Time in therapeutic range is lower in women than in men and is not explained by differences in age or comorbidity.

BACKGROUND: Time in therapeutic range (TTR) measures the stability of the international normalized ratio in patients on vitamin K antagonists (VKA). Low values are associated with poor outcomes. Women were shown to have lower TTR than men, but the causes are poorly defined. It was suggested that women on VKA are older and more morbid than men, and this could affect the stability of anticoagulation. We aimed to identify variables that affect TTR differently in women and men. MATERIALS AND METHODS: This is a retrospective study in patients referred to a University hospital anticoagulant clinic. Age, sex, comorbidities, number of daily medications, indication and type of anticoagulant, weekly dosage and distribution, were derived from electronic records. Differences by sex and regression analysis to identify significant modulators of TTR were computed. RESULTS: 1182 women and 1281 men on VKA were studied. Women were older than men (81.5 yrs. ± 11.2 vs 78.4 yrs. ± 12.2), and had lower TTR (65% ± 20.3 vs 69% ± 19.8). Comorbidity was similar between sexes and negatively affected TTR in both. Mechanical valves as an indication to anticoagulation and acenocoumarol as an anticoagulant as opposed to warfarin had a strong negative influence on TTR, while age increased TTR. Being a man rather than a woman afforded more than three TTR points. Number of medications and average anticoagulant dose were equal between sexes. DISCUSSION: Women have a lower TTR than men, on average below the safety threshold. They were indeed older, but age positively influenced TTR. Since women and men were equally comorbid, neither age nor disease explains differences in TTR. None of the other variables included in the study could explain the gender gap in TTR. Since women are at increased risk of cardioembolic stroke in atrial fibrillation, an effort at defining other causes for the observed differences, closer monitoring and switching to direct anticoagulants whenever possible is warranted.

Sex, gender and venous thromboembolism: do we care enough?

: The role of sex and gender in determining clinical presentation, diagnostic approach and outcomes of venous thromboembolism is not fully and systematically addressed, except for hormone-related events in women. A lack of knowledge is also apparent regarding drug prescription patterns, physician bias, enrolment in clinical studies and analysis of sex-related confounders in preclinical and clinical studies. As was shown for cardiovascular disease, ignoring sex and gender in medicine can have important impact on outcomes, including mortality. In this review, we seek to address some aspects of venous thromboembolism such as epidemiology and clinical presentation, recurrence, risk factors, animal studies, safety and efficacy of antithrombotic drugs, highlighting what is known and what is not regarding the role of sex and gender, and hoping to focus some interest and to promote the inclusion of these variables in all future studies on venous thromboembolism.

To be or not to be: the patient's view of thrombophilia testing.

INTRODUCTION: The literature on the psychological effects of thrombophilia testing is unclear. Little is known about the complex world of significance subjects construct around the test. OBJECTIVE: The study explored the peculiar network of implicit meanings that may be linked to the experience of being tested. MATERIALS AND METHODS: The research was designed according to Interpretative Phenomenological Analysis (IPA). 19 patients were interviewed. Integral verbatim reports of the interviews were analyzed through an inductive process aimed at gaining a holistic understanding of the narratives. RESULTS: Two main issues were identified, each with sub-issues: (1) the clinical problem: (1.1) unhealthy blood and (1.2) the family issue; (2) the test: (2.1) knowing for the sake of knowing; (2.2) knowing for the sake of doing; (2.3) not knowing. CONCLUSIONS: The thrombophilia test is part of a larger network of meanings, where information about the test and its results seem to be lost. PRACTICE IMPLICATION: The study suggests the importance of paying greater attention to the process of doctor-patient communication at the time of the test. The theme of being informed is important for patients, yet often they are not able to understand or retain the information they receive, increasing the risk of misunderstandings.

Why does ticagrelor induce dyspnea?

In studies that compared the reversible P2Y12 inhibitor ticagrelor with the irreversible inhibitor clopidogrel, dyspnea was observed more frequently among ticagrelor-treated patients than among clopidogrel-treated patients. Because dyspnea was not associated with acidosis, pulmonary or cardiac dysfunction, alterations in the mechanisms and pathways of the sensation of dyspnea may be involved in its pathogenesis. It has been hypothesised that the sensation of dyspnea in ticagrelor-treated patients is triggered by adenosine, because ticagrelor inhibits its clearance, thereby increasing its concentration in the circulation. However, dipyridamole, a much stronger inhibitor of adenosine clearance than ticagrelor, usually does not cause dyspnea. We hypothesise that inhibition of P2Y12 on sensory neurons increases the sensation of dyspnea, particularly when reversible inhibitors are used. We base our hypothesis on the following considerations: 1) cangrelor and elinogrel, which, like ticagrelor, are reversible P2Y12 inhibitors, also increase the incidence of dyspnea; 2) it is biologically plausible that inhibition of P2Y12 on sensory neurons increases the sensation of dyspnea; 3) inhibition of P2Y12 on platelets (which do not have a nucleus) by clopidogrel is permanent, despite the once daily administration and the short plasma half-life of the inhibitor; 4) in contrast, inhibition of P2Y12 on neurons by clopidogrel may be temporary and transient, because neurons have a nucleus and can therefore rapidly replace the inhibited receptors with newly synthetised ones; 5) inhibition of P2Y12 on neurons by reversible inhibitors is permanent, because the plasma drug concentration is maintained high by repeated dosing, in order to ensure permanent inhibition of platelet P2Y12.

Screening for thrombophilia and antithrombotic prophylaxis in pregnancy: Guidelines of the Italian Society for Haemostasis and Thrombosis (SISET).

The term thrombophilia describes an increased tendency to develop thrombosis and many laboratory markers with different strengths of association with thrombosis have been identified. The main causes of maternal mortality and morbidity in developed countries is venous thromboembolism (VTE) and obstetric complications. During pregnancy and puerperium the risk for VTE increases due to hemostatic imbalance towards a prothrombotic state, and it is further increased in women carriers of thrombophilia; recent studies have also demonstrated an association between thrombophilia and obstetric complications. These complications are, therefore, considered potentially preventable with the prophylactic administration of anticoagulant drugs, although their efficacy is not proven by data from randomized controlled trials. After a systematic comprehensive literature review and using a rigorous methodology, the expert panel formulated recommendations regarding the usefulness of screening for thrombophilia in pregnancy to identify high-risk women and for the management of antithrombotic prophyalxis. When evidence is lacking, consensus-based recommendations are provided.

ProCMD: a database and 3D web resource for protein C mutants.

BACKGROUND: Activated Protein C (ProC) is an anticoagulant plasma serine protease which also plays an important role in controlling inflammation and cell proliferation. Several mutations of the gene are associated with phenotypic functional deficiency of protein C, and with the risk of developing venous thrombosis. Structure prediction and computational analysis of the mutants have proven to be a valuable aid in understanding the molecular aspects of clinical thrombophilia. RESULTS: We have built a specialized relational database and a search tool for natural mutants of protein C. It contains 195 entries that include 182 missense and 13 stop mutations. A menu driven search engine allows the user to retrieve stored information for each variant, that include genetic as well as structural data and a multiple alignment highlighting the substituted position. Molecular models of variants can be visualized with interactive tools; PDB coordinates of the models are also available for further analysis. Furthermore, an automatic modelling interface allows the user to generate multiple alignments and 3D models of new variants. CONCLUSION: ProCMD is an up-to-date interactive mutant database that integrates phenotypical descriptions with functional and structural data obtained by computational approaches. It will be useful in the research and clinical fields to help elucidate the chain of events leading from a molecular defect to the related disease. It is available for academics at the URL http://www.itb.cnr.it/procmd/.

Abnormalities of homocysteine and B vitamins in the nephrotic syndrome.

INTRODUCTION: The nephrotic syndrome is associated with heightened risk for arterial and venous thrombosis. Multiple derangements of hemostasis and acquired risk factors such as hyperlipidemia and hypertension contribute to this risk. The prevalence in the nephrotic syndrome of high circulating levels of homocysteine and of low levels of the B vitamins that are involved in its metabolism, which may play a role in thrombosis, is not well defined. MATERIALS AND METHODS: In 84 patients with nephrotic syndrome and 84 sex- and age-matched controls, hemostasis variables and the circulating levels of total homocysteine (tHcy), vitamin B(6), B(12) and folates were measured. RESULTS: tHcy levels were higher, vitamin B(6) and vitamin B(12) levels were lower in nephrotic patients than in controls. The association of low vitamin B(6) levels with the nephrotic syndrome was independent of any other alteration associated with the disease. Eighty-two percent of patients with the nephrotic syndrome had vitamin B(6) levels falling in the lowest quartile of the normal distribution. Antithrombin deficiency, factor V Leiden, antiphospholipid antibodies, hypertension, dyslipidemia, were more frequent in patients with the nephrotic syndrome than in controls. CONCLUSIONS: Patients with the nephrotic syndrome have multiple risk factors for thrombosis. We report that they frequently have low circulating levels of vitamin B(6), which associate with a heightened risk for venous and arterial thrombosis.

Identification and computationally-based structural interpretation of naturally occurring variants of human protein C.

Protein C (PC) is a key regulator of blood clotting and inflammation. Its inherited deficiency is associated with venous thromboembolism, and recombinant activated PC is currently used to increase survival in severe sepsis. The molecular basis of inherited PC deficiency is heterogeneous. Due to its multiple physiologic interactions and functions, and its modular structure, natural variants aid in the understanding of the relationship between critical residues and discrete functions. This knowledge has important therapeutic implications in the planning of a recombinant activated PC with a specific therapeutic target and devoid of major collateral effects. A way of predicting important functional consequences of residue variation is the use of molecular modeling and structural interpretation of amino acidic substitutions. A study of 21 out of 32 identified PC gene (PROC) variants is presented. For three of them, localized in the active site, electrostatic potential variation was calculated. For more than half of the studied variants, an explanation for the functional impairment could be derived from computational analysis, allowing a focused choice of which variants it is worthwhile pursuing.

Association of estrogen receptor-alpha genepolymorphisms with venous thrombosis.

XbaI and PvuII polymorphisms of the estrogen receptor-alpha gene (ERalpha) have been associated with several multifactorial diseases. We studied the distribution of ERalpha polymorphisms in patients with deep vein thrombosis. PvuII PP and XbaI XX genotypes may be associated with an approximately 2-fold increased DVT risk of deep vein thrombosis in men.

Molecular diversity and thrombotic risk in protein S deficiency: the PROSIT study.

The Protein S Italian Team (PROSIT) enrolled 79 protein S (PS) deficient families and found 38 PROS1 variations (19 novel) in 53 probands. Of these, 23 variants were selected for expression in'vitro, to evaluate their role as possible causative variants. Transient expression showed high secretion levels (>75%) for three variants, which were considered neutral. Seven missense and five nonsense variants showed low (

Expression of endothelial protein C receptor and thrombomodulin in the intestinal tissue of patients with inflammatory bowel disease.

OBJECTIVE: Inflammatory bowel diseases are characterized by disorders of immunity, thrombosis of large vessels, and microthrombosis of mucosal vessels. The expression of endothelial protein C receptor (EPCR) and thrombomodulin-two receptors of the protein C pathway involved in thrombin scavenging and inflammation-was studied in intestinal resection specimens or mucosal biopsies from patients with inflammatory bowel disease and from controls. The soluble forms of the receptors in plasma were measured. DATA SOURCE: This study involved patients from two large university hospitals. After surgery or biopsy, tissue samples were either frozen or fixed in formalin and embedded in paraffin. Sections for immunohistochemistry examination were cut and tested with the specific antibodies to EPCR and thrombomodulin. RNA was extracted from frozen tissue for amplification via reverse-transcriptase polymerase chain reaction. Normal intestinal and diverticulitis tissue was used as a control. Resection samples from 36 patients with ulcerative colitis, 38 with Crohn's disease, 38 with colonic cancer, and 32 with diverticulitis were studied by immunohistochemistry, and frozen sections from the same patients were studied by immunofluorescence. Twelve biopsy specimens of adjacent intestinal areas from six patients with inflammatory bowel disease were included in the study for reverse-transcriptase polymerase chain reaction. Soluble receptors were measured in the plasma of 52 inflammatory bowel disease patients and 52 controls. DATA SUMMARY: EPCR and thrombomodulin were expressed on the mucosal endothelium of controls, and the intensity of the signal decreased in inflammatory bowel disease patients. EPCR was expressed by dendritic-like cells in controls, which also stained positive for CD21. The EPCR/CD21 dendritic-like cells were not as commonly observed in sections from ulcerative colitis patients as they were in sections from control patients (12.0 +/- 3.6 cells per high-power field vs. 23.8 +/- 10.4 cells per high-power field, p =.03), and this decrease was less evident in sections from Crohn's disease patients. Levels of messenger RNA for EPCR paralleled protein expression. Soluble thrombomodulin and EPCR levels were both higher in patients than in controls: 41.5 vs. 26.0 ng/mL (p <.0001) and 141 vs. 130 ng/mL (p <.05), respectively. CONCLUSIONS: EPCR expression on dendritic-like cells that bear the key complement receptor CD21 suggests a role for EPCR in innate immunity. The reduced expression of thrombomodulin and EPCR in the mucosal vessels in inflammatory bowel disease impairs protein C activation, favoring microthrombosis.

Expression of endothelial protein C receptor and thrombomodulin in human coronary atherosclerotic plaques.

BACKGROUND: The expression of selected genes in human coronary atherosclerotic plaques may help to clarify the evolution of atherogenesis and the causes of thrombogenesis on some fissured plaques. The aim of this study was to analyze the expression of the genes known to participate in inflammation and hemostasis: thrombomodulin and endothelial protein C receptor, E- and P-selectin, intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), tissue factor and plasminogen activator inhibitor-1 (PAI-1). METHODS: RNA was extracted and reverse-transcribed from 27 atherectomized human coronary atherosclerotic plaques. The genes were specifically amplified together with a housekeeping gene. RESULTS: Thrombomodulin was not expressed in the 8/27 plaques from which RNA could be obtained. The levels of expression of tissue factor, ICAM-1, P- and E-selectin, and PAI-1 were low, whereas those of endothelial protein C receptor and VCAM-1 were high. CONCLUSIONS: RNA may be extracted from ex vivo atherosclerotic plaques. In addition to anticoagulation, endothelial protein C receptor may play an important inflammation-related role in plaque development.

Association of factor V deficiency with factor V HR2.

BACKGROUND AND OBJECTIVES: Factor V HR2 possesses decreased co-factor activity to activated protein C and an increased ratio of factor V1 to factor V2. Factor V HR2 is associated with a mild increase in the risk of venous thromboembolism although not all studies concur on this point. DESIGN AND METHODS: Inconsistencies in results of the epidemiological studies may stem from a failure to identify other variables in factor V which might contribute to an increased risk of thrombosis in selected HR2 carriers. The aim of this study was to establish whether factor V deficiency increases the risk of venous thromboembolism when associated with HR2. RESULTS: Four hundred and ninety-seven patients with venous thromboembolism and 498 controls were studied. HR2 was present in 12.5% of patients and 10.4% of controls. Factor V deficiency was associated with HR2 in 4.6% of patients and 1.0% of controls. The OR for venous thromboembolism in individual with HR2 alone was 1.2 (95% CI 0.8-1.8), while it was 4.7 (95% CI 1.8-12.5) for those with HR2 plus factor V deficiency. INTERPRETATION AND CONCLUSIONS: Patients with HR2 and factor V deficiency developed a thrombotic event earlier (median age 35 years) than patients with HR2 alone (median age 43 years, p = 0.018). Double heterozygosity for HR2 and a factor V defect, including factor V deficiency, increased the thrombotic risk afforded by HR2.

The factor V HR2 haplotype and the risk of venous thrombosis: a meta-analysis.

BACKGROUND AND OBJECTIVES: A complex haplotype of factor V gene (FV HR2) has been recently reported. FVHR2 possesses decreased co-factor activity to APC in the degradation of FVIIIa, and an increased ratio of the more procoagulant isoform FV1 compared to FV2. Contrasting results on whether the haplotype induces a significant risk of venous thromboembolism (VTE) have been reported. DESIGN AND METHODS: It has been surmised that FVHR2 enhances the risk of VTE carried by FV Leiden. We carried out a meta-analysis of the reported studies on the role of HR2 haplotype in inducing a risk of VTE and the influence of the polymorphism on the risk carried by patients with FV Leiden. RESULTS: Eight studies were analyzed for the estimation of the risk of VTE. A total of 338 out of 2,696 cases (12.5%; range 7.8 to 18.5%) and 885 out of 7,710 controls (11.5%; range 8.1 to 12.1%) were HR2 positive. The odds ratio for VTE associated with HR2 haplotype was not statistically significant (OR 1.15; 95% C.I. 0.98-1.36). The OR for the association between FV Leiden and FV HR2 and the risk of VTE in cases and controls was largely heterogeneous as to OR and 95% C.I. and no statistical significant difference was observed. INTERPRETATION AND CONCLUSIONS: The data from the present meta-analysis suggests that FVHR2 could be a very mild prothrombotic factor. The association of FV Leiden and HR2 haplotype seems not to increase significantly the risk of VTE carried by isolated heterozygosity for FV Leiden. However, well-designed clinical studies are needed to clarify this issue definitely.

A novel G-to-A mutation in the intron-N of the protein S gene leading to abnormal RNA splicing in a patient with protein S deficiency.

BACKGROUND AND OBJECTIVES: Hereditary protein S (PS) deficiency is a rare autosomal disorder of the coagulation pathway associated with familial thrombophilia. DESIGN AND METHODS: We investigated a young propositus with recurrent deep vein thrombosis, a positive family history for thrombotic episodes, and low plasma concentrations of free, but not total PS antigen (12% and 70%, respectively). RESULTS: Sequence analysis of the PS gene showed a heterozygous G-to-A mutation at the first nucleotide of intron N. The patient's father, who had suffered from deep vein thrombosis and had reduced total and free PS antigen (59% and 28%, respectively) was a heterozygote. The G-to-A change predicts the disappearance of a donor splice site. After transfection with a construct, containing either the wild-type or the mutated sequence, cells with the mutant construct showed an aberrant mRNA, consistent with exclusion of exon 14, but not the expected mRNA. Sequencing of the abnormal mRNA showed the complete absence of exon 14. Exclusion of exon 14 predicts the deletion of the amino acid sequence from residue 508 to residue 582, and the shift of the reading frame of the following 8 amino acids with a premature stop codon within exon 15 at position 591. Thus, the truncated PS gene product would not contain the terminal portion of the sex hormone binding globulin-like domain. INTERPRETATION AND CONCLUSIONS: We have identified a mutation in a highly conserved intronic region of PS gene. The mutation affects in vitro mRNA processing and efficiency of normal splicing.