Clauss Method Research Articles

Investigation of the mechanism(s) involved in decreasing increased fibrinogen activity in hyperglycemic conditions using L-lysine supplementation

IntroductionFibrinogen is a plasma glycoprotein that participates in the hemostasis system. Its malfunction has been reported as a consequence of diabetic complications. In this study, the inhibitory effect of L-Lysine (Lys) on the nonenzymatic glycation of fibrinogen was investigated in both in vitro and in vivo conditions. Materials and MethodsFibrinogen was incubated with glucose in the presence or absence of Lys. Then, its structure was studied by fluorescence spectroscopy, circular dichroism, and electrophoresis. The Clauss method was used to determine fibrinogen activity. In addition, one of the two groups of type 2 diabetic patients receiving ordinary treatment was additionally treated with Lys for 3months. Fibrinogen activity and some other parameters were evaluated in their plasma. ResultsThe results indicated increases in the activity of glycated fibrinogen in both of the in vivo and in vitro experiments. Advanced glycation end products were increased by time, as shown using fluorometry in both the plasma of the diabetic patients and the incubation medium of protein with glucose. The circular dichroism spectra showed some changes in the fibrinogen secondary and tertiary structures after glycation. The electrophoretic mobility of the glycated fibrinogen changed and the cross-link formation between the fibrinogen subunits due to glycation was observed. Lys inhibited all of the mentioned fibrinogen changes both in the in vitro experiments and after its administration to the diabetic patients. ConclusionLys, as an inhibitor of protein glycation, improved fibrinogen's structure and function, both in vitro and in vivo.

Compound heterozygous mutations of a family with inherited hypofibrinogenemia

Objective To analyze the phenotype and genotype of a Chinese family with inherited hypofibrinogenemia, and to investigate its molecular mechanism. Methods Peripheral blood was collected from seven people of this family and then plasma was separated.Activated partial thromboplastin time (APTT), prothrombin time (PT), thrombin time (TT), reptilase time (RT), the activities of antithrombin (AT∶A), protein C (PC∶A) and protein S (PS∶A) were tested.The activity and antigen of plasma fibrinogen were analyzed by Clauss method and immunoturbidimetry method, respectively.The fibrinogen peptide chain of the proband was semiquantitatively assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE).Thrombin generation test was performed by calibrated automated thrombogram.The dynamic process of blood coagulation was evaluated by the thrombelastography (TEG).Genomic DNA was extracted from the peripheral blood.The sequences of all the exons and exon-intron boundaries of the three fibrinogen genes FGA, FGB and FGG were amplified by polymerase chain reaction（PCR） and analyzed by direct sequencing. Results The activity and the antigen levels of the proband′s plasma fibrinogen were reduced to 0.48 g/L and 0.68 g/L, respectively.TT prolonged to 29.2 s and RT prolonged to 75.8 s.The assays of SDS-PAGE showed no abnormal molecular weight of fibrinogen.Peak height of thrombin generation was reduced to 249.93 nmol/L and endogenous thrombin potential was reduced to 1007.0 nmol·L-1·min.Hypocoagulability state of the whole blood was found by TEG test.The coagulation index was -8.6.The proband was diagnosed as inherited hypofibrinogenemia by phenotype analysis.Two mutations (Gln143Pro and g.4642delC) were found in the proband′s fibrinogen Aa-chain gene, Gln143Pro came from her mother and g.4642delC came form her father. Conclusion Compound Heterozygous Mutations (Gln143Pro and g.4642delC) of fibrinogen Aa-chain causes the proband congenital hypofibrinogenemia.（Chin J Lab Med,2012,35:322-327） Key words: Afibrinogenemia; Fibrinogen; Pedigree; Heterozygote; Mutation

The Impact of Hematocrit on Fibrin Clot Formation Assessed by Rotational Thromboelastometry

Rotational thromboelastometry (ROTEM®)-based FIBTEM is used perioperatively to assess the extent of fibrin polymerization in whole blood. In FIBTEM, cytochalasin D eliminates the contribution of platelets to whole blood clotting, but changing levels in fibrin(ogen) and erythrocytes may differently affect clot formation. Because dynamic changes of hematocrit are not reflected in plasma fibrinogen measurements, we hypothesized that the lack of erythrocytes in isolated plasma measurements would affect the relationship between the Clauss method and whole blood-based FIBTEM during cardiac surgery. Therefore, in the current study we investigated the influence of perioperative hematocrit changes on FIBTEM and fibrinogen measurements. Blood samples were collected from 6 consenting healthy volunteers. FIBTEM tests were run before and after serial in vitro dilutions of whole blood with saline or autologous plasma (5:1, 2:1, and 1:1 v/v). We then evaluated the relationship between FIBTEM-maximal clot firmness (MCF) and the Clauss fibrinogen method in relation to hematocrit values before and after cardiac surgery. Pearson correlation coefficients were determined between laboratory test results and ROTEM variables. Upon in vitro hematocrit reduction, FIBTEM-MCF was progressively decreased depending on the extent of saline dilution, but it was increased by 31% after 1:1 volume replacement with autologous plasma (P < 0.05). In samples from cardiac patients (150 measurements in 50 patients), the overall correlation coefficient between FIBTEM-MCF and plasma fibrinogen was 0.80 (P < 0.001). In hemodiluted blood samples (during surgery or at intensive care unit), FIBTEM-MCF 10 mm corresponded to plasma fibrinogen levels of 200 mg/dL. In the subgroup analysis (n = 50 each), according to hematocrit levels (<25%, ≥25% to 30%, ≥30%), plasma fibrinogen levels of 200 mg/dL corresponded to 11 mm, 10 mm, and 8 mm of FIBTEM-MCF, respectively. The correlation between FIBTEM-MCF and plasma fibrinogen was higher at lower hematocrit (<25%) than at higher hematocrit (>30%) (r = 0.88 and 0.67, respectively). Perioperative changes in hematocrit affect the correlation between plasma fibrinogen levels and FIBTEM-MCF values. The higher correlation between FIBTEM-MCF and plasma fibrinogen with lower hematocrit (<25%) indicates that FIBTEM is a practical method to determine the need for fibrinogen replacement in bleeding patients who typically develop perioperative anemia.

Fibrinogen Šumperk II: Dysfibrinogenemia in an individual with two coding mutations

Fibrinogen—a 340-kDa glycoprotein—plays a crucial role in blood coagulation, platelet aggregation, wound healing, and other physiological processes. A mutation in fibrinogen may lead to congenital dysfibrinogenemia,a rare disease characterized by the functional deficiency of fibrinogen. About 580 cases of abnormal fibrinogens have been reported worldwide; thereof 335 cases in the fibrinogen Aa chain[1]. To our knowledge, only five cases of abnormal fibrinogens with two mutations [2–6] and one case of two different mutations in the same family [7] have been described earlier. A 52-year-old female was examined for bleeding. Routine hemostasis screening resulted in a diagnosis of dysfibrinogenemia. Functional testing revealed prolonged fibrin polymerization, prolonged lysis of the clot, abnormal fibrin morphology,and fibrinopeptides release. Genetic analysis showed two heterozygous nonsense mutations—previously described mutation AaGly13Glu and a novel mutation Aa Ser314Cys. The mutation Aa Gly13-Glu was found in her brother and niece, but there was no evidence in either of the mutation Aa Ser314Cys. While mutation Aa Gly13Glu is responsible for abnormal fibrinopeptide release and prolonged thrombin time, the novel mutation Aa Ser314Cys seems to affect fibrin morphology and fibrinolysis.

Fibrinogen, Lp(a), Microalbuminuria and Left Ventricular Mass Index: Cardiovascular Disease Risk factors in Diabetes.

The present work was undertaken to study interrelation of Fibrinogen, Lp(a) and LVMI in Type II diabetes patients with or without nephropathy. 100 Type II Diabetic patients attending OPD/IPD of DMC&H, Ludhiana were included. They were divided in two groups. Group I: 50 patients without microalbuminuria (MAU). Group II: 50 patients with MAU. Fibrinogen (Clauss method), Lp(a) and MAU were estimated on Multichannel Autoanalyzer Hitachi-911 (Roche). LVMI was estimated by echocardiography using formula of Devereux and Reicheck. Type II diabetes patients with MAU had significantly raised levels of Fibrinogen, Lp(a), and LVMI as compared to normoalbuminuric diabetics (P<0.01). Group II patients had positive correlation between Lp(a) and LVMI but no relation between Fibrinogen and LVMI. MAU, marker of microangiopathy, is associated with higher Fibrinogen and Lp(a) levels. This becomes basis of increase cardiovascular risk as demonstrated by higher mean LVMI in Group II patients.

A study of plasma fibrinogen level in type-2 diabetes mellitus and its relation to glycemic control.

The high prevalence of classic cardiac risk factors in patients with type 2 diabetes mellitus does not explain the increased cardiovascular related morbidity and mortality in these patients. Fibrinogen may have a role in this excess risk. This study is undertaken to know the fibrinogen levels in type 2 diabetes mellitus and its relations to glycemic control. In the present study fibrinogen levels (Clauss method) were estimated in 100 type 2 diabetic subjects and 100 age and sex matched controls. Fibrinogen was correlated with various parameters like glycosylated hemoglobin (cation exchange resin method), age, sex, smoking, body mass index (kg/m(2)), hypertension and ischemic heart disease. Higher plasma fibrinogen levels were found in type 2 diabetes mellitus patients (656±130mg/dl) as compared to controls (324±139mg/dl) which were statistically significant. Fibrinogen levels were associated with age (P<0.01), hypertension (P<0.01), body mass index (P<0.01), smoking (P<0.01), ischemic heart disease (P<0.01), and glycosylated hemoglobin (r=0.49) in diabetics in a significant manner. But no correlation was found with sex (P<0.05) in diabetes. In controls, association was found between fibrinogen levels and smoking (P<0.01) and body mass index (P<0.01). Patients with type 2 diabetes mellitus had a high prevalence of hyperfibrinogenemia. Fibrinogen levels were independently associated with hemoglobin A1c values, which suggests that fibrinogen may be involved in the increased cardiovascular risk of patients with type 2 diabetes mellitus.

Comparison of functional fibrinogen assessment using thromboelastography with the standard von Clauss method

To compare assessment of fibrinogen by thromboelastography with the standard von Clauss method. Observational prospective study. Thromboelastography provides direct and complex evaluation of the entire coagulation cascade based upon changes in blood viscosity. It affects both platelets and plasma components. New application of this method measures fibrinogen contribution to coagulation as opposed to fibrinogen antigen levels measured by immunoassay. Paired samples from 117 patients before cardiopulmonary bypass were compared. A moderate correlation was found between fibrinogen and functional fibrinogen with a Spearman correlation coefficient of 0.476. The functional fibrinogen test is a valid point-of-care method for fibrinogen assay with a moderate correlation to the standard method.

Rotational thromboelastometry (ROTEM) – the future of point of care testing in obstetrics?

IntroductionROTEM is a point of care testing (POCT) device that provides rapid specific coagulation pathway assessment. The use of ROTEM is well established in liver and cardiac surgery, but not...

Gerinnungsoptimierung mit ROTEM®– Pro

Coagulation and haemostasis in critical ill patients, either in massively bleeding patients in the emergency room and the operating theatre (TIC = trauma induced coagulopathy) or in septic patients suffering from multiorgan failure (DIC = disseminated intravascular coagulopathy) have to be identified and monitored promptly to initiate and monitor an effective therapy - either aggressive therapy using coagulation factor concentrates and blood products or a keen anticoagulation. Under this circumstances, standard coagulation monitoring using PT (INR), aPTT and fibrinogen (Clauss method) is limited. In critical ill patients, these parameters are neither able to predict risk of bleeding nor the risk of thrombosis. New viscoelastic methods like the ROTEM® may help to improve coagulation monitoring and management even in the emergency situation or under critical care conditions.

Clauss assay and fibrinogen protein estimated by capillary zone electrophoresis

The clottability and the amount of total protein in fibrinogen provide information about qualitative or quantitative alterations. We aimed to evaluate whether capillary zone electrophoresis (CZE) Capillarys II analyzer with the protein 6 buffer is able to estimate the amount of fibrinogen antigen. Citrated plasmas were assayed for clottable fibrinogen, and any relationship with the β(2)-globulin fraction (percentage of the area under the curve) was evaluated. The integration method used was "tangent skimming" in order to reduce the overestimation due to high protein background. Linearity was optimized according to the ICH Q2R1 recommendations and evaluated using polynomial regression. The precision was computed in accordance with the Clinical and Laboratory Standards Institute EP5-A2 protocol. In patients, clottable fibrinogen (Clauss method) was compared to its protein CZE amount by Passing and Bablok regression and the Bland-Altman plot. The correlation was linear y=0.0744+0.8991x (R(2)=0.9707) within the range of 2.26-17.26 μmol/L. The repeatability and the within-device precision were <15% for three levels of percentage of the β(2)-globulin fraction (1.61%, 3.51%, and 9.24%). In patients, clottable fibrinogen and its protein amount were similar (-0.1779+0.9654x). The ratio activity/protein was 1.08 ± 0.32 (mean ± 2 SD). CZE with the Capillarys II and the buffer protein 6 is an easy method. It is a good candidate for estimation of the concentration of fibrinogen antigen, which may have diagnostic utility for the screening of quantitative or qualitative fibrinogen abnormalities.

Increased nitric oxide production in platelets from severe chronic renal failure patients

Nitric oxide (NO) production occurs through oxidation of the amino acid L-arginine by NO synthase (NOS). NO inhibits platelet activation by increasing the levels of cyclic guanosine monophosphate (cGMP), thus maintaining vascular homeostasis. Our group previously demonstrated (da Silva et al. 2005) an enhancement of the L-arginine-NO-cGMP pathway in platelets taken from chronic renal failure (CRF) patients on haemodialysis associated with reduced platelet aggregation. We investigate the platelet L-arginine-NO-cGMP pathway, platelet function, and inflammation from patients in CRF on conservative treatment. A total of 42 CRF patients and 42 controls (creatinine clearance = 27 ± 3 vs. 93 ± 1 mL per min per 1.73 m2, respectively) participated in this study. NOS activity and expression and cGMP concentration were measured in platelets. Platelet aggregation induced by collagen or ADP was evaluated and plasma levels of fibrinogen were determined by the Clauss method. A marked increase in basal NOS activity was seen in undialysed CRF patients compared with controls, accompanied by an elevation of fibrinogen plasma levels. There were no differences in expression of NOS and in cGMP levels. In this context, platelet aggregation was not affected. We provide the first evidence of increased intraplatelet NO biosynthesis in undialysed CRF patients, which can be an early marker of future haemostatic abnormalities during dialysis treatment.

Dextran and hydroxyethyl starch do not interfere with fibrinogen measurement if Clauss method with mechanical clot detection is used

In an updated European guideline for the management of bleeding following major trauma, published in a recent issue of Critical Care, Rossaint and colleagues [1] recommended that thrombelastometry instead of other laboratory methods be used to measure fi brinogen. Th e authors stated that the Clauss method [2] overestimates fi brino gen concentration when colloids are given. Th is claim is supported by studies of patients receiving colloids in which fi brinogen was measured by an automated Clauss assay performed on an ACL-300R coagulometer (Instrumentation Laboratory, Bedford, MA, USA), which detects clot formation photo-optically [3]. Automated coagulometers use diff erent technologies to detect fi brin clot endpoints. Some coagulometers employ mechanical principles involving the movement of a metal hook or the clot impedance of a ball bearing in a magnetic fi eld; others use photo-optical devices to measure changes in light transmission [2]. Recently, Adam and colleagues [4] analyzed plasma diluted with diff erent proportions of saline or hydroxyethyl starch (HES). Fibrinogen concentration was overestimated if the clotting time was measured photooptically in samples diluted with HES. But fi brinogen measurements in analyzers that used mechanical clot detection endpoints were accurate. Fenger-Eriksen and colleagues [5] assessed the accuracy of fi brinogen measure ments in diff erent analyzers and diff erent methods in plasma diluted in diff erent proportions with saline, HES, and albumin. Th e authors found that the photo-optical measurement overestimated fi brinogen concentration but that mechanical detection did not. Th romboelastometry seems to be a rapid and accurate estimation of the level and function of fi brinogen but is not available in most centers. However, most laboratories can easily measure fi brinogen by the Clauss method using mechanical detection. We recommend that the European guidelines specifying the use of thromboelastometry over the Clauss method be revised and qualifi ed accordingly.

The role of fibrinogen: a new paradigm in the treatment of coagulopathic bleeding

Fibrinogen is involved in both primary and secondary hemostasis, playing an important role in platelet aggregation and the establishment of a fibrin network. Recent evidence suggests that very high levels of fibrinogen act as antithrombin and can reduce endogenous thrombin potential and compromise clot stability, particularly following a low tissue factor stimulus. Several laboratory methods for measuring plasma fibrinogen concentrations are available, but results vary depending on the type of method and the use of artificial colloid plasma expanders. Adopting only the Clauss method can provide erroneously high levels when used in patients who have received colloid plasma expanders. This may contribute to a hazardous delay or complete lack of treatment. Multiple in vitro experiments, animal studies, and proof-of-principle randomized, clinical studies have recently suggested that hemostatic intervention with a fibrinogen concentrate may be efficient and safe in controling perioperative bleeding. In particular, fibrinogen concentrate has a key role in improving clotting function and reducing blood loss in settings such as trauma and cardiothoracic surgery. However, prospective studies are needed to determine the clinical efficacy and safety of fibrinogen concentrate when used as a hemostatic intervention for patients with massive bleeding due to trauma or surgery.

The role of fibrinogen plasma levels, the –455G&gt;A fibrinogen and the factor XIII A subunit (FXIII-A) Val34Leu polymorphism in cancer-associated venous thrombosis

Venous thromboembolism (VTE) is a life-threatening complication in cancer patients. Identification of risk factors has been in focus in the past years. Functional single nucleotide polymorphisms (SNP) of coagulation factors known to influence the concentration or function may be considered to influence the risk of VTE in cancer patients. We evaluated the influence of fibrinogen plasma levels, the -455G>A SNP in the fibrinogen beta gene and the Val34Leu (163G>T) SNP in the factor XIII A-subunit (FXIII-A) gene on the risk of VTE. In 1,079 tumour patients recruited for the prospective Vienna Cancer and Thrombosis Study (CATS) fibrinogen levels were determined by the Clauss method. The FXIII-A Val34Leu and the fibrinogen -455G>A SNPs were tested by allele-specific PCR. The median follow-up time was 604 days, 83 thrombotic events occurred. The median fibrinogen level was 381 mg/dl (25th-75th percentile: 312 to 467). In a multivariable Cox model adjusted to chemotherapy, surgery, radiotherapy, age and sex, neither the fibrinogen concentration (hazard ratio [HR] =1.05, confidence interval [CI] 0.839-1.310 p=0.68), nor the -455G>A SNP (HR=0.77, 95%CI 0.491-1.197; p=0.24), nor the Val34Leu SNP (HR=0.99, 95%CI 0.646-1.542 p=0.99) were associated with occurrence of VTE. The fibrinogen concentration was not significantly different among the fibrinogen -455G or A genotype carriers (p = 0.33). Disseminated intravascular coagulation was observed in only five patients, none of these developed VTE. In conclusion, fibrinogen plasma levels, the fibrinogen -455G>A and the FXIII-A Val34Leu polymorphisms were not associated with VTE in our study.

Rivaroxaban differentially influences ex vivo global coagulation assays based on the administration time

It was the objective of this study to quantify the effects of rivaroxaban administration on global coagulation parameters associated with routine clinical procedures, we collected plasma samples from patients undergoing major orthopaedic surgery receiving rivaroxaban at various time points after drug administration. Forty-seven patients received rivaroxaban (10 mg daily) for venous thromboembolism prophylaxis. Blood samples were collected at four different time points: A) before surgery; B) before drug administration at day 4-5 after surgery (steady state of rivaroxaban); C) 2 hours (h) after drug administration and D) 12 h after drug administration. The prothrombin time (PT), activated partial thromboplastin time (aPTT), thrombin time (TT), antithrombin (AT) level, fibrinogen level by Clauss method (FibC), and derived fibrinogen (dFIB) level were assessed with various reagents. At 2 h after rivaroxaban administration, the PT and aPTT clotting times were significantly prolonged to different extents up to 1.4 fold, whereas 12 h after drug administration, no significant effect was observed. Rivaroxaban administration had no influence on the TT or the FibC concentration. The dFIB assay was differentially affected by rivaroxaban when different reagents were tested. The AT assay dependent on thrombin activity was not influenced by rivaroxaban, whereas the AT levels dependent on factor Xa activity were significantly increased by rivaroxaban. Clinicians should be aware of the time-dependent influence of rivaroxaban on factor Xa-dependent routine coagulation assays. Therefore, routine coagulation parameters should be assessed directly before drug administration to keep the interaction of rivaroxaban low.

Fibrinogen concentrate for management of bleeding

Fibrinogen concentrate for management of bleeding

In Vitro Effects of Detergent Sclerosants on Clot Formation and Fibrinolysis

Objective To investigate the in vitro effects of detergent sclerosants sodium tetradecyl sulphate (STS) and polidocanol (POL) on clot formation and lysis. Materials and methods clot kinetics were assessed in whole blood by thromboelastography (TEG ®) and rotational thromboelastometry (ROTEM ®). Fibrinogen was measured by the Clauss method in plasma and factor XIII (FXIII) by enzyme-linked immunosorbent assay (ELISA). Turbidity measurements were used to assess clot lysis in plasma, and fibrinolysis in non-cross-linked and cross-linked fibrin. D-dimer was measured by VIDAS ®, STA ®Liatest ® and AxSYM ® assays. Results Strong clots were formed at low sclerosant concentrations (0.075–0.1%). At midrange concentrations (0.15% STS, 0.15–0.3% POL), both agents inhibited the contribution of platelets to clot firmness and formed weak clots prone to lysis. At higher concentrations (STS ≥0.3% and POL ≥0.6%), clot formation was inhibited. STS destroyed FXIII at ≥0.15% and fibrinogen at ≥0.6%. Neither sclerosant had a significant effect on cross-linked fibrin, but STS had a lytic effect on non-cross-linked fibrin. STS caused an artefactual elevation of D-dimer in the VIDAS ® assay when fibrinogen was present. Conclusion Detergent sclerosants demonstrated a trimodal effect on clot formation, initiating strong clots at low concentrations, weak clots at midrange concentrations and preventing clot formation at higher concentrations. Neither agent had fibrinolytic activity.

Fibrinogen estimates are influenced by methods of measurement and hemodilution with colloid plasma expanders

Measurement of plasma fibrinogen is often required in critically ill patients or massively bleeding patients being resuscitated with colloid plasma expander. This study aimed at evaluating different assays of plasma fibrinogen after in vitro dilution with commonly used plasma expanders and challenged the hypothesis that levels of fibrinogen are estimated significantly higher in plasma diluted with colloid plasma expander compared with isotonic saline. Fibrinogen measurements were established in plasma samples each diluted in vitro to 30 or 50% with isotonic saline, hydroxyethyl starch (HES) 130/0.4, and human albumin. Fibrinogen levels were assessed using an antigen determination, three photo-optical Clauss methods, one mechanical Clauss method, a prothrombin-derived method, and viscoelastic measurement through thromboelastometry. Measurement of fibrinogen levels was significantly different when performed on alternate analytical platforms. By 30 and 50% dilution with HES 130/0.4 coagulation analyzers using the photo-optical Clauss methods significantly overestimated levels of fibrinogen. Dilution with human albumin did not affect fibrinogen levels except from one analyzer by 50% dilution level. Viscoelastic measurement of fibrin polymerization was reduced at both dilution levels and appeared to reflect the impairment of fibrin polymerization induced by HES 130/0.4 and to a lesser extent human albumin. This study demonstrated that different automated coagulation analyzers revealed significantly different levels of fibrinogen. The presence of colloid plasma expander gave rise to erroneous high levels of fibrinogen returned from some coagulation analyzers employing the method of Clauss.

Bleeding and Thrombosis In 90 Iranian Patients with Inherited Afibrinogenemia.

AbstractAbstract 3667 Background:Afibrinogenemia is a rare coagulation disorder transmitted as an autosomal recessive trait. In Iran, where consanguineous marriages are common coagulation disorders occur approximately ten times more frequently than in western countries. Afibrinogenemia is characterized by abnormal thrombin time, prothrombin time, activated partial thromboplastin time and absent plasma fibrinogen levels. We investigated the clinical manifestations of bleeding and thrombosis in a series of patients with Afibrinogenemia in Iran. To our knowledge this is the largest series ever reported. Patients & Methods:All patients in our cohort were diagnosed and registered in the Iranian National Registry of Inherited Bleeding Disorders. Every patient with an inherited blood disorder is reported to this registry because it is the only source of free treatment for bleeding disorders in the country. Afibrinogenemia has been tracked in Iran since 1965. Diagnoses in all patients was confirmed when plasma fibrinogen levels were absent both as functional activity (the Clauss method) and using the antigen level (electroimmunoassay). We retrospectively evaluated data from all 90 Afibrinogenemia patients available in the registry. Results:There were 46 males and 44 females in our cohort. Ages ranged from 1 to 71 years. 82% of patients were born from consanguineous marriages with asymptomatic parents. The most frequent symptoms in our patients were umbilical cord bleeding (87%), hematomas (74%) epistaxis (70%) and hemarthrosis (52%). Central nervous system bleeding was relatively rare (2.2%). In women of reproductive age, menorrhagia occurred in 75% of cases. Recurrent spontaneous abortions (more than 2) were seen in 28% of women (range 2 to 6). With prophylactic treatment (cryoprecipitate and limited fibrinogen) two (16.6%) of our patients with recurrent spontaneous abortion out of the 12 treated were able to carry pregnancies to term. A total of 7 patients in our population developed arterial or venous thromboses (3 males and 4 females). Four cases of the thromboses observed occurred after infusion of fibrinogen concentrate. Three episodes of spontaneous thrombosis were observed including a case of mesenteric thrombosis. Discussion:Afibrinogenemia is associated with severe and life threatening bleeding symptoms. Treatment can help women afflicted with this disease carry pregnancies to term. As reported elsewhere, thrombotic complications both spontaneous and acquired from treatment were also seen in our population. Disclosures:No relevant conflicts of interest to declare.

Abdominal Obesity, IL-6 Levels and the Risk of Venous Thromboembolism Among Women: a Case-Control Study

AbstractAbstract 477 Background:abdominal obesity is one of the components of the metabolic syndrome (MS), which is a cluster of cardiovascular risk factors. Recent prospective studies demonstrated that among the components of the MS, abdominal obesity was the only independent risk factor for venous thromboembolism (VTE). However, it remains unclear to what extent inflammation contributes to the risk of VTE from abdominal obesity. The aim of this study was to investigate the association of abdominal obesity and VTE in women and the effect of inflammation on this association. Material and methods:94 female patients were included with a first objectively confirmed episode of VTE, with a median age of 39 years (range: 21–60). Exclusion criteria were malignancy, autoimmune diseases, chronic renal or liver diseases and arterial thrombosis. Patients were seen at least 1 month after the discontinuation of the anticoagulant treatment and > 7 months after the event of VTE. Controls were comprised of 100 healthy women, with a median age of 37 years (range: 18–63), recruited via the patients from the same geographic region and ethnic background. Controls were matched for age. Waist circumference (WC) was measured at the umbilical line. A WC ≥ 88 cm in women defined abdominal obesity according to the National Cholesterol Education Program. Plasma level of IL-6 was measured by a highly sensitive ELISA, fibrinogen by Clauss method and high-sensitive C-reactive protein (CRP) by immunoturbidimetry. High levels of IL-6 (> 2.01 pg/mL), CRP (> 6.66 mg/L) and fibrinogen (> 3.59 g/L) were dichotomized at the 90th percentile of the control values. Result:median WC was higher in patients (89 cm; range: 65–126) than in controls (84 cm; range: 62–131), P<0.01. In both groups, WC significantly correlated (P<0.01) with body mass index as expected, but also with age, fibrinogen, CRP and IL-6. Fifty-five patients and 43 controls had abdominal obesity, yielding an odds ratio (OR) of 1.87 [95% Confidence Interval (CI) 1.06 – 3.31]. The association of VTE and abdominal obesity was not affected after adjustment for potential confounders, as high levels of CRP (OR 1.86; 95% CI 1.05 – 3.30) and fibrinogen (OR 1.94; 95% CI 1.08 – 3.48), and it was of borderline significance when adjusted for age (OR 1.76; 95% CI 0.97–3.20). However, the risk was no longer significant after adjustment for high levels of IL-6 (OR 1.57; 95% CI 0.87–2.84). Simultaneous adjustment for age and high levels of CRP, fibrinogen and IL-6 yielded an OR of 1.60 (95% CI 0.85 – 3.00). The impact of abdominal obesity on the risk of VTE was more pronouncedly affected when IL-6 entered into the logistic regression model as a continuous variable (OR 1.33; 95% CI 0.72 – 2.47). Conversely, high IL-6 levels increased the risk of VTE even after adjustment for age, abdominal obesity and high levels of CRP and fibrinogen (OR 3.65; 95%CI 1.56 – 8.50). Conclusion:in this study abdominal obesity was associated with VTE in crude analysis in a relatively young population composed of women. However, the association was no longer significant after adjustment for IL-6. There is evidence that visceral adipose tissue, assessed by WC, is metabolic active, releasing cytokines. To our knowledge this is the first study that evaluated in the same population the effect of IL-6 and abdominal obesity on the risk of VTE. More data, including prospective studies, are required to elucidate the role of inflammation in the association of VTE and abdominal obesity.This study was supported by FAPESP (2005/56799-0). Disclosures:No relevant conflicts of interest to declare.