Franco Manzato

Polymorphisms in the factor VII gene and the risk of myocardial infarction in patients with coronary artery disease

BACKGROUND High plasma levels of coagulation factor VII have been suggested to be predictors of death due to coronary artery disease. Since polymorphisms in the factor VII gene contribute to variations in factor VII levels, such polymorphisms may be associated with the risk of myocardial infarction, which is precipitated by thrombosis. METHODS We studied a total of 444 patients, 311 of whom had severe, angiographically documented coronary atherosclerosis. Of these 311 patients, 175 had documentation of a previous myocardial infarction. As a control group, 133 patients with normal coronary arteriograms were also included. We measured the levels of activated factor VII and assessed three polymorphisms in the factor VII gene, one involving the promoter (A1 and A2 alleles), one involving the catalytic region (R353Q), and one involving intron 7. RESULTS Each of the polymorphisms influenced factor VII levels. Patients with the A2A2 and QQ genotypes had the lowest levels of activated factor VII (66 percent and 72 percent lower, respectively, than the levels in patients with the wild-type genotypes). The frequencies of the various genotypes in the patients free of coronary artery disease were similar to those in the entire population of patients with coronary artery disease. In the latter group, there were significantly more heterozygotes and homozygotes for the A2 and Q alleles among those who had not had a myocardial infarction than among those who had had an infarction (P=0.008 for the presence of the promoter polymorphism and P=0.01 for the presence of the R353Q polymorphism by chi-square analysis). The adjusted odds ratio for myocardial infarction among the patients with the A1A2 or RQ genotype was 0.47 (95 percent confidence interval, 0.27 to 0.81). CONCLUSIONS Our findings suggest that certain factor VII genotypes have a role in protection against myocardial infarction. This may explain why some patients do not have myocardial infarction despite the presence of severe coronary atherosclerosis.

Resistance to activated protein C in healthy women taking oral contraceptives.

Summary. Resistance to activated protein C (APC) is at present considered the most frequent laboratory abnormality in patients with deep‐vein thrombosis. An increased risk for venous thrombosis is associated to the use of oral contraceptives (OC).

Expression of plasminogen activator inhibitor-1 in human adipose tissue : a role for TNF-α?

Elevated plasminogen activator inhibitor-1 (PAI-1) plasma levels, responsible for reduced fibrinolysis, are associated with animal and human obesity and with increased cardiovascular disease. The expression of PAI-1 has been found recently in animal and human adipose tissue. Factors and mechanisms regulating such an expression remain to be elucidated. In omental and/or subcutaneous biopsies from obese non-diabetic patients, incubated in Medium 199, we have confirmed that human adipose tissue expresses PAI-1 protein and mRNA; furthermore we have demonstrated that such an expression is clearly evident also in collagenase isolated human adipocytes and that it is stimulated by incubation itself and enhanced by exogenous human tumor necrosis factor-alpha (h-TNF-alpha). Since human adipose tissue produces TNF-alpha, to further characterize the relationship of PAI-1 to TNF-alpha, human fat biopsies were also incubated with Pentoxifylline (PTX) or Genistein, both known to inhibit endogenous TNF-alpha through different mechanisms. PTX caused a dose-dependent decrease of basal PAI-1 protein release, reaching 80% maximal inhibitory effect at 10(-3)M, the same inhibitory effect caused by Genistein at 100 microg/ml. This was associated to a marked inhibition of PAI-1 mRNA and of endogenous TNF-alpha production. Furthermore, when human fat biopsies were incubated in the presence of polyclonal rabbit neutralizing anti-human TNF-alpha antibody (at a concentration able to inhibit 100 UI/ml human TNF-alpha activity), a modest but significant decrease of the incubation induced expression of PAI-1 mRNA was observed (19.8+/-19.0% decrease, P = 0.04, n = 7). In conclusion, the results of this study demonstrate that PAI-I expression is present in human isolated adipocytes and that it is enhanced in human adipose tissue in vitro by exogenous TNF-alpha. Furthermore our data support the possibility of a main role of endogenous TNF-alpha on human adipose tissue PAI-1 expression. This cytokine, produced by human adipose tissue and causing insulin resistance, may be a link in the clinical relationship between insulin-resistance syndrome and increased PAI-1 plasma levels.

Recent acquisitions in the pathophysiology, diagnosis and treatment of disseminated intravascular coagulation

Disseminated intravascular coagulation (DIC) is a disorder characterized by both acute generalized, widespread activation of coagulation, which results in thrombotic complications due to the intravascular formation of fibrin, and diffuse hemorrhages, due to the consumption of platelets and coagulation factors. Systemic activation of coagulation may occur in a variety of disorders, including sepsis, severe infections, malignancies, obstetric or vascular disorders, and severe toxic or immunological reactions.In this review, we briefly report the present knowledge about the pathophysiology and diagnosis of DIC. Particular attention is also given to the current standard and experimental therapies of overt DIC.

Study of platelet adhesion in patients with uncomplicated hypertension.

Objective To evaluate platelet function in patients with essential hypertension by sensitive methods investigating platelet adhesion and expression of some platelet glycoproteins (GP), namely GPIIb/llla (CD41/α2β3) and GMP- 140 (CD62/P-selectin/PADGEM). Other markers of platelet (β-thromboglobulin) and endothelium activation (von Willebrand factor) were also measured. Methods We studied 21 uncomplicated essential hypertensive patients and 20 healthy normotensive control subjects, non-smokers, matched for age and sex. Resting and stimulated platelet adhesion was performed with a colorimetric method using the activity of platelet acid phosphatase for the determination of the number of platelets adhering to human plasma- or fibrinogen-coated microwells. Platelet activation was characterized by flow cytometric measurement of GPIIb/llla and GMP-140 in whole blood and washed platelets suspensions, with antihuman fluorescent monoclonal antibodies. Results Thrombin-stimulated platelet adhesion to human plasma-coated microwells was significantly higher in hypertensive patients than in control subjects (0.05 U/ml thrombin: 13.4 ± 1.0 versus 7.7 ± 0.6% adhesion; 0.1 U/ml thrombin: 19.4 ±2.3 versus 12.6 ± 1.8%; means ± SEM), whereas platelet adhesion to fibrinogen-coated wells did not differ in the two groups. Flow-cytometry analysis of whole blood demonstrated a significantly increased expression of GMP-140 in hypertensive patients compared with normal subjects (percentage of CD62+ platelets: 7.3 ± 1.2 versus 3.7 ± 1; means ± SEM), whereas the expression of GPIIb/llla did not differ in the two groups (percentage of CD41a+ platelets: 72.5 ± 4.5 versus 70.4 ± 3.9). Moreover, flow cytometry showed an increased size of platelets in hypertensive patients compared with that in control subjects (forwards scattering: 46.5 ± 1.5 versus 38.9 ±1.1; means ± SEM). Flow-cytometric evaluation of washed platelet suspensions showed no statistically significant differences between the expression of GMP-140 and GPIIb/llla in the two groups. β-Thromboglobulin plasma levels were higher in hypertensive patients than they were in normal subjects (36.3 ± 2.0 versus 28.2 ± 1.3 ng/ml; means ± SEM). Von Willebrand factor plasma levels were not significantly different in the two groups (101.2 ± 10.3 versus 86.3 ± 5.6 U/dl). Conclusions These findings provide further evidence that there is a significant, albeit weak, platelet activation in hypertensive patients compared with normal subjects.

Pathogenesis, clinical and laboratory aspects of thrombosis in cancer

The relationship between increased clotting and malignancy is well recognized, though the bidirectional development of this association is often overlooked. In the challenging cancer biology, transforming genes often act in concert with numerous epigenetic factors, including hypoxia, inflammation, contact between blood and cancer cells, and emission of procoagulant vesicles from tumors, to determine a net imbalance of the hemostatic potential which is detectable by a variety of laboratory tests. Procoagulant factors, in particular, are intimately involved in all aspects of hemostatic, cell proliferation and cellular signalling systems. However, the biggest as yet unresolved question is why cancer patients develop thrombosis? Since the thrombus itself does not apparently contributes directly to the tumor biology, enhanced hemostasis activation in cancer patients may be interpreted according to the most recent biological evidences. Coagulation and cancer biology interact bidirectionally in a “vicious cycle”, in which greater tumor burden supplies greater procoagulants (tissue factor, cancer procoagulant) and thrombin, which would in turn act as strong promoters of cancer growth and spread. In this perspective, thrombosis may be interpreted as a epiphenomenon of an intricate an effective biological feedback to maintain or promote cancer progression. In this review article, we briefly analyze the pathogenesis, laboratory, clinical and therapeutic features of cancer and thrombosis.

Hemostatic abnormalities in endocrine and metabolic disorders

The hemostatic balance is a complex system where the delicate equilibrium is regulated by several factors, including hormones. This review summarizes current knowledge of the effects of most frequent endocrine and metabolic diseases (such as hypothyroidism, hyperthyroidism, Cushings syndrome, GH-related pituitary dysfunctions, pituitary prolactin-producing adenomas, polycystic ovary syndrome, primary hyperparathyroidism, and metabolic syndrome) on coagulation and fibrinolysis. Overt hypothyroidism appears to be associated with a bleeding tendency, whereas all other endocrine diseases appear to be associated with a thrombotic tendency. Globally, the disorders of coagulation and fibrinolysis usually range from mild to moderate, and, rarely, to severe laboratory abnormalities (for example, bleeding diathesis in overt hypothyroidism mainly due to an acquired von Willebrands disease type 1). Further larger and high-quality studies are needed to provide more definitive information on the effects of endocrine disorders on coagulation and fibrinolysis.

Interrelation between angiographic severity of coronary artery disease and plasma levels of insulin, C-peptide and plasminogen activator inhibitor-1

Plasma insulin, C-peptide and plasminogen activator inhibitor-1 (PAI-1) levels were measured in 64 men with coronary artery disease (CAD) documented by angiography. Coronary arteriograms were analyzed, and the severity and diffusion of coronary lesions were quantified by score systems. C-peptide and PAI-1 levels in patients with CAD were significantly higher than in 30 control subjects. Insulin, C-peptide and PAI-1 showed a highly significant correlation with the severity scores for coronary lesions (C-peptide more than insulin), but only a weak correlation with diffusion scores. Highly significant correlations were found between insulin and PAI-1, and even greater ones between C-peptide and PAI-1. It has been proposed that hyperinsulinemia may be involved in the etiology of atherosclerotic cardiovascular disease by dysregulating lipoprotein metabolism and blood pressure. These findings support that hypothesis and suggest that insulin secretion may be an index of the severity of CAD. Because a direct effect of insulin on the cells that synthesize PAI-1 has been shown, the present data further indicate that the effect of insulin on fibrinolysis may be another way by which hyperinsulinemia accelerates atherogenesis.

G20210A Prothrombin Gene Polymorphism and Prothrombin Activity in Subjects With or Without Angiographically Documented Coronary Artery Disease

Background—G20210A prothrombin mutation has been associated with high prothrombin levels and an increased risk of venous thrombosis. The role of this common polymorphism, as well as that of prothrombin levels, in determining the risk of arterial disease is still somewhat controversial. Methods and Results—We determined the prevalence of the G20210A mutation and prothrombin activity in 660 individuals, of whom 436 had angiographically documented severe coronary artery disease (CAD patients) and 224 had normal coronary angiography (CAD-free control subjects). Heterozygosity for the 20210A allele was found in 5.3% of the CAD patients versus 3.1% of the CAD-free subjects (P =0.21). Similarly, no statistically significant difference was found between CAD patients with or without previous myocardial infarction (4.5% versus 5.3%, respectively;P =0.73). The genotype-phenotype correlation study showed a significant influence of the 20210A allele on prothrombin activity, with higher levels in carriers compared with noncarriers (153.2% versus 122.2%, respectively;P <0.001). Prothrombin activity was significantly higher in CAD patients than in CAD-free subjects (132.8% versus 123.3%, respectively;P <0.005). By multiple logistic regression, prothrombin activity in the upper tertile of the control distribution was significantly associated with CAD compared with prothrombin activity in the lower tertile (adjusted odds ratio 1.86, 95% CI 1.01 to 3.4). Conclusions—In a population with a clear-cut definition of the phenotype, the G20210A prothrombin mutation was not significantly associated, per se, with either angiographically documented CAD or myocardial infarction, whereas it significantly influenced prothrombin activity. In our population, high prothrombin activity itself was independently associated with CAD but not with the presence or absence of previous myocardial infarction.

Thyroid Dysfunction and Hemostasis: An Issue Still Unresolved

Thyroid hormones exert various effects on the hemostatic system, as documented by the fact that subclinical or overt thyroid dysfunctions may be associated with hypocoagulable or hypercoagulable states. In this review, the hemostatic balance (primary hemostasis, coagulation factors, and fibrinolytic system) in different thyroid disorders is analyzed from a laboratory, pathogenic, and clinical point of view. Although limited, the published studies suggest that patients with hyperthyroidism or subclinical hypothyroidism have an increased thrombotic risk, whereas patients with overt hypothyroidism have a bleeding tendency. Further trials on larger series of patients are needed to confirm these preliminary findings and to elucidate the pathogenic mechanisms regulating the complex interaction between thyroid disorders and hemostasis.