers administered demographic, general health and physical activity questionnaires in the participants’ home language. Anthropometric measurements included weight, height and waist circumference, while body mass index (BMI) was calculated in kg/m2. Cardiovascular measurements included brachial and central systolic blood pressure (SBP), brachial diastolic blood pressure (DBP), brachial and central pulse pressure (PP) as well as CIMT and carotid cross-sectional wall area (CSWA). Blood pressure measurements were performed on the right arm with the participant in the sitting position. Blood was drawn after an overnight fasting period. We performed biochemical analyses from serum and plasma samples for follicle stimulating hormone (FSH), PTH, 25(OH)D3, and CTX. HIV testing was performed according to standardised procedures. Since interactions existed for BMI with regards to associations of CIMT and cPP with PTH:25(OH)D3, the study population was divided into the lean (BMI <25 kg/m2) and overweight/obese (BMI ≥25 kg/m2) groups. We performed independent T-tests to compare means and used the chi-square test to compare proportions. Single and multiple regression analyses were performed to investigate the associations of markers of vascular structure and function with CTX and calciotropic hormones. Results - In this study, 90% of the women displayed an FSH concentration exceeding the cut-off value of 35 mIu/mL, indicating a postmenopausal state. When comparing lean and overweight/obese African women, we found that lean women had higher levels of CTX and 25(OH)D3 (both p<0.001), while the overweight/obese group was older (p=0.007) and presented with higher PTH and PTH:25(OH)D3 levels (both p<0.001). Brachial and central pressures did not differ between the groups (p≥0.23), except for DBP being higher in the overweight/obese group (p=0.017). Overweight/obese women had higher CIMT (p<0.001) and CSWA (p=0.001) as compared to their lean counterparts. A larger proportion of lean women smoked (63%) and self-reported on alcohol use (37%) than overweight/obese women (41% and 18%, respectively) (both p<0.001). Forty-one percent of overweight/obese women used antihypertensive medication, opposed to 25% in the lean group (p=0.001). In multivariate regression analyses, an independent positive association existed between CIMT and PTH:25(OH)D3 (R2=0.22; β=0.26; p=0.003) in lean women. In the overweight/obese group independent positive associations were confirmed between brachial SBP and PTH (p=0.013) and CTX (p=0.038), and between DBP and PTH (p=0.030). Brachial PP and central SBP remained positively associated with CTX (p=0.016 and p=0.024, respectively), while cPP was independently associated with PTH:25(OH)D3 (R2=0.20; β=0.17; p=0.017) and CTX (R2=0.20; β=0.17; p=0.025). Conclusion - Our results indicate that in older African women, large artery structure and function are associated with calciotropic hormones and bone resorption, suggesting that altered bone metabolism and associated calciotropic hormones play a role in the development of vascular calcification. The different associations in lean and overweight/obese women suggest different mechanisms at work regarding arterial calcification in states of low and high adiposity. These findings need confirmation in larger prospective and experimental studies. / MSc (Physiology), North-West University, Potchefstroom Campus, 2014

Motivation - The burden of cardiovascular diseases (CVD) is increasing in developing countries worldwide, but even more so in sub-Saharan Africa. Due to rapid urbanisation, black populations experience lifestyle changes (e.g. unhealthy diet, increased access to alcohol and tobacco) that predispose them to increased obesity and cardiovascular risk. In this study, attention will be given to cardiovascular alterations, specifically arterial calcification, in lean and overweight/obese women nearing or already experiencing menopause. These include elevated blood pressure, large artery stiffness (indicated by increased central pulse pressure (cPP)) and carotid intima-media thickness (CIMT). Other factors linked to arterial calcification include the level of obesity as well as low bone mineral density. Ectopic calcification plays a significant role in cardiovascular morbidity and mortality, especially in renal failure patients, osteoporotic and elderly women. Factors contributing to the development and progression of arterial calcification include calciotropic hormones and altered bone metabolism, particularly in older postmenopausal women. This is due to the lack of protective effects of oestrogen against vascular alterations and bone loss after menopause. Previous studies have shown that increased bone resorption indicated by elevated levels of c-telopeptide of type I collagen (CTX), parathyroid hormone (PTH), low 25- hydroxycholecalciferol (25(OH)D3) and parathyroid hormone to 25-hydroxycholecalciferol ratio (PTH:25(OH)D3) are independently linked to arterial stiffening, CIMT and vascular calcification. Knowledge on the contribution of altered bone metabolism and associated calciotropic hormones on cardiovascular health in Africans is limited. Previous studies on ectopic calcification in South Africans focused on men and renal failure patients. This study will explore the possible role of altered calcium regulation and bone metabolism in the development of arterial calcification and CVD in older African women. Aim - The aim of this study was to investigate the associations of brachial and central pressures and CIMT with PTH, PTH:25(OH)D3 and CTX, a marker of bone resorption, in lean and overweight/obese African women older than 46 years. Methodology - This sub-study forms part of the Prospective Urban Rural Epidemiology (PURE) study. A total of 434 urban and rural women older than 46 years were included in the study. Women infected with the human immunodeficiency virus (HIV) were excluded from the study. The study was reviewed and approved by the Ethics Committee of the North-West University (Potchefstroom campus) and all participants signed an informed consent form prior to enrolment into the project. Field workers administered demographic, general health and physical activity questionnaires in the participants’ home language. Anthropometric measurements included weight, height and waist circumference, while body mass index (BMI) was calculated in kg/m2. Cardiovascular measurements included brachial and central systolic blood pressure (SBP), brachial diastolic blood pressure (DBP), brachial and central pulse pressure (PP) as well as CIMT and carotid cross-sectional wall area (CSWA). Blood pressure measurements were performed on the right arm with the participant in the sitting position. Blood was drawn after an overnight fasting period. We performed biochemical analyses from serum and plasma samples for follicle stimulating hormone (FSH), PTH, 25(OH)D3, and CTX. HIV testing was performed according to standardised procedures. Since interactions existed for BMI with regards to associations of CIMT and cPP with PTH:25(OH)D3, the study population was divided into the lean (BMI <25 kg/m2) and overweight/obese (BMI ≥25 kg/m2) groups. We performed independent T-tests to compare means and used the chi-square test to compare proportions. Single and multiple regression analyses were performed to investigate the associations of markers of vascular structure and function with CTX and calciotropic hormones. Results - In this study, 90% of the women displayed an FSH concentration exceeding the cut-off value of 35 mIu/mL, indicating a postmenopausal state. When comparing lean and overweight/obese African women, we found that lean women had higher levels of CTX and 25(OH)D3 (both p<0.001), while the overweight/obese group was older (p=0.007) and presented with higher PTH and PTH:25(OH)D3 levels (both p<0.001). Brachial and central pressures did not differ between the groups (p≥0.23), except for DBP being higher in the overweight/obese group (p=0.017). Overweight/obese women had higher CIMT (p<0.001) and CSWA (p=0.001) as compared to their lean counterparts. A larger proportion of lean women smoked (63%) and self-reported on alcohol use (37%) than overweight/obese women (41% and 18%, respectively) (both p<0.001). Forty-one percent of overweight/obese women used antihypertensive medication, opposed to 25% in the lean group (p=0.001). In multivariate regression analyses, an independent positive association existed between CIMT and PTH:25(OH)D3 (R2=0.22; β=0.26; p=0.003) in lean women. In the overweight/obese group independent positive associations were confirmed between brachial SBP and PTH (p=0.013) and CTX (p=0.038), and between DBP and PTH (p=0.030). Brachial PP and central SBP remained positively associated with CTX (p=0.016 and p=0.024, respectively), while cPP was independently associated with PTH:25(OH)D3 (R2=0.20; β=0.17; p=0.017) and CTX (R2=0.20; β=0.17; p=0.025). Conclusion - Our results indicate that in older African women, large artery structure and function are associated with calciotropic hormones and bone resorption, suggesting that altered bone metabolism and associated calciotropic hormones play a role in the development of vascular calcification. The different associations in lean and overweight/obese women suggest different mechanisms at work regarding arterial calcification in states of low and high adiposity. These findings need confirmation in larger prospective and experimental studies. / MSc (Physiology), North-West University, Potchefstroom Campus, 2014