Differential vascular effects of aspirin in people with Type 2 diabetes without cardiovascular disease and matched controls without diabetes.

AIM: We investigated whether the effect of low-dose aspirin on endothelium-dependent vasodilation and arterial stiffness in people with Type 2 diabetes is different from a matched control group. We examined acute and chronic effects, and effects over the 24h dosing interval. METHODS: In an open-label parallel group intervention study, we included 21 participants with Type 2 diabetes and 21 age- and sex-matched controls. Endothelium-dependent vasodilation was assessed as the reactive hyperaemia index (lnRHI) measured by peripheral arterial tonometry (EndoPAT® ). Arterial stiffness was assessed as pulse wave velocity (PWV) measured by applanation tonometry (SphygmoCor® ). Measurements were performed prior to aspirin intake and 1h after aspirin administration (75 mg). Participants were then treated for 6 days, and measurements were repeated at 24 h and 1 h after aspirin intake. RESULTS: Baseline lnRHI did not differ between groups. The controls had an immediate increase in lnRHI after the first aspirin tablet. This was not observed in participants with diabetes (difference between groups; P < 0.05). After 1 week, both groups demonstrated increased lnRHI compared with baseline (P < 0.01). In participants with diabetes, lnRHI was significantly lower 24 h after aspirin administration compared with 1 h after administration (P < 0.05). This difference was not observed in controls (P = 0.84, difference between groups; P = 0.12). The effect on PWV did not differ between groups. CONCLUSION: Aspirin had a reduced immediate effect on endothelium-dependent vasodilation in participants with diabetes. Both groups had improved endothelial function after 1 week of treatment. Further, the effect of aspirin on endothelial function may be declining during a 24 h dosing interval in people with Type 2 diabetes. (Clinical Trial Registry No: 2016-000515-32).

Low risk of venous thromboembolism following early pregnancy loss in pregnancies conceived by IVF.

STUDY QUESTION: What is the risk of venous thromboembolism (VTE) in the 12 weeks following early pregnancy loss in pregnancies conceived by IVF? SUMMARY ANSWER: The VTE risk is low in the 12 weeks following early pregnancy loss in pregnancies conceived by IVF. WHAT IS KNOWN ALREADY: There is an excess VTE risk during first trimester in complete IVF pregnancies leading to delivery. It is unknown whether this excess VTE risk also is present in IVF pregnancies terminated by early pregnancy loss (implantation failure, missed abortion, or spontaneous or induced abortion before 10 completed gestational weeks). STUDY DESIGN, SIZE, DURATION: A nationwide registry-based cohort study including all Danish IVF pregnancies registered in the Danish IVF Register between 1995 and 2005. Women who underwent frozen embryo replacement or oocyte donation were not included. PARTICIPANTS/MATERIALS, SETTING, METHODS: We included all 24 931 IVF treatments leading to pregnancy among 19 260 women. We identified 16 701 complete IVF pregnancies, 7567 IVF pregnancies with early pregnancy loss, and 663 IVF pregnancies terminated by late abortion (≥gestational weeks 10). We followed women for 12 weeks after termination of pregnancy and calculated the absolute risk of VTE during follow-up with 95% CI. As a relative risk estimate, we calculated the risk ratio for VTE following IVF pregnancies with early loss compared to the VTE risk following complete IVF pregnancies. MAIN RESULTS AND THE ROLE OF CHANCE: During the 12 weeks of follow-up, only one case of VTE occurred in the group of early pregnancy loss, none in the late abortion group, and 13 VTE cases occurred in complete IVF pregnancies. The VTE risk per 10 000 pregnancies was 1.3 [0.03-7.4] for IVF pregnancies with early loss and 7.8 [4.1-13.3] for complete pregnancies; the corresponding risk ratio was 0.17 [0.02-1.3]. Thus, we found a low absolute VTE risk in the 12 weeks following early pregnancy loss in IVF pregnancies. The relative VTE risk was low in comparison to the VTE risk in early gestation reported for complete IVF pregnancies and for Danish background pregnancies. LIMITATIONS, REASONS FOR CAUTION: Despite the use of complete nationwide data, only a few VTE events were available for the statistical analyses thereby limiting the precision of our estimates. We included both inpatient and outpatient VTE hospital diagnoses, but we cannot rule out the occurrence of VTE cases not diagnosed at hospital and hence not registered in the National Patient Registry. No information was available on the use of prophylactic low molecular weight heparin: access to such might have helped to explain our findings, but would not have changed our conclusions. WIDER IMPLICATIONS OF THE FINDINGS: The results of the present study do not indicate a need for prophylactic anticoagulant therapy in women suffering early IVF pregnancy loss without other risk factors. STUDY FUNDING/COMPETING INTEREST(S): No external funding was obtained for this study. There are no conflicts of interest to declare.

The effect of fresh frozen plasma in critically ill patients.

BACKGROUND: Critically ill patients often receive fresh frozen plasma (FFP) if they have abnormal conventional coagulation tests. The aim of this study was to investigate the effect of FFP transfusion judged by a wide range of coagulation tests. METHODS: We included 30 critically ill patients receiving FFP and 30 critically ill patients who did not receive FFP. For patients receiving FFP, blood samples were obtained before and 1 h after FFP transfusion. Conventional coagulation tests, thromboelastometry (ROTEM® , EXTEM, INTEM and FIBTEM) and thrombin generation were performed. Systematic recording of vital signs was performed for all patients. RESULTS: The median values of the conventional coagulation tests were abnormal before and after FFP (PT: (normal > 0.6) median 0.3 before vs. 0.3 after; INR: (normal < 1.2) median 2 before vs. 1.7 after; APTT: (normal < 38 s) median 45 s before vs. 42 s after). Eight of nine ROTEM® parameters were within the reference interval judged by median values before FFP transfusion, and all median parameters were within the reference interval after FFP transfusion. Median in three of four thrombin generation parameters was within the reference interval both before and after FFP transfusion. CONCLUSION: Patients presented abnormal conventional coagulation tests both before and after FFP transfusion. In contrast, ROTEM® and thrombin generation parameters were mainly within the reference interval both before and after FFP transfusion. FFP transfusions caused only negligible, although statically significant, improvements on coagulation measurements judged by conventional coagulation tests, ROTEM® and thrombin generation.

Post-cardiac arrest level of free-plasma DNA and DNA-histone complexes.

BACKGROUND: Plasma DNA-histone complexes and total free-plasma DNA have the potential to quantify the ischaemia-reperfusion damages occurring after cardiac arrest. Furthermore, DNA-histone complexes may have the potential of being a target for future treatment. The aim was to examine if plasma DNA-histone complexes and the levels of total free-plasma DNA were elevated in post-cardiac arrest patients compared with healthy individuals, and to examine if these biomarkers were capable of predicting mortality. METHODS: We included 42 comatose out-of-hospital cardiac arrest patients and collected blood samples after 22, 46 and 70 h. Samples for DNA-histone complexes were quantified by Cell Death Detection ELISAplus . The total free-plasma DNA analyses were quantified with qPCR by analysing the Beta-2 microglobulin gene. The control group comprised 40 healthy individuals. RESULTS: We found no difference in the level of DNA-histone complexes between the 22-h sample and healthy individuals (P = 0.10). In the 46-h sample, there was an increased level of DNA-histone complexes in non-survivors compared with survivors 30 days after the cardiac arrest (P < 0.01) and the area under the ROC curve was 0.78 (95% confidence interval: 0.59;0.96). The level of total free-plasma DNA was increased in the 22-h sample compared with healthy individuals (P < 0.001) but no significant difference was found between non-survivors and survivors 30 days after the cardiac arrest (all P ≥ 0.06). CONCLUSION: An increased level of DNA-histone complexes was associated with increased mortality and that the level of total free-plasma DNA was elevated post-cardiac arrest.

Increased venous thrombosis incidence in pregnancies after in vitro fertilization.

STUDY QUESTION: Is venous thrombosis risk increased in pregnancies after in vitro fertilization? SUMMARY ANSWER: The venous thrombosis incidence was significantly increased in pregnancies after in vitro fertilization; especially in the first trimester and in the first 6 weeks post-partum. WHAT IS KNOWN ALREADY: In vitro fertilization without pregnancy is not associated with increased venous thrombosis incidence. STUDY DESIGN, SIZE, DURATION: This national register-based cohort study covered the period from 1995 to 2005. PARTICIPANTS/MATERIALS, SETTING, METHODS: All Danish pregnancies conceived by in vitro fertilization (n = 18 787) were included. Venous thrombosis incidence rates in pregnancies after in vitro fertilization were compared with venous thrombosis incidence rates in reference pregnancies, by calculating incidence rate ratios. MAIN RESULTS AND THE ROLE OF CHANCE: In total, 48 cases were identified. In pregnancies after in vitro fertilization, the overall venous thrombosis incidence rate was 28.6 per 10 000 pregnancy-years (95% confidence interval (CI) 20.6-39.6) in comparison to 10.7 per 10 000 woman-years in reference pregnancies. Post-partum, the venous thrombosis incidence rate was 27.9 per 10 000 woman-years (95% CI 15.8-49.1) after in vitro fertilization in comparison to 17.5 per 10 000 woman-years in reference pregnancies. The overall venous thrombosis incidence rate ratio during in vitro fertilization (IVF) pregnancies compared with reference pregnancies was 3.0 (95% CI 2.1-4.3). The venous thrombosis incidence rate ratios during pregnancy were 2.8 (95% CI 1.9-4.1) in singleton IVF pregnancies and 4.4 (95% CI 2.4-8.3) in multiple IVF pregnancies, compared with reference pregnancies. The venous thrombosis incidence rate ratio post-partum was 1.2 (95% CI 0.6-2.8) for singleton IVF pregnancies and 3.9 (95% CI 1.7-8.8) for multiple IVF pregnancies compared with reference pregnancies. The post-partum venous thrombosis risk was higher in multiple IVF pregnancies compared with singleton IVF pregnancies. Maternal age, smoking and parity did not significantly affect the venous thrombosis risk. Ovarian hyperstimulation syndrome and polycystic ovarian syndrome did increase the risk of venous thrombosis during pregnancy. Caesarean section also increased the post-partum venous thromboembolism risk, but the increase was not significant. LIMITATIONS, REASONS FOR CAUTION: Other known confounders in our reference population could have contributed to the results. Access to such data may have helped to explain the observations, but would not have changed the conclusion that IVF pregnancies have an increased risk of venous thrombosis compared with other pregnancies. WIDER IMPLICATIONS OF THE FINDINGS: Our study adds new insights by demonstrating an excess venous thrombosis incidence post-partum after in vitro fertilization. The high venous thrombosis incidence in first trimester after in vitro fertilization supports previous studies. Our findings are generalizable to other Western Countries. STUDY FUNDING/COMPETING INTERESTS: Expenses for the acquirement of data were covered by a grant from The Secretary of Doctors further education, Central Denmark Region. None of the authors have any competing interests to declare. TRIAL REGISTRATIONS NUMBER: Not applicable.

Thromboelastometry as a supplementary tool for evaluation of hemostasis in severe sepsis and septic shock.

BACKGROUND: Sepsis leads to disruption of hemostasis, making early evaluation of coagulation essential. The aim of this study was to provide a detailed investigation of coagulation and the use of blood products in patients with severe sepsis or septic shock, admitted to a multidisciplinary intensive care unit. METHODS: Thirty-six patients with severe sepsis or septic shock were included in this prospective observational study. Blood samples and information on transfusion of blood products were obtained for up to 3 consecutive days, and day 7 if the patient was still in the intensive care unit. Thromboelastometry (ROTEM(®)), analyses of thrombin generation, and conventional coagulation tests were performed. RESULTS: ROTEM(®) revealed an overall normo-coagulable state among patients with severe sepsis or septic shock. Conventional coagulation analyses showed divergent results with hypercoagulable trends in terms of reduced antithrombin and acute phase response with increased fibrinogen and fibrin d-dimer, and on the other hand, coagulation disturbances with a decreased prothrombin time and prolonged activated partial thromboplastin time. This hypocoagulabe state was supported by a delayed and reduced thrombin generation. Twelve patients experienced 21 independent transfusion episodes with fresh frozen plasma. Of these, only five (22%) transfusions were performed because of active bleeding. CONCLUSION: ROTEM(®) demonstrated an overall normo-coagulation, whereas the conventional coagulation tests and thrombin generation analyses mainly reflected hypocoagulation. Given the dynamic and global features of ROTEM(®), this analysis may be a relevant supplementary tool for the assessment of hemostasis in patients with severe sepsis or septic shock.

No evidence that assisted reproduction increases the risk of thrombosis: a Danish national cohort study.

BACKGROUND: Case reports have reported venous and arterial thromboses in women undergoing assisted reproduction. No large systematic studies on the risk of thrombosis have been published. The objective of our study was to investigate whether the risk of thrombosis is increased in women undergoing assisted reproduction. METHODS: A national register-based cohort study was conducted on all women undergoing IVF or ICSI treatment in Denmark from 1994 to 2005. Data were obtained from the National Patient Registry and the IVF Registry. Women with prior malignant or cardiovascular disease were excluded. Thrombosis occurring within the first 6 and 12 months after assisted reproduction was considered potentially related to the treatment. Thromboses during pregnancy as well as the pregnancy-related diagnoses were excluded from the statistical analysis. The incidence rates of venous and arterial thromboses were compared with previously published estimates of the risk of thrombosis among young Danish women. RESULTS: We analyzed 30 884 Danish women undergoing 75 141 treatments from 1994 to 2005. The mean age of the women at first treatment was 32.3 years. The delivery rate per cycle was 22%. The incidence rate ratio, with 95% confidence interval (CI), of venous thrombosis within 6 months was 0.95 (CI: 0.38-1.95). The incidence rate ratio of arterial thrombosis within 6 months was 0.36 (CI: 0.04-1.30). CONCLUSIONS: Our study showed no evidence that assisted reproduction increases the risk of thrombosis.

Urgent reversal of vitamin K antagonist therapy.

In the developed world, an increasing number of patients receive therapy with vitamin K antagonists (VKA). This group of patients poses an additional challenge in the perioperative management of emergency surgery and trauma. The present review offers a detailed description of some treatment options for reversal of VKA therapy. Optimal treatment of the anticoagulated patient requires a well-balanced intervention securing a reduced risk of haemorrhagic surgical complications as well as optimal anticoagulation post-operatively without exposing the patient to an increased risk of thromboembolic complications. The following factors must be considered in VKA-treated patients scheduled for emergency surgery: (1) the indication for VKA therapy, including the risk of thromboembolic events when the International normalized ratio (INR) is reduced, (2) type of surgery, including the risk of haemorrhagic complications and (3) the pharmacodynamic/-kinetic profile of the therapy used to revert the VKA therapy. Therapeutic options for acute reversal of VKA therapy include: vitamin K, fresh frozen plasma (FFP), prothrombin complex concentrate (PCC) and perhaps activated recombinant factor VII. PCC is a relatively new drug in some European countries and clinical experience is limited compared with the use of FFP. Reversal of VKA anticoagulation with PCC is faster and more efficient compared with FFP, but there are currently no randomized studies demonstrating an improved clinical outcome.

Assay discrepancy in mild haemophilia A: entire population study in a National Haemophilia Centre.

Assay discrepancy in mild haemophilia, here defined by a significantly higher factor VIII (FVIII):C response by the one-stage procoagulant assay as compared with a two-stage enzymatic method, has repeatedly been reported in literature. The purpose of this study was to determine the overall prevalence of this phenomenon amongst mild haemophilia families from a population of 2.95 million inhabitants in the Western Danish region. Information was collected retrospectively through a thorough search of archives of the National Haemophilia Centre in Aarhus. We identified 109 patients with mild haemophilia A amongst whom 92 were eligible to enter the study. These represent a total of 53 unrelated families. Our data illustrate that this assay discrepancy pattern is found quite frequently amongst our mild haemophilia A families. While the ratio of FVIII:C chromogenic/FVIII:C clot values was quite consistent amongst patients belonging to same family pattern, ratios in the entire cohort of families ranged from 0.18 to 1.00. Selecting a cut-off level for the FVIII:C chromogenic/FVIII:C clot ratios at 0.7, 0.6 and 0.5, respectively, we found that 38 (72%), 27 (51%) and 19 (36%) of families, respectively, displayed this assay discrepancy. In 10 patients, the FVIII:C chromogenic level was inside the category of moderate haemophilia at >0.01-<0.05 IU mL(-1), pointing to a class-shift in the biochemical phenotype. In conclusion, our data illustrate a substantial prevalence of the assay discrepancy phenomenon amongst mild haemophilia A patients in our geographical area.

Tranexamic acid combined with recombinant factor VIII increases clot resistance to accelerated fibrinolysis in severe hemophilia A.

BACKGROUND: Most patients with severe hemophilia A suffer from a profoundly compromised hemostatic response. In addition to both the delayed and slow development of a clot, previous studies have documented that severe hemophilia A is also associated with reduced clot stability. OBJECTIVES: We examined whether the clot stability in hemophiliacs could be improved by treatment with tranexamic acid (TXA) in combination with recombinant factor VIII (rFVIII). PATIENTS/METHODS: Baseline blood samples were obtained from eight males with severe hemophilia A. Thereafter, a bolus injection of rFVIII was administered to increase the functional level of FVIII to approximately 50%. After 10 min, blood was collected followed by an intravenous injection of TXA. A third blood sample was obtained after a further 10 min. Whole blood clotting profiles were determined by thrombelastography using minimal tissue factor activation. Clot formation was assessed by both clot initiation and clot propagation. At clot termination, the maximum clot firmness and area under the elasticity curve were used to illustrate clot stability. Tissue-plasminogen activator was included in those experiments designed to assess clot stability. RESULTS: As expected, rFVIII increased clot formation, whereas TXA had no effect upon this parameter. Assays including tissue-plasminogen activator revealed that rFVIII increased the maximum clot firmness 3-fold; whereas the presence of TXA induced an additional 4-fold increase. The area under the elasticity curve increased 5-fold after rFVIII and 24-fold after addition of TXA. CONCLUSIONS: The study demonstrates that simultaneous treatment with TXA and rFVIII significantly improves the clot stability in patients with hemophilia A.

Measurement of platelet aggregation, independently of patient platelet count: a flow-cytometric approach.

Essentials Platelet function may influence bleeding risk in thrombocytopenia, but useful tests are needed. A flow cytometric platelet aggregation test independent of the patient platelet count was made. Platelet aggregation was reduced in thrombocytopenic patients with hematological cancer. High platelet aggregation ruled out bleeding tendency in thrombocytopenic patients. SUMMARY: Background Methods for testing platelet aggregation in thrombocytopenia are lacking. Objective To establish a flow-cytometric test of in vitro platelet aggregation independently of the patient's platelet count, and examine the association of aggregation with a bleeding history in thrombocytopenic patients. Patients/methods We established a flow-cytometric assay of platelet aggregation, and measured samples from healthy individuals preincubated with antiplatelet drugs, and samples from two patients with inherited platelet disorders. Then, we included 19 healthy individuals and 20 patients with platelet counts of ≤ 50 × 109 L-1 , diagnosed with acute myeloid leukemia or myelodysplastic syndrome. We measured platelet aggregation and platelet activation by platelet surface expression of activated glycoprotein IIb-IIIa, P-selectin and CD63 after addition of agonists: collagen-related peptide, thrombin receptor-activating peptide (TRAP), and ADP. Results The platelet aggregation assay showed a low intraserial coefficient of variation of ≤ 3%. Similar results were obtained for platelet-rich plasma and isolated platelets at platelet counts of > 10 × 109 L-1 ; otherwise, platelet isolation was required. The platelet aggregation percentage decreased with increasing antiplatelet drug concentration. Platelet aggregation in patients was reduced as compared with healthy individuals: 42% (interquartile range [IQR] 27-58) versus 66% (IQR 60-67) for TRAP; 41% (IQR 25-48) versus 70% (IQR 69-72) for collagen-related peptide; and 44% (IQR 30-53) versus 65% (IQR 46-72) for ADP. Platelet activation after stimulation was reduced in patients and correlated with platelet aggregation (e.g. r = 0.78-0.81 when stimulated with collagen-related peptide). Platelet aggregation had a negative predictive value of 100% for a bleeding tendency among patients. Conclusion The established platelet aggregation assay was applicable for thrombocytopenic patients, and improved the identification of bleeding risk.

PO-20 - Crosstalk between the lectin pathway and haemostasis in patients with pulmonary cancer.

INTRODUCTION: Recent research has focused on the complement system in cancer, including the lectin pathway of complement activation. Mannose-binding lectin (MBL), a key activator of the lectin pathway, can bind to tumor cell surfaces in vitro, and lectin pathway activation is increased in several types of cancer. The exact role of the complement system in cancer is currently discussed. However, one possible consequence of the increased complement activation could be contribution to the increased thrombosis risk which cancer patients experience. Proteins of the lectin pathway can activate coagulation and impair fibrinolysis in vitro, but the significance of this in a clinical setting is not well understood. AIM: We aim to investigate associations between lectin pathway and haemostatic activation in patients with lung cancer undergoing thoracoscopic surgery. MATERIALS AND METHODS: Patients with lung cancer (n=60) eligible for thoracoscopic tumor resection were included as part of a randomized controlled trial, the COPPVATS project (EudraCT no: 2012-002409-23), conducted at the Department of Thoracic Surgery, Aarhus University Hospital. Exclusion criteria were arterial or venous thrombosis within the last three months, other systemic disease than cancer, and anticoagulant treatment prior to inclusion. Blood samples were obtained the day before surgery, perioperatively, and on the 1st and 2nd postoperative days. Laboratory analyses on the complement system include MBL, MBL-associated protease (MASP)-1 and -2, MBL-MASP complex, ficolin-1, -2, and -3 and complement factor C3b. Haemostasis was evaluated with routine coagulation parameters (INR, APTT, fibrinogen, fibrin d-dimer), platelet function, and tissue factor-induced thrombin generation. RESULTS: Recruitment of the study subjects is concluded, and laboratory work is in progress. The complement analyses and data processing will be performed during early spring 2016, so that results will be ready for presentation on the conference. CONCLUSIONS: The present project will provide new knowledge on lectin pathway activation and the pathogenesis of thrombosis in cancer patients. In the long term, this will help improve the individual risk assessment and lead to new studies on thromboprophylaxis and treatment in conditions with increased complement activation.

PO-62 - Remote ischemic preconditioning in head and neck cancer reconstruction - a randomized controlled trial.

INTRODUCTION: In reconstructive head and neck cancer surgery, tissue flaps are transferred to the surgical defect and revascularized by anastomosis of small vessels. Cancer patients are in a hypercoagulable state with high risk of peri- and postoperative thrombotic events. Thrombosis to the tissue flap anastomoses or microcirculation is the main reason for total flap necrosis with potential fatal consequences for the patient. Remote ischemic preconditioning (RIPC), where brief cycles of upper extremity ischemia are induced with an inflatable tourniquet, triggers a global protection of tissues subjected to ischemia-reperfusion injury. RIPC has also been shown to impact the coagulation system. AIM: The aim of the trial is to investigate, if RIPC attenuates platelet aggregation during reconstructive head and neck cancer surgery. MATERIALS AND METHODS: Sixty patients with head and neck cancer will be included in the trial. The subjects will be randomized to RIPC or sham during surgery. RIPC is administered by four 5-minute cycles of upper extremity ischemia, each separated by five minutes of reperfusion. Blood samples will be drawn preoperatively, before RIPC/sham, 3 hours after RIPC/ sham, 6 hours after RIPC/sham, and on the first postoperative day. Platelet aggregation will be measured with the Multiplate® analyzer using collagen, ADP and TRAP as agonists. Furthermore, platelet count, mean platelet volume, immature platelet fraction, and von Willebrand factor antigen are determined. RESULTS: The trial is ongoing. Preliminary results will be presented at ICTHIC 2016. CONCLUSIONS: If RIPC proves to attenuate platelet aggregation, it could become a novel antithrombotic treatment in oncologic reconstructive surgery. Hence, morbidity and mortality related to surgery is reduced, and adjuvant oncologic therapy can be initiated in timely fashion.

Platelet count is associated with cardiovascular disease, cancer and mortality: A population-based cohort study.

INTRODUCTION: Platelet count is used to determine bleeding risk and monitoring thrombopoiesis. While abnormal platelet counts are associated with mortality and morbidity, it is unclear whether it also apply to platelet counts within reference range. We investigated the relationship between platelet count (100-450×109/L) and mortality, development of future cardiovascular disease (myocardial infarction, ischaemic stroke, or peripheral vascular disease), venous thromboembolism, bleeding or cancer in the general population. MATERIAL AND METHODS: We conducted a register-based cohort study of 21,252 adults (≥20years) from the Danish General Suburban Population Study (GESUS). Laboratory results from GESUS were linked to information from national registers regarding morbidity and death. Cox proportional hazard regression was conducted with adjustment for age, sex, smoking status, haemoglobin, leukocyte count, C-reactive protein and Charlson comorbidity index. RESULTS: We found a U-shaped relationship between mortality and platelet count. Mortality was significantly increased for platelet count <175×109/L or >300×109/L. When categorizing platelet count using the interval 201-250×109/L as reference group, platelet count 301-450×109/L was associated with mortality, adjusted hazard ratio (HR)=1.42(95% CI 1.06-1.90) and cardiovascular disease, adjusted HR=1.32 (95% CI 1.03-1.69). Platelet count 100-200×109/L was associated with future cancer, adjusted HR=1.28(95% CI 1.05-1.57), but not with future bleeding or venous thromboembolism. CONCLUSIONS: Platelet count is associated with mortality, future cardiovascular disease, and future cancer.

Increased plasma methylmalonic acid level does not predict clinical manifestations of vitamin B12 deficiency.

BACKGROUND: The prevalence of vitamin B(12) deficiency, defined as an elevated concentration of plasma methylmalonic acid (P-MMA), has been estimated to be 15% to 44% in the elderly. However, we do not know whether an increased P-MMA level actually indicates or predicts a clinical condition in need of treatment. PARTICIPANTS AND METHODS: In a follow-up study, 432 individuals not treated with vitamin B(12) were examined 1.0 to 3.9 years after initial observation of an increased P-MMA concentration (>0.28 micromol/L). The examination included laboratory tests, a structured interview to disclose symptoms, a food frequency questionnaire, and a clinical examination including a Neurological Disability Score. RESULTS: Variation in P-MMA levels over time was high (coefficient of variation, 34%). In only 16% of participants, P-MMA levels increased substantially, whereas 44% showed a decrease. Level of P-MMA was significantly but not strongly associated with levels of plasma cobalamins (r = -0.22, P<.001) and plasma total homocysteine (r = 0.37, P<.001). After adjustment for age and sex, we found no associations between P-MMA concentration and the total symptom score (P =.61), the total Neurological Disability Score (P =.64), or other clinical manifestations related to vitamin B(12) deficiency. CONCLUSIONS: An increased level of P-MMA did not predict a further increase with time and clinical manifestations related to vitamin B(12) deficiency. We therefore challenge the use of an increased P-MMA concentration as the only marker for diagnosis of vitamin B(12) deficiency.

Preconception venous thromboembolism and placenta-mediated pregnancy complications.

BACKGROUND: Placenta-mediated complications are leading causes of maternal and fetal morbidity and mortality. We hypothesized that a preconception history of venous thromboembolism (VTE) is associated with increased risk of placenta-mediated pregnancy complications. METHODS: A nationwide population-based cohort study of all singleton pregnancies leading to delivery from 1997 to 2012 (n = 964 967). We obtained data on placenta-mediated pregnancy complications from the Danish Medical Birth Registry and data on VTE before pregnancy from the Danish National Patient Registry. We computed absolute risks, crude and adjusted risk differences (RDs) using a binomial regression model, and crude and adjusted risk ratios (RRs) from a modified Poisson regression model. RESULTS: Overall, 1419 women had a preconception history of VTE, while 578 112 did not. Preeclampsia occurred in 4.2% of pregnancies in the VTE group and in 2.7% of pregnancies in a comparison cohort (adjusted RD = 1.3%, 95% confidence interval (CI) 0.6-2.0%; adjusted RR = 1.5, 95% CI 1.3-1.8). Stillbirth occurred in 0.7% of pregnancies in the VTE group and in 0.4% of pregnancies in the comparison cohort (adjusted RD = 0.3%, 95% CI 0.02-0.6%; adjusted RR = 1.8, 95% CI 1.1-3.0). Placental abruption occurred in 0.8% of pregnancies in the VTE group and in 0.5% of pregnancies in the comparison cohort (adjusted RD = 0.3%, 95% CI - 0.05-0.6%; adjusted RR = 1.6, 95% CI 1.1-2.4). Small-for-gestational-age infants accounted for 10.9% of live births in the VTE group and 9.8% of live births in the comparison cohort (adjusted RD = 0.6%, 95% CI - 0.5-1.7%; adjusted RR = 1.1, 95% CI 0.9-1.3). CONCLUSION: Women with a history of VTE were at increased risk of placenta-mediated complications.

The influence of type 2 diabetes on fibrin clot properties in patients with coronary artery disease.

Type 2 diabetes mellitus (T2DM) increases the risk of coronary thrombosis and both conditions are associated with altered fibrin clot properties. However, the influence of T2DM on fibrin clot properties in patients with coronary artery disease (CAD) remains unclear. We aimed to investigate the influence of T2DM on fibrin clot properties in patients with CAD. Fibrin clot structure and fibrinolysis were investigated in 581 CAD patients (148 with T2DM) using turbidimetric assays, confocal and scanning electron microscopy. Clots made from plasma and plasma-purified fibrinogen were studied, and plasma levels of inflammatory markers were analysed. T2DM patients had increased clot maximum absorbance compared with non-diabetic patients (0.36 ± 0.1 vs 0.33 ± 0.1 au; p=0.01), displayed longer lysis time (804 [618;1002] vs 750 [624;906] seconds; p=0.03) and showed more compact fibrin structure assessed by confocal and electron microscopy. Fibrinogen levels were elevated in T2DM (p< 0.001), but clots made from purified fibrinogen showed no differences in fibrin properties in the two populations. Adjusting for fibrinogen levels, T2DM was associated with C-reactive protein and complement C3 plasma levels, with the former correlating with clot maximum absorbance (r=0.24, p< 0.0001) and the latter with lysis time (r=0.30, p< 0.0001). Independent of fibrinogen levels, females had more compact clots with prolonged lysis time compared with males (all p-values< 0.001). In conclusion, T2DM is associated with prothrombotic changes in fibrin clot properties in patients with CAD. This is related to quantitative rather than qualitative changes in fibrinogen with a possible role for inflammatory proteins.

Reduced antiplatelet effect of aspirin is associated with low-grade inflammation in patients with coronary artery disease.

Inflammation has been proposed to modify platelet function. This may lead to increased platelet reactivity and reduced antiplatelet drug efficacy in patients with coronary artery disease (CAD). However, this hypothesis has not been investigated in stable CAD patients receiving aspirin as mono antiplatelet therapy. It was the objective of this study to investigate the association between platelet reactivity, the inflammatory markers high-sensitive C-reactive protein (hs-CRP) and interleukin-6 (IL-6), and platelet activation. We performed a cross-sectional study on 524 stable high-risk CAD patients. Among these, 91% had a history of myocardial infarction, 23% had type 2 diabetes, and 13% had both. All patients received 75 mg aspirin daily as mono antiplatelet therapy. Platelet reactivity was assessed by multiple electrode aggregometry (Multiplate®, MEA) and VerifyNow®. Inflammation was evaluated by hs-CRP and IL-6. Platelet activation was assessed by soluble P-selectin (sP-selectin), and cyclooxygenase-1 inhibition was evaluated by measurement of serum thromboxane B2. Hs-CRP levels were significantly higher in upper platelet reactivity tertile patients than in lower platelet reactivity tertile patients (p≤0.02). Similar results were obtained with IL-6, though not statististically significant (p≥0.15). Platelet activation evaluated by sP-selectin was significantly higher in patients with MEA reactivity levels in the upper tertile than in the lower tertile (p=0.0001). Optimal compliance was confirmed by low serum thromboxane B2 levels in all patients. In conclusion, increased levels of hs-CRP were associated with augmented platelet reactivity in stable high-risk CAD patients receiving aspirin as mono antiplatelet therapy. These findings may suggest that chronic low-grade inflammation reduce the antiplatelet effect of aspirin.

Effect of platelet turnover on whole blood platelet aggregation in patients with coronary artery disease.

BACKGROUND: Previous studies have demonstrated considerable variation in the antiplatelet effect of aspirin. OBJECTIVES: To investigate the impact of platelet turnover on the antiplatelet effect of aspirin in patients with stable coronary artery disease (CAD) and to identify determinants of platelet turnover. METHODS: Platelet turnover was evaluated by measurements of immature platelets and thrombopoietin in 177 stable CAD patients on aspirin monotherapy, including 85 type 2 diabetics and 92 non-diabetics. Whole blood platelet aggregation was determined using the VerifyNow(®) Aspirin test and multiple electrode aggregometry (MEA, Multiplate(®) ) induced by arachidonic acid (AA) (1.0 mm), adenosine diphosphate (ADP) (10 µm) and collagen (1.0 µg mL(-1) ). RESULTS: Immature platelet levels significantly correlated with MEA (r = 0.31-0.36, P-values < 0.0001) and the platelet activation marker sP-selectin (r = 0.19, P = 0.014). Contrary to the VerifyNow(®) test, MEA significantly correlated with variations in platelet count (r = 0.45-0.68, P-values < 0.0001). Among patients with residual platelet reactivity according to AA, there were significantly more diabetics (61% vs. 41%, P = 0.027) and higher levels of sP-selectin (77.7 ± 29 vs. 70.2 ± 25 ng mL(-1) , P = 0.070) and serum thromboxane B(2) (0.81 [0.46; 1.70] vs. 0.56 [0.31; 1.12] ng mL(-1) , P = 0.034). In a multivariate regression analysis, immature platelet levels were determined by thrombopoietin levels (P < 0.001), smoking (P = 0.020) and type 2 diabetes (P = 0.042). CONCLUSIONS: The antiplatelet effect of aspirin was reduced in CAD patients with an increased platelet turnover. Once-daily dosing of aspirin might not suffice to adequately inhibit platelet aggregation in patients with an increased platelet turnover.