Factor VII (fVII) deficiency is a rare congenital bleeding disorder in which fVII activity level and bleeding tendency do not completely correlate. Pregnancy and delivery present a significant haemostatic challenge to women with fVII deficiency. Treatment with recombinant factor VIIa (rfVIIa) carries a thrombotic risk and the literature is not clear whether prophylaxis is necessary prior to delivery. The aim of this study was to define management, haemorrhagic and thrombotic complications of pregnant women with fVII deficiency through a systematic review. Medical databases (PubMed, MEDLINE, CINAHL, Academic Search Premier, Cochrane Library, Web of Science and Scopus) were searched using "factor VII deficiency" and "pregnancy" or "surgery." Overall 34 articles, four abstracts, and three institutional cases were reviewed. Literature from 1953 to 2011 reported 94 live births from 62 women with fVII deficiency. The median fVII activity was 5.5%. Haemostatic prophylaxis was used in 32% of deliveries. Without prophylaxis, 40 vaginal deliveries and 16 caesarean sections were completed. The odds of receiving prophylaxis were 2.9 times higher in women undergoing caesarean section compared to vaginal delivery. Post-partum haemorrhage occurred in 10% of deliveries with prophylaxis and 13% of deliveries without prophylaxis. The fVII level did not significantly differ between women who did and did not receive prophylaxis. We present the only systematic review of the management of pregnancy in fVII deficient women. No difference in post-partum haemorrhage was seen in deliveries with and without prophylaxis. Therefore, we recommend that rfVIIa be available in the case of haemorrhage or surgical intervention, but not as mandatory prophylaxis.

To investigate the effect of tissue-type plasminogen activator (t-PA) on blood coagulation, we examined the effects of the addition of t-PA to normal pool plasma (NPP) on clotting times such as diluted prothrombin time (PT) and kaolin clotting time (KCT). The diluted PT but not the KCT was significantly shortened by the addition of t-PA to NPP compared with the normal controls, suggesting a t-PA-induced activation of blood coagulation through factor VII (FVII) activation. The activated factor VII (FVIIa) concentration in the NPP was significantly increased by the addition of t-PA. Although the FVIIa formation was not observed following the incubation of purified FVII with only t-PA or plasminogen, an increase in the FVIIa level was observed after the incubation of purified FVII with t-PA together with plasminogen, or only plasmin. This plasmin-mediated FVIIa formation was also confirmed by Western blotting. These findings suggest that t-PA enhances the activation of the coagulation system through FVII activation.

As yet, there have been neither systematic reviews nor reports of randomized, controlled trials involving factor VII (FVII) deficiency. Hence, a picture of this disorder can only be drawn by reviewing and summarizing the data that is available. This article provides an overview of the understanding of this rare, inherited disorder of coagulation. In particular, the status of current knowledge of the disorder's prevalence, clinical presentation, diagnostic characteristics and molecular genetics is reviewed, followed by a summary of currently available options for its treatment and management.

It has been shown that coagulation factor VII (FVII) has an increased coagulant activity (FVIIc) in cardiovascular high risk patients and that it is a important risk factor for the development of ischaemic heart disease (IHD) and cardiovascular death. In this study, we measured FVII coagulant (FVIIc) and immunological (FVIIag) activities during the acute phase of unstable angina (UA) and acute uncomplicated and complicated myocardial infarction (AMI). We have also studied its changes in relation to thrombin formation and coagulation activation, as assessed by determination of thrombin-antithrombin circulating complexes (T-AT) at the same time. Our results show a marked increase in FVIIc in all patients, with highest significant levels in complicated AMI. In fact, this increase was also different between groups, complicated AMI showing a significant degree of increase in FVIIc in relation to UA and uncomplicated AMI. FVIIag did not vary between groups and controls, implicating a progressive activation of FVII. As expected, we found comparable levels of T-AT in UA and in AMI patients, suggesting that a common thrombotic process is involved in both situations. FVIIc was strongly correlated to T-AT in all patients (r = 0. 750; p less than 0.001) and also within groups. This study underlines the important positive contribution of FVIIc to IHD and to the prognosis of its thrombotic acute events, and shows that the increase in FVII activity is associated with an increase of a thrombotic marker (thrombin-antithrombin). Further studies are needed to evaluate if FVII activation is the cause or the consequence of the thrombotic processes.

Factor VII activating protease (FSAP) is a circulating serine protease with high homology to fibrinolytic enzymes. A role in the regulation of coagulation and fibrinolysis is suspected based on in vitro studies demonstrating activation of FVII or pro-urokinase plasminogen activator (uPA). However, considering the paucity of any studies in animal models or any correlative studies in humans the role of FSAP in haemostasis remains unclear. In relation to vascular remodeling processes or inflammation it has been convincingly shown that FSAP interacts with growth factors as well as protease activated receptors (PAR). Against this sparse background there are a plethora of studies which have investigated the linkage of single nucleotide polymorphisms (SNP) in the FSAP gene (HABP2) to various diseases. The G534E SNP of FSAP is associated with a low proteolytic activity due to an amino acid exchange in the protease domain. This and other SNPs have been linked to carotid stenosis, stroke as well as thrombosis in the elderly and plaque calcification. These SNP analyses indicate an important role for FSAP in the regulation of the haemostasis system as well as fibroproliferative inflammatory processes.

On the basis of a crystal structure of a phenylpyrrolidine lead and subsequent molecular modeling results, we designed and synthesized a novel series of macrocyclic FVIIa inhibitors. The optimal 16-membered macrocycle was 60-fold more potent than an acyclic analog. Further potency optimization by incorporation of P1' alkyl sulfone and P2 methyl groups provided a macrocycle with TF/FVIIa Ki = 1.6 nM, excellent selectivity against a panel of seven serine proteases, and FVII-deficient prothrombin time EC2x = 1.2 μM. Discovery of this potent, selective macrocyclic scaffold opens new possibilities for the development of orally bioavailable FVIIa inhibitors.

We investigated the mechanisms responsible for severe factor VII (FVII) deficiency in homozygous Italian patients with either Gly97Cys or Gln100Arg mutations in the second epidermal growth factor domain of FVII. Transient expression of complementary DNA coding for the mutations in COS-1 cells showed impaired secretion of the mutant molecules. Using stably transfected Chinese hamster ovary (CHO) cells, we performed pulse-chase labeling studies, immunohistochemistry, and experiments with inhibitors of protein degradation, showing that FVII-Cys97 did not accumulate intracellularly but was degraded in a pre-Golgi, nonlysosomal compartment by a cysteine protease. In stably transfected CHO cells expressing FVII-Arg100, the level of intracellular FVII was not increased by several inhibitors of protein degradation, but FVII-Arg100 was retained in the endoplasmic reticulum for a longer period of time than wild-type FVII. FVII-Arg100 had a lower apparent molecular weight than did wild-type FVII under nondenaturing conditions, which is attributable to misfolding due to abnormal disulfide bond formation.

Activation of stromal response pathways in cancer is increasingly viewed as both a local and systemic extension of molecular alterations driving malignant transformation. Rather than reflecting passive and unspecific responses to anatomical abnormalities, the coagulation system is a target of oncogenic deregulation, impacting the role of clotting and fibrinolytic proteins, and integrating hemostasis, inflammation, angiogenesis and cellular growth effects in cancer. These processes signify, but do not depend on, the clinically manifest coagulopathy and thrombosis. In this regard, the role of driver mutations affecting oncoprotein coding genes such as RAS, EGFR or MET and tumour suppressors (PTEN, TP53) are well described as regulators of tissue factor (TF), protease activated receptors (PAR-1/2) and ectopic coagulation factors (FVII). Indeed, in both adult and pediatric brain tumours the expression patterns of coagulation and angiogenesis regulators (coagulome and angiome, respectively) reflect the molecular subtypes of the underlying diseases (glioblastoma or medulloblastoma) as defined by their oncogenic classifiers and clinical course. This emerging understanding is still poorly established in relation to the transforming effects of non-coding genes, including those responsible for the expression of microRNA (miR). Indeed, several miRs have been recently found to regulate TF and other effectors. We recently documented that in the context of the aggressive embryonal tumour with multilayered rosettes (ETMR) the oncogenic driver miR (miR-520g) suppresses the expression of TF and correlates with hypocoagulant tumour characteristics. Unlike in adult cancers, the growth of pediatric embryonal brain tumour cells as spheres (to maintain stem cell properties) results in upregulation of miR-520g and downregulation of TF expression and activity. We postulate that oncogenic protein and miR coding genes form alternative pathways of coagulation system regulation in different tumour settings, a property necessitating more personalised and biologically-based approaches to anticoagulation.

It was recently reported that the circadian clock machinery controls plasma levels of factor (F) VII, the serine protease triggering blood coagulation. Here, by exploiting the mouse model, this study showed that variations of photoperiod (i.e., winter or summer conditions or simulated chronic jetlag conditions) have a strong impact on plasma FVII activity levels. Under conditions mimicking summer or winter photoperiods, FVII activity showed a clear 24 h rhythmicity. Interestingly, mean daily FVII activity levels were significantly reduced in mice exposed to summer photoperiods. Behavioral activity rhythms under both photoperiods were synchronized to LD cycles, and the amount of activity per 24 h was comparable. The authors also investigated the influence of chronic jetlag (CJL) on the FVII activity rhythms, which can be easily mimicked in mice through continuous abrupt shifts in the lighting schedule. The exposure of mice to simulated CJL of either consecutive westward or consecutive westward and eastward flights for 15 days did not abolish the behavioral activity rhythms but was associated with a period significantly different from 24 h. Intriguingly, both types of CJL exerted a strong influence on FVII activity rhythms, which were virtually suppressed. Moreover, the mean daily FVII activity was significantly lower in the CJL than in the winter photoperiod condition. Taken together, these findings in mice provide novel insights into the modulation of FVII activity levels, which might have implications for human pathophysiology.

Elevated plasma homocyst(e)ine is a risk factor for cardiovascular disease (CVD) in many populations, but the relationship between homocyst(e)ine and CVD in Japanese subjects has been unclear. It has been hypothesized that the link between homocyst(e)ine and CVD may be mediated in part by activation of coagulation and endothelial cell injury in the elderly Japanese subjects. To further evaluate this hypothesis, the present cross-sectional study was designed to assess the relationships among plasma homocyst(e)ine concentrations, risk of CVD, and markers of coagulation (fibrinogen, FVII, F1+2, FVIIa and FXIIa) and endothelial cell damage (vWF and thrombomodulin) in 146 elderly Japanese subjects (79 healthy controls and 67 patients with CVD). The geometric mean (range) of plasma homocyst(e)ine concentrations was 10.2 (3.2--33) micromol/l in 79 Japanese healthy elderly subjects. As expected, healthy female and male elderly subjects had homocyst(e)ine levels that were 2.5 and 5.3 micromol/; higher, respectively, compared to healthy young control subjects (n=62). Healthy young and elderly men had homocyst(e)ine levels that were 1.7 and 4.5 micromol/l higher, respectively, compared to values in women. This higher plasma homocyst(e)ine levels in the elderly subjects were negatively correlated with levels of folic acid, albumin and total cholesterol, but were not significantly related to markers of coagulation or endothelial cell-damage. The results of multiple logistic regression analyses suggested that high homocyst(e)ine levels were independently related to CVD risk. In addition, levels of FVIIa, and F1+2 were significantly higher in elderly Japanese patients with CVD compared to elderly subjects without CVD, but were unrelated to plasma homocyst(e)ine concentrations. In summary, elevated plasma concentrations of homocyst(e)ine, FVIIa, and F1+2 were associated with increased risk of CVD in elderly male and female Japanese subjects, but the association between homocyst(e)ine and CVD was unrelated to abnormalities in markers of coagulation and endothelial cell damage in this population.

The purpose this study was to determine whether Arg353Gln and -323Del/Ins polymorphisms of factor VII (FVII) are related to blood pressure levels and hypertension. Subjects were drawn from the Stanislas Cohort, a longitudinal, familial French cohort examined twice since 1994. The "blood pressure study" included 1342 subjects free of medication use that could affect blood pressure. The "hypertension study" included 645 normotensive and 77 hypertensive adult subjects. Association with hypertension was also studied in 547 hypertensives enrolled in a clinical trial and in 624 normotensives drawn from the Stanislas Cohort. In the "blood pressure study," parents with the 353Gln or -323Ins allele had lower blood pressures than did noncarriers at each examination, independent of covariates (0.01< or =P< or =0.05, except for diastolic blood pressure [DBP] at baseline, where P=0.103). Similarly significant relations were observed in their offspring (P< or =0.05, except for systolic blood pressure [SBP] at 5 years, where P=0.186). In a representative subgroup of 267 individuals, the -323Del/Ins polymorphism was significantly associated with plasma FVII levels in both parents and offspring (P<0.001). FVII levels in plasma were significantly correlated with SBP and DBP in parents but not in offspring. After inclusion of both FVII levels and the -223Del/Ins in the same model in parents, only FVII levels remained significantly associated with SBP and DBP. The "hypertension study" revealed that the 353Gln and -323Ins alleles were related to decreased risk (odds ratio [OR]=0.554, 95% confidence interval [CI], 0.362 to 0.848, and OR=0.475, 95% CI, 0.299 to 0.755, respectively). These results suggest that the FVII gene may be a susceptibility locus for hypertension.

Coagulation factors VII (FVII), IX (FIX), X (FX), and protein C share the same domain organization but display very different plasma half-lives. It is plausible that the half-life is influenced by the activation peptide, differing in length and glycosylation and missing in FVII. To test this hypothesis, the influence of activation peptides on the plasma half-life of human FVII was studied by administering human FVII variants containing activation peptide motifs to mice. Insertion of the activation peptide from FX gave 4-fold longer terminal half-life (5.5 hours vs 1.4 hours for FVII), whereas the activation peptide from FIX and protein C resulted in half-lives of 4.3 and 1.7 hours, respectively. Using FX's activation peptide we identified the N-linked glycans as structural features important for the half-life. The peptide location within the FVII molecule appeared not to be critical because similar prolongation was obtained with the activation peptide inserted immediately before the normal site of activation and at the C-terminus. However, only the latter variant was activatable, yielding full amidolytic activity and reduced proteolytic activity with preserved long half-life. Our data support that activation peptides function as plasma retention signals and constitute a new manner to extend the half-life of FVII(a).

After menopause the haemostatic balance shifts towards a latent hypercoagulable state. To evaluate the effects of two regimens of transdermal estradiol (E2) combined with progestin on the balance between procoagulant factors and inhibitors, 255 women in physiological menopause for 1-5 years were randomly allocated to 1 year of treatment with cyclic transdermal E2 (50 micrograms/day for 21 days) plus medroxyprogesterone acetate (MPA) (10 mg/day from days 10 to 21), continuous transdermal E2 (50 micrograms/day for 28 days) plus MPA (10 mg/day from days 14 to 25), or placebo. Fibrinogen, factor VII (FVII), factor VIII:C (FVIII:C), antithrombin III (ATIII), protein C, protein S, heparin cofactor II (HCII) and plasminogen activator inhibitor (PAI-1) levels were measured at baseline and after 6 and 12 cycles. 167 women who took the treatment for at least 6 cycles were evaluable. The continuous treatment group had significantly lower final values of fibrinogen, FVII, ATIII, protein S and HCII than the placebo group; the cyclic treatment reduced fibrinogen in comparison with placebo but the difference was not significant. In conclusion, both regimens produce a clinically relevant decrease of fibrinogen levels; the continuous regimen affects also the levels of FVII and inhibitors suggesting that the haemostatic balance is shifted to a more physiological state.

BACKGROUND

With buffy coat (BC) processing of whole blood (WB) donations, increase in WB storage time to facilitate overnight holding before the separation of blood components would be a logistically attractive development. This study undertakes a comparative in vitro characterization of blood components prepared from WB samples that were either processed within 8 hours or stored overnight at room temperature before processing by the BC method.

METHODS

The WB units (400 mL) collected were either processed within 8 hours (fresh blood) or stored overnight (overnight blood) at room temperature. WB units were separated into individual-component red blood cells (RBCs), BC, and plasma. The in vitro quality of these blood components (RBCs, pooled platelet concentrates [PCs], and plasma) was analyzed during storage.

RESULTS

Levels of 2,3-diphosphoglycerate (2,3-DPG) were found to be significantly lower immediately after processing, compared with the fresh WB samples, in RBCs that had been separated from an overnight-hold sample. However, this difference was not apparent after 14 days of storage. In pooled PCs, measurements for glucose, lactate, PO(2), PCO(2), extent of shape change, and hypotonic shock response were similar. The platelet yield in PCs prepared from an overnight-hold WB sample was significantly higher, while CD62P expression and annexin V binding were lower (p < 0.05). For frozen plasma (FP), no significant differences were observed for the coagulation factors (F)II, FVII, FV, F IX, FX, and FXI; fibrinogen; and von Willebrand factor content between the 8- and 24-hour FP. The FVIII was the component that was most sensitive to the prolongation of production time and it only had 80% of the activity of the 8-hour FP.

CONCLUSIONS

These data suggest that blood components (RBCs, pooled PCs, and FP) separated from WB that has been stored overnight at room temperature by the BC method are of acceptable quality.

The aim of this study was to evaluate the association of prothrombotic gene polymorphisms [factor V Leiden (FVL) 1691GA, factor VII (FVII) 10976GA, FVII HVR4, platelet membrane glycoproteins GP1BA 1018CT, GP1BA VNTR, integrin ITGB3 1565TC, integrin ITGA2 807CT and methylenetetrahydrofolate reductase (MTHFR) 677C/T], plasma factors (fibrinogen and homocysteine) and traditional risk factors with acute myocardial infarction (AMI) in 184 patients ≤ 40 years of age and 350 controls (≤ 40 years) from north India. Multiple logistic-regression analysis showed that hypertension (OR 1.9, 95 % CI 1.1-3.8, p = 0.042), diabetes mellitus (OR 10.5, 95 % CI 2.0-56.7, p = 0.006), smoking (OR 7.1, 95 % CI 3.7-13.6, p < 0.001), low socio-economic status (OR 13.5, 95 % CI 2.3-78.4, p = 0.004), high waist-hip ratio (OR 35.6, 95 % CI 11.1-53.7, p < 0.001) and FVL 1691GA (OR 6.0, 95 % CI 1.2-13.4, p = 0.03) were independent risk predictors of AMI in young. Elevated plasma fibrinogen also showed association with increased AMI risk. ITGA2 807C/T polymorphism showed protection against AMI in univariate analysis only, while GP1BA VNTR-ac (OR 0.4, 95 % CI 0.2-0.9, p = 0.033) showed significant protection even after adjusting for age and sex. Multinominal logistic-regression analysis showed gene-gene (GP1BA 1018C/T with GP1BA VNTR and ITGA2 807C/T with ITGB3 1565T/C polymorphisms) and gene-environment interactions (gene polymorphisms with smoking) operating in the occurrence of AMI in young. In conclusion, the role of inherited predisposition to thrombosis in complex, polygenic and multifactorial disease like AMI is limited to certain genetic factors, in combination with environmental factor like smoking.

A link has been suggested between blood lipids and hemostatic activation. Factor VII (FVII) is a coagulation factor which plays a pivotal role in fibrin generation and thrombus formation. Clinical trials have demonstrated that inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase greatly reduce cardiovascular events in patients with and without coronary artery disease but few data, at this time, are available on the effects of lipid-lowering treatment on factor VII levels. We studied thirty-six IIA and IIB type hyperlipidemic patients who, after a preliminary period of lipid-lowering diet, added atorvastatin (20 mg/daily) or continued dietary treatment alone until they achieved LDL-C recommended levels (<4 mmol/L). Four to six weeks of lipid lowering treatment with diet plus atorvastatin, produced a significant reduction in FVII coagulant activity (FVIIc) and antigen (FVIIAg). No significant changes were observed in activated FVII (FVIIa). The lipid-lowering treatment with diet alone induced an improved lipid pattern, but no significant changes in FVII profile. Our study suggests a significant effect of lipid-lowering treatment on FVII levels. A possibile nonlipid mechanism that modifies FVII pathway may be suggested.

Investigation of the molecular basis of a severe factor VII (fVII) deficiency revealed compound heterozygosity in the fVII gene. On the paternal allele the patient had 3 structural gene abnormalities frequently associated with fVII deficiency. A new mutation, a C to T transition at position -55 relative to the translational start site, was found on the maternal allele. The study demonstrates that this mutation partially impeded binding of the transcriptional activator, hepatic nuclear factor 4, to the fVII promoter while greatly reducing reporter gene expression in hepatic cells. (Blood. 2000;96:4370-4372)

Greenland Inuit are a population with a low risk of cardiovascular disease. Recently, we stated that frequencies of potentially high risk alleles of the apolipoproteins, fibrinogen, factor V, glycoprotein IIIa and factor VII (FVII) genes have different allele frequencies in the Inuit when compared with Caucasian populations. We have extended this study and evaluated whether or not this was also true for the genetic polymorphisms of tissue-type plasminogen activator (t-PA), plasminogen activator inhibitor-1 (PAI-1), angiotensin-converting enzyme (ACE) and angiotensinogen in a group of 133 Greenland Inuit, aged 30-34 gamma. In addition, we compared the plasma levels of these factors and those of C-reactive protein (CRP) and D-Dimer in Inuit and in Danes, comparable for age and gender. Frequencies (f) were assessed of the alleles that are known as the potential high risk alleles in Caucasians. In the Inuit, the f(insertion allele) of the t-PA intron8ins311 polymorphism was 0.37 (CI 0.32-0.43), the f(4G allele) of the PAI-1 promoter polymorphism was 0.88 (CI 0.83-0.91), the f(deletion allele) of the ACE intron16ins287 polymorphism was 0.40 (CI 0.33-0.47) and the f(M-allele) of the angiotensinogen M/T353 polymorphism was 0.30(CI 0.25-0.38). As for fibrinogen and FVII polymorphisms, these frequencies are all significantly different from what is reported for Caucasian populations. In the Inuit, plasma levels of fibrinogen and D-Dimer were higher than in the Danes, the PAI-1 levels were lower and FVII, t-PA and CRP levels were comparable. The observed allele frequencies of the polymorphisms of t-PA, fibrinogen, FVII, ACE, angiotensinogen and the plasma levels of PAI-1 and D-Dimer were in accordance with the low CVD risk in the Inuit, considering the observed associations between these measures and CVD risk in Caucasian populations, but for other measures this was not the case (allele frequencies of the PAI-1 polymorphism, and plasma levels of fibrinogen, FVII and t-PA). In conclusion there are clear differences in genetic background and plasma levels of risk factors in Greenland Inuit compared with Caucasian populations, and these differences were sometimes, but not always, in accordance with the observed low cardiovascular disease risk of the Inuit population.

Human coagulation factor VII (FVII) plays an important role in the blood coagulation process and exists in micro amounts in human plasma; therefore, any attempt at the large-scale production of FVII in significant quantities is challenging. The purpose of this study was to express and obtain biologically active recombinant FVII (rFVII) from Chinese hamster ovary K1 (CHO-K1) cells. The full-length FVII cDNA was isolated from a HepG2 cell line and then subcloned in pcDNA3.1 to construct an expression vector, pcDNA-FVII. CHO-K1 cells were transfected with 1 µg pcDNA-FVII. The cell line that stably expressed secretory FVII was screened using 900 µg/mL G418. The FVII copy number in CHO-K1 cells was detected by quantitative polymerase chain reaction (qPCR). The rFVII was purified in ligand affinity chromatography medium. The purified protein was detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis. The biological activity of the purified FVII protein was determined by a prothrombin time assay. Three cell lines that permanently expressed rFVII were screened. The qPCR results demonstrated that each CHO-K1 cell harbored two FVII DNA copies. The SDS-PAGE and Western blot analysis showed that the purified protein was about 50 kDa. The purity of the target protein was 95%. The prothrombin time assay indicated that the FVII-specific activity of rFVII was 2573 ± 75 IU/mg. This method enabled the fast preparation of high-purity rFVII from CHO-K1 cells, and the purified protein had good biological activity.

Eight common polymorphisms of known myocardial infarction (MI) risk factors (factor V Leiden (FVL), factor V HR2 (FVHR2), factor II 20210G>> A (FII), factor VII IVS7 (FVII IVS7), factor VII Arg353Gln (FVII), factor XIII Va134Leu (FXIII), Methylenetetrahydrofolate reductase C677T (MTHFR), Angiotensin Converting Enzyme (ACE)) and environmental risk factors were analyzed in a MI patients of Costa Rica. This case-control study included 186 MI subjects, 95 of them < or = 45 years and 201 age and sex matched controls. With the use of PCR method the polymorphisms were detected and through interviews additional information was collected. Hypercholesterolemia and smoking were associated with a significant risk in younger patients. High fibrinogen level was an important risk factor and interaction with smoking was detected. Mainly, the genotype 34LeuLeu of FXIII showed significant protective effect, (OR 0.32, 95% CI 0.13-0.80) while the other polymorphisms showed no significant difference between the cases and the controls. Carriers of FVII (OR 2.75, 95% CI 1.07-7.02) and FXIII (OR 4.20, 95% CI 2.03-8.67) polymorphisms showed interaction with fibrinogen in the statistical analysis. It was concluded that there was an important interaction between the common risk factors and the polymorphisms (FVII; FXIII) in the development of MI. This is one of the first reports in a Latin-American population dealing with these molecular markers and MI.

BACKGROUND

Elevated levels of coagulation factor VII (FVII) have been associated with increased risk for myocardial infarction (MI). The R353Q polymorphism of the FVII gene has been shown to modify plasma levels of FVII, and has in some studies also been associated with reduced risk for MI.

OBJECTIVE

To examine the R353Q polymorphism of the FVII gene and the relation to myocardial infarction (MI), cardiovascular disease (CVD), and diabetes, and furthermore, to elucidate the association between the polymorphism and plasma levels of FVII coagulant activity (FVIIc), FVII antigen (FVIIag), activated FVII (FVIIa), and serum choline-containing phospholipids (PC).

METHODS

In 560 elderly men characterised as hypercholesterolemic in 1972, we examined the R353Q polymorphism by melting curve analysis after real-time PCR. In a subgroup of 205 individuals, FVIIc, FVIIag, FVIIa, and PC were analysed.

RESULTS

There were no significant associations between genotype and the disease states, although we observed a lower number of MI cases among subjects with the Q allele, compared to the RR individuals (14% vs. 19%). FVIIag and FVIIc levels were lower in RQ compared to RR subjects, whereas for FVIIa the opposite was observed (p<0.001 for all). PC correlated positively with FVIIag (r=0.24, p<0.001), but negatively with FVIIa (r=-0.25, p<0.001). No genotype specific interactions were found for the association between FVII and PC.

CONCLUSIONS

No significant associations between the R353Q polymorphism and MI, CVD, or diabetes were observed, although the polymorphism strongly influenced plasma levels of FVII. Serum PC correlated significantly with FVIIag and inversely with FVIIa, independently of genotype.

A 37-year-old man was found to have an inherited abnormal factor VII (FVII) (designated FVIINagoya) that has a lower FVII procoagulant activity when tested in a one-stage assay using rabbit, simian, or human tissue factor (TF), but surprisingly has a nearly normal activity using bovine TF. DNA sequence analysis of the propositus' FVII gene revealed a C to T mutation that results in a novel amino acid substitution of Arg304 to Trp. Restriction enzyme analysis of the amplified fragment obtained from each family member demonstrated that the patient is homozygous for the point mutation, and the mother and sister of the patient are heterozygous for this mutation. To elucidate the abnormality in FVIINagoya, we compared the recombinant mutant FVII with normal FVII. In the presence of human TF, kinetic analyses demonstrated that the apparent Km values of FVIINagoya for factor Xa-mediated FVII activation and for FVIIa-mediated factor X (FX) activation were 3.6- and 4.5-fold higher than those of normal FVII, respectively. The ligand binding assay to human TF also showed that FVIINagoya had a 5.2-fold larger apparent Kd than normal. In the presence of bovine TF, however, these values showed no difference between normal and mutant FVII. These findings indicate that the major abnormality in FVIINagoya is not a catalytic dysfunction but an impaired complex-formation of human TF.FVII.FX, suggesting the importance of Arg304 in the interaction with human TF.

Hypertension and diabetes are known risk factors for obesity-related thrombosis, but several studies have shown that obesity is characterised by a potentially prothrombotic inflammatory state because of activated coagulation and impaired fibrinolysis. In order to verify if obese patients-unaffected by hypertension, diabetes, dyslipidemia, cigarette smoking or inflammatory diseases-show increased prothrombotic markers and whether the weight loss induced by gastric banding normalises such parameters. Plasma levels of C reactive protein (CRP), fibrinogen, plasminogen activator inhibitor-1 (PAI-1), von Willebrand factor (VWF) and factor VII (FVII) were measured in 25 women with isolated obesity prior to, as well as 3, 6 and 12 months subsequent to gastric banding. Twenty-five healthy women served as a baseline control group. The obese women had higher CRP (p = 0.0001), fibrinogen (p = 0.014), PAI-1 (p = 0.003), VWF (p = 0.004) and FVII levels (p = 0.0001) than the normal controls, and their body mass index (BMI) positively correlated with CRP (r = 0.462, p = 0.02), fibrinogen (r = 0.426, p = 0.04) and PAI-1 (r = 0.468, p = 0.02). Twelve months after gastric banding, the median BMI had decreased from 40.0 to 34.9 (p = 0.0001); CRP from 4.18 to 1.69 μg/ml (p = 0.01); fibrinogen from 389 to 318 mg/dl (p = 0.0001); PAI-1 from 32.1 to 12.0 UI/ml (p = 0.003); VWF from 144 to 120% (p = 0.0001); and FVII from 134 to 112% (p = 0.002). Even in the absence of major cardiovascular risk factors, obese patients are characterised by a prothrombotic state. The weight loss induced by gastric banding decreases the parameters of inflammation, coagulation and impaired fibrinolysis, thus potentially reducing the thrombotic risk.

Contrary to low-fat meals, high-fat meals are known to cause postprandial factor VII (FVII) activation, but the mechanism is unknown. To study the postprandial FVII activation in detail, 18 young men consumed in randomized order high-fat or low-fat test meals. Fasting and non-fasting blood samples were collected. The high-fat test was associated with an increase in plasma triglyceride and kallikrein concentrations and postprandial FVII activation (p<0.001). Plasma kallikrein was strongly associated with triglycerides in fasting and non-fasting samples (r2=0.74-0.87, p<0.0001), suggesting that triglyceride-rich lipoproteins may activate prokallikrein. Neither plasma triglycerides nor kallikrein and activated FVII were statistically associated. This may suggest that additional factors are involved in the postprandial FVII activation. No clear evidence for a role of tissue factor expression by monocytes, factor XII or insulin in postprandial FVII activation was observed. Tissue factor pathway inhibitor and prothrombin fragment 1+2, a marker of thrombin generation, were not affected postprandially after either the high-fat or the low-fat meals. Our findings indicate that triglyceride-rich lipoproteins activate prokallikrein postprandially, which might form an important initial event in FVII activation after consumption of high-fat meals.