The prothrombotic state resulting from the elevation of certain clotting factors and suppression of fibrinolysis contribute to the risk of atherothrombotic diseases. Peripheral artery disease (PAD) has been associated with elevated plasma fibrinogen concentration, increased plasminogen activator inhibitor (PAI-1) level and decreased tissue plasminogen activator (tPA) activity.1–3 Coagulation factor XIII (FXIII) circulates in association with fibrinogen and plays a central role in fibrin stabilization and fibrinolysis.4 We have recently shown that in women with coronary stenosis and history of myocardial infarction adjusted plasma FXIII levels are elevated as compared to clinical controls, and in women, but not in men, elevated FXIII level represents a 2.5–3.0-fold increased risk of myocardial infarction.5 In this study we investigated FXIII levels in patients with PAD and determined the risk of PAD conferred to patients by elevated FXIII level.
The study included 302 consecutive patients with PAD recruited over a 2-year period from the 3 Department of Medicine, University of Debrecen. Patients with history of myocardial infarction, stroke, symptoms of angina and renal insufficiency were excluded. The patients were exempt of acute inflammation during the last two months before blood drawing and did not have any chronic inflammatory state, other than PAD. Finally, 278 patients (173 males and 105 females) with ankle brachial pressure index (ABPI) ≤0.9 remained in the study group (173 with Fontaine classification II and 105 with Fontaine classification III). Patients were compared to sex-matched clinical controls (n=278) who also presented at the hospital, but no significant health problem, other than diabetes mellitus in some of the patients, was diagnosed and ABPI was in the range of 0.91–1.3. The mean age ± SD was 59.0±9.5 in the control and 64.3±12.2 in the patient group. The occurrence of diabetes mellitus was 20% and 25% among controls and patients respectively. Thirty percent of controls and 36% of patients were smokers. The differences were not statistically significant. Two hundred and seventeen PAD patients received aspirin and 33 patients received statins; since in the FXIII levels there was only a non-significant difference (<1%) between PAD patients on treatment and the rest of the patients, these groups were not analyzed separately. Eighty-eight percent of women were menopausal; none of them on hormonal replacement therapy. Ethical approval was obtained from the Ethics Committee of the University of Debrecen, and subjects gave their informed consent.
Plasma FXIII activity and antigen were measured by established methods6,7 using commercially available reagent kits (REA-chrom FXIII assay and R-ELISA FXIII, Reanal-ker, Budapest, Hungary). In the measuring range, the CV for both assays was below 3%. Serum total cholesterol, LDL-cholesterol, HDL-cholesterol, triglyceride, apo AI, apo B, lipoprotein (a) and high sensitivity C-reactive protein (hsCRP), plasma fibrinogen, homocysteine, folic acid and vitamin B12 were determined by routine laboratory methods and used for the determination of parameters independently associated with FXIII levels.
Neither FXIII activity nor FXIII antigen levels of clinical controls differed significantly from the reference interval established in our laboratory.6,7 Mean FXIII activity and antigen levels, both non-adjusted and adjusted for the respective independently associated parameters, were moderately, but significantly higher in patients than in the clinical control group (Table 1). In a small (n=50) early study elevation of non-adjusted FXIII levels has also been observed in PAD patients.8 However, in this case differences according to gender and severity of the disease were not analyzed. In our study PAD was associated with a statistically significant elevation of FXIII activity and antigen levels in women. In men with PAD, elevations were somewhat more moderate and in the case of FXIII antigen concentrations were not statistically significant. No difference in adjusted FXIII levels were found in PAD patients with Fontaine II and Fontaine III stage disease (data not shown). As determined by Pearson’s method, ABPI values showed no significant correlation with FXIII activity or antigen level. One may conclude from these results that the existence of PAD and not its severity is associated with the elevation in FXIII levels. As opposed to controls, in the patient group there was a statistically significant correlation between fibrinogen and FXIII activity (r=0.273, p<0.001) or FXIII antigen (r=0.148, p=0.014) level. However, the calculated coefficients of determination (r) indicate that changes in fibrinogen level contributed to only 7% and 2% to the changes in FXIII activity and antigen levels. Gender had no effect in this respect. FXIII activity in the upper tertile (>120%) conferred a more than two-fold risk of PAD on females (Table 2). FXIII antigen in the upper tertile (>25.5 mg/L) also represented a two-fold risk of PAD in women, however in this case the p value was somewhat above the limit of statistical significance. In males elevated FXIII activity or antigen level did not increase the risk of PAD to a significant extent, although in the case of adjusted FXIII activity a tendency of increased risk was observed.
The reason for the elevation of FXIII levels in PAD is not known. Plasma FXIII consists of two potentially active A and two inhibitor/carrier B subunits (FXIII-A and FXIII-B).4 FXIII-B is in excess to FXIII-A, the actual amount of FXIII A2B2 complex is determined by the amount of FXIII-A. Megakaryocytes are a major source of FXIII-A synthesis and circulating platelets contain a huge amount of FXIII-A.4 Platelets may become damaged when pressed through occluded arteries and release their FXIII-A content into the circulation.
Impaired fibrinolysis caused by elevated PAI-1 level and decreased tPA activity increases the risk of PAD.1,9–11 FXIII is a major regulator of fibrinolysis. Activated FXIII (FXIIIa) mechanically strengthens fibrin clot and makes it more resistant to shear forces and to fibrinolysis by cross-linking fibrin α-βand γ-chains and α2-plasmin inhibitor to fibrin. At elevated FXIII levels this mechanism could be more forceful and in PAD, could participate in impairing the fibrinolytic potential. Although it remains to be seen why the effect of elevated FXIII level is more prominent in women, the results support the suggestion that in atherothrombotic diseases the clotting/fibrinolytic system plays a more prominent role in females than in males.12
- Funding: this work was supported by grants from the Hungarian National Research Fund (OTKA-NKTH NI 69238), from the Hungarian Academy of Sciences (MTA 2006TKI227) and from the Hungarian Ministry of Health (ETT 406/2006).
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