Recurrent Thrombotic Events after Discontinuation of Vitamin K Antagonist Treatment for Splanchnic Vein Thrombosis: A Multicenter Retrospective Cohort Study.

It is generally recommended that patients with splanchnic vein thrombosis (SVT) should receive a minimum of 3 months of anticoagulant treatment. However, little information is available on the long-term risk of recurrent thrombotic events. The aim of this study was to evaluate the risk of venous and arterial thrombosis after discontinuation of vitamin K antagonist (VKA) in SVT patients. Retrospective information from a cohort of SVT patients treated with VKA and followed by 37 Italian Anticoagulation Clinics, up to June 2013, was collected. Only patients who discontinued VKA and did not receive any other anticoagulant drug were enrolled in this study. Thrombotic events during follow-up were centrally adjudicated. Ninety patients were included: 33 unprovoked SVT, 27 SVT secondary to transient risk factors, and 30 with permanent risk factors. During a median follow-up of 1.6 years, 6 venous and 1 arterial thrombosis were documented, for an incidence of 3.3/100 patient-years (pt-y). The recurrence rate was highest in the first year after VKA discontinuation (8.2/100'pt-y) and in patients with permanent risk factors (10.2/100'pt-y). Liver cirrhosis significantly increased the risk of recurrence. In conclusion, the rate of recurrent vascular complications after SVT is not negligible, at least in some patient subgroups.

Clonal populations of hematopoietic cells with paroxysmal nocturnal hemoglobinuria phenotype in patients with splanchnic vein thrombosis.

INTRODUCTION: Splanchnic vein thrombosis (SVT) is a serious complication in patients with paroxysmal nocturnal hemoglobinuria (PNH). Mutant PNH clones can be associated with an increased risk of SVT even in the absence of overt disease, but their prevalence in non-selected SVT patients remains unknown. MATERIALS AND METHODS: Patients with objective diagnosis of SVT and without known PNH were tested for the presence of PNH clone using high-sensitivity flow cytometric analysis. RESULTS: A total of 202 SVT patients were eligible, 58.4% were males, mean age was 54.6years (range 17-94), site of thrombosis was portal in 103 patients, mesenteric in 67, splenic in 37, and supra-hepatic in 10. SVT was associated with JAK2 V6167F in 28 of 126 (22.2%) screened patients, liver cirrhosis in 15.3% patients, recent surgery in 10.9%, and myeloproliferative neoplasm in 10.6%, whereas in 34.6% of patients neither permanent nor transient risk factors were detected. None of the patients had a clearly demonstrable PNH clone, but in two patients (0.99%, 95% CI 0.17-3.91) we observed very small PNH clones (size 0.014% and 0.16%) confirmed in two independent samples. One patient had portal vein thrombosis and no associated risk factors, the second had superior mesenteric vein thrombosis and inflammatory bowel disease. CONCLUSIONS: Very small PNH clones can be detected in patients with SVT and no clinical manifestations of disease. Future studies are needed to explore the potential role of this finding in the pathogenesis of SVT.

In retinal vein occlusion platelet response to thrombin is increased.

INTRODUCTION: Retinal vein occlusion is a major cause of ocular morbidity. The precise mechanism leading to thrombosis in retinal vein occlusion has not yet been clearly elucidated. Several risk factors have been identified, including hypertension diabetes, history of cardiovascular disease, hypercholesterolemia, hyperhomocysteinaemia, increased ocular pressure and glaucoma. Although thrombus formation in the vein plays a significant role in the onset of retinal vein occlusion, the relationship between platelet aggregation and retinal vein occlusion remains to be clarified. MATERIALS AND METHODS: In the present study the platelet response to thrombin in a selected group of retinal vein occlusion patients was investigated. Retinal vein occlusion patients were compared to a group of healthy subjects matched for age, sex, clinical and metabolic characteristics. In resting and activated platelets of both groups of subjects total protein tyrosine phosphorylation, p38MAPK and cytosolic phospholipase A(2) phosphorylation, arachidonic acid release, intracellular calcium levels, thromboxane B(2) and superoxide anion formation were measured. RESULTS: Results show that platelets of patients were more responsive to thrombin than healthy subjects. In resting or in thrombin stimulated platelets of patients total protein tyrosine phosphorylation, p38MAPK and cytosolic phospholipase A(2) phosphorylation were increased. Also arachidonic acid release, thromboxane B(2) and superoxide anion formation were higher in patients than in healthy subjects. In addition intracellular calcium rise induced by thrombin was increased in patients. CONCLUSIONS: Altogether data suggest that platelet hyperaggregability inducing thrombus formation might be an important factor in the onset and/or development of retinal vein occlusion.

Quinolizidinyl-benzimidazoles as platelet-antiaggregating agents.

Two series of (+/-)-2-phenyl-1-(quinolizidin-1 alpha-yl)benzimidazoles, 12A-26A, and (+/-)-2-phenyl-1-(quinolizidin-1 beta-yl)benzimidazoles, 12B-26B, were prepared and tested for the inhibition of human platelets aggregation induced by ADP, collagen, and adrenaline. All epimers A, i.e., 12A-26A, were devoid of any activity against the three agonists, while the epimers B, i.e., 12B-26B, exhibited different degrees of activity, though practically confined against the ADP-induced aggregation. The best compounds were 19B, 24B, and 26B, which inhibited for 69-67% at 260 microM and for 40-29% at 65 microM concentration the ADP (2 microM)-induced aggregation. The observed agonist and spatial structure selectivity warrant further investigations of this kind of benzimidazole derivatives.

Survival of atherosclerotic patients as related to oxidative stress and gene polymorphisms.

A prospective molecular epidemiology study was implemented in a cohort of 98 subjects suffering from severe atherosclerotic lesions requiring removal of an abdominal aorta fragment. We previously published the results relative to detection, in the aorta medium layer, of bulky DNA adducts and fluorescent polycyclic aromatic hydrocarbon-related DNA adducts, oxidative DNA damage, and mitochondrial DNA 4977 common deletion, as well as GSTM1 and GSTT1 gene polymorphisms. We report herein new data, relative to oxidative stress biomarkers, including oxidative DNA damage in both inner and medium aorta layers, malondialdehyde in the medium layer, homocysteine and reduced glutathione in plasma, and those relative to additional gene polymorphisms, including NAT1, NAT2, OGG1, MTHFR, Leiden factor V, and prothrombin. The results of biochemical and molecular analyses were related to survival of the patients, whose average age was 70 at the start of the follow up. During the following 14 years, 71.4% of them died. The results obtained provide evidence for the crucial impact of oxidative stress and certain gene polymorphisms on clinical and biochemical patterns as well as on survival of patients. Survival was significantly affected not only by traditional risk factors for atherosclerosis but also by molecular end-points and adverse gene polymorphisms, and by their combinations.

Platelet activation by collagen is increased in retinal vein occlusion.

Retinal vein occlusion (RVO) is the most common retinal vascular disorder second to diabetic retinopathy. The main risk factors in patients with RVO are hypertension, diabetes, hyperlipidemia, increased blood viscosity and glaucoma. The pathogenesis of RVO has not yet been clarified. In these events platelets could play a very important role. In the present study the platelet response to collagen was deeply investigated. Experiments were carried out on a selected group of RVO patients, which were compared to a group of healthy subjects matched for age, sex, clinical and metabolic characteristics. In resting and activated platelets of both groups of subjects p72syk phosphorylation, phospholipase Cgamma2 phosphorylation, protein kinase C activation, intra-cellular calcium levels and nitric oxide formation were measured. Results show that platelets of patients were more responsive to collagen or ADP than healthy subjects and that the response was significantly different (p < 0.0005) at low concentrations of these agonists. In platelets of patients stimulated with collagen increased phosphorylation of p72syk and phospholipase Cgamma2 was found. Also protein kinase C was more activated in patients. In addition intracellular calcium rise induced by collagen was significantly higher in patients than in healthy subjects. RVO patients showed a lower basal level of nitric oxide both in resting and stimulated platelets compared to healthy subjects. Altogether these results suggest that the platelet hyperaggregability described in patients might be an important factor in the development of RVO contributing to the thrombogenic effects.

2-Phenyl-3-(quinolizidin-1-yl)-5-substituted indoles as platelet antiaggregating agents.

A set of ten 2-phenyl-3-(quinolizidin-1-yl)-5-substituted indoles was prepared through the Fischer cyclization of lupinyl- and epi-lupinylphenylketone 4-substituted phenylhydrazones. Compounds were tested for antiaggregating activity on human platelets activated by adenosine diphosphate (ADP), collagen and adrenaline. At 2.5 x 10(-4) M concentration most compounds strongly inhibited the aggregation induced by all the agonists considered and many of them still displayed good activity at 0.625 x 10(-4) M concentration. The least active (1c) and one of the most active (1d) compounds were also tested for antiaggregating activity on rabbit platelets activated by ADP, PAF and sodium arachidonate. Both the compounds were active against ADP and PAF, but only 1d inhibited the arachidonate-induced aggregation (100% at 8 x 10(-6) M concentration) and increased the bleeding time in mice. The same compounds were subjected to a general pharmacological screening and found to display several activities; of particular interest was the dose dependent reduction of serum cholesterol and heparin precipitating betalipoproteins in hypercholesterolemic mice exerted by 1c, which was still significant at the oral dose of 10 mg/kg.

4-substituted 1,5-diarylpyrazole, analogues of celecoxib: synthesis and preliminary evaluation of biological properties.

A number of 5-aryl-1-[4-(methylsulfonyl)-phenyl]-1H-pyrazoles and 4-(5-aryl-1H-pyrazol-1-yl)benzenesulfonamides 3, 4, 5, 6, analogues of the COX-2 selective inhibitor celecoxib (celebrex), were synthesized. In order to verify the effects on the biological properties of certain substituents put on position 4 of the pyrazole nucleus, some of these compounds were screened in vivo for their anti-inflammatory and analgesic activities. Moreover, sodium salts of carboxylic acids 4 were tested in vitro for their platelet anti-aggregating properties. The results of these preliminary biological assays showed that new derivatives are not endowed with improved anti-inflammatory and analgesic properties, in comparison with celecoxib. In addition, docking studies were carried out on the most significative compounds to evaluate their interaction mode at the active site of both COX-1 and COX-2. Some remarks about the SAR of this class of COX-inhibitors are drown out.