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C0233: Prothrombin Fragment 1 + 2 in Urine in Patients with Clinical Suspected Venous Thromboembolism
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POSTERS / Thrombosis Research 133S3 (2014) S35–S123
an effective method to investigate the molecular mechanism of the hereditary diseases. Methods: Here we report nine hereditary PC deficiency pedigrees which involving one newly reported mutation sites (c.1157T>C, p.Leu386Pro) and manifested mutant PC 3D construction by homology modeling. Results: The results showed that mutation (c.565C>T, p.Arg189Trp; c.740G>A, p.Trp247Terminal and c.1157T>C, p.Leu386Pro) induced significant conformational changes on PC construction. c.565C>T increased the distance between Trp and former Arg’s adjacent amino acids and destroyed the hydrogen bonds between Arg189 and His411; c.1157T>C substantially increased the VDW’s interaction energy from −5.70 kcal/mol to 31986.45 kcal/mol and further decreased the stability of PC’s construction; c.740G>A made large fragment of heavy chain disappeared and formed an unknown PC’s construction. But the mutation (c.508G>T, p.Asp170Tyr) made no contribution on PC’s construction. Conclusions: Our research concluded that the molecular mechanism of c.565C>T (p.Arg189Trp), c.740G>A (p.Trp247Terminal) and c.1157T>C (p.Leu386Pro) were the misfolding of PC’s conformation. The molecular mechanism of c.508G>T (p.Asp170Tyr) needs further research. C0205 D-DIMER LEVELS AND THROMBOPROPHYLAXIS AFTER DELIVERY J. Samonikov-Tosevska1 , T. Makarovska-Bojadzieva1 , E. Todorovska1 , I. Nikoloska1 , V. Neceva1 , R. Apostolovska1 , M. Blagoesvska1 , E. Velkova1 . 1 Institute of Transfusion medicine, Vodnjanska, 1000 Skopje, Macedonia Background: The risk of thrombosis is increased throughout pregnancy and is particularly high after delivery. Our aim was to assess the value of D-dimer (DD) as a coagulation activation marker during the postpartum period in order to target women who are at particular risk of venous thromboembolic event (VTE). Methods: We evaluated data of screening coagulation testing and quantitative D-dimer measurements in blood samples from 170 women with normal, uncomplicated pregnancy who delivered at term either vaginally (n = 33) or by cesarean section (n = 137). We determined the median and the standard deviation (SD) of the platelet count (150–450 x 109 /L), activated partial thromboplastin time-aPTT (26–38 seconds) and D-dimer (<500 ng/mL) in the period from 10 to 20 days after delivery. Results: There was no significant difference between the vaginally and the cesarean section delivered women in the platelet count (356±207 and 394±130) and the aPTT value (28±3 and 27±3) respectively. Levels of DD were significantly higher (p3030 ng/mL) was measured in 22 (16%) of the women in the cesarean group. None of them had clinically and ultrasonographically signs of venous thromboembolism (VTE). However, in 21 (12%) of the delivered women in both of the groups the DD levels were below the 500 ng/mL (the established cut-off for non-pregnant women). Conclusions: The great individual DD variations observed, as well as the lack of correlation with the clinical signs of thrombosis, indicate that DD measurements are of little value for excluding VTE during the period of postpartum. However, it is recommended to give a prophylaxis with LMWH at least 4 weeks in the postpartum period in women at high risk of thromboembolic events especially when delivered by cesarean section.
C0233 PROTHROMBIN FRAGMENT 1 + 2 IN URINE IN PATIENTS WITH CLINICAL SUSPECTED VENOUS THROMBOEMBOLISM F. Wexels1 , A.H. Pripp1 , I. Seljeflot1 , A. Haslund1 , T.E. Gudmundsen1 , O.E. Dahl1 . 1 Fru Kroghs Brygge, 0252 Oslo, Norway Background: Prothrombin fragment 1 + 2 (F1 + 2) is generated when prothrombin is converted to thrombin. F1 + 2 is excreted in urine due to a low molecular weight. Levels of uF1 + 2 can be measured with enzyme-linked immuno-sorbent assay (ELISA). We evaluated levels of uF1 + 2 and the association with radiological confirmed venous thromboembolism (VTE) in symptomatic nonselected patients. Methods: Patients with clinical suspected VTE and age ≥18 years were consecutively included after informed consent. A urine sample was collected before imaging with either B-mode compression ultrasound or contrast-enhanced computer tomography (CT) angiography depending on their clinical symptoms. uF1 + 2 was quantified with an ELISA kit. In order to detect new thromboembolic events or other conditions that modified the thrombin activity, all patients were followed up for at least three months. Data analyses were conducted using SPSS version 21. Due to asymmetric data distribution, the non-parametric Mann–Whitney U test was used to assess differences in uF1 + 2 levels in the VTE negative and positive patients. A p-value ≤0.05 was considered statistically significant. Results: VTE was diagnosed in 151 (20.8%) of the 725 (360 men and 365 women) included patients. 44 (6.1%) patients had pulmonary embolism (PE) and 108 (14.9%) had deep vein thrombosis (DVT). One patient was diagnosed with both PE and DVT. Median age was 60 years (range 18–93). uF1 + 2 levels [median (25th , 75th percentiles)] in the total population was 31.9 (20.0–101.6) pmol L−1 . The VTE positive patients had statistically significant higher uF1 + 2 levels than those without VTE (60.4 (20.0–179.1) vs. 26.4 (20.0– 78.4) pmol L−1 , p < 0.001). Conclusions: In our study statistically significantly higher urine F1 + 2 levels were found in patients with confirmed VTE versus those without VTE. This indicates that F1 + 2 in urine reflects procoagulant activity and radiological confirmed VTE. C0234 ASSESSMENT OF THE DIAGNOSTIC ALGORITHM FOR HEPARIN-INDUCED THROMBOCYTOPENIA M. Farm Stephanie1 , T. Bakchoul1 , T. Frisk1 , A. Odenrick1 , K. Althaus1 , E.-M. Norberg1 , M. Berndtsson1 , A. Greinacher1 , J. Antovic1 . 1 Klinisk Kemi L7:00, Karolinska Universitetssjukhuset Solna, 17176 Stockholm, Sweden Background: Heparin-Induced Thrombocytopenia (HIT) is a rare but serious adverse reaction to treatment with Heparin. HIT is caused by acquired antibodies that bind to platelet factor 4 (PF4), resulting in a transient acute thrombocytopenia potentially combined with a prothrombotic state. Such functional antibodies are present in 1–20% of heparin treated patients, whereas clinical HIT affects only 0.25–3% of patients. Diagnosis is always a combination of laboratory results and clinical presentation. Our laboratory uses an algorithm which includes the 4T-score for estimation of clinical pretest probability of HIT. According to the algorithm samples with low 4T score are not analysed, but answered as negative. Samples with intermediary 4T risk are analysed by PF4 particle-gel immunoassay (PaGIA), where a negative PaGIA gives a negative answer, but a positive result warrants further analysis by IgG-specific ELISA. If that ELISA is negative, the sample is negative for HIT, but a positive ELISA needs to be confirmed by a Heparin-induced platelet activation assay (HIPA). Samples with high 4T risk are answered as positive if the PaGIA and/or ELISA are positive. Methods: We assessed the validity of this diagnostic algorithm against a reference method, namely the HIPA performed in the
POSTERS / Thrombosis Research 133S3 (2014) S35–S123
an effective method to investigate the molecular mechanism of the hereditary diseases. Methods: Here we report nine hereditary PC deficiency pedigrees which involving one newly reported mutation sites (c.1157T>C, p.Leu386Pro) and manifested mutant PC 3D construction by homology modeling. Results: The results showed that mutation (c.565C>T, p.Arg189Trp; c.740G>A, p.Trp247Terminal and c.1157T>C, p.Leu386Pro) induced significant conformational changes on PC construction. c.565C>T increased the distance between Trp and former Arg’s adjacent amino acids and destroyed the hydrogen bonds between Arg189 and His411; c.1157T>C substantially increased the VDW’s interaction energy from −5.70 kcal/mol to 31986.45 kcal/mol and further decreased the stability of PC’s construction; c.740G>A made large fragment of heavy chain disappeared and formed an unknown PC’s construction. But the mutation (c.508G>T, p.Asp170Tyr) made no contribution on PC’s construction. Conclusions: Our research concluded that the molecular mechanism of c.565C>T (p.Arg189Trp), c.740G>A (p.Trp247Terminal) and c.1157T>C (p.Leu386Pro) were the misfolding of PC’s conformation. The molecular mechanism of c.508G>T (p.Asp170Tyr) needs further research. C0205 D-DIMER LEVELS AND THROMBOPROPHYLAXIS AFTER DELIVERY J. Samonikov-Tosevska1 , T. Makarovska-Bojadzieva1 , E. Todorovska1 , I. Nikoloska1 , V. Neceva1 , R. Apostolovska1 , M. Blagoesvska1 , E. Velkova1 . 1 Institute of Transfusion medicine, Vodnjanska, 1000 Skopje, Macedonia Background: The risk of thrombosis is increased throughout pregnancy and is particularly high after delivery. Our aim was to assess the value of D-dimer (DD) as a coagulation activation marker during the postpartum period in order to target women who are at particular risk of venous thromboembolic event (VTE). Methods: We evaluated data of screening coagulation testing and quantitative D-dimer measurements in blood samples from 170 women with normal, uncomplicated pregnancy who delivered at term either vaginally (n = 33) or by cesarean section (n = 137). We determined the median and the standard deviation (SD) of the platelet count (150–450 x 109 /L), activated partial thromboplastin time-aPTT (26–38 seconds) and D-dimer (<500 ng/mL) in the period from 10 to 20 days after delivery. Results: There was no significant difference between the vaginally and the cesarean section delivered women in the platelet count (356±207 and 394±130) and the aPTT value (28±3 and 27±3) respectively. Levels of DD were significantly higher (p3030 ng/mL) was measured in 22 (16%) of the women in the cesarean group. None of them had clinically and ultrasonographically signs of venous thromboembolism (VTE). However, in 21 (12%) of the delivered women in both of the groups the DD levels were below the 500 ng/mL (the established cut-off for non-pregnant women). Conclusions: The great individual DD variations observed, as well as the lack of correlation with the clinical signs of thrombosis, indicate that DD measurements are of little value for excluding VTE during the period of postpartum. However, it is recommended to give a prophylaxis with LMWH at least 4 weeks in the postpartum period in women at high risk of thromboembolic events especially when delivered by cesarean section.
C0233 PROTHROMBIN FRAGMENT 1 + 2 IN URINE IN PATIENTS WITH CLINICAL SUSPECTED VENOUS THROMBOEMBOLISM F. Wexels1 , A.H. Pripp1 , I. Seljeflot1 , A. Haslund1 , T.E. Gudmundsen1 , O.E. Dahl1 . 1 Fru Kroghs Brygge, 0252 Oslo, Norway Background: Prothrombin fragment 1 + 2 (F1 + 2) is generated when prothrombin is converted to thrombin. F1 + 2 is excreted in urine due to a low molecular weight. Levels of uF1 + 2 can be measured with enzyme-linked immuno-sorbent assay (ELISA). We evaluated levels of uF1 + 2 and the association with radiological confirmed venous thromboembolism (VTE) in symptomatic nonselected patients. Methods: Patients with clinical suspected VTE and age ≥18 years were consecutively included after informed consent. A urine sample was collected before imaging with either B-mode compression ultrasound or contrast-enhanced computer tomography (CT) angiography depending on their clinical symptoms. uF1 + 2 was quantified with an ELISA kit. In order to detect new thromboembolic events or other conditions that modified the thrombin activity, all patients were followed up for at least three months. Data analyses were conducted using SPSS version 21. Due to asymmetric data distribution, the non-parametric Mann–Whitney U test was used to assess differences in uF1 + 2 levels in the VTE negative and positive patients. A p-value ≤0.05 was considered statistically significant. Results: VTE was diagnosed in 151 (20.8%) of the 725 (360 men and 365 women) included patients. 44 (6.1%) patients had pulmonary embolism (PE) and 108 (14.9%) had deep vein thrombosis (DVT). One patient was diagnosed with both PE and DVT. Median age was 60 years (range 18–93). uF1 + 2 levels [median (25th , 75th percentiles)] in the total population was 31.9 (20.0–101.6) pmol L−1 . The VTE positive patients had statistically significant higher uF1 + 2 levels than those without VTE (60.4 (20.0–179.1) vs. 26.4 (20.0– 78.4) pmol L−1 , p < 0.001). Conclusions: In our study statistically significantly higher urine F1 + 2 levels were found in patients with confirmed VTE versus those without VTE. This indicates that F1 + 2 in urine reflects procoagulant activity and radiological confirmed VTE. C0234 ASSESSMENT OF THE DIAGNOSTIC ALGORITHM FOR HEPARIN-INDUCED THROMBOCYTOPENIA M. Farm Stephanie1 , T. Bakchoul1 , T. Frisk1 , A. Odenrick1 , K. Althaus1 , E.-M. Norberg1 , M. Berndtsson1 , A. Greinacher1 , J. Antovic1 . 1 Klinisk Kemi L7:00, Karolinska Universitetssjukhuset Solna, 17176 Stockholm, Sweden Background: Heparin-Induced Thrombocytopenia (HIT) is a rare but serious adverse reaction to treatment with Heparin. HIT is caused by acquired antibodies that bind to platelet factor 4 (PF4), resulting in a transient acute thrombocytopenia potentially combined with a prothrombotic state. Such functional antibodies are present in 1–20% of heparin treated patients, whereas clinical HIT affects only 0.25–3% of patients. Diagnosis is always a combination of laboratory results and clinical presentation. Our laboratory uses an algorithm which includes the 4T-score for estimation of clinical pretest probability of HIT. According to the algorithm samples with low 4T score are not analysed, but answered as negative. Samples with intermediary 4T risk are analysed by PF4 particle-gel immunoassay (PaGIA), where a negative PaGIA gives a negative answer, but a positive result warrants further analysis by IgG-specific ELISA. If that ELISA is negative, the sample is negative for HIT, but a positive ELISA needs to be confirmed by a Heparin-induced platelet activation assay (HIPA). Samples with high 4T risk are answered as positive if the PaGIA and/or ELISA are positive. Methods: We assessed the validity of this diagnostic algorithm against a reference method, namely the HIPA performed in the