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Background: Von Willebrand factor (VWF) levels are regulated by genetic and acquired factors. The acquired factors are mostly related to age and could be mediators of the age effect on VWF levels., Objectives: To disentangle the role of genetic (sex, blood group) and acquired factors (comorbidities, body mass index, reduced kidney function, hormone use, and inflammation) in regulating von Willebrand factor antigen (VWF:Ag) and factor VIII activity (FVIII:C) levels in the normal population., Methods: Analysis were performed in a large population sample (2923 individuals) from the Multiple Environmental and Genetic Assessment of risk factors for venous thrombosis (MEGA study), after exclusion of individuals with active cancer and women who were pregnant or within nine months postpartum. The increase of VWF:Ag and FVIII:C with age was evaluated by linear regression after the age of 40 years. Analyses were adjusted for acquired factors and stratified for sex and blood group., Results: VWF:Ag and FVIII:C increased with age: increase per decade of age for VWF:Ag 18 IU/dL (95%CI 15-20) and for FVIII:C 12 IU/dL (95%CI 10-14). After adjustment for acquired factors, the increase per decade was 13 IU/dL (95%CI 10-16) for VWF:Ag and 9 IU/dL (95%CI 6-11) for FVIII:C. The stratified analysis for blood group showed higher increase in the non-O group, but these differences were annulled after adjustment for acquired factors., Conclusions: VWF:Ag and FVIII:C increase with age. Carriers of blood group non-O present a steeper increase of VWF:Ag and FVIII:C with age, that is mediated by acquired factors., (Copyright © 2020. Published by Elsevier Ltd.)

Background: Acquired von Willebrand Syndrome is a rare bleeding disorder, which arises in individuals with no personal or family history of bleeding, associated with lymphoproliferative and myeloproliferative disorders or other diseases., Aim: To develop a two-step approach assay to detect autoantibodies against VWF and to verify their prevalence in AVWS., Methods: AVWS definition: negative personal or family history of bleeding diathesis, VWF below normal range and recent history of bleeding manifestations. Twenty-three consecutive patients affected by AVWS were enrolled. An ELISA assay (first step) using recombinant VWF protein immobilized on plates and sheep/goat polyclonal anti-human IgG or IgM labelled with peroxidase was developed. A group of 40 healthy subjects was tested to calculate the floating cut point value. A confirmation assay (with addition of purified VWF vs buffer) was performed (second step)., Results: Twenty-one patients (93%) had an associated disease, two patients had idiopathic AVWS. Anti-VWF autoantibodies were detected in 9 patients (39%). Of these, eight (89%) had VWF:RCo levels <10%, but none of them resulted positive using Bethesda assay (neutralizing antibodies). The confirmation test confirmed the positive results obtained with ELISA and resulted negative in those patients with negative results and in the controls., Conclusion: With the present two-step approach assay nine out of 23 (39%) patients affected with AVWS resulted positive for anti-VWF autoantibodies. This ELISA assay might be used as an additional confirmation tool in the diagnostic procedure in patients affected by AVWS or in the follow-up of congenital and acquired patients exposed to replacement therapy., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Introduction: Antithrombin (AT), protein C (PC) and protein S (PS) deficiencies are risk factors for venous thromboembolism. Overlapping values between heterozygous carriers and normal individuals often make a correct classification of a deficiency difficult. The aim of this study was to investigate the effect of sex, age, menopause and hormone therapy on natural anticoagulant plasma levels in a large group of healthy individuals, and to evaluate the need of separate reference ranges., Materials and Methods: AT and PC were measured with a chromogenic assay, antigenic free PS with an ELISA test. To evaluate the effect of sex, age, oral contraception, hormonal status (and their interaction) on AT, PC and PS levels, linear regression models were used. Biological relevance and the value of the normal deviate z were chosen as rules to decide for separate reference ranges., Results: The study population consisted of 1837 healthy adult individuals (741 men, 1096 women), aged 18-85 years (median age: 44 years). In men AT levels decreased after the age of 50 years. Men had higher levels of PS than women, particularly at young ages. In women, after correction for menopause, only PC levels increased with age. Menopause affected AT and PS, but not PC levels. Oral contraceptive intake was associated with a decrease of AT and PS, and an increase of PC levels., Conclusions: For AT, PC and PS, sex- and age-specific normal reference ranges can be useful, in order to better discriminate true carriers of a natural anticoagulant deficiency., (© 2013.)

Introduction: Postpartum hemorrhage is responsible for 25% of maternal pregnancy-related deaths and it is the first cause of maternal morbidity and mortality worldwide., Objective: To define the prevalence of postpartum hemorrhage and associated risk factors after vaginal birth and to develop a risk model that improves postpartum hemorrhage prediction., Patients and Methods: All women who underwent a vaginal delivery at the Obstetric Unit of a large University hospital in Milan (Italy) between July 2007 and September 2009 were enrolled. Postpartum hemorrhage was defined as ≥ 500 mL blood loss. A nomogram tailored to predict postpartum hemorrhage was developed, summarizing the impact of each covariate on the probability of postpartum hemorrhage., Results: 6011 women were studied (24% had blood loss ≥ 500 mL and 4.8% ≥ 1000 mL). Nulliparity, episiotomy, retained placenta and high neonatal body weight were confirmed as risk factors for postpartum hemorrhage. The odds ratio of postpartum hemorrhage was 0.86 (95%CI 0.78-0.90) for each 1 gr/dL increase in ante-partum hemoglobin. An extensive internal validation of the developed nomogram demonstrated a good stability of the risk model., Conclusions: Low ante-partum hemoglobin is a new potentially modifiable risk factor for postpartum hemorrhage. A nomogram to predict the probability of postpartum hemorrhage is now available for external validation., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background: The usefulness of coagulation tests performed before epidural analgesia for surgery or to alleviate labour pain is controversial. The aims of this study were: (1) to evaluate the prevalence of abnormal tests in a large cohort of healthy pregnant women and their association with epidural hematoma; (2) to assess the approach of the anesthesiologists to women with abnormal tests; (3) to evaluate the cost of performing coagulation tests before epidural analgesia in all healthy pregnant women., Methods: Data regarding epidural analgesia, epidural hematoma, PT, APTT, fibrinogen and platelet count were extracted from medical charts., Results: There was no case of epidural hematoma in 2546 pregnant women undergoing epidural analgesia. PT and APTT results were obtained in 2871 women; fibrinogen in 4063 women; platelet count in 5090 women. Three of them (0.1%) had a prolonged PT, 4 (0.14%) had a prolonged APTT, 27 (0.53%) had platelets ≤ 100 × 10(9)/L and 37 (0.91%) had plasma fibrinogen levels <3 g/L. No further tests were requested by the anesthesiologists in these women. Only women with platelets <80 × 10(9)/L were denied epidural analgesia. Based on the data from the literature on the frequencies of epidural hematoma after epidural analgesia, a total cost ranging from 4.5 to 40 million Euros to perform coagulation tests would be necessary to avoid one case of epidural hematoma., Discussion: Unselected coagulation tests before epidural analgesia are not recommended, because epidural hematoma is extremely rare in healthy pregnant women and the cost of screening is not justified., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background: Results for lupus anticoagulant (LA) are currently expressed as ratio of patient-to-normal clotting times (LA-ratio). Yet, numerical results do vary according to the method used for testing, thus making difficult the between-method comparison of results. We hypothesized that the standardization model currently used for the INR for patients on oral-anticoagulants (OAT) would be of value also for LA standardization., Patients and Methods: To test this hypothesis we determined a sensitivity index valid for LA (called LASI) for six LA-detection methods against a common-standard using two sets of calibration-plasmas: (i)normal-plasmas spiked with IgG derived from patients strongly-positive for LA or (ii)plasmas from LA-positive patients. The LASI was then used to convert the LA-ratio into the standardized-LA-ratio (SLA-ratio) according to the equation: SLA-ratio = (LA-ratio)(LASI)., Results: We demonstrate that (i)the model is feasible because calibration plots of log-transformed clotting times obtained for the LA-detection methods-vs.-the common-standard gave acceptable LASI values; (ii)the model is effective because between-method variability expressed as coefficient of variation, which was 42.8% with results expressed as LA-ratio, decreased to 7.8% with results expressed as SLA-ratio; (iii)the LASI value calculated with the LA-positive plasmas is more effective in minimizing between-method variability than the LASI value calculated with IgG-spiked plasmas., Conclusions: A model of LA calibration similar to the INR for patients on OAT is feasible by using plasmas from LA-positive patients instead of patients on OAT. Potential application of the model are:(i)to compare the relative responsiveness of different LA-detection methods,(ii)to minimize differences between their results and (iii)to quantify LA potency., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The HemosIL ThromboPath assay (Instrumentation Laboratory) is a new chromogenic assay designed to globally evaluate the functionality of the protein C (PC) pathway. It is based on the ability of endogenous APC generated after activation of PC by a snake venom extract (Protac) to reduce the thrombin generation induced by a reagent containing tissue factor. The aim of this multicenter study involving three laboratories was to evaluate the test sensitivity to PC pathway abnormalities by retrospectively testing frozen plasma samples obtained in the different laboratories. Test results were significantly lower (p < 0.0001) in subjects who presented with any confirmed PC pathway abnormality than in those without. The cut-off value, defined in each participating center as the mean value minus one standard deviation of test results obtained in 30 normal samples, was found to provide a sensitivity-to-specificity ratio similar to that obtained using ROC-analysis. The assay performed well in carriers of the factor V Leiden mutation (n = 81), patients with PC deficiency (n = 40), combined defects (n = 55) or lupus anticoagulant (n = 44), with test results below the locally defined cut-off values in 97.5%, 95.0%, 100% and 100% of the tested subjects, respectively. The assay sensitivity for PS deficiency (n = 62) was 87.1%. Only 13.6% of the 272 subjects without any PC pathway abnormality had a decreased test result. So, using the locally defined cut-off values, the overall test sensitivity to all tested PC pathway abnormalities was 95.0% (95%CI = 91.8-97.3), its specificity 86.4% (95%CI = 81.8-90.2), its negative predictive value 94.4% (95%CI = 90.8-96.9) and its positive predictive value 87.9% (95%CI = 83.7-91.3).

Introduction: Thrombomodulin (TM) and endothelial protein C receptor (EPCR) are two transmembrane endothelial receptors involved in the protein C pathway, that regulates coagulation and inflammation processes. We postulated that soluble thrombomodulin and EPCR are plasmatic markers of progression to hepatocellular carcinoma (HCC) and prognostic indicators in cirrhotic patients., Materials and Methods: Plasma levels of TM and EPCR were measured in 104 patients affected by different stages of liver diseases (66 patients with HCC, and 38 without HCC), and in 52 healthy controls., Results: EPCR levels were higher in patients than in controls (239+/-1.8 ng/mL vs. 127+/-1.5 ng/mL, p<0.0001). TM levels were higher in patients with HCC than in those without (42.1+/-2.0 ng/mL vs. 28.3+/-2.1 ng/mL; p=0.039), while EPCR levels were similar in the two groups. No association between TM and clinical outcome was found, while high levels of EPCR were associated with death and thrombosis of the portal vein., Conclusions: We surmise a possible role for high levels of TM as a marker of HCC development in patients with cirrhosis, whereas high levels of EPCR are a possible marker of worse HCC prognosis, being a sign of endothelial damage of large vessels.