Increased Microparticle-Linked Procoagulant Activity In Patients with Primary Immune Thrombocytopenia.

Abstract 3707

Primary immune thrombocytopenia (ITP) is an acquired immune-mediated disorder characterized by isolated thrombocytopenia (platelet count less than 100,000/μL) and the absence of any obvious initiating and/or underlying cause for the thrombocytopenia. In spite of the low platelet number, some thrombocytopenic patients seldom bleed, indicating the existence of other factors that regulate haemostasis in these patients. Elevated levels of plasma microparticles (MPs) had been observed in IPT patients. MPs are vesicles with a size less than 0.5 micrometers, derived from cell membranes after their activation or apoptosis. Most MPs are highly procoagulant, expressing annexin V binding sites and tissue factor. However, relatively little is known of their specific functions in ITP.

In the present study we aim to elucidate if a relationship exists between microparticle-linked procoagulant activity and haemostasis in ITP patients.

Twenty-two ITP patients, 3 male and 19 female, aged between 25 to 92 years, were included. Sixteen age- and sex-matched healthy individuals were used as control group.

Platelet-related primary haemostasis was evaluated with an automated platelet function analyzer (PFA-100®, Siemens Healthcare Diagnostics). Samples of citrated blood were aspirated under a shear rate of 4,000–5,000/s through a 150-micrometer aperture cut into a collagen-ADP (COL-ADP) or collagen-epinephrine (COL-EPI) coated membrane. The platelet haemostatic capacity is indicated by the time required for the platelet plug to occlude the aperture (closure time), which is expressed in seconds.

MP procoagulant activity was determined with ZYMUPHEN MP-Activity kit (Aniara, Mason, Ohio) and by calibrated automated thrombography (CAT) in plasma samples obtained after 2 centrifugations at room temperature (first: 15 min at 1,500 g, second: 2 min at 13,000 g). These methods measure endogenous thrombin generation. CAT evaluates four parameters of thrombin generation: the endogenous thrombin potential (ETP), lag time, time to peak (TTP) and peak height.

PFA-100® determinations with COL-EPI and COL-ADP cartridges in blood samples from ITP patients with less than 50,000/μL showed longer closure times than control group (p<0.05), whereas samples from ITP patients with a platelet count between 50,000/μL and 100,000/μL showed closure times of the same order of magnitude as control ones (platelet count ranging from 162,000 to 368,000)μL).Plasma from these patients had higher MP-mediated procoagulant activity evaluated with ZYMUPHEN kit (control 6.1+3.9 nM, ITP group 10,1±8.2 nM, p<0.05) as well as with CAT (ETP (nM*min): control: 1692.6±341.9, ITP: 2191,8±398.9, p<0.01; lag time (min): control: 19.9±8.2, ITP: 14.3±4.3, p<0.05; TTP (min): control: 22.0±8.3, ITP:16.3±4.4, p<0.05; peak height (nM): control: 389.7±70.6, ITP: 498,8±97.5, p<0.01).

Our results indicate that increased MP procoagulant activity in ITP patients may be protective against bleeding events that should be observed in those thrombocytopenic conditions.

Three of the ITP patients included in this study had been splenectomyced and we consider of interest to point out that two of them in spite of recovering a normal platelet count still maintain a high MP procoagulant activity. This observation agrees with a recent work that postulates that MPs might contribute to an increased risk of thrombosis, progression of atherosclerosis and cardiovascular disease following splenectomy (Fontana et al, Thromb Research, 2008;122:59).