Mild peripheral neuropathy prevents both leg muscular ischaemia and activation of exercise-induced coagulation in Type 2 diabetic patients with peripheral artery disease.

AIM: To study the influence of peripheral neuropathy on intermittent claudication in patients with Type 2 diabetes (T2DM). METHODS: Twenty-five patients with T2DM were grouped according to the ankle/brachial index (ABI): 10 with ABI > 0.9 without peripheral artery disease (PAD; group T2DM) and 15 with ABI < 0.9 with PAD (group T2DM + PAD). Twelve individuals without T2DM with PAD (group PAD without T2DM) were also enrolled. Tests for peripheral neuropathy were performed in all patients. ABI, rate pressure product, prothrombin fragments 1 + 2 (F1+2), thrombin-anti-thrombin complex (TAT), and d-dimer were measured before and after a treadmill test. During exercise both initial and absolute claudication distance and electrocardiogram readings were recorded. RESULTS: We found mild peripheral neuropathy in 20% of group T2DM and 46.7% of group T2DM + PAD (P < 0.01). After exercise, the rate pressure product increased in each group; ABI fell in T2DM + PAD (P < 0.0001) and in PAD without T2DM (P = 0.0005); the fall was greater in the latter group. Initial and absolute claudication distances were similar in PAD patients. In group T2DM + PAD, absolute claudication distance was longer in the subgroup without peripheral neuropathy (P < 0.05), whereas ABI and rate pressure products were similar. F1+2 values at rest were higher in group T2DM + PAD. After exercise, F1+2 values and TAT increased only in group PAD without T2DM. CONCLUSION: Only group PAD without T2DM experienced muscular ischaemia, whereas group T2DM + PAD did not. Mild peripheral neuropathy may have prevented them from reaching the point of muscular ischaemia during the treadmill test, because they stopped exercising with the early onset of pain. Reaching a false absolute claudication distance may induce ischaemic preconditioning. These findings suggest a possible protective role of mild peripheral neuropathy in T2DM patients with intermittent claudication, by preventing further activation of coagulation during treadmill testing.

Homozygous patients with the 20210 G to A prothrombin polymorphism remain often asymptomatic in spite of the presence of associated risk factors.

Patients who are homozygous for the G to A nontranslated prothrombin polymorphism only occasionally have venous thrombosis. An evaluation of all published papers on the subjects has disclosed that nine patients of the 35 so far reported remained asymptomatic in spite of the presence of associated congenital or acquired thrombotic risk factors. We saw an additional patient recently, bringing the total to 10 of 36 patients. Some of these patients remained asymptomatic in spite of multiple or repetitive risk factors (e.g., five pregnancies in the case of one patient). Twelve patients who were homozygous and who had this polymorphism developed symptoms only in the presence of the same risk factors. This may suggest that this abnormality played a small role, if any, in both groups of patients. The finding that several patients with this abnormality remained asymptomatic in spite of associated risk factors casts serious doubts about the prothrombotic significance of this polymorphism. Until this problem is clarified, the clinician must abstain from attributing a prothrombotic effect to this polymorphism.

Symptomatic combined homozygous factor XII deficiency and heterozygous factor V Leiden. luscaber@tin.it.

A family with a combined deficiency of factor XII and factor V Leiden is presented. The proposita is a 72-year-old who showed a mild to moderate thrombotic tendency characterized by two episodes of deep venous thrombosis and superficial phlebitis between the age of 50 and 71. She was shown to be carrier of homozygous factor XII deficiency and heterozygous FV Leiden mutation. A sister of the proposita showed the same pattern but remained asymptomatic. Other family members showed either isolated heterozygous factor XII deficiency or combined heterozygous factor XII deficiency and heterozygous FV Leiden mutation but were all asymptomatic. These data lend support to those who maintain that FV Leiden is a mild genetic determinant for thrombosis. The role of FXII deficiency as an additional risk factor remains questionable.

Serological markers of autoimmunity in patients with hemophilia A: the role of hepatitis C virus infection, alpha-interferon and factor VIII treatment in skewing the immune system toward autoreactivity.

The aim of this study was to investigate whether the immune system of patients with hemophilia A is skewed toward an aspecific activation and to identify the causative factors. It is well known that an immune derangement does exist in patients with hemophilia A. At least three factors potentially play a role: hepatitis C virus (HCV) infection, alpha-interferon therapy and the administration of factor VIII (FVIII). Sixty human immunodeficiency virus (HIV)-negative patients with severe or moderate hemophilia A were studied retrospectively. The serological markers of autoimmunity were evaluated and the results correlated with anti-HCV antibodies, FVIII treatment and alpha-interferon therapy. The role of these factors in the development of the anti-FVIII antibody was estimated concomitantly. The prevalence of autoantibodies and anti-FVIII antibodies was higher in HCV-positive than in HCV-negative patients before any treatment, although the difference was not statistically significant. The administration of FVIII further influenced the development of autoantibodies both in HCV-negative and HCV-positive patients, with no difference being observed between the two groups. As expected, fewer HCV-negative than HCV-positive patients developed anti-FVIII antibodies after administration of FVIII (31.8% versus 38%, respectively). Therapy with alpha-interferon did not seem to enhance significantly the risk of developing autoantibodies nor anti-FVIII antibodies. We observed a high prevalence of humoral signs of autoimmunity among patients with hemophilia A. Treatment with FVIII concentrate is probably the most important triggering factor. Monitoring these patients for autoimmune manifestations is recommended.

Failure of soluble fibrin polymers in the diagnosis of clinically suspected deep venous thrombosis.

A new diagnostic test has recently become available which is highly specific for plasma soluble fibrin polymers, the thrombus precursor protein (TpP) enzyme immunoassay. In order to evaluate the diagnostic accuracy of this test and that of a new rapid and automated test for the determination of D-dimers, the BC D-dimer test, in patients with clinically suspected deep vein thrombosis (DVT), 70 consecutive symptomatic patients underwent laboratory analysis with both tests and with the classic enzyme-linked immunosorbent assay (ELISA) D-dimer test, followed by the execution of a compression ultrasound (CUS) test of the affected limb. Patients with a positive CUS test were considered to have DVT (20 of 70), whereas those with a serially negative test and an uneventful 3-month follow-up test were regarded as not having DVT (50 of 70). The sensitivity of the TpP test (45.0%) was significantly lower than that of both the BC D-dimer test (80.0%; P = 0.02) and the classic ELISA test (90.0%; P = 0.002). The specificity of the TpP test (66.0%) did not differ from that of either D-dimer test (60.0 and 64.0%, respectively). The negative predictive value of the TpP test (75.0%) was significantly lower than that of the classic ELISA D-dimer test (94.1%; P = 0.02), which in turn did not differ from that of the BC D-dimer test (88.2%). The positive predictive value was similarly low for each investigated test (34.6, 44.4, and 50.0%, respectively). In conclusion, the TpP test can neither be used to detect a DVT nor to exclude its development in patients with the clinical suspicion of this disease. By contrast, the BC D-dimer might safely replace the classic ELISA test for excluding DVT in symptomatic patients.

Prothrombin and the prothrombin 20210 G to A polymorphism: their relationship with hypercoagulability and thrombosis.

Polymorphisms of several clotting factors have been associated during the past few years with an increased risk of both venous or arterial thrombosis. However, final proof for the existence of a pathogenetic relationship between a given polymorphism and an increased risk for thrombosis is still lacking. Particular emphasis has been placed recently on a 20210 G to A prothrombin polymorphism. A critical review of available data indicates that such an abnormality may be associated with an increased risk of venous thrombosis but not arterial thrombosis (with a possible exception for myocardial infarction). However, this conclusion is based only on retrospective cohort studies which compared the prevalence of the abnormality in a group of patients with past venous or arterial thrombosis with a normal group (with no thrombosis). No prospective study has yet to show that patients with the abnormality, given similar additional acquired risk factors, have a higher incidence of thrombotic complications as compared with controls. The mechanism whereby the abnormality might cause thrombosis has been assumed to be an increase in prothrombin levels. Since an association between two phenomena does not necessarily mean that a causal relationship exists between the same events, it is important to be cautious before claiming that such abnormality is responsible for thrombosis. Therefore, although included commonly in the investigation profile, the search for the 20210 G to A prothrombin abnormality should not be considered yet to be an essential component in the routine study of hypercoagulable and/or thrombotic conditions.

Congenital deficiencies and abnormalities of prothrombin.

Prothrombin (factor II) deficiency was first described in 1947 by Quick et al., although the first prothrombin abnormality was reported in 1969 by Shapiro et al. The condition is still considered very rare. In spite of its rarity, the defect has allowed important improvements in our understanding of both congenital and acquired prothrombin deficiencies. The diagnosis of prothrombin deficiency or abnormality can be made using a combination of clotting, chromogenic and immunological assays. In cases of true deficiency, a parallel decrease in all these assays is observed, regardless of the activating agent. If discrepancies among the clotting assays are noted, particularly using viper venoms, a dysprothrombinemia should be suspected. Usually, activity levels less than 10% of normal are found in homozygotes, and between 40 and 60% in heterozygotes. Factor II levels in congenital dysprothrombinemias are more variable since one may encounter homozygotes, heterozygotes and compound heterozygotes between a heterozygous abnormality and heterozygous 'true' deficiency or between two distinct abnormalities. Usually the levels of factor II vary between 1 and 50% of normal. Antigen levels in congenital dysprothrombinemias will be normal, near normal or slightly decreased but always higher than the clotting counterpart. Cases with a parallel decrease in prothrombin activity and antigen should not be considered as examples of hypoprothrombinemia. The gene involved in the synthesis of prothrombin is located in chromosome 11. It is composed of 10 exons and 8 introns. Molecular biology studies have discovered several point mutations in some of the dysprothrombinemias. Bleeding manifestations may be severe in homozygous 'true' deficiency and may be more variable in dysprothrombinemias. Heterozygotes are usually asymptomatic. Prognosis is variable and generally in agreement with the prothrombin activity level. In homozygous true deficiency, hemarthroses and intracranial bleeding have been described. Substitution therapy is based on the administration of prothrombin complex concentrates or of plasma. The long half-life of prothrombin injected, about 70 h, allows the achievement of hemostatically effective levels (about 50% of normal) without difficulty.

D-dimer testing as an adjunct to ultrasonography in patients with clinically suspected deep vein thrombosis: prospective cohort study. The Multicentre Italian D-dimer Ultrasound Study Investigators Group.

OBJECTIVE: To investigate the efficacy of using a rapid plasma D-dimer test as an adjunct to compression ultrasound for diagnosing clinically suspected deep vein thrombosis. DESIGN: D-dimer concentrations were determined in all patients with a normal ultrasonogram at presentation. Repeat ultrasonography was performed 1 week later only in patients with abnormal D-dimer test results. MAIN OUTCOME AND MEASURES: Patients with normal ultrasonograms were not treated with anticoagulants and were followed for 3 months for thromboembolic complications. SETTING: University research and affiliated centres. SUBJECTS: 946 patients with clinically suspected deep vein thrombosis. RESULTS: Ultrasonograms were abnormal at presentation in 260 (27.5%) patients. Of the remaining 686 patients tested for D-dimer, 88 (12.8%) had abnormal concentrations. During follow up venous thromboembolic complications occurred in one of the 598 patients who were not treated with anticoagulants and who had an initial normal ultrasonogram and D-dimer concentration, whereas thromboembolic complications occurred in two of the 83 untreated patients who had abnormal D-dimer concentrations but a normal repeat ultrasonogram. The cumulative incidence of venous thromboembolic complications during follow up was 0.4% (95% confidence interval 0% to 0.9%). The rapid plasma D-dimer test used as an adjunct to compression ultrasonography resulted in a reduction in the mean number of repeat ultrasound examinations and additional hospital visits from 0.7 to 0.1 per patient. CONCLUSIONS: Testing for D-dimer as an adjunct to a normal baseline ultrasound examination decreased the number of subsequent ultrasound examinations considerably without any increased risk of venous thromboembolic complications in patients not receiving anticoagulants. The use of ultrasound and testing for D-dimer enabled treatment decisions to be made at the time of presentation in most patients.

Use of an algorithm for administering subcutaneous heparin in the treatment of deep venous thrombosis.

BACKGROUND: Despite the widespread use of subcutaneous heparin in the initial treatment of deep venous thrombosis, there are no guidelines for achieving adequate anticoagulation with this drug. OBJECTIVE: To implement a weight-based algorithm for the administration of subcutaneous unfractionated heparin after an intravenous loading dose. DESIGN: Prospective cohort study. SETTING: University hospital. PARTICIPANTS: 70 outpatients with proximal venous thrombosis. INTERVENTION: An intravenous bolus of heparin followed by a subcutaneous injection of heparin in doses adjusted for body weight. Subsequent adjustments of the subcutaneous heparin dose were scheduled twice daily according to the algorithm; the activated partial thromboplastin time (aPTT) was measured in the mid-interval (target range, 50 to 90 seconds). RESULTS: The therapeutic threshold aPTT (> or = 50 seconds) was achieved in 61 patients (87%) within 24 hours and in 69 patients (99%) within 48 hours. In 7 patients (10%), a supratherapeutic aPTT lasted more than 12 hours. No major bleeding episodes or cases of heparin-induced thrombocytopenia were seen. Three patients (4.3% [95% CI, 0.9% to 12.0%]) had recurrent thromboembolism during 3 months of follow-up. CONCLUSION: The administration of subcutaneous heparin according to a weight-based algorithm allows the rapid achievement of effective and safe anticoagulation in patients with deep venous thrombosis.

Hemorrhagic and thrombotic disorders due to factor V deficiencies and abnormalities: an updated classification.

The recent description of a factor V abnormality (factor V Leiden) associated with an increased incidence of thrombosis has considerably increased interest in this clotting factor. The discovery of this new clinical entity indicated the need for an updated classification of factor V defects. These should be divided into hemorrhagic and thrombotic disorders. A proper classification of hemorrhagic disorders should include: 1) homozygous and heterozygous 'true' factor V deficiency; and 2) combined factor V and factor VIII deficiencies. The latter should be subdivided in Type I (association type) and Type II (common defect). A suitable classification of the thrombotic factor V defects should include: 1) homozygous and heterozygous factor V Leiden; and 2) combined heterozygous factor V Leiden and heterozygous 'true' factor V deficiency. The presence of thrombosis in these latter patients, often as severe as those seen in homozygous patients with activated protein C (APC) resistance, allows important considerations on the functions of factor V. It would seem that half the normal level of factor V activity and antigen is unable to protect against thrombosis in patients with heterozygous APC resistance. An accurate evaluation of factor V activity and antigen is indicated in all patients with suspected factor V defects. The first suspicion may be obtained by the presence of a mild prolongation of prothrombin time and of partial thromboplastin time. The suspicion should then be immediately confirmed by specific factor V activity and antigen assays. This approach is of great importance even for the presumptive diagnosis of pseudohomozygosis for APC resistance. In fact, in these cases, factor V activity is about 50% of normal, whereas factor V antigen is 100% of normal. In heterozygous 'true' factor V deficiency both activity and antigen are about 50% of normal.

Accuracy of two newly described D-dimer tests in patients with suspected deep venous thrombosis.

Accumulating evidence suggests that the ELISA determination of D-Dimer might be a useful tool for the exclusion of deep vein thrombosis (DVT) of lower extremities, because of its high sensitivity and negative predictive value. However, conventional ELISA assay is time-consuming and, therefore, is not suitable for emergency use. To evaluate the accuracy of two rapid assays recently described, 126 consecutive outpatients with the clinical suspicion of DVT underwent the NycoCard D-Dimer and the Instant I.A. D-Dimer determination, using venography as the reference test. In all patients, the conventional ELISA assay was also performed. Venography confirmed the presence of DVT in 30 patients (23.8%), and ruled out the diagnosis in the remaining 96. Instant I.A D-Dimer was positive in 28 patients with DVT (sensitivity, 93.3%), and negative in 90 subjects free from thrombosis (specificity, 93.8%). Nycocard D-Dimer correctly identified 27 patients with DVT (sensitivity 90.0%), and was negative in 77 subjects free from thrombosis (specificity, 80.2%). Sensitivity, specificity, negative and positive predictive values of both tests did not differ from those found with the classic ELISA method. In conclusion, both Instant I.A. D-Dimer and Nycocard D-Dimer assays show a great potential for clinical use.

Venous and arterial thrombophilia.

BACKGROUND AND OBJECTIVE: The thrombophilic state may be defined as a condition which predisposes to thrombosis. It is well know that the pathogenesis of venous and arterial thrombosis may be different. However, such differences may be more apparent than real. In fact, both venous and arterial thrombosis may occur in any given patient with a thrombophilic state. The aim of this review is to critically analyze the thrombophilic state and define a rational approach to the patient with overt or suspected venous and/or arterial thrombosis. EVIDENCE AND INFORMATION SOURCES: The material examined in the present review includes personal papers in this field, and articles and abstracts published in journals covered by the Science Citation Index. STATE OF ART AND PERSPECTIVES: Both venous and arterial thrombosis may occur in thrombophilic states such as APC resistance, protein C or S defects, and antiphospholipid antibody syndrome. Venous thrombosis is surely more frequent than arterial thrombosis in such conditions but, fortunately, it is usually less severe. Antithrombin deficiency is almost exclusively associated with venous thrombosis. In foreseeing the occurrence of venous or arterial thrombosis in a given thrombophilic patient, one must explore the state of the vessels carefully. Often venous or arterial thrombosis occurs only because a vessel injury is present. Severely decreased blood flow, such as that seen in policythemia vera, may be responsible for arterial or venous thrombosis without any other predisposing cause. From a laboratory stand point there is no sure demonstration that some changes may indicate a more likely occurrence of arterial or venous thrombosis. The same alteration of one or more than one test may be accompanied by either arterial or venous thrombosis or both. One exception to this rule is represented by increased blood viscosity, which is usually associated with arterial thrombosis. The hypercoagulable or thrombophilic state is a single clinical entity that cannot be divided into arterial and venous thrombophilia, although the unfortunate outcome, namely thrombosis, tends to manifest itself in just one district. The preexisting condition of the vessels, together with sudden triggering factors, plays an important role in the transformation of the "sol" into the "gel" that is a thrombosis in any given district.

Deep venous thrombosis and lupus anticoagulant. A case-control study.

BACKGROUND: A definite evidence in favour of an association of deep-vein thrombosis (DVT) with lupus anticoagulant (LA) in patients free from systemic lupus erythematosus is still lacking. METHODS: In a case-control study, LA was determined in 176 consecutive outpatients who underwent phlebography because of the first episode of clinically suspected DVT of lower limbs. The association between DVT and LA was described using odds ratios (OR). RESULTS: Contrast venography confirmed the clinical suspicion in 59 patients (33.5%). LA was detected in 5 of the 59 patients with DVT (8.5%), and in none of the 117 subjects with normal venogram (P = 0.007). The OR for having an acute DVT in patients with LA was 10.7 (95% CI: 1.2-94.2). CONCLUSIONS: LA is significantly associated with DVT in symptomatic patients. Further studies are needed to establish the clinical implications of this association.

Prothrombin fragment 1+2 and thrombin-antithrombin complex levels in patients with inherited APC resistance due to factor V Leiden mutation.

Inherited activated protein C (APC) resistance is a newly described pathological condition associated with familial thrombophilia. A recent report on family with APC resistance showed increased levels of prothrombin fragment 1 + 2 (F1 + 2) in the affected individuals. No data concerning thrombin-antithrombin complex (TAT) levels in patients with inherited APC resistance are presently available. The aim of this study was to assess the plasma levels of F1 + 2 and TAT in patients with inherited APC resistance due to factor V (F.V) Leiden mutation and to evaluate F1 + 2 and TAT levels in symptomatic and asymptomatic patients with the defect ('carriers') as compared to their family members having no evidence of F.V Leiden mutation ('non-carriers'). One hundred and twenty-nine individuals belonging to 30 families with inherited APC resistance due to F.V Leiden mutation were studied. F1 + 2 and TAT levels were determined using two commercially available ELISA kits and cut-off values were defined as the higher limits of normal ranges obtained in healthy volunteers. Out of the 129 family members investigated, 36 were non-carriers, 85 were heterozygous and eight homozygous for F.V Leiden mutation. Thrombosis had occurred in 2/36 (6%) non-carriers, in 36/85 (42.3%) heterozygous and in 5/8 (63%) homozygous. Median levels of F1 + 2 and TAT were above cutoff values in carriers, whereas they were below in non-carriers. An overall percentage of 68.8% of carriers exhibited F1 +2 levels above the cut-off value as compared to 38.9% of non-carriers. For TAT, an overall percentage of 63.4% of carriers presented with levels above the cut-off compared with 28% of non-carriers. In conclusion, patients with inherited F.V Leiden mutation may exhibit increased levels of F1 + 2 and TAT. There are no differences in F1 + 2 and TAT median levels among symptomatic and asymptomatic carriers. The percentage of carriers of F.V Leiden with levels of F1 + 2 and TAT above cut-off appears to be higher than that found in other clotting inhibitors defects and in this respect the defect might be considered different. However, these findings and the presence of high percentage of non-carriers presenting with increased F1 + 2 and TAT levels may suggest the possible coexistence in these families of other unknown defects predisposing to thrombosis.

Recombinant thromboplastin inhibition assay for the detection of lupus anticoagulant.

Tissue thromboplastin inhibition assay (TTI) is a sensitive test for lupus anticoagulant (LA). We have performed TTI in 12 LA positive patients using a new recombinant human tissue factor (Innovin, IN) and compared it with Thromborel S (TH), a commonly used human placenta thromboplastin. The effect of using two different dilutions of each thromboplastin (1:10 & 1:26) was investigated. A 1:26 dilution discriminated better than the 1:10 and this was more evident for Innovin. The mean value obtained with a 1:26 IN dilution was statistically different from that observed with TH at the same dilution. Furthermore, when PT and TTI ratios were considered, differences were statistically significant and seemed to increase depending on thromboplastin dilutions. When we used IN at 1:26 all LA positive patients had a value > 1.2. Then we compared TTI at a 1:26 dilution with dilute Russell's Viper Venom Time (dRVVT) in 50 consecutive patients with suspected lupus anticoagulant not treated with warfarin or heparin. In these patients the diagnosis of lupus anticoagulant was carried out using dilute APTT mixing studies and a platelet neutralization procedure: four out of 50 patients thus tested were LA positive. When dRVVT or TTI-I 1:26 were used, five patients were positive for lupus anticoagulant. Innovin showed similar sensitivity of dRVVT for detection of lupus anticoagulant. It is likely that higher dilutions of thromboplastins could further improve the specificity of this method.