Platelet dysfunction and other hemostatic disorders in women with menorrhagia: the utility of whole blood lumi-aggregometer

Abstract

Menorrhagia is a very common clinical problem among women of reproductive age and is unexplained in more than 50% of cases [1]. The widely accepted clinical definition of menorrhagia is blood loss of 80 ml, or more, per period [2]. Underlying bleeding disorders, such as von Willebrand disease (VWD) and platelet abnormalities may present as menorrhagia. However, the relevance of a systemic screening for hemostasis in women with menorrhagia remains controversial [3]. Here, we describe the findings of a prospective study investigating the frequency of platelet dysfunction and other hemostatic defects among women with unexplained menorrhagia using whole blood lumi-aggregometer. Sixty-seven Turkish white women aged between 17 and 50 years, with a physician diagnosis of menorrhagia were screened after the study was approved by the local ethics committee. Women with known bleeding or other systemic disorders such as renal, hepatic and endocrine diseases were excluded initially. All cases were examined at the gynecology department and had pelvic ultrasonography; those with submucous uterine fibroids, fibroids more than 2 cm in diameter, uterine polyps, ovarian tumors and intrauterine device were excluded. Women were required to have not ingested combined oral contraceptives and other hormonal based therapy at least one cycle prior to sampling. We also attempted to prevent any medication especially non-steroidal anti-inflammatory drugs, aspirin as well as alcohol in the last 10 days prior to the aggregation studies. All subjects were evaluated on days 3–7 of their menstrual cycle to minimize interindividual variation. Sixteen age-matched women with no menorrhagia served as controls. Both patients and controls underwent the following laboratory tests: hemoglobin, platelet count, activated partial thromboplastin time (aPTT), factors (F) VIII, IX, XI, ristocetin cofactor activity (RCof), platelet aggregation and ATP release. Platelet function testing was performed on whole blood platelet lumi-aggregometer (Chronolog Corporation, Model 560-Ca) using luminescence method in diluted blood (1:1 blood normal saline ratio). Citrated blood was collected under light tourniquet through 19 gauge needles into 4.5-ml vacutainers (Becton Dickinson) containing 3.2% trisodium citrate in a 9:1 blood anticoagulant ratio. The citrate tubes were thoroughly mixed by gentle inversion before dispensing 450 ll of citrated whole blood into cuvettes (chronolog No: 367) each containing 450 ll normal saline and a disposable siliconized stir bar. After the tubes were warmed at 37 C and stirred in the incubation wells, 100 ll luciferin (chronolog No: 395 chrono Lume Reagent) was added to the cuvette and the luminesence callibrated 5 min later; the impedance electrode was placed into the cuvette and calibration checked. After agonist addition platelet aggregation and ATP release tracings were measured over 6 min. The agonist used and their final concentrations were; ADP (Chrono Par 384) 5.0 mM, Arachidonic acid (AA) (Chrono Par 390) 0.5 mM, Ristocetin (Chrono Par 396) 1.0 mg/ml, and Collagen (Chrono Par 385) 2 lg/ml. Calculated platelet aggregation (ohms) and ATP release (nmol) normal ranges (as mean ± standard deviation) were 8–21 ohms and 0.4–2.9 nmol for ADP, 13–23 ohms and 0.8–3.5 nmol for AA, 17–28 ohms and 0.5–2.7 nmol for collagen, 5–16 ohms for ristocetin. O. M. Akay (&) Z. Gulbas Department of Hematology, Eskisehir Osmangazi University Medical School, 26480 Eskisehir, Turkey e-mail: melhak@hotmail.com