Factor VII coagulant activity (FVIIc) is associated with an increased risk of fatal ischaemic heart disease (IHD). Several reports have suggested that dietary fat intake or hypertriglyceridaemia are associated with elevated levels of FVII. This study demonstrates that an intake of long-chain fatty acids sufficient to induce postprandial lipaemia in healthy subjects leads to a substantial elevation in both FVIIc and the concentration of FVII circulating in the activated form. Such an increase in FVIIc could not be induced by medium-chain triglycerides. These results suggest that the consumption of a sufficient amount of long-chain triglycerides to induce postprandial lipaemia induces the activation of FVII.

We investigated the molecular basis of factor VII deficiency in a Japanese patient and identified a novel missense mutation in the signal sequence of the gene. Factor VII activity and antigen level measured in the patient were 10.7% and 11% of normal, respectively. All exons except 1B and the 5'-flanking region containing promoter region were amplified by polymerase chain reaction (PCR) from genomic DNA. Sequencing analysis of the PCR fragments revealed that the patient was a homozygote for a T to C substitution at nucleotide position 38. This mutation predicts an amino acid replacement of leucine to proline at codon -26 in the hydrophobic core of the signal peptide, which probably affects translocation of the protein into endoplasmic reticulum and subsequently causes reduction in plasma factor VII level.

BACKGROUND

Factor VII (FVII) is a vitamin K-dependent glycoprotein secreted into the blood circulation from hepatic cells. We investigated the molecular basis of the congenital FVII deficiency found in a Japanese patient.

METHODS

We analyzed the F7 gene of the patient, who was diagnosed with a FVII deficiency at pregnancy. We expressed a carboxyl-terminal truncated FVII (Arg462X FVII) corresponding to the identified mutation in CHO-K1 cells. To study roles of the carboxyl-terminus in the secretion of FVII, we also expressed a series of recombinant FVIIs deleted of limited numbers of carboxyl-terminal amino acids (462Arg-466Pro).

RESULTS

We identified a nonsense mutation (c.1384C>T: p.Arg462X) in F7, leading to a lack of five amino acids in the carboxyl-terminus. In expression experiments, Arg462X FVII was undetectable not only by Western blotting, but also by ELISA. A Western blot analysis of the truncated FVIIs revealed that all mutants were expressed in the cells the same as the wild type, but were secreted into the culture medium in lesser amounts than the wild type depending on the length of the deletion, which was confirmed by ELISA. Arg462X FVII did not colocalize with the Golgi on immunofluorescence staining, suggesting that it might be retained in the ER and degraded in the cell.

CONCLUSIONS

The carboxyl-terminal amino acids of FVII play an important role in its secretion, and the p.Arg462X mutation was likely to have caused the FVII deficiency in this patient.

Knowledge about species compatibility is crucial for proper interpretation of data from in vivo experiments with human proteins in pharmacological models and of data from cross-species in vitro experiments. Information about the cross-species compatibility of tissue factor (TF) and coagulation factor (F) VII (FVII) has accumulated since the early history of coagulation research. Many observations were connected to the introduction and development of the prothrombin time (PT) assay where fibrin clot formation was observed when tissue extracts of different origins were added to recalcified human or non-human plasmas. Studies on cross-species TF-FVIIa compatibility entered into a new area with the cloning and recombinant expression of TF and FVII from a number of species as well as with the possibility of specific amino acid substitution. TF and/or FVIIa from cattle, dog, rabbit, mouse, rat and zebrafish have been purified and characterized in varying detail. In addition to adding knowledge about the species-specific TF-FVIIa interactions, cross-species studies often reveal information which adds to the general view of the structural and functional properties of the human TF-FVIIa complex. This review briefly outlines the features of human TF and FVIIa, their intermolecular interactions, and the biological effects of TF-FVIIa complex formation and compares this information to findings obtained in studies addressing TF or FVIIa of non-human origin. By examples we point to difficulties which may arise from the transcendence across species borders and how some cross-species data have advanced our understanding of the structure and function of the human TF-FVIIa complex.

We studied the relationships among different polymorphisms of FVII gene in determining FVII levels, in a sample of 335 male and female Italian volunteers. The hypervariable region 4 (HVR4), the promoter decanucleotide insertion (-323 0/10 bp) and the R353Q polymorphisms of FVII gene were evaluated. The association of HVR4 or -323 0/10 bp polymorphism with plasma FVII levels differed between gender (Interaction term: p = 0.02 and p = 0.03, respectively), showing stronger effect in males than in females. In males, the R353Q and the HVR4 polymorphisms showed an incremental influence on FVII variance (F = 8.9, p <0.001 and F = 4.4, p = 0.01, respectively). Moreover, the effects of Q and 10 bp alleles on the reduction of FVII activity levels were significantly potentiated by the presence of H7 allele of HVR4 (Interaction term p = 0.03 for R353Q*HVR4 and p = 0.03 for -323 0/10 bp*HVR4). In conclusion, the effect of FVII polymorphisms on FVII levels was gender dependent and derived from a complex interaction among them. The HVR4 polymorphism seems to add an independent, albeit small, contribution to the regulation of FVII plasma levels.

Hemorrhagic events due to production of antibodies directed against coagulation factors are rarely observed in systemic lupus erythematosus (SLE). We report the case of a patient with clinically quiescent SLE who developed factor VIII inhibitor in acquired hemophilia presenting as hemarthrosis. Initial treatment with FVII, FVIII and FIX plasma concentrate, metilprednisolone and immunoglobulins i.v. were started but new hemorrhagic manifestation occurred. Plasma exchange was also administered, but it was discontinued early due to partial efficacy. In addition, pulse cyclophosphamide 0.5 g/m(2) was started. Eight weeks later, FVIII and FIX activity returned within normal ranges, FVIII and FIX inhibitors decreased significantly and hemorrhagic manifestations disappeared. The rare occurrence of acquired hemophilia due to the presence of anti-factor VIII antibodies associated to SLE, which was reviewed, might explain the lack of therapeutic guide-lines; indeed therapeutic options are available but the outcome in each single patient is not predictable.

The tissue factor (TF) coagulation pathway is initiated when circulating factor (F)VII(a) encounters TF, a cell surface glycoprotein, as a result of vascular injury or pathological perturbation. TF-induced coagulation plays a primary role in hemostasis and also in the pathogenesis of various thrombotic disorders. Recent studies suggest that activation of the TF-pathway may also contribute to other pathophysiological processes by altering intracellular responses, either directly or via activated factor X (FXa) and thrombin generation. Therefore, suppression of the aberrant expression of TF/FVIIa on cell surfaces not only prevents thrombotic disorders but may also provide other protective effects. Recent ex-vivo and in-vivo experiments document the effectiveness of active site-blocked activated factor VII (FVIIai) in inhibiting TF-mediated injury. It is generally believed that FVIIai exerts its effects by limiting the formation of functional TF/FVIIa complexes by directly competing with plasma FVII(a) for limited available TF sites on cell surfaces. Although such competition can explain the effectiveness of FVIIai immediately after administration, it is not clear how it exerts its prolonged effects. In this manuscript, we summarize the use of FVIIai as an antithrombotic agent in various model systems and discuss potential mechanisms by which FVIIai may exert protective effects.

BACKGROUND

The aim of this study was to investigate the effect of gamma irradiation with 30 Gy on the coagulation system in leukoreduced fresh-frozen plasma (FFP).

METHODS

In 74 FFP units that had been stored for 352 +/- 103 days below -30 degrees C, the following variables were determined in parallel in an irradiated and not irradiated half: prothrombin time (PT); activated partial thromboplastin time (APTT); thrombin time; antithrombin III; protein C; protein S; von Willebrand factor antigen; ristocetin cofactor; plasminogen-alpha(2)-antiplasmin; the coagulation factors fibrinogen, factor (F)II, FV, FVII, VIII, F IX, FX, FXI, FXII, FXIII, and activated factor XII (FXIIa); D-dimer; fibrin monomer; thrombin-antithrombin complex; prothrombin fragment 1 + 2 (F1+2); plasmin-alpha(2)-antiplasmin complexes (PAPs); and platelet factor 4. The FVII activity ratio was assayed to quantify activation of FVII.

RESULTS

Irradiation with 30 Gy resulted in a reduction of APTT (35.0 +/- 4.1 sec vs. 34.4 +/- 4.1 sec; p = 0.00000006) and PT (89.8 +/- 8.2% vs. 90.7 +/- 8.0%; p = 0.002) and a significant increase of the activities of the coagulation factors FII, FV, FVII, F IX, FX, and FXII. FVIII activity decreased from 118 +/- 31 to 116 +/- 32 percent (p = 0.02). Activation of the coagulation system was shown by an increase in the FVII activity ratio (1.19 +/- 0.29 vs. 1.31 +/- 0.34; p = 0.0000001), FXIIa (0.81 +/- 0.50 ng/mL vs. 0.90 +/- 0.51 ng/mL; p = 0.006), and F1+2 (1.19 +/- 0.20 nmol/L vs. 1.24 +/- 0.20 nmol/L; p = 0.000005) after irradiation with 30 Gy, whereas an increase of PAP (16.2 +/- 11.5 ng/mL vs. 20.2 +/- 12.0 ng/mL; p = 0.0004) demonstrated activation of the fibrinolytic system. No negative influence of irradiation with 30 Gy on inhibitors of coagulation was observed.

CONCLUSIONS

Gamma irradiation of leukoreduced FFPs with 30 Gy results in a significant but very weak activation of the coagulation and fibrinolytic system in FFPs.

BACKGROUND

A relationship has been reported between blood concentrations of coagulation factor VII (FVII) and obesity. In addition to its role in coagulation, FVII has been shown to inhibit insulin signals in adipocytes. However, the production of FVII by adipocytes remains unclear.

OBJECTIVE

We herein investigated the production and secretion of FVII by adipocytes, especially in relation to obesity-related conditions including adipose inflammation and sympathetic nerve activation.

METHODS

C57Bl/6J mice were fed a low- or high-fat diet and the expression of FVII messenger RNA (mRNA) was then examined in adipose tissue. 3T3-L1 cells were used as an adipocyte model for in vitro experiments in which these cells were treated with tumor necrosis factor-α (TNF-α) or isoproterenol. The expression and secretion of FVII were assessed by quantitative real-time PCR, Western blotting and enzyme-linked immunosorbent assays.

RESULTS

The expression of FVII mRNA in the adipose tissue of mice fed with high-fat diet was significantly higher than that in mice fed with low-fat diet. Expression of the FVII gene and protein was induced during adipogenesis and maintained in mature adipocytes. The expression and secretion of FVII mRNA were increased in the culture medium of 3T3-L1 adipocytes treated with TNF-α, and these effects were blocked when these cells were exposed to inhibitors of mitogen-activated kinases or NF-κB activation. The β-adrenoceptor agonist isoproterenol stimulated the secretion of FVII from mature adipocytes via the cyclic AMP/protein kinase A pathway. Blockade of secreted FVII with the anti-FVII antibody did not affect the phosphorylation of Akt in the isoproterenol-stimulated adipocytes.

CONCLUSIONS

Obese adipose tissue produced FVII. The production and secretion of FVII by adipocytes was enhanced by TNF-α or isoproterenol via different mechanisms. These results indicate that FVII is an adipokine that plays an important role in the pathogenesis of obesity.

Synthetic peptides corresponding to portions of the wild-type and variant sequences of the human factor VII gamma-carboxyglutamic acid (Gla)-containing domain have been prepared by direct peptide synthesis using the Fmoc-based protection strategy. Peptides were purified by ion-exchange and reversed-phase chromatography and characterized as the correct products. A peptide comprising residues 1-49 (GP 1-49) inhibited the activation of factor X (FX) by soluble tissue factor (sTF) and recombinant activated factor VII (rFVIIa). In the absence of phospholipid, no inhibition by this peptide was observed. GP 1-49 did not inhibit the hydrolysis of a peptidyl substrate by rFVIIa in the presence of either sTF or relipidated TF apoprotein in the presence or absence of phospholipid. A similar peptide (residues 1-38, GP 1-38) that did not contain the aromatic stack region was also inhibitory. Two variant peptides, one identical to GP 1-49 but lacking the N-terminal alanine residue (GP 2-49) and one identical to GP 1-38 but with an arginine to alanine substitution at position 9 (GP 1-38 R9A), showed substantially reduced inhibitory activity. Kinetic analysis of the inhibition of Xa generation by GP 1-49 revealed a noncompetitive mode of inhibition, probably via a substrate-depletion mechanism. GP 1-49 does not inhibit by preventing FX binding to phospholipid surfaces. This indicates that the N-terminal residues of the FVII Gla domain are important for the structural integrity of the peptide, and implicates the Gla domain per se in a direct interaction with phospholipid-bound FX.

OBJECTIVE

Recent data have shown that the hemostatic system may play a role in cancer development and progression. To test whether factor VII (FVII) can be a candidate factor for breast cancer, we have evaluated the distribution of FVII gene polymorphisms in breast cancer patients and healthy subjects.

METHODS

The nested case-control study consisted of 92 women with breast cancer (group 1) and 80 control subjects (in age-matched women) (group 2). Genotyping of the -323ins10-bp, -401GT, and -402GA polymorphisms of the FVII gene was performed by the method of single-strand conformation polymorphism analysis and sequencing.

RESULTS

A significant difference was observed in the distribution of the -402GA genotype and allele frequencies in breast cancer and control cases (p < 0.05). For other polymorphisms of the FVII gene, the distributions of genotypes and allele frequencies were not significantly different between two groups (p>> 0.05). There was also a significant difference between the distributions of the haplotypes in breast cancer patients and control subjects (p < 0.05).

CONCLUSIONS

Although the number of cases in this study was small, the preliminary findings revealed a possible contribution of the FVII -402GA polymorphism in the development of breast cancer. However, further case-control studies with larger series are needed to confirm our findings.

BACKGROUND

Recombinant activated factor VIIa (rFVIIa) is used to treat bleeds in hemophilia patients with inhibitors. A subcutaneous formulation could potentially improve its half-life and make it suitable for prophylactic treatment.

OBJECTIVE

A study was conducted to determine the safety of subcutaneously administered rFVIIa in patients with hemophilia and the pharmacokinetic profile (including bioavailability).

METHODS

This was a multicenter, open-label, cross-over comparison of single doses of intravenous rFVIIa 90μgkg(-1) and a new formulation of rFVIIa for subcutaneous injection at dose levels of 45, 90, 180, 270 and 360μgkg(-1) . Sixty subjects (12 per dose cohort) with hemophilia A or B were enrolled.

RESULTS

Subcutaneously administered rFVIIa showed lower mean peak plasma concentrations and prolonged FVII activity (C(max) , 0.44-5.16IU mL(-1) [across doses]; t(1/2) , 12.4h; t(max) , 5.6h) compared with intravenously administered rFVIIa (C(max) , 51.7IUmL(-1) ; t(1/2) , 2.7h; t(max) , <10min). The absolute bioavailability of subcutaneous rFVIIa ranged from 21.1 to 30.1% across dose levels. Dose proportionality was observed within a 2-fold dose increase but not across the full dose range. No thromboembolic events, drug-related serious adverse events, severe injection-site reactions or neutralizing antibodies were reported (primary endpoint). Mild and moderate injection-site reactions were more frequent with subcutaneous than with intravenous injections.

CONCLUSIONS

This phase I clinical trial did not identify safety concerns of prolonged exposure to rFVIIa administered subcutaneously in single doses to hemophilia patients.

The tumor microenvironment is generally hypoxic because of the limited oxygen supply from inefficient or insufficient vasculature. Hypoxic tumor tissues are also poorly supplied with serum components. We have previously demonstrated that expression of the FVII gene is induced in response to hypoxia in ovarian clear cell carcinoma (CCC) cells. This gene activation is synergistically enhanced when cells are simultaneously subjected to serum starvation, and is dependent on the transcription factor Sp1 directly associating with the FVII promoter. We have identified additional genes activated via a similar Sp1-dependent mechanism by conducting cDNA microarray analysis (GSE55565). ICAM1, which encodes intercellular adhesion molecule-1 (ICAM-1), is one such gene. ICAM-1 confers an anti-apoptotic effect upon CCC cells in vitro and promotes growth of CCC tumors. Here we describe the transcriptome analysis performed in our recently published study (Koizume et al., 2015). We further show that autonomous activation of the TNFα-NFκB axis is responsible for the synergistic activation of ICAM1 under hypoxic and serum starvation conditions. This study provides additional information as to how CCC cell survival can be facilitated under conditions of serum starvation and hypoxia.

BACKGROUND

Normal pregnancy is associated with a local hypercoagulable state that becomes more profound in certain obstetric complications such pre-eclampsia (P-EC). Current literature on the levels of individual haemostatic factors in women with P-EC is limited and results are inconsistent. In this study we provide detailed investigation on the tissue factor (TF)-dependent pathway in women with P-EC.

METHODS

Enzyme-linked immunosorbent assays (ELISA) were used to measure plasma factor (F) FVII, FVIIa, TF and tissue factor pathway inhibitor (TFPI) in healthy non-pregnant women (n = 22), normal pregnant women (n = 15), and women with P-EC (n = 20). All subjects were age matched. In addition, pregnant women were matched for gestational age, parity and were all at the third trimester.

RESULTS

Plasma FVII levels were significantly higher in women with P-EC compared to the healthy non-pregnant (P<0.001) or the normal pregnant groups (P<0.001). No such significant trends were observed for plasma FVIIa, TF or TFPI levels. Plasma FVII levels can distinguish women with P-EC from healthy non-pregnant women or normal pregnant women at the third trimester, with high sensitivity (90%), specificity (80%), positive and negative predictive values (86%).

CONCLUSIONS

Plasma FVII levels are significantly elevated in women with P-EC, in the absence of comparable changes in other TF-dependent pathway factors (FVIIa, TF and TFPI). We propose the use of plasma FVII as a marker for P-EC.

Individuals belonging to six different Amerindian tribes and two African groups of Costa Rica were genotyped for factor V Leiden (FV), factor V haplotype HR2 (FV HR2), Factor II 20210G>A (FII), the methylenetetrahydrofolate reductase (MTHFR), factor VII polymorphisms (FVII IVS7, FVII R353Q), factor XIII (FXIII V34L), and the insertion/deletion (I/D) polymorphism of the gene of angiotensin converting enzyme (ACE). Clear differences in the prevalence were found and are first reported. The prevalence of some of the established genetic risk factors was low in Amerindians of Costa Rica (ACE) or even absent (FVL, FII), and others (MTHFR, FVHR2) had an extremely high prevalence. People of African origin carried very rare FVL or FII polymorphisms, but the DD genotype of ACE is the highest reported. Concerning the protective factors, the QQ genotype of FVII R353Q was absent in Amerindians, but the protective 7/7 genotype of FVII IVS7 frequently found. Novel alleles of FVII IVS7 (4, 8, and 9 monomers) were found. Intertribal heterogeneity was observed that may reflect the evolutionary history of these tribal groups and their admixture with other populations.

The first epidermal growth factor-like domain (EGF-1) from blood coagulation factor VII (FVII) contains two unusual O-linked glycosylation sites at Ser-52 and Ser-60. We report here a detailed study of the effect of O-fucosylation at Ser-60 on the structure of FVII EGF-1, its Ca2+-binding affinity, and its interaction with tissue factor (TF). The in vitro fucosylation of the nonglycosylated FVII EGF-1 was achieved by using O-fucosyltransferase purified from Chinese hamster ovary cells. Distance and dihedral constraints derived from NMR data were used to determine the solution structures of both nonglycosylated and fucosylated FVII EGF-1 in the presence of CaCl2. The overall structure of fucosylated FVII EGF-1 is very similar to the nonfucosylated form even for the residues near the fucosylation site. The Ca2+ dissociation constants (Kd) for the nonfucosylated and fucosylated FVII EGF-1 were found to be 16.4 +/- 1.8 and 8.6 +/- 1.4 mM, respectively. The FVII EGF-1 domain binds to the extracellular part of TF with a low affinity (Kd approximately 0. 6 mM), and the addition of fucose appears to have no effect on this affinity. These results indicate that the FVII EGF-1 alone cannot form a tight complex with TF and suggest that the high binding affinity of FVIIa for TF requires cooperative interaction among the four domains in FVII with TF. Although the fucose has no significant effect on the interaction between TF and the individual FVII EGF-1 domain, it may affect the interaction of full-length FVIIa with TF by influencing its Ca2+-binding affinity.

Three commercial prothrombin complex concentrates (PCC) were compared in vitro. Differences in the activities or contents, respectively, of the PCC factors FII, FVII, FIX, FX and the proteins C, S, and Z (antigen) in particular were found. Global tests of activated factors did not reveal marked differences between the products. Neither free thrombin nor elevated levels of activated FX were detected. In contrast, analyses of FVIIa, of residual amidolytic activities, and of concentrations of unwanted ingredients demonstrated considerable differences between the products. While antithrombin III was detected only in two of three concentrates, heparin was found in all PCCs but in markedly different concentrations. Although the homogeneity of the products has been clearly improved in comparison with former comparative investigations, the products can be distinguished by their compound profile and by a couple of in vitro assays.

OBJECTIVE

The aim of this study was to determine the potencies of factor VII (FVII) and of activated FVII (FVIIa) in prothrombin complex concentrates (PCC).

METHODS

We examined 56 lots of PCC from 5 manufacturers. Three brands were licensed preparations, and 1 product series had been involved in thromboembolic complications. FVII and FVIIa were measured using a two-stage amidolytic assay and a specific clotting assay, respectively. We also quantified FVII clotting activity by a one-stage assay reflecting a mixture of FVII zymogen and FVIIa.

RESULTS

All PCC contained substantial amounts of FVII, and FVIIa could be detected in all lots. There were marked differences between manufacturers and some significant variabilities between batches. The two lots involved in thromboembolic events contained considerably more FVIIa than the PCC still licensed. The lowest FVIIa potencies were observed in an experimental product series, indicating that PCC can be produced without activation of FVII during the manufacturing process.

CONCLUSIONS

FVIIa is present in all PCC containing FVII. High FVIIa potencies may contribute to the thrombogenic potential of these preparations, and determination of FVIIa potencies should be included in the in vitro characterization of PCC.

BACKGROUND

The D allele of angiotensin-converting enzyme (ACE) insertion/deletion (I/D) polymorphism and coagulation activity play important roles in cardiovascular events, however, the precise association between these two risk factors remains unclear.

METHODS

We identified the ACE I/D genotype and measured the plasma coagulation factor VII and X (FVII and FX) activities and serum lipids in 172 patients (110 men and 62 women, mean age 56.7+/-13.3 years) undergoing coronary angiography.

RESULTS

The frequency of the D allele was significantly higher in those with a history of myocardial infarction (MI) than in those with normal coronary arteries, but there was no significant association between FVII and FX activities and the stage of coronary disease. Plasma coagulation factor VII and FX activities were significantly lower in the DD genotype (n=42) than in the II genotype (n=67, P<0.001 and P<0.001, respectively) or the ID genotype (n=63, P<0.01 and P<0.05, respectively). The association of the ACE D allele with lower activities of FVII and FX was also seen in patients with coronary artery disease (CAD). There was a significant association between serum triglyceride levels with FVII and FX, but not with the ACE I/D genotype.

CONCLUSIONS

We concluded that the ACE I/D polymorphism may contribute more to the onset of MI than the activities of FVII and FX and that the ACE D allele might be associated with lower plasma activities of FVII and FX. The potential link between ACE I/D polymorphism and the plasma activities of FVII and FX is probably independent of triglyceride metabolism.

Genetic factors play a role in determining the variability of plasma factor VII (FVII) levels in healthy individuals. There is also evidence that high serum lipids are associated with high FVII levels in plasma. In the promoter region of the human FVII a DNA polymorphism has been described, originating from a decanucleotide insert present in the less frequent allele. This biallelic system, reflecting the absence (AA) or presence (Aa) of the decanucleotide, can be detected by a DNA enzyme immunoassay of PCR products. We evaluated the association between the polymorphic alleles and the levels of FVII:Ag and FVII:C in 100 healthy individuals and in 19 hypertriglyceridemic individuals. Among healthy individuals, mean FVII:Ag and FVII:C levels of those with the homozygous genotype (A/A; mean FVII:Ag 112%, mean FVII:C 109%) were significantly higher (P < 0.001) than the mean levels of those with the heterozygous genotype (A/a, mean FVII:Ag 80%, mean FVII:C 90%; P < 0.001). Similar genotype-associated differences for FVII:Ag and FVII:C were found in individuals with triglycerides above 250 mg/dl (P < 0.05). FVII:C and FVII:Ag levels were positively related to triglycerides only in individuals without the insert (P < 0.01); there was no significant relationship in those carrying the allele with the insert (A/a; P = 0.43 and 0.08). Our findings of genotype-associated differences in FVII levels and interactions with triglycerides are similar to those obtained with the amino acid dimorphism at position 353 of the factor VII protein.

OBJECTIVE

To elucidate the association of plasma factor VII coagulant activity (FVIIc) with the risk of myocardial infarction (MI) and to assess the influence of factor VII gene MspI polymorphism and lipid metabolism on FVIIc in the Chinese.

METHODS

A total of 137 patients with angiographically confirmed MI and 125 healthy individuals were evaluated retrospectively. Plasma FVIIc was measured by one-stage prothrombin time, and FVII genotype was determined after MspI digestion of polymerase chain reaction-amplified genomic DNA. Serum lipid levels were assessed by routine methods.

RESULTS

MI patients had significantly higher levels of FVIIc (119.5% +/- 22.7% vs 99.9% +/- 21.8%, P < 0.01) and total serum cholesterol (5.80 +/- 1.06 mmol/L vs 5.53 +/- 1.08 mmol/L, P < 0.05) than controls, but only FVIIc independently correlated with the risk of MI (OR = 1.04, P < 0.01). There were no significant differences in FVII genotype or allele frequency between patients and controls (P>> 0.05). Subjects with the Gln353 allele were associated with significantly lower FVIIc levels than Arg353 homozygotes (99.7% +/- 19.3% vs 111.4% +/- 24.6%, P < 0.05). Serum triglyceride was positively correlated with plasma FVIIc in both control (r = 0.25, P < 0.01) and case (r = 0.87, P < 0.01) groups, but this correlation was restricted to Arg/Arg genotype (r = 0.68, P < 0.01). A significant correlation of total serum cholesterol with FVIIc only appeared in Arg/Arg homozygotes (r = 0.17, P < 0.01).

CONCLUSIONS

Our findings support the role of plasma FVIIc as a risk factor for MI in Chinese. Plasma triglyceride and FVII gene MspI polymorphism are two independent determinants of FVIIc. Assay of this polymorphism will be helpful in determining who will benefit most from lipid-lowing therapy.

The arginine/glutamine (Arg/Gln) polymorphism of the factor VII (FVII) gene is associated with variation in coagulation activity (FVII:C) and antigen concentration (FVII:Ag) of the FVII protein. We estimated frequency distributions of the Arg and Gln alleles and respective genotypes in North Karelia, and evaluated the utility of this polymorphism, serum lipids, and body mass index (BMI) in the prediction of the distributions of FVII:C and FVII:Ag in a cross-sectional study and in a prospective cohort study. The sample comprised 203 males and 262 females (aged 45-64 years) who were seen twice, in 1992 and 1995. The Arg/Arg genotype and the Arg allele frequencies were among the highest reported so far (86 and 93% respectively, in men; and 89 and 94% respectively, in women). Intragenotypic means of both FVII:C and FVII:Ag were significantly higher in the Arg/Arg genotype than in the Arg/Gln genotype in both genders. Also, intragenotypic variances were different in different genotypes in females. Regression relationships between the FVII:C and FVII:Ag and serum triglyceride, and total cholesterol levels and BMI were positive in both genotypes in both genders, which has not been found in other populations. In prospective analyses, average changes in the FVII:C and FVII:Ag were genotype specific in both genders, as were also regression relationships between these changes and changes in triglyceride level in females (P = 0.065 for FVII:C and P = 0.061 for FVII:Ag). A consequence of these complex genetic architectures is that predictive utility of the Arg/Gln genotypes depends on population, gender, serum lipid levels, and BMI, and changes in these factors over time.

BACKGROUND

During fluid infusion therapy, plasma proteins are diluted and leak from the intravascular space, which alters the colloid osmotic pressure (COP) and potentially affects coagulation. We hypothesised that acetated Ringer's and starch solution, alone or in combination, influence these mechanisms differently.

METHODS

On different occasions, 10 male volunteers were infused with 20 ml/kg acetated Ringer's and 10 ml/kg 6% hyroxyethyl starch 130/0.4 (Voluven(®) ) alone or in combination (first with starch solution followed by Ringer's solution). Blood samples were collected every 30-min for measurements of COP, blood haemoglobin, platelets, and plasma concentrations of albumin, immunoglobulins (IgG and IgM), coagulation factor VII (FVII), fibrinogen, cystatin C, activated partial thromboplastin time (APTT) and prothrombin international normalised ratio (PT-INR). Changes were compared with the haemoglobin-derived plasma dilution.

RESULTS

The COP increased by 8.4% (SD 3) with starch and decreased by 26.2% (7.9) with Ringer's. These infusions diluted the plasma by 23.4% (5.3) and 18.7% (4.9) respectively. The COP changes in the combined experiment followed the same pattern as the individual infusions. Albumin and IgG changes in excess of the plasma dilution were very subtle. The intravascular contents of the IgM and platelets decreased, whereas FVII, fibrinogen and cystatin C increased. PT-INR increased by 1/3 of the plasma dilution, whereas changes in APTT did not correlate with the plasma dilution.

CONCLUSIONS

The starch increased COP and only minor capillary leak occurred in healthy volunteers. The fluid-induced plasma dilution correlated with mild impairment of the extrinsic coagulation pathway but not of the intrinsic pathway.

Both purified plasma-derived FVIIa (pFVIIa) and recombinant FVIIa (rFVIIa) were shown to shorten the in vitro APTT in haemophilic plasma significantly. After addition of 3.8 micrograms/ml of pFVIIa or rFVIIa, corresponding to 110-140 FVII U/ml, the APTT was normalized in both haemophilia A and haemophilia B plasma. Furthermore, the prolonged APTT in plasma from a patient with acquired inhibitors against FVIII:C was normalized by addition of pFVIIa or rFVIIa in vitro. These results indicate an alternative pathway to the FVII or FVIIa-TF complex formation involving phospholipids (present in the APTT reagent). No signs of systemic activation of the coagulation system in rabbits following infusion of 500, 1000, 5000 U pFVIIa or rFVIIa/kg body weight, corresponding to about 150 micrograms/kg body weight, were seen. This corresponded to 2-3 times the clinical dose of FVIIa.