Ian Mackie

Frequency and prevention of symptomless deep-vein thrombosis in long-haul flights: a randomised trial

BACKGROUND The true frequency of deep-vein thrombosis (DVT) during long-haul air travel is unknown. We sought to determine the frequency of DVT in the lower limb during long-haul economy-class air travel and the efficacy of graduated elastic compression stockings in its prevention. METHODS We recruited 89 male and 142 female passengers over 50 years of age with no history of thromboembolic problems. Passengers were randomly allocated to one of two groups: one group wore class-I below-knee graduated elastic compression stockings, the other group did not. All the passengers made journeys lasting more than 8 h per flight (median total duration 24 h), returning to the UK within 6 weeks. Duplex ultrasonography was used to assess the deep veins before and after travel. Blood samples were analysed for two specific common gene mutations, factor V Leiden (FVL) and prothrombin G20210A (PGM), which predispose to venous thromboembolism. Asensitive D-dimer assay was used to screen for the development of recent thrombosis. FINDINGS 12/116 passengers (10%; 95% CI 4.8-16.0%) developed symptomless DVT in the calf (five men, seven women). None of these passengers wore elastic compression stockings, and two were heterozygous for FVL. Four further patients who wore elastic compression stockings, had varicose veins and developed superficial thrombophlebitis. One of these passengers was heterozygous for both FVL and PGM. None of the passengers who wore class-I compression stockings developed DVT (95% CI 0-3.2%). INTERPRETATION We conclude that symptomless DVT might occur in up to 10% of long-haul airline travellers. Wearing of elastic compression stockings during long-haul air travel is associated with a reduction in symptomless DVT.

Remission in acute refractory and relapsing thrombotic thrombocytopenic purpura following rituximab is associated with a reduction in IgG antibodies to ADAMTS-13

Thrombotic thrombocytopenic purpura (TTP) is a life‐threatening disorder and plasma exchange (PEX) remains the primary treatment modality. Twenty‐five patients with acute refractory/relapsing idiopathic TTP received rituximab in conjunction with PEX because of progressive clinical disease or deterioration in laboratory parameters, despite intensive standard therapy. In relapsing TTP, rituximab was started if antibody to ADAMTS‐13 (a disintegrin and metalloproteinase with thrombospondin motif‐13) was demonstrated during previous episodes. All 25 patients attained complete clinical and laboratory remission in a median of 11 d after initiating rituximab. In 21 cases, ADAMTS‐13 activity was within the normal range following rituximab. Inhibitors were detected in 24/25 patients by mixing studies and/or immunoglobulin G (IgG) antibodies to ADAMTS‐13 pre‐rituximab. There was no evidence of inhibitors and/or IgG activity <10% in 23/25 patients following rituximab. In acute refractory cases, the median number of PEX pre‐rituximab and following the first rituximab infusion was 13 and 9, respectively. There have been no infectious complications, despite low CD 19 levels and no relapses. In patients with acute refractory/relapsing idiopathic TTP, rituximab appears to be a safe, effective, targeted therapy with a significant reduction in the requirement for PEX.

Increased circulating platelet–leucocyte complexes and platelet activation in patients with antiphospholipid syndrome, systemic lupus erythematosus and rheumatoid arthritis

It is possible that platelet activation may play a pathogenic role in the increased risk of thrombosis associated with antiphospholipid antibodies (APA). In this study, levels of in vivo platelet activation were measured in 20 patients with primary antiphospholipid syndrome (PAPS) and 30 systemic lupus erythematosus (SLE) patients (14 of whom had secondary APS) using sensitive flow cytometry. Soluble P‐selectin levels were also assayed. Platelet CD63 expression was significantly higher in PAPS than normal controls (P = 0·007), as well as SLE patients with and without secondary APS (P = 0·03 and P = 0·002 respectively). PAC‐1 binding was significantly higher in PAPS than the control group (P = 0·007) and SLE patients without APS (P = 0·015). Platelet–leucocyte complexes were significantly higher in SLE patients than both PAPS and the control group, and platelet–monocyte complexes were significantly increased in PAPS compared with the control group. (Platelet–leucocyte complexes were also significantly higher than controls in 10 rheumatoid arthritis (RA) patients without APA). Soluble P‐selectin levels were significantly higher in PAPS and SLE patients than the control group. Platelet CD62p expression, annexin V binding and platelet microparticle numbers were not increased in PAPS or SLE patients. We conclude that there is evidence of increased platelet activation in PAPS and SLE, and this is important to note as it may have potential therapeutic implications with respect to use of antiplatelet agents in these patients.

Guidelines for the laboratory investigation of heritable disorders of platelet function

The guideline writing group was selected to be representative of UK‐based medical experts. MEDLINE was systematically searched for publications in English up to the Summer of 2010 using key words platelet, platelet function testing and platelet aggregometry. Relevant references generated from initial papers and published guidelines/reviews were also examined. Meeting abstracts were not included. The writing group produced the draft guideline, which was subsequently revised and agreed by consensus. Further comment was made by members of the Haemostasis and Thrombosis Task Force of the British Committee for Standards in Haematology. The guideline was then reviewed by a sounding board of approximately 40 UK haematologists, the British Committee for Standards in Haematology (BCSH) and the British Society for Haematology Committee and comments incorporated where appropriate. Criteria used to quote levels and grades of evidence are as outlined in appendix 7 of the Procedure for Guidelines Commissioned by the BCSH [http://www.bcshguidelines.com/BCSH_PROCESS/EVIDENCE_LEVELS_AND_GRADES_OF_RECOMMENDATION/43_GRADE.html]. The objective of this guideline is to provide healthcare professionals with clear guidance on platelet function testing in patients with suspected bleeding disorders. The guidance may not be appropriate to patients receiving antiplatelet therapy and in all cases individual patient circumstances may dictate an alternative approach.

Guidelines on fibrinogen assays

Haematology departments in the UK have traditionally performed fibrinogen assays to detect decreased levels and abnormalities of fibrinogen, and to assess haemorrhagic risk. It has also been shown that elevated fibrinogen levels are a predictor of a variety of arterial cardiovascular events, and fibrinogen assays are sometimes recommended with this in mind. The Clauss fibrinogen assay (based on the thrombin clotting time) is the most popular technique in UK hospital laboratories, although many other methods are also in use. There appears to be great variability in both the source of reagents and the exact method used for the Clauss assay. Most laboratories are now equipped with automated coagulation analysers, and many of these perform a fibrinogen estimation derived from the degree of change of light scatter or optical density during the prothrombin time (PT-Fg). A number of problems have been described in the use of the PT-Fg method: it generally gives higher values than the Clauss technique, but the exact degree of discrepancy seems to depend on a number of different variables. International and National standards are available for fibrinogen, but do not appear to be universally used. These guidelines have been prepared against this background and recommend which methods should be used in various clinical settings, as well as highlighting a variety of problems with fibrinogen assays.

Regional UK TTP registry: correlation with laboratory ADAMTS 13 analysis and clinical features.

Thrombotic thrombocytopenic purpura (TTP) is an acute, rare, life‐threatening disorder. This report presents the South East (SE) England registry for TTP, from April 2002 to December 2006, which included 176 patients and 236 acute episodes; 75% of patients were female and 25% were male, overall median age at presentation was 42 years. Mortality was 8·5%, most cases died before treatment was instigated. The main ethnic groups were Caucasian (64%) and Afro Caribbean (27%). Seventy‐seven percent of cases were idiopathic, 5% were congenital and the remaining cases had a defined precipitant. Neurological features were the most prevalent, but cardiac involvement accounted for 42% of presenting features. The overall median number of plasma exchanges (PEXs) to remission was 15; between April 2002 and December 2003, the median number of PEXs was 19 and it was 12 between January 2004 and December 2006 (P < 0·0001). In the latter period, adjuvant therapies were reduced, but Rituximab was increased. ADAMTS 13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) activity was <10% in 74% and 95% of these cases had positive IgG antibodies to ADAMTS 13. Renal impairment and delayed normalisation of platelet count were the main differences between idiopathic and secondary TTP.

Performance of the platelet function analyser PFA-100 in testing abnormalities of primary haemostasis.

The PFA-100 device is a new instrument for the in-vitro testing of platelet function. Primary haemostasis is stimulated by recording the closure time taken for platelets to seal a 150 microm aperture in the centre of a membrane coated with collagen and either epinephrine or ADP. Patients with type 3 von Willebrands disease (n = 4) all had infinitely prolonged closure times (> 200 s) with both types of cartridge. A patient with afibrinogenemia exhibited only slightly prolonged closure times of 111 and 166 s for the ADP and epinephrine membranes, respectively. Patients with Glanzmanns thrombasthenia (n = 6) and Bernard Soulier syndrome (n = 2) had grossly prolonged closure times (> 200 s) with both types of cartridges. These results confirmed that the PFA-100 system was highly dependent on normal von Willebrand factor, glycoprotein Ib and glycoprotein IIb/IIIa levels but not on plasma fibrinogen. Patients with storage pool disease (n = 6) and Hermansky Pudlak syndrome (n = 7) had prolonged closure times with the epinephrine cartridge. There was no evidence of enhanced platelet function in patients with antiphospholipid syndrome, in sickle-cell disease or thalassemia. However, ingestion of aspirin resulted in a near consistent and significant prolongation of the closure time for the epinephrine cartridge but not for the ADP cartridge in both normal subjects and patients. The test offers a reliable, reproducible, rapid and simple means of assessing high-shear platelet function in vitro.

Microparticle sizing by dynamic light scattering in fresh-frozen plasma

Background   We have previously shown that fresh‐frozen plasma (FFP) contains red blood cell‐derived procoagulant microparticles (MPs) that are removable by 0·2 µm filtration. Given the limitations of current methods for accurately sizing MPs, we have applied the novel approach of dynamic light scattering (DLS) to characterize the size distributions of these MPs within FFP.

Microvesicles in haemoglobinopathies offer insights into mechanisms of hypercoagulability, haemolysis and the effects of therapy.

Levels of circulating red blood cell (RBC)‐derived vesicles are increased in sickle cell anaemia (SCA) and thalassaemia intermedia (TI) but the mechanisms, effects and controlling factors may differ. This study found that levels of vesicles and intravascular haemolysis were linked as shown by the correlation between levels of vesicles and plasma Hb. Vesicle levels were 6‐fold greater in SCA and 4‐fold greater in TI than in controls. The proportion of plasma Hb within vesicles was increased in SCA and TI with a significantly higher proportion in TI. We examined whether subpopulations of RBC expressing phosphatidylserine (PS) were a source of PS(+) vesicles and observed a significant association. Thrombin generation was promoted by the vesicles in which 40–50% expressed PS. In TI, markers of thrombin generation were significantly related to PS(+) RBC. Splenectomy in TI had significant effects including greater increases in vesicle levels, plasma Hb, PS(+) RBCs and thrombin generation markers than in unsplenectomised patients. In hydroxycarbamide (HC)‐treated SCA patients these measures were decreased compared with untreated controls. The relationship between vesicle levels and plasma Hb suggests a mechanism linking vesiculation to haemolysis and consequently nitric oxide (NO) bioavailability and suggests a means by which HC treatment improves NO bioavailability.

Platelet degranulation and monocyte-platelet complex formation are increased in the acute and convalescent phases after ischaemic stroke or transient ischaemic attack.

Flow cytometric studies suggest that platelets are activated in ischaemic stroke or transient ischaemic attack (TIA). However, few studies have measured circulating leucocyte–platelet complexes in this patient population. Whole blood flow cytometry was used to quantify the expression of CD62P‐, CD63‐, and PAC1‐binding, and the percentages of leucocyte–platelet complexes in acute (1–27 d, n = 79) and convalescent (79–725 d, n = 70) ischaemic cerebrovascular disease (CVD) patients compared with controls without CVD (n = 27). We performed a full blood count, and measured plasma levels of soluble P‐selectin, soluble E‐selectin, and von Willebrand factor antigen (VWF:Ag) as additional markers of platelet and/or endothelial cell activation. The median percentage CD62P expression and the median percentage monocyte–platelet complexes were higher in both acute and convalescent CVD patients than controls (P ≤ 0·02). The mean white cell count and mean VWF:Ag levels were significantly elevated in the acute and convalescent phases after ischaemic stroke or TIA (P ≤ 0·02). Otherwise, there was no significant increase in any other marker of platelet or endothelial activation in CVD patients. There was a positive correlation between the percentage expression of CD62P and the percentages of both neutrophil–platelet and monocyte–platelet complexes in the acute phase, and the percentages of all leucocyte–platelet complexes in the convalescent phase after ischaemic CVD. This study provides evidence for ongoing excessive platelet and/or endothelial activation in ischaemic CVD patients despite treatment with antithrombotic therapy.