Balkan Journal of Medical Genetics

RISK FACTORS OF VENOUS THROMBOEMBOLISM IN SUDANESE PREGNANT WOMEN
Abdalhabib EK, Alfeel A, Ali EI, Ibrahim IK, Mobarki AA, Dobie G, Hamali HA, Saboor M,
*Corresponding Author: Dr. Muhammad Saboor, Department of Medical Laboratory Technology, Faculty of Applied Medical Science, Jazan University, Jazan, Saudi Arabia. Tel.: +966-54-495-9029. E-mail: msaboor@ jazanu.edu.sa
page: 49

MATERIAL AND METHODS

Study Design. This prospective case-control study was conducted between January 2018 and August 2019 at the Khartoum Teaching Hospital in Khartoum State, Sudan, recruiting 396 pregnant women. Ethics approval was obtained from the University of Science and Technology, Sudan, and the study was conducted according to the Declaration of Helsinki. Informed consent was obtained from all women before the collection of blood samples. Blood samples were collected before the initiation of the anticoagulant therapy. The 396 participants were equally divided into pregnant women diagnosed with VTE (n = 198) and normal pregnant women (n = 198) without any history or diagnosis of VTE as age-matched controls. The diagnosis of VTE was based on the presence of clinical signs suggestive of VTE and confirmed by venous compression ultra-sonography and/or multidetector-row (spiral) computed tomography. Data were collected using a predesigned and pretested questionnaire. Participants were asked about demographic data and VTE risk factors, such as familial history of thrombosis, fractures, recent surgery, kidney disease, malignant tumors, immobilization, history of hypertension, diabetes mellitus, smoking habits, and use of oral contraception or hormone replacement therapy. The BMI was calculated to evaluate obesity [body weight (kg) divided by the square of height (m)]. Plasma Extraction and Analysis. Blood samples were collected in EDTA-containing vacutainers for DNA analysis and in sodium citrate-anticoagulated tubes for PC, PS and AT measurements. Citrated plasma was obtained by centrifugation of blood samples at 3000 rpm for 30 min. Plasma was extracted and stored at –80 °C until analysis. Protein C and PS levels were measured by a clot-based assay using the reagents of TECHNOCLOT® (Technoclone Herstellung von Diagnostika und Arzneimitteln GmbH, Vienna, Austria). These levels were determined from a calibration curve in which prolongation of clotting time is proportional to the protein concentration. The AT level was determined using an automated coagulation analyzer (Sysmex CA-1500) using the TECHNOCHROM® AT III Kit (Technoclone Herstellung von Diagnostika und Arzneimitteln GmbH). Reference values of PC, PS and AT in the range of 55.0-125.0, 50.0-130.0 and 62.0-120.0 ng/mL, respectively, were considered as normal based upon the preestablished normal levels. A decreased level of any of these natural anticoagulants from the reference values were considered to be deficient. DNA Extraction and Analysis. DNA was extracted from the blood samples collected in the EDTA vacutainers and purified using the QIAamp DNA Blood Mini Kit according the manufacturer’s standard protocol (Qiagen Sciences Inc., Germantown, MD, USA). The FVL G1691A polymorphism and prothrombin G20210A mutation were detected by allele-specific polymerase chain reaction (ASPCR) using the Bio-Rad DNA Engine Dyad Peltier Thermal Cycler (Bio-Rad Laboratories, Foster City, CA, USA) as described previously [10]. Statistical Analysis. Qualitative data, which are expressed as proportions, were analyzed using the χ2 test. Quantitative data are expressed as mean ± SD. The z-test was used for testing the significance of quantitative data. The association between two quantitative parameters was evaluated by correlation analysis, and adjusted odds ratios (ODs) were calculated for determining the strength of association between qualitative parameters. Statistical significance was set at p <0.05.

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