Procoagulant mutations and venous thrombosis in Behcet's disease

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(1)Letters to the Editor. 1298. Procoagulant mutations and venous thrombosis in Behçet’s disease S, Behçet’s disease (BD) is a vasculitis of unknown aetiology characterized by involvement of both arteries and veins and vessels of all sizes [1]. Thrombosis of superficial and deep veins is more frequent than arterial aneurysms and thrombotic occlusions. Deep vein thrombosis of the lower extremities is seen in about one-fifth of Turkish patients with BD [2]. Thrombotic occlusions of inferior and superior venae cavae, hepatic veins and cerebral sinuses can also be observed, albeit less frequently. We previously reported that coagulation factor V gene G1691A mutation (factor V Leiden), the most common inherited coagulation defect associated with venous thrombosis, could be detected in 37.5% of BD patients with a history of deep vein thrombosis in a case–control study [3]. The association of factor V Leiden with venous thrombosis in BD was later confirmed in different ethnic groups [4, 5]. Herein, we present our results of screening the same study group for the presence of a new mutation identified in the 3∞-untranslated region of the prothrombin gene (20210 GA). This mutation is associated with elevated plasma prothrombin levels and a 3-fold increased risk of venous thrombosis [6 ], and it has been reported as the second most common inherited hypercoagulable state [7]. The prevalence of heterozygous 20210A carriers varied from 1 to 5% in healthy Caucasians and was found to be 2.2–2.7% in healthy Turkish controls [7–9]. The study group consisted of 64 patients, 32 with a history of deep vein thrombosis ( T + ) of the lower extremities, and 32 age- and sex-matched patients without any thrombotic events ( T − ) during a minimum of 8 yr disease duration. All patients fulfilled three or more International Study Group (ISG) criteria for BD [2]. In the T+ group, three patients also had inferior vena cava, two had superior vena cava and three had sagittal sinus thromboses. Arterial involvement in pulmonary, brachial, iliac and femoral arteries was present in four patients. Clinical details of the patients were given previously [3]. The G20210A mutation in the 3∞ untranslated region of the prothrombin gene was detected by allele-specific polymerase chain reaction (PCR) as described previously [10]. Briefly, it was demonstrated by using a. forward consensus primer and two reverse primers, one of which is specific for the wild type and the other for the mutated alleles. The 340 bp PCR products were visualized on ethidium bromide-stained agarose gels. A second amplification was performed to confirm the presence of the mutation. We compared the frequencies of the prothrombin and/or factor V gene mutations in the T+ and T − groups by x2 test, and calculated the odds ratio (OR) for estimation of thrombosis risk in the presence of procoagulant mutations. Heterozygous prothrombin gene G20210A mutation was found in 10 (nine male, one female) of the T+ (31.3%) and in one (male) of the T − group (3.1%) (P = 0.003, OR = 14.1, 95% CI 1.7–118.2; Table 1). One of these 10 patients had sagittal sinus thrombosis, one had sagittal sinus + inferior vena cava and one superior vena cava thrombosis in addition to thrombosis of the lower extremities. The prothrombin gene mutation was found in two male patients, one with femoral + popliteal artery thrombosis and the other with brachial artery aneurysm + thrombosis. Four patients (three male, one female) in the T+ group were carrying both factor V Leiden and prothrombin gene G20210A mutations, and one of them was the patient with femoral + popliteal artery thrombosis. A total of 18 patients (56.3%) with BD and deep vein thrombosis were carrying factor V Leiden and/or the prothrombin gene G20210A mutation compared with only four of the BD patients without any thrombotic event (P = 0.0002, OR = 9, 95% CI 2.6–31.7). The detection of the two most frequent hereditary prothrombotic mutations known in more than half of BD patients with deep vein thrombosis supports the important role of procoagulant mutations in the development of venous thrombosis in BD. We previously suggested that the tendency to thrombosis in BD could be explained by endothelial activation induced by immune-mediated vasculitis. Endothelial changes may trigger venous (and arterial ) thrombosis in patients carrying factor V Leiden, the prothrombin G20210A mutation, or other yet unidentified mutations of the coagulation system, if the inflammatory response is strong enough. Cerebral vein and arterial thrombosis has been reported in individuals heterozygous for the prothrombin gene mutation [11, 12]. We identified two patients with sagittal sinus thrombosis, one of whom. T 1. Prevalence of the factor V Leiden and prothrombin gene G20210A mutations in Behçet’s disease patients with ( T + ) and without (T − ) deep vein thrombosis Factor V Leiden (%). T+ T−. Prothrombin gene mutation (%). n. Sex (M/F ). G,G. G,A*. G,G. G,A†. 32 32. 26/6 26/6. 20 (62.5) 29 (90.6). 12 (37.5)a 3 (9.4)b. 22 (68.7) 31 (96.9). 10 (31.3)c 1 (3.1)d. *G,A, heterozygous for the factor V gene G1691A mutation (factor V Leiden); †G,A, heterozygous for the prothrombin gene G20210A mutation; G,G, no mutation. a vs b, P = 0.008; c vs d, P = 0.003..

(2) Letters to the Editor. also had inferior vena cava thrombosis, and two males with arterial thrombosis carrying the prothrombin gene mutation in this series of patients with BD. Further studies are needed to show any predilection for the site of thrombosis in patients carrying a particular procoagulant mutation or a combination of mutations in larger series of patients. A. G, A. B. A1, T. T1, M. K, ¨ 1 T. O Division of Rheumatology, Department of Internal Medicine, Istanbul School of Medicine, Istanbul University, Istanbul and 1Department of Molecular Biology and Genetics, Faculty of Science, Bilkent University, Ankara, Turkey Accepted 21 June 1999 Correspondence to: A. Gu¨l, Division of Rheumatology, Department of Internal Medicine, Istanbul School of Medicine, Capa 34390, Istanbul, Turkey. 1. Lie JT. Vascular involvement in Behçet’s disease: Arterial and venous and vessels of all sizes. J Rheumatol 1992;19:341–3. 2. International Study Group for Behçet’s Disease. Evaluation of diagnostic (‘Classification’) criteria in Behçet’s disease—towards internationally agreed criteria. Br J Rheumatol 1992;31:299–308. ¨ zbek U, O ¨ ztu¨rk C, Inanç M, Koniçe M, Ozçelik T. 3. Gu¨l A, O Coagulation factor V gene mutation increases the risk of venous thrombosis in Behçet’s disease. Br J Rheumatol 1996;35:1178–80. 4. Mammo L, Al-Dalaan A, Bahabri SS, Saour JN. Association of factor V Leiden with Behçet’s disease. J Rheumatol 1997;24:2196–8. 5. Verity DH, Vaughan RW, Madanat W et al. Factor V Leiden mutation is associated with ocular involvement in Behçet disease. Am J Ophthalmol 1999;128:352–6. 6. Poort RS, Rosendaal FR, Reitsma PH, Bertina RM. A common genetic variation in 3∞-untranslated region of prothrombin gene is associated with elevated plasma prothrombin levels and an increase in venous thrombosis. Blood 1996;88:3698–703. 7. Bertina RM. The prothrombin 20210 G to A variation and thrombosis. Curr Opin Hematol 1998;5:339–42. 8. Akar N, Misirlioglu M, Akar E, Avcu F, Yalcin A, So¨zuö¨z A. Prothrombin gene 20210 G-A mutation in the Turkish population. Am J Hematol 1998;58:249. 9. Gu¨rgey A, Hicso¨nmez G, Parlak H, Balta G, Celiker A. Prothrombin gene 20210 G-A mutation in Turkish patients with thrombosis. Am J Hematol 1998;59:179–80. 10. He´zard N, Cornillet-Lefebvre P, Gillot L, Potron G, Nguyen P. Multiplex ASA PCR for a simultaneous determination of factor V Leiden gene, GA 20210 prothrombin gene and CT 677 MTHFR gene mutations. Thromb Haemostasis 1998;79:1054–5. 11. Arruda VR, Annichino-Bizzacchi JM, Gonçalves MS, Costa FF. Prevalence of the prothrombin gene variant (nt20210A) in venous thrombosis and arterial disease. Thromb Haemostasis 1997; 78:1430–3. 12. Martinelli I, Sacchi E, Landi G, Taioli E, Duca F, Mannucci PM. High risk of cerebral vein thrombosis in carriers of a prothrombingene mutation and in users of oral contraceptives. N Engl J Med 1998;338:1793–7.. Coronary dissection associated with hepatitis C virusrelated cryoglobulinaemia S, Mixed cryoglobulinaemia is a multisystemic disease, characterized by chronic angiitis, whose main symptoms are purpura, arthritis, peripheral neuropathy and glom-. 1299. erulonephritis. Cardiac involvement is extremely rare. Since the discovery of the relationship of hepatitis C virus (HCV ) to mixed cryoglobulinaemia, >90% of type 2 and type 3 cryoglobulinaemias have been attributed to this infectious agent [1–3]. Accordingly, interferon alpha (IFN-a) therapy has been used with promising results [4, 5]. Although the association between HCV-related cryoglobulinaemia and ischaemic disease, including coronary ischaemic disease, has been reported previously [6–9], our case is, to our knowledge, the first report of HCV-related cryoglobulinaemia with symptomatic coronary dissection. A 27-yr-old female was admitted to our hospital with prolonged chest pain and a 5 yr history of intermittent arthralgia in the knees and hands. Pulmonary and cardiac auscultation were unremarkable. Slight hepatomegaly and splenomegaly were noted. Non-pruritic purpura was seen in the lower limbs with severe involvement around the malleoli (Fig. 1, upper right). The purpura developed 24 h prior to the chest pain. ECG showed a typical pattern of an acute lateral myocardial infarction. Laboratory findings on admission were haematocrit 30%, platelet count 900 000 mm3, ASAT 174 U/l (normal: 5–40), ALAT 376 U/l (normal: 5–40), LDH 250 U/l (normal: 60–225), creatine clearance 55 ml/min. The creatinine kinase (CK ) peak 6 h after the onset of the chest pain was 700 U/l (normal: 40–150). Immunological studies showed normal levels of IgG and IgA, and elevated levels of IgM (600 mg/dl; normal: 50–200). Complement study showed diminished levels of C3 (62 mg/dl; normal: 70–120) and C4 (4 mg/dl; normal: 15–25) and elevated levels of C3d (26 U/ml; normal: 0–20). Antinuclear antibodies and antineutrophil cytoplasmic antibodies were negative. Rheumatoid factor was positive (625 IU/ml; normal: 0–60) and antiHCV antibodies ( ELISA and immunoblot) were found. Cryoglobulins were discovered and immunoelectrophoresis confirmed a mixed type 2 cryoglobulinaemia (polyclonal IgG and monoclonal IgM kappa). HCV RNA was detected by polymerase chain reaction in both the cryoprecipitate and the serum. Skin biopsy of purpuric lesions confirmed the presence of dermal vasculitis, with endothelial damage, extravasation of red blood corpuscles and leucocytoclasis (Fig. 1, bottom right). Immunofluorescence of the skin biopsy revealed the presence of immune reactants (IgM, IgG and C3) in the endothelium. The hepatic biopsy was consistent with chronic active hepatitis. Coronary angiography revealed an image of coronary dissection in the left anterior descending and first diagonal branch ( Fig. 1, upper left). Treatment with steroids, anticoagulants and nitrates was started. The possibility of plasmapheresis was considered, but eventually rejected due to the cardiac instability. Subcutaneous IFN-a was given; the treatment included 1 month at a dose of 2 million U/24 h and 6 months at 2 million U/48 h, and following 36 months of follow-up the patient is still on IFN-a at a dose of 2 million U/96 h. Three months after IFN-a was started,.

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