CARDIOVASCULAR JOURNAL OF AFRICA • Volume 33, No 2, March/April 2022 66 AFRICA interpreted as a sign of long-term endothelial dysfunction for those women.18 Additionally, interleukin-6 levels are known to be elevated in certain cardiovascular diseases. Higher plasma and myocardium levels of interleukin-6 were present in patients with end-stage heart failure compared to recent-onset heart failure.19 Its expression was proved to be induced in ischaemic and reperfused areas during myocardial infarction,20 and it was also able to predict future coronary incidents.21 On the other hand, changes in cardiac structure and function, as assessed echocardiographically, appear to be more pronounced during the course of hypertensive pregnancies compared to normotensive ones.22-24 This suggests a degree of abnormal cardiovascular response of the female organism during pre-eclampsia and gestational hypertension. In this study we aimed to determine serum levels of interleukin-6 in women with pre-eclampsia and gestational hypertension, and compare them with those of healthy pregnant controls. Additionally, we examined correlations of interleukin-6 levels with some characteristics of the women and echocardiographic findings as a potential link between hypertensive disorders of pregnancy and cardiovascular diseases. Methods Between August 2018 and January 2020, a prospective, singlecentre, clinical, epidemiological study was conducted at the Clinic of Cardiology at the University multi-profile hospital Sveti Georgi, Plovdiv, Bulgaria, and 123 pregnant women over 18 years of age were enrolled, 37 with the diagnosis of pre-eclampsia, 36 with gestational hypertension. Fifty healthy pregnant controls were also enrolled. The women were recruited from the Clinic of Obstetrics and Gynecology in the same hospital and some of the controls were referred by local obstetrics and gynecology practices. The study included the analysis of certain biomarkers as well as echocardiographic assessment of the women. The study was carried out according to the Declaration of Helsinki and approved by the ethics committee of the Medical University – Plovdiv. All of the participants signed a written, informed consent after a detailed explanation about the study and the required procedures. One hundred and sixteen of the women had singleton pregnancies and nine had bigeminal pregnancies, four in the pre-eclampsia group, two in the gestational hypertension group and three controls. Current weight and height of the women were measured with standardised equipment. Weight before the pregnancy was self-reported. A diagnosis of pre-eclampsia was established if the women had high blood pressure [office-measured systolic blood pressure (SBP) ≥ 140 mmHg and/or diastolic blood pressure (DBP) ≥ 90 mmHg at least twice over the course of a minimum of four hours] registered for the first time after the 20th gestational week, and proteinuria of ≥ 300 mg/l for 24 hours. Gestational hypertension was diagnosed if high blood pressure was registered for the first time after the 20th gestational week, and proteinuria was < 300 mg/l for 24 hours. Both hypertensive forms were considered early if the hypertension was first discovered before the 34th gestational week.21 Both hypertensive conditions were considered severe if the women had registered SBP ≥ 160 mmHg and/or DBP ≥ 110 mmHg or levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) twice the upper reference limit of the laboratory. No women in the study had thrombocytopaenia, corresponding to the criterion for severe forms of hypertensive disorder of pregnancy (< 100 × 109 cells/l). In order to ensure that the changes in biomarker levels were not influenced by other conditions, women with recent infections and any serious systemic diseases or organ failure were not asked to participate in the study. Women with chronic arterial hypertension, diabetes mellitus and any known or significant heart diseases discovered in the course of the study were also not included. For ethical reasons women who had pulmonary congestion, encephalopathy, epigastric pain, or HELLP syndrome (all considered forms of severe HDP) or any other medical emergency were not asked to participate in the study as their participation could delay urgent medical interventions. For the control group, women with diagnosed intrauterine retardation of the foetus were not included either. Venous blood was collected from the women in certified monovettes with a cloth activator. Serum was separated via centrifugation at 3 000 rpm for 10 minutes and then stored at –20°C as recommended by the test kit manufacturer. Serum interleukin-6 levels were determined with solid-phase sandwich ELISA (Diaclone, Besançon, France) with a biotinylated human interleukin-6 antibody. Electrocardiograms were performed on all women in order to exclude any significant rhythm or conduction disturbances, which could compromise the results of the study. A thorough transthoracic echocardiographic examination according to a protocol was performed with the cardiovascular ultrasound systemGeneral Electric Vivid 9.5 and the echographic recordings were analysed using EchoPAC clinical workstation software version 201 (General Electric Medical System, Milwaukee, WI, USA). Measurements were performed according to the current recommendations of the guidelines of the European Association of Cardiovascular Imaging (EACVI) and the American Society of Echocardiography (ASE).25,26 Statistical analysis Statistical analysis was performed using IBM SPSS statistics 25.0 (IBM SPSS Statistics for Windows, SPSS Inc, Chicago, IL, USA) and MedCalc Version 14.8.1 (MedCalc Software, Mariakerke, Belgium). Continuous variables were tested for normality with Kolmogorov–Smirnov and Shapiro–Wilk tests. The Student’s t-test, analysis of variance (ANOVA) test and Bonferroni post hoc test were used to compare the continuous variables that had normal distribution and more than two independent groups with homogeneity of variances. Continuous variables with non-normal distribution were compared with the Kruskal– Wallis test and the Mann–Whitney U-test. The relationship between categorical variables in cross tables was analysed using the χ2 and Fisher’s exact tests. Correlation analysis was performed using either Pearson’s correlation coefficient or Spearman’s rho according to the normality of the continuous variables. Receiver operating characteristic (ROC) curve analysis was carried out to determine discriminative abilities of interleukin-6. Logistic regression was performed to explain the relationship between variables. Findings with p < 0.05 were considered statistically significant.
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