Plasminogen activator inhibitor-1 in black South Africans, methodological and clinical considerations

Abstract

Introduction and aim Black South Africans are experiencing an increase in the prevalence of cardiovascular disease (CVD). Obesity and abnormal haemostasis are among the underlying risk factors associated with CVD development in the African populations. Fibrinolysis and, in particular, one of its main inhibitors, plasminogen activator inhibitor-1 (PAI-1), have been found to play an important regulating role in the development of abnormal haemostasis and consequently, increase the risk of CVD development. Most of the studies investigating PAI-1 provide data on individuals of European descent. Much less information is available regarding PAI-1 in other ethnicities. From these initial studies, two issues were identified that requires investigation in African participants and formed the aim of this thesis. The first is a methodological issue relating to the influence of platelets in plasma on PAI-1 and PAI-1 related assays as a possible explanation for inconsistencies in ethnic differences reported for PAI-1. The second is a clinical issue focusing on the relationship between PAI-1 and body fat distribution in Africans. Preliminary evidence suggests this relationship may differ from what has been observed in European populations. The specific objectives were to determine the effect of residual platelets in plasma on different plasma PAI-1 and PAI-1 related assays, to investigate the relationship of PAI-1 with body fat distribution patterns in Africans and also to review the literature reporting on the association between body fat distribution and plasma PAI-1 levels. Participants and methods Methodological issues were addressed in the Sympathetic activity and Ambulatory Blood Pressure in Africans (SABPA) study that included 151 teachers. In addition, data was also used from a follow-up study where 20 academic staff members of a tertiary institution were recruited. As for the clinical issues, available data from the Sarcopenic Obesity and Non-communicable Disease Risk in African Adults (SONDRAA) study were used in which 246 African women were included. Fasting blood samples were collected from the participants’ antebrachial vein branches with a sterile winged infusion set before 10:00. Samples were centrifuged at different centrifugation speeds, as stipulated by the respective study protocols and stored at -82°C until further analysis. The following variables were analysed in the SABPA and follow-up study: PAI- 1 activity (PAI-1act), antigen (PAI-1ag), tissue plasminogen activator (tPA)/PAI-1 complex, clot lysis time (CLT), beta thromboglobulin (βTG), and plasma platelet count and size. In the SONDRAA study, PAI-1act, insulin, glucose, HIV status, high sensitivity C-reactive protein, urinary albumin and creatinine were analysed. In order to determine different body fat distribution patterns in the SONDRAA study population, measurements of body weight, waist circumference, abdominal and supraspinal skinfolds, as well as body composition by means of dual-energy x-ray absorption were analysed. The identified clinical issues were also address by undertaking an overview of ex vivo and in vivo studies investigating the association between body fat distribution and plasma PAI-1 levels in order to contextualise apparent disparate findings. Results Results from the methodological issues that were addressed indicated that a marker of platelet α-granule release (plasma βTG) associated significantly with total PAI-1 content (PAI-1ag) levels and demonstrated weak associations only with active PAI-1 (PAI-1act) and the functional marker, clot lysis time (CLT). In the follow-up study it was indicated that plasma PAI-1ag was also strongly affected by platelet count in a concentrated-dependent manner, and that plasma PAI- 1ag levels increased even further after complete platelet degradation. Regarding the association of PAI-1 with different body fat distribution patterns (clinical considerations), the research determined that the PAI-1act of sarcopenic obese women did not differ significantly from that of the non-sarcopenic obese women (p=0.8) in this study population. Body fat distribution patterns and degree of obesity influenced the relationship of PAI-1act with body fat percentage, insulin, triglycerides and appendicular skeletal mass (ASM). It was also established that PAI-1act was higher (1.65 vs 0.16 U/ml; p=0.001) in women with proportionally higher visceral adipose tissue (VAT), compared to women with proportionally higher abdominal subcutaneous adipose tissue (SCAT) in the total study population. This was, however, not the case in the centrally obese sub-group (1.72 vs 0.83 U/ml; p=0.5). By applying multiple regression models, it was established that body fat percentage as such, did not contribute markedly to the PAI-1act variance in women with increased fat mass, but that other factors associated with obesity such as inflammation and endothelial damage contributed to a larger extent. Additionally, in the overview of the literature, factors that influence the relationship between body fat distribution and plasma PAI-1 levels were identified, while the relative contribution of adipose tissue compared to other PAI-1 source tissue was also put into perspective. Conclusion With regard to methodological issues, it was concluded that residual platelets in plasma significantly influence plasma PAI-1ag levels mainly by increasing latent PAI-1 levels with limited effects on PAI-1act, tPA/PAI-1 complex or CLT and that this is done in a platelet concentration dependent manner. Platelet concentration should therefore be strictly monitored specifically when measuring PAI-1ag. In terms of the clinical data, it was found that patterns of fat distribution and the degree of obesity influenced the association of PAI-1act with insulin, triglycerides, ASM and body fat percentage in African women and that in conditions of extreme obesity, abdominal SCAT contributes equally to plasma PAI-1act than does VAT. It was furthermore established that the relationship between VAT and plasma PAI-1 levels is not fixed; it rather seems to be regulated by a number of other factors such as the size of the subcutaneous adipose tissue depot, ethnicity, possibly genetics and other obesity-related metabolic abnormalities. Lastly, it was also determined that body fat percentage per se contributes less to PAI-1act variance in African women than other obesity related derangements such as endothelial dysfunction and inflammation which should be taken into account when investigating the relationship between body composition and PAI-1.