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Patients with lower extremity peripheral artery disease (PAD) have a substantially increased risk for mortality as compared to healthy individuals. We aimed to evaluate the risk for all-cause mortality in PAD patients and in healthy controls during a 10-year follow-up period. Our hypothesis was that the mortality rates at 10 years would differ in diabetic and non-diabetic PAD patients. Our study group consisted of 331 consecutive patients with symptomatic PAD

Objectives: Mortality rates at 10 years are higher in diabetic patients with chronic lower extremity peripheral arterial disease than in non-diabetic peripheral arterial disease patients. We tested the hypothesis that the predictors of mortality differ between diabetic and non-diabetic peripheral arterial disease patients. Methods: We studied 331 consecutive patients who were

Background We aimed to compare head-to-head the diagnostic and prognostic capabilities of galectin-3, soluble ST2 (sST2) and B-type natriuretic peptide (BNP) for heart failure (HF) in an emergency setting. Methods We studied 251 consecutive patients with dyspnoea as a chief compliant presenting to an emergency department. The diagnosis of HF was based on the Framingham score for HF plus echocardiographic evidence of systolic or diastolic dysfunction. All-cause mortality was assessed at one year. Plasma concentrations of galectin-3 and BNP were measured with two commercially available assays from Abbott Diagnostics, plasma concentrations of sST2 were quantified with the Presage ST2 assay. The diagnostic and prognostic accuracies of galectin-3, sST2 and BNP were assessed by receiver operating characteristic (ROC) curve analysis. Results Of the 251 patients, 137 had dyspnoea attributable to acute HF and 114 had dyspnoea attributable to other reasons. BNP had a higher area under the curve (AUC) for the diagnosis of HF (0.92; 95% CI, 0.87–0.95) than galectin-3 (0.57; 95% CI, 0.51–0.64) and sST2 (0.63; 95% CI, 0.56–0.69). Of the 137 patients with acute HF, 41 died and 96 survived during follow up. The AUC of BNP for the prediction of one-year all-cause mortality in HF patients (0.72; 95% CI, 0.63–0.79) was not different from the AUCs of galectin-3 (0.70; 95% CI, 0.62–0.78) and sST2 (0.75; 95% CI, 0.67–0.82). Conclusions In this study, galectin-3, sST2 and BNP were equally useful for the prediction of one-year all-cause mortality in patients with acute HF. However, in contrast to BNP, galectin-3 and sST2 were not useful as an aid in the diagnosis of acute HF in short of breath patients presenting to an emergency department.

Soluble ST2 (sST2) has emerged as a prognostic biomarker in patients with heart disease. We tested the hypothesis that sST2 is an independent predictor of mortality in patients admitted to an intensive care unit (ICU).We performed measurements of sST2 plasma concentrations in 530 consecutive patients admitted to a medical ICU of a tertiary care hospital during a study period of one year. The patients recruited during the first six months were used for the derivation cohort (n=274) and the patients recruited during the second six months were used for the validation cohort (n=256). The endpoint was defined as 90-day all-cause mortality.In the derivation cohort, sST2 was higher among decedents (n=56; median, 146 U/mL) than survivors (n=218; median 42 U/mL, p0.001). In multivariate Cox proportional-hazard regression analysis (offering age, sex, BMI, APACHE II score, SAPS II, CRP, IL-6, PCT, creatinine, total cholesterol, albumin, hs-cTnT, BNP and sST2 as independent variables), sST2 was a significant predictor of mortality (risk ratio 1.48, 95% CI 1.15-1.90; p=0.002 per 1 SD increase in log transformed values). In this statistical model, only sST2 and SAPS II contributed independently to mortality prediction. We further observed an additive effect of an sST2 plasma concentration of84 U/mL and an increased SAPS II for mortality prediction. The findings from the derivation cohort were confirmed in the independent validation cohort. In those patients with a length of stay of48 h at the ICU (n=225), sST2 obtained two days after baseline measurement had a better capability than baseline sST2 to predict mortality.In an unselected cohort of patients admitted to the ICU, sST2 was an independent predictor of 90-day all-cause mortality and added prognostic information to the SAPS II.

Acute dyspnea is a common cause for emergency department visits. The aim of this study was to evaluate the prognostic value of established and novel biomarkers in patients with acute dyspnea.We measured 10 biomarkers [B-type natriuretic peptide (BNP), midregional pro-A-type natriuretic peptide (MR-proANP), midregional-proadrenomedullin (MR-proADM), copeptin, C-terminal endothelin-1 precursor fragment (CT-proET-1), soluble ST2 (sST2), chromogranin A (CgA), adiponectin, proguanylin, and prouroguanylin] in 251 consecutive patients with acute dyspnea presenting to the emergency department of a tertiary care hospital. Outcome measure was all-cause mortality at 1 year.At baseline decedents (n=62) had significantly higher median plasma concentrations of all 10 biomarkers than survivors (n=189). Applying univariate Cox proportional-hazard regression analyses, all biomarkers were significant outcome predictors displaying risk ratios (RR) from 1.4 to 2.4 (per 1 SD increase in log transformed values). In multivariate Cox proportional-hazard regression analysis, however, only MR-proANP (RR 1.6; 95% CI, 1.1-2.2; p=0.008), sST2 (RR 1.7; 95% CI, 1.3-2.3; p0.001), and CgA (RR 1.5; 95% CI, 1.2-1.9, p0.001) were independently associated with 1-year mortality. We provide a possible explanation for the complementary prognostic value of those three biomarkers in our cohort, where coincidence of heart failure and inflammatory pulmonary disease was common and also related to worse outcome.Our evaluation of biomarkers in patients with acute dyspnea suggests that MR-proANP, sST2, and CgA are strong, independent and complementary outcome predictors. MR-proANP is considered a specific marker of cardiac stretch, sST2 might reflect both inflammation and cardiac stretch, and CgA obviously indicates neuroendocrine activation in various diseases.

We have previously demonstrated that adiponectin is associated with amino terminal pro-B-type natriuretic peptide (NT-proBNP) in patients with peripheral artery disease (PAD). Furthermore, we have shown that NT-proBNP is a strong predictor of mortality in these patients. The aim of this study was therefore to evaluate the value of adiponectin as long-term prognostic marker in patients with atherosclerotic PAD in the same cohort.We measured adiponectin serum concentrations in 487 consecutive patients with symptomatic PAD admitted to a tertiary care hospital. The endpoint was defined as all-cause mortality, and the study participants were followed for 5 years.Of the 487 patients enrolled, 114 died and 373 survived during follow-up. The median adiponectin concentration was higher among decedents than survivors (11.3 vs. 9.1mg/L; p0.001). Univariate Cox proportional-hazard regression analysis revealed that adiponectin concentrations were associated with 5-year mortality in PAD patients (risk ratio 1.05, 95% CI 1.03-1.07; p0.001 per 1mg/L increase). Even after adjustment for age, sex, body mass index, estimated glomerular filtration rate, clinical stage of PAD, cardiovascular comorbidity, and other potential confounders, the predictive value of adiponectin serum concentrations remained statistically significant (risk ratio 1.03, 95% CI 1.00-1.05; p=0.030 per 1mg/L increase). However, adiponectin lost its independent association with mortality in symptomatic PAD after additional adjustment for NT-proBNP.In this study, adiponectin serum concentrations predicted 5-year all-cause mortality in patients with symptomatic PAD independently of other established and emerging outcome predictors. Only after adjustment for NT-proBNP, adiponectin lost its independent predictive value.

Objectives To evaluate the prognostic value of adiponectin in patients with acute destabilized heart failure. Design and methods Adiponectin was measured in 137 consecutive heart failure patients attending an emergency department. The endpoint was 1-year all-cause mortality. Results In Cox proportional-hazards regression, an adiponectin plasma concentration > 24.1 mg/L had a risk ratio of 2.46 (95% CI, 1.24–4.87), independently of classical risk factors and B-type natriuretic peptide. Conclusions Adiponectin predicts mortality in patients with acute destabilized heart failure.

Background: Increased concentrations of lipoprotein(a) [Lp(a)] have been considered a genetically determined risk factor for coronary artery and cerebrovascular disease. Only 2 small and conflicting studies have investigated the possibility of an association of peripheral arterial disease (PAD) with high serum Lp(a) concentrations and low molecular weight (LMW) phenotypes of apolipoprotein(a) [apo(a)].Methods: We measured serum concentrations of Lp(a) and apo(a) phenotypes in 213 patients with symptomatic PAD and 213 controls matched for sex, age (within 2 years), and presence of diabetes.Results: Patients with PAD showed significantly higher median serum concentrations of Lp(a) (76 vs 47 mg/L; P = 0.003) and a higher frequency of LMW apo(a) phenotypes (41% vs 26%; P = 0.002) than controls. After adjustment for several potential confounders, increased Lp(a) concentrations (>195 mg/L, i.e., 75th percentile of the entire study sample) and LMW apo(a) phenotypes were significant predictors of PAD, with odds ratios of 3.73 (95% CI 2.08–6.67; P Conclusions: In this study sample, both increased serum concentrations of Lp(a) and the presence of LMW apo(a) phenotypes were associated with the presence of symptomatic PAD independent of traditional and nontraditional cardiovascular risk factors. Because PAD is considered an indicator of systemic atherosclerotic disease, our results suggest a possible role of Lp(a) as a genetically determined marker for systemic atherosclerosis.

Non-responsiveness to anti-platelet therapy has been reported and has been linked to the occurrence of adverse events. No standard method to monitor clopidogrel efficacy is available at present. We aimed at comparing the utility of whole blood impedance aggregometry for the assessment of clopidogrel action using the novel Multiplate analyzer with two flow cytometric methods.Platelet function was determined before and after the initiation of clopidogrel therapy (300 mg loading dose, followed by 75 mg qd) in 40 patients (observational study). Furthermore, 77 patients and 77 referents with and without clopidogrel treatment (75 mg qd) were evaluated (case control study). Platelet function was assessed by Multiplate ADP and ADP+PG tests, by P-selectin (CD62P) expression, and by vasodilator stimulated phosphoprotein (VASP) phosphorylation status.The observational study revealed that platelet reactivity decreased significantly after clopidogrel administration with all 4 methods (p0.001 for each). In the case control study the median values of all 4 tests were significantly higher in the referents without clopidogrel treatment than in the patients on clopidogrel therapy (p0.001 for each). Applying test specific lower reference limits as criterion for the differentiation between responders and non-responders to clopidogrel treatment, 57% of the patients on clopidogrel therapy were classified as non-responders with the Multiplate ADP test, 38% with the Multiplate ADP+PG test, 55% with the P-selectin assay, 9% with the PLT VASP/P2Y12 assay.The VASP phosphorylation assay appeared to be advantageous for the assessment of clopidogrel action compared to the Multiplate ADP+PG test, the P-selectin assay, and the Multiplate ADP test (listed in descending order). However, our method comparison study underscores the critical nature of the dependence of results on the techniques used in specific studies, and it remains to be elucidated which method correlates best with the occurrence of adverse events.

Purpose The aim of this study was to compare the prognostic value of interleukin 6 (IL-6), galectin 3, growth differentiation factor 15 (GDF-15), and soluble ST2 (sST2) in an unselected cohort of critically ill patients. Methods During a study period of 1 year, we recruited 530 consecutive patients admitted to a medical intensive care unit of a tertiary care hospital. We examined a combination of inflammatory, renal, and cardiac biomarkers for the prediction of 90-day all-cause mortality. Results During follow-up, 118 patients died (22%). In univariate analyses, increased IL-6, galectin 3, GDF-15, and sST2 plasma concentrations at baseline were strong prognostic markers. However, in the multivariate models, only IL-6 and sST2 remained independent biomarkers adding additional prognostic information to the routinely used Simplified Acute Physiology Score (SAPS) II. Using a simple multimarker approach, patients with increased SAPS II, IL-6, and sST2 (ie, SAPS II > 35, IL-6 > 32.3 pg/mL, and sST2 > 103 ng/mL) had the poorest outcome. Conclusions In this heterogeneous group of critically ill patients, only SAPS II, IL-6, and sST2 remained independent and additive prognostic markers for 90-day all-cause mortality. A combination of the SAPS II with the 2 complementary biomarkers might provide a valuable tool for risk stratification of critically ill patients.

Background: The aim of the present investigation was to test the hypothesis that pregnancy-associated plasma protein-A (PAPP-A), a zinc-binding metalloproteinase implicated in acute coronary syndrome, is associated with atherosclerotic peripheral arterial disease (PAD). Methods: The study comprised 433 patients with symptomatic atherosclerotic PAD (i.e., chronic limb ischemia) and 433 controls matched to the patients with PAD in a 1:1 design by sex, age (±2 years), and diabetes mellitus status. Serum PAPP-A concentrations were measured with an enzymatically amplified 2-step sandwich-type immunoassay. Results: The entire study sample included 612 male and 254 female patients with a median age of 68 years. The median PAPP-A value was higher in the patients with PAD than in the referents (0.81 vs 0.64 mU/L; P Conclusions: PAPP-A was associated with atherosclerotic PAD in the elderly sample studied. Because atherosclerotic PAD is considered an indicator of systemic atherosclerotic disease in elderly patients, the present results indicate that circulating PAPP-A may be a marker for systemic atherosclerotic disease.

Background: The aim of the present study was to assess the utility of amino-terminal pro-A-type natriuretic peptide (NT-proANP) measurements for the emergency diagnosis of acute destabilized heart failure (HF), using a novel sandwich immunoassay covering midregional epitopes (MR-proANP). Methods: The retrospective analysis comprised 251 consecutive patients presenting to the emergency department of a tertiary care hospital with dyspnea as a chief complaint. The diagnosis of acute destabilized HF was based on the Framingham score for HF plus echocardiographic evidence of systolic or diastolic dysfunction. A commercially available immunoluminometric assay was used for measurement of MR-proANP plasma concentrations. Results: Median MR-proANP plasma concentrations were significantly higher in patients with dyspnea attributable to acute destabilized HF (338 pmol/L; n = 137) than in patients with dyspnea attributable to other reasons (98 pmol/L; n = 114; P Conclusions: MR-proANP measurements may be useful as an aid in the diagnosis of acute destabilized HF in short-of-breath patients presenting to an emergency department. The diagnostic value of MR-proANP appears to be comparable to that of BNP and NT-proBNP.

Background: The aim of the present study was to prospectively evaluate the diagnostic utility of B-type natriuretic peptide (BNP) and amino-terminal proBNP (NT-proBNP) measurements for the detection of cardiac structural disease in asymptomatic patients with systemic arterial hypertension and to test the hypothesis that the 2 analytes are equally useful in this clinical setting. Methods: We studied a consecutive series of 149 asymptomatic patients referred for echocardiographic evaluation of the cardiac effects of systemic arterial hypertension. Diagnosis of cardiac structural disease was based on the presence of systolic or diastolic dysfunction, left atrial dilatation, left ventricular dilatation or hypertrophy, pulmonary hypertension, and wall motion or valvular abnormalities. Blood concentrations of BNP and NT-proBNP were measured by 2 commercially available assays (Abbott AxSYM and Roche Elecsys, respectively). Diagnostic accuracies of BNP and NT-proBNP were assessed by ROC curve analysis. Areas under the curves were compared by analysis of equivalency. Results: In distinguishing between hypertensive patients with cardiac structural disease (n = 118) and hypertensive patients without (n = 31), areas under the curves were 0.740 (95% confidence interval, 0.662–0.808) for BNP and 0.762 (0.685–0.828) for NT-proBNP and were significantly equivalent (P = 0.015). Cutoff values with a 90% sensitivity for cardiac structural disease were 17 ng/L for BNP and 39 ng/L for NT-proBNP, with 29% and 32% specificity, respectively. Conclusions: BNP and NT-proBNP have similar capabilities for detecting cardiac structural disease in asymptomatic patients with systemic arterial hypertension. However, in the setting evaluated, a screening strategy relying on measurement of BNP or NT-proBNP may be of limited value because of the low specificity at the selected cutoff values.

The amino-terminal fragment of the B-type natriuretic peptide prohormone (NT-proBNP) is a marker for functional cardiac impairment and is increased in heart disease with or without symptoms of heart failure (HF) (1). There are indications that currently used assays for NT-proBNP may differ in their cross-reactivity with circulating NT-proBNP split products and may also be affected by breakdown products of NT-proBNP produced after blood collection (2). A newer generation assay, a commercially available competitive enzyme immunoassay (EIA) for NT-proBNP (Biomedica Gruppe) (3) that does not require sample pretreatment, has been used in various methodologic and clinical studies (4)(5)(6), but noncompetitive immunoassays may offer advantages of better speed, sensitivity, precision, and possibly specificity over competitive immunoassays (7). Recently, a noncompetitive immunoassay for NT-proBNP has been developed (8). A fully automated version of this assay (Roche Diagnostics) has now been cleared by the US Food and Drug Administration. Our aim was to compare the Biomedica and Roche NT-proBNP assays, addressing whether the predictive values of both assays are similar with respect to structural heart disease with or without symptoms of HF. The present study, carried out at the Division of Internal Medicine, St. John of God Hospital (Linz, Austria), was approved by the local ethics committee in accordance to the Helsinki Declaration. We prospectively recruited 157 consecutive patients admitted for extensive cardiac evaluation (including performance of bicycle ergometry) and 23 consecutive patients with symptomatic HF admitted for inpatient treatment; all participants gave written informed consent. Study participants were classified according to the American College of Cardiology/American Heart Association guidelines for the evaluation and management of chronic HF in the adults (9) to one of the four following categories: ( a ) healthy individuals (n = 42); ( b ) patients at high risk for developing HF but without structural disorders of the …

Elevated erythrocyte mean corpuscular volume (MCV) may be a risk factor for peripheral arterial disease (PAD). The aim of the present study was to evaluate whether MCV was associated with the severity of atherosclerotic findings in the lower limbs of PAD patients, as measured by an angiographic scoring system based on vessel lumen reduction. One hundred male patients with symptomatic PAD were studied. MCV was significantly correlated with the angiographic score (rs = 0.247, p = 0.013). PAD patients with an angiographic score in the lower third were compared to those with values in the upper third using a logistic regression model with age, smoking, hypertension, MCV, homocysteine, and total cholesterol and triglycerides as independent variables. This model revealed significant odds ratios (OR) for MCV (OR = 2.02 for an increment of 5 fl, 95% CI = 1.08-3.8) and for age (OR = 2.41 for an increment of 10 years, 95% CI = 1.21-4.81) and facilitated classification of 71% of all subjects correctly. In conclusion, MCV may be associated with angiographically determined disease severity in patients with PAD. This finding supports the hypothesis that MCV is a risk factor for PAD, although the mechanism by which MCV may contribute to the presence and severity of the disease is not yet determined.

High total serum homocysteine (tHcy) concentrations are associated with an increased risk of carotid artery disease in the general population. Since patients with peripheral arterial disease (PAD) have a threefold risk of cerebrovascular morbidity compared to individuals free of PAD, and since the total neurological event rate is associated with aor = 50% lumen reduction in extracranial carotid arteries, it was tested whether tHcy is a predictor of internal carotid artery stenosis in patients with symptomatic PAD. A total of 443 consecutive male PAD patients without previous carotid surgery/stenting were studied. In all, 100 patients with PAD had an internal carotid artery stenosisor = 50%. Of the remaining 343 patients, 100 individuals matched for age (+/- 2 years) and diabetes served as controls. The extent of carotid stenosis was evaluated with color duplex measurement; tHcy was determined by high-performance liquid chromatography. Cases displayed a significantly higher median fasting tHcy level (17.0 micromol/l) than controls (13.7 micromol/l, p=0.001). Multivariate analysis showed that tHcy (p=0.036) was an independent predictor of internal carotid artery stenosisor = 50% in PAD patients, representing an odds ratio of 1.32 (95% CI, 1.02-1.72) for an increment of 5 micromol/l. In the present study, high tHcy was an independent risk factor for an internal carotid artery stenosisor = 50% in patients with PAD. Since PAD patients suffer a threefold risk of stroke compared to healthy individuals, a simple vitamin substitution in PAD patients may reduce the occurrence of internal carotid artery stenosis and therefore diminish the relatively high rate of cerebrovascular events in this population.

Blood measurements of B-type natriuretic peptide (BNP), a cardiac neurohormone, are proposed as a diagnostic and prognostic aid in congestive heart failure (CHF) and as a prognostic marker in acute coronary syndromes (ACS) (1). In addition, there are preliminary data indicating a possible role of BNP as an aid in guiding medical therapy in patients with CHF (2)(3). The growing interest in clinical determination of BNP has led to the development of immunoassays for the determination of this analyte that are suitable for fully automated, high-throughput clinical instruments with random access, e.g., the ADVIA Centaur BNP (Bayer Diagnostics) and AxSYM BNP (Abbott Laboratories). We aimed to perform a preliminary analytical evaluation of the recently developed AxSYM BNP assay and to compare its clinical performance with that of the ADVIA Centaur BNP assay. The properties of the ADVIA Centaur BNP assay have been reported previously (4). As indicated by the manufacturer, the AxSYM BNP assay, produced by Axis-Shield plc for Abbott Laboratories, is a fully automated microparticle enzyme immunoassay that uses two monoclonal mouse antibodies in a two-step sandwich format. The analyte is captured on anti-BNP-coated latex microparticles with detection via alkaline phosphatase and measurement of fluorescence produced from the breakdown of 4-methylumbelliferyl phosphate substrate. The solid-phase antibody is directed at the NH2 terminus of the BNP peptide, whereas the conjugate antibody is directed at the COOH terminus. Six calibrators are used for the AxSYM BNP assay. Calibrator A is an acetate buffer with protein stabilizers. Calibrators B–F contain increasing concentrations of synthetic BNP (100, 400, 1000, 2000, and 4000 ng/L) in acetate buffer with protein stabilizers. These calibrators are traceable to a reference standard that was prepared gravimetrically with synthetic BNP; the reference standard underwent a value assignment to align with the Biosite Triage BNP assay …

BACKGROUND Soluble ST2 (sST2) plasma concentrations are significantly higher in healthy men than in healthy women. The reason for the sex-specific difference of sST2 plasma concentrations is not established. The aim of this study was to evaluate the association of sST2 with sex-hormones in healthy males and females separately. METHODS We recruited 528 consecutive blood donors and measured plasma concentrations of sST2 and several sex-hormones (i.e., total testosterone, estradiol, sex hormone-binding globulin, follicle-stimulating hormone, and luteinizing hormone). Of the 528 blood donors, 338 were male and 190 were female. For data analysis, we further divided the group of females into the subgroups of pre- and postmenopausal women using the age of 50 years as a proxy for menopause. RESULTS In non-parametric Spearman's correlation analyses, we found a weak association between sST2 and total testosterone (r(s)+0.126, p=0.021) and also between sST2 and estradiol (r(s)+0.117, p=0.032) in males. In females

Background: Pregnancy-associated plasma protein-A (PAPP-A) has been associated with peripheral artery disease (PAD). The aim of this study was to evaluate the utility of PAPP-A as a marker for long-term mortality in patients with atherosclerotic PAD. Methods: PAPP-A serum concentrations were measured using an enzymatically amplified two-step sandwich-type immunoassay in 487 consecutive patients admitted to a tertiary care hospital with symptomatic PAD. The main outcome measure was all-cause mortality at 5 years. Results: During follow-up, 114 patients died and 373 survived. The median PAPP-A concentration was higher among decedents compared with survivors (0.96 vs. 0.78 mU/L, p=0.024). The area under the receiver operating characteristic curve for the prediction of 5-year mortality by PAPP-A was 0.57 [95% confidence interval (CI), 0.53–0.61; p=0.026]. Survival probability was not significantly associated with PAPP-A concentrations using Kaplan-Meier curve analysis. However, univariate Cox proportional-hazards regression analysis revealed that PAPP-A was associated with 5-year mortality [risk ratio 1.25; 95% CI, 1.05–1.50; p=0.013 per one standard deviation (SD) increase in log transformed values]. In the multivariate model using a bootstrapping method, the predictive value of PAPP-A remained significant (risk ratio 1.31; 95% CI, 1.01–1.73; p=0.024 per 1 SD increase in log transformed values), even after adjustment for clinical confounders and other biomarkers, such as high-sensitivity C-reactive protein and amino terminal pro-B-type natriuretic peptide. Conclusions: In this study, PAPP-A was an independent predictor of 5-year all-cause mortality in patients with symptomatic PAD. However, based on the weak association between PAPP-A and outcome in our cohort, we consider PAPP-A measurements to not be useful in clinical practice for prognostic purposes in patients with PAD. Clin Chem Lab Med 2010;48:537–42.

The protein ST2 is a member of the interleukin-1 receptor family. Blood concentrations of the soluble isoform of ST2 (sST2) are increased in inflammatory diseases and in heart disease and are considered a prognostic marker in both. The aim of this study was the analytical and clinical evaluation of the novel Presage ST2 assay for the determination of sST2 in human plasma.We evaluated precision and linearity of the assay, analyte stability, and biological variability, determined reference values, performed a method comparison with an established ELISA, and quantified sST2 concentrations in various diseases.Within-run and total coefficients of variation were2.5% and4.0%. The method was linear across the whole measurement range of the assay. The analyte was stable for 48 h at room temperature, for 7 days at 4 degrees C, and for at least 2 months at -20 degrees C and -80 degrees C. The reference change value for healthy individuals was 30%. Age-independent reference values were 3-28 U/mL in males, and 2-16 U/mL in females. The method comparison revealed a high proportional bias. sST2 plasma concentrations were increased modestly in heart failure and moderately in pneumonia and chronic obstructive pulmonary disease. Patients with sepsis exhibited highly elevated sST2 values. In patients with chronic renal disease, however, there was no difference compared to healthy individuals.The Presage ST2 assay meets the needs of quality specifications of laboratory medicine. The results of the clinical assay evaluation are novel with respect to sST2 in various diseases and should initiate further studies.

This study evaluated the utility of the PFA-100 and the Multiplate analyzer for the assessment of aspirin and clopidogrel effects on platelet function in patients with cardiovascular disease. Platelet function was determined with the PFA-100 using collagen+epinephrine (CEPI) and collagen+adenosine-5’-diphosphate (CADP) cartridges, and with whole blood impedance aggregometry using the Multiplate ASPI and ADP+PG tests (aggregation triggered with arachidonic acid and ADP+ prostaglandin E1, respectively). Four study groups were identified from the 154 patients enrolled: patients without antiplatelet therapy, patients with 100 mg aspirin daily but without clopidogrel treatment, patients with 75 mg clopidogrel daily but without aspirin treatment, and patients with both 100 mg aspirin daily plus 75 mg clopidogrel daily. It was found that the PFA-100 instrument is useful for detection of aspirin but not for detection of a clopidogrel effect, while the Multiplate analyzer is useful for specific detection of both aspirin and clopidogrel effects on platelet function.

Background: The soluble isoform of the interleukin-1 receptor family member ST2 (sST2) has been implicated in heart failure. The aim of the present study was to evaluate the capability of sST2 as a prognostic marker in patients with acute destabilized heart failure. Methods: sST2 plasma concentrations were obtained in 137 patients with acute destabilized heart failure attending the emergency department of a tertiary care hospital. The endpoint was defined as all-cause mortality, and the study participants were followed up for 365 days. Results: Of the 137 patients enrolled, 41 died and 96 survived during follow-up. At baseline the median sST2 plasma concentration was significantly higher in the patients who died than in those who survived (870 vs 342 ng/L, P 700 ng/L; 25 deaths vs 21 survivors) of sST2 plasma concentrations compared with the first tercile (sST2, ≤300 ng/L; 5 deaths vs 41 survivors). In multivariable Cox proportional-hazards regression analyses, an sST2 plasma concentration in the upper tercile was a strong and independent predictor of all-cause mortality. Conclusions: Increased sST2 concentrations determined in plasma samples drawn from patients with acute destabilized heart failure at their initial presentation indicate increased risk of future mortality. Increased sST2 plasma concentrations are independently and strongly associated with one-year all-cause mortality in these patients.

Background: The chemical inertness of hydroxyethyl starch (HES) might cause interferences of the colloid with a variety of laboratory tests. We aimed to evaluate potential influences of HES 130/0.4, the newest HES type, on several common hematology and clinical chemistry parameters. Methods and results: A convenient sample of 25 patients scheduled for rheological therapy with 500 mL 6% HES 130/0.4 was evaluated. Blood samples were drawn before and after colloid application. Comparing pre- and post-infusion values of a battery of laboratory tests (i.e., hematology and hemostasis parameters, electrolytes, enzymes, kidney and metabolic parameters, lipids, etc.) in time course, a median difference greater than the reference change value for a specific parameter was considered clinically relevant. Among all parameters tested, only serum amylase activity displayed a clinically relevant difference between pre- and post-infusion values (median increase of 85% due to HES administration). By applying in vitro experiments, we demonstrated that serum amylase values obtained in the samples diluted in a 1:1 ratio with HES 130/0.4 and in samples diluted in a 1:1 ratio with 0.9% NaCl displayed a negligible median difference of 3%. Conclusions: The in vivo effect of HES 130/0.4 administration on serum amylase activity observed in our study was pharmacological (real) in nature. With the exception of the influence of HES 130/0.4 on amylase activity, the effects of HES 130/0.4 on other parameters tested in this study can be interpreted as having no clinical relevance.

The aim of the present study was to investigate the relationship of adiponectin, a novel adipocytokine, and amino terminal proBNP (NT-proBNP) in patients with peripheral arterial disease (PAD).Serum concentrations of adiponectin and NT-proBNP were measured in 487 patients with symptomatic PAD from the Linz Peripheral Arterial Disease (LIPAD) study.Correlation analysis revealed an association of adiponectin and NT-proBNP (r, +0.47; p0.001). Even after adjustment for age, sex, body mass index, diabetes mellitus, smoking, arterial hypertension, estimated glomerular filtration rate (eGFR), fasting glucose, LDL-cholesterol, HDL-cholesterol, triglycerides, high-sensitivity C-reactive protein, and total homocysteine the relationship of adiponectin and NT-proBNP remained significant (r, +0.35; p0.001). Furthermore, a subgroup analysis of patients with first manifestation of symptomatic PAD (n=287) demonstrated that disease severity (classified by Fontaine stages) was positively related to adiponectin (r, +0.13; p=0.003) and NT-proBNP (r, +0.28; p0.001).Adiponectin was positively associated with NT-proBNP in symptomatic atherosclerotic PAD, independent of traditional and non-traditional risk factors. Moreover, adiponectin and NT-proBNP were related to disease severity, indicating a possible role for assessment of future morbidity and mortality in patients with PAD.

There is growing evidence that adiponectin, an adipocytokine with anti-inflammatory and antiathero-genic properties, is involved in the development of atherosclerosis. The aim of the present study was to examine whether serum levels of adiponectin were associated with symptomatic atherosclerotic peripheral arterial disease (PAD). Serum concentrations of adiponectin were measured in 433 patients with symptomatic PAD and 433 controls from the Linz Peripheral Arterial Disease (LIPAD) study. Cases and controls were matched for age, sex and diabetes mellitus. The median serum level of adiponectin was significantly lower in PAD patients than in control subjects (9.5 vs. 10.8mg/L; p=0.014). After adjustment for several possible confounding variables using multivariable logistic regression, odds ratios for symptomatic PAD were 0.95 (95% CI, 0.64–1.42; p=0.080) and 0.59 (95% CI, 0.36–0.97; p=0.037) in the second and third tertiles for adiponectin serum concentrations, respectively, compared with the first tertile. Low serum levels of adiponectin were associated with the presence of symptomatic atherosclerotic PAD, independent of traditional and non-traditional risk factors, suggesting that hypoadiponectinemia may be a marker for systemic atherosclerotic disease.Clin Chem Lab Med 2006;44:830–3.

Background The aim of the present study was to evaluate the capability B-type natriuretic peptide (BNP) as a prognostic marker in patients with acute destabilized heart failure in comparison with mid-regional pro-A-type natriuretic peptide (MR-proANP), mid-regional pro-adrenomedullin (MR-proADM), and the C-terminal part of the arginine vasopressin prohormone (Copeptin). Methods and Results BNP, MR-proANP, MR-proADM, and Copeptin plasma concentrations were obtained in 137 patients with acute destabilized heart failure attending a tertiary care hospital. The end point was defined as all-cause mortality, and the study participants were followed for 365 days. Of the 137 patients enrolled, 41 died and 96 survived during follow-up. ROC curve analysis showed that the areas under curve for the prediction of 1-year mortality were similar for BNP (0.716; 95% CI 0.633–0.790), MR-proANP (0.725; 95% CI 0.642–0.798), MR-proADM (0.708; 95% CI 0.624–0.782), and Copeptin (0.688; 95% CI 0.603–0.764). Using tercile approaches, Kaplan-Meier curve analyses demonstrated that the predictive value of all four analytes for survival probability was comparable (log-rank test for trend, P < .001 for each). In multivariable Cox proportional-hazards regression analyses, increased BNP, MR-proANP, MR-proADM, and Copeptin plasma concentrations were the strongest predictors of mortality. Conclusion BNP is considered an established prognostic marker for heart failure patients. The present study provides evidence that MR-proANP, MR-proADM, and Copeptin measurements might have similar predictive properties compared with BNP determinations for one-year all-cause mortality in acute destabilized heart failure.

Background The aim of the present study was to demonstrate the capability of B-type natriuretic peptide (BNP) and amino terminal proBNP (NT-proBNP) as prognostic markers in patients with dyspnoea as a chief complaint. Methods BNP and NT-proBNP plasma concentrations were obtained from 251 short of breath patients presenting to the emergency department of a tertiary care hospital. Patients with acute coronary syndromes or trauma were excluded. The endpoint was defined as all-cause mortality, and the study participants were followed up for 365 days from the time they attended the emergency department. Results Of the 251 patients, 62 died and 189 stayed alive during follow-up. In the present study, optimal cut off levels for the prediction of survival were 454 ng/L for BNP, and 2060 ng/L for NT-proBNP. Mortality was higher in patients with baseline BNP and NT-proBNP concentrations above these cut off levels (log rank p

To address the value of plasma B-type natriuretic peptide (BNP) concentrations as a diagnostic tool for determining the cardiac etiology of pleural effusions, and to determine possible differences of plasma BNP concentrations before and after pleurocentesis in patients with congestive heart failure (CHF).Observational study.Tertiary care hospital.Consecutive series of 64 patients with indications for diagnostic pleurocentesis. The final diagnosis of the underlying disease was assessed by clinical criteria. Seven patients were excluded due to pleural effusions of equivocal origin or due to obvious hemothorax secondary to trauma.Pleurocentesis attempting to drain effusions dry. Plasma BNP concentrations were measured directly before pleurocentesis and 24 h after the intervention. During these 24 h, the dosages of patients' medications were held constant.In distinguishing between patients with pleural effusions caused by CHF (n = 31) and patients with pleural effusions attributable to other causes (n = 26), the area under the curve was 0.974 (SE, 0.021; 95% confidence interval, 0.892 to 0.997) for plasma BNP. A BNP cutoff concentration of 2,201 ng/L had a sensitivity of 77% and a specificity of 100% in the diagnosis of CHF. The median plasma BNP concentrations in patients with pleural effusions caused by CHF (n = 31) did not change within 24 h after pleurocentesis compared with the concentrations obtained before the procedure (before pleurocentesis, 3,227 ng/L; 24 h after pleurocentesis, 2,759 ng/L; p = 0.189), despite a median removal of 1,100 mL pleural fluid.Plasma BNP concentrations of patients with pleural effusions of unknown origin may be an aid in the diagnosis of CHF as the underlying cause. If plasma BNP is used as a surrogate marker of global cardiac function, there is no indication of hemodynamic improvement caused by pleurocentesis alone in patients with CHF and pleural effusions.

Objective: To compare head to head the diagnostic accuracy of B type natriuretic peptide (BNP) and the amino terminal fragment of its precursor hormone (NT-proBNP) for congestive heart failure (CHF) in an emergency setting. Methods: 251 consecutive patients presenting to the emergency department with dyspnoea as a chief complaint were prospectively studied. Patients with acute coronary syndromes were excluded. The diagnosis of CHF was based on the Framingham score for CHF plus echocardiographic evidence of systolic or diastolic dysfunction. Blood concentrations of BNP and NT-proBNP were measured by two commercially available assays (Abbott and Roche methods). The diagnostic accuracies of BNP and NT-proBNP were assessed by receiver operating characteristic curve analysis. Results: Areas under the curve for BNP and NT-proBNP in patients with dyspnoea caused by CHF (n = 137) and in patients with dyspnoea attributable to other reasons (n = 114) did not differ significantly (area under the curve 0.916 v 0.903, p = 0.277, statistical power 94%). Cut off concentrations with the highest diagnostic accuracy were 295 ng/l for BNP (sensitivity 80%, specificity 86%, diagnostic accuracy 83%) and 825 ng/l for NT-proBNP (sensitivity 87%, specificity 81%, diagnostic accuracy 84%). Evaluation of discordant false classifications at these cut off concentrations showed no advantage for either BNP nor NT-proBNP in the biochemical diagnosis of CHF (17 misclassifications by BNP and 14 by NT-proBNP, p = 0.720). In the population studied, age, sex, and renal function had no impact on the diagnostic utility of both tests when compared by logistic regression models. Conclusions: BNP and NT-proBNP may be equally useful as an aid in the diagnosis of CHF in short of breath patients presenting to the emergency department.

A new fully automated chemiluminescence assay for total homocysteine (tHcy) determination (ADVIA Centaur homocysteine, Bayer) was evaluated in comparison with a previously established fluorescence polarization assay (AxSYM homocysteine, Abbott). Linearity could be demonstrated in a concentration range up to 50 micromol/l for both methods. The detection limit was 0.92 micromol/l for the AxSYM and 0.61 micromol/l for the ADVIA Centaur analyzer. Within-run coefficients of variation (%CV) ranged from 1.7% to 1.8% for the AxSYM, and from 2.2% to 2.7% for the ADVIA Centaur analyzer, total CV ranged from 2.5% to 3.5% for the AxSYM, and from 3.6% to 4.5% for the ADVIA Centaur analyzer. Passing and Bablock regression analysis of 180 samples with the AxSYM assay as reference method revealed an intercept of -0.41 micromol/l (95% CI -1.17 to 0.20 micromol/l) and a slope of 1.11 (95% CI 1.05 to 1.18), the Bland-Altman difference plot showed a mean difference of -0.9 micromol/l between tHcy measurements with wide 95% limits of agreement (-3.6 to 1.7 micromol/l). At thresholds of 10 and 15 micromol/l there was a considerable proportion of discordant classifications of study subjects by the AxSYM and ADVIA Centaur method. When evaluating case-control status for vascular disease both assays showed similar characteristics (i.e., significant difference of tHcy in 71 CAD patients and 109 control subjects, and non-significant odds ratios for tHcy in the multivariate model). In conclusion, both methods are reliable for routine tHcy determination in clinical laboratories, as they are fast and completely automated systems with good accuracy and precision allowing sample random access, automatic dilution and stored calibration capabilities. However, results of both assays may not be used interchangeably since the ADVIA Centaur method tends to overestimate tHcy values compared to the AxSYM method.

The short-term infusion of levosimendan (Simdax™), a calcium sensitizer, improves the hemodynamic function in patients with decompensated heart failure (1)(2)(3). Blood concentrations of B-type natriuretic peptide (BNP) and the amino-terminal fragment of its precursor hormone (NT-proBNP) have been reported to reflect the severity of heart failure (4), and BNP concentrations have been shown to decrease with improving hemodynamic function during tailored intravenous treatment of decompensated heart failure (5). Because of the shorter biological half-life of BNP compared with NT-proBNP (6), we hypothesized that BNP would show a faster response to hemodynamic improvement during intravenous levosimendan therapy. This could be of relevance considering the possible role of natriuretic peptides for guiding therapy in acutely decompensated heart failure. The present observational study, carried out prospectively at the Division of Internal Medicine, St. John of God Hospital (Linz, Austria), was approved by the local ethics committee in accordance to the Helsinki Declaration. Eligible patients were those admitted with acute decompensation of chronic heart failure who were judged to require hemodynamic monitoring and intravenous treatment. Inclusion criteria were defined as follows, based on a previous report (1): documented left ventricular ejection fraction ≤30% by echocardiogram and a pulmonary artery catheter placed for clinical purposes that demonstrated a pulmonary capillary wedge pressure (PCWP) ≥15 mmHg along with a cardiac index (CI) ≤2.5 L · min−1 · m−2. Exclusion criteria were angina-limited exercise; unstable angina or acute myocardial infarction with urgent need for invasive procedure; obstructive cardiomyopathy; uncorrected primary stenotic valve; history of ventricular flutter, fibrillation, …

The aim of the present study was to assess the long-term stability of endogenous B-type natriuretic peptide (BNP) and amino terminal proBNP (NT-proBNP) in plasma samples stored at -20 degrees C without addition of protease inhibitors (e.g., aprotinin). Stability of BNP and NT-proBNP was tested in 60 EDTA plasma samples with BNP values between 30 and 420 pg/ml. Initial BNP and NT-proBNP plasma concentrations were determined within four hours after blood collection using the AxSYM BNP and the Elecsys NT-proBNP assays. Subsequently, all samples were stored at -20 degrees C and were thawed for the second BNP and NT-proBNP determination on the two instruments after one day, 30 days, 60 days, 90 days and 120 days, respectively. Mean recovery (i.e., residual immunoreactivity) of BNP and NT-proBNP expressed in percent of the initial value for the given time interval of storage was calculated. Mean recovery of BNP was less than 70% after one day of storage at -20 degrees C and decreased to less than 50% after two to four months of storage (e.g., recovery of endogenous BNP after three months of storage at -20 degrees C ranging from 0% to 71%). In contrast, mean recovery of NT-proBNP was generally greater than 90%, irrespective of the duration of storage at -20 degrees C (e.g., recovery of endogenous NT-proBNP after three months of storage at -20 degrees C ranging from 91% to 112%). In conclusion, the determination of endogenous BNP with the AxSYM assay using frozen plasma samples may not be valid under the conditions tested. In contrast, NT-proBNP as measured by the Elecsys assay may be stored at -20 degrees C for at least four months without a relevant loss of the immunoreactive analyte.

Background: B-type natriuretic peptide (BNP) and the amino-terminal fragment of the BNP prohormone (NT-proBNP) are markers for functional cardiac impairment and are elevated in heart failure (HF). Aim of the present study was to perform a head-to-head comparison of the diagnostic utility of BNP and NT-proBNP in symptomatic and asymptomatic structural heart disease. Methods: We prospectively classified 180 consecutive subjects according to ACC/AHA guidelines. Blood concentrations of BNP and NT-proBNP were determined by two fully automated chemiluminescent assays (Bayer and Roche method). Diagnostic utilities were tested by ROC analyses and logistic regression. Results: ROC curves of BNP and NT-proBNP in patients with symptomatic HF (n=43) and asymptomatic subjects (n=137) did not differ significantly (AUC 0.930 vs. 0.918, p=0.650), but comparison of patients with asymptomatic structural heart disease (n=56) and subjects without structural disorder of the heart (n=81) revealed different AUCs for the respective assays (0.735 vs. 0.839, p=0.009). In the population studied, age, sex and renal function had no impact on the diagnostic performance of both tests when compared by logistic regression models. Conclusions: Both assays facilitate diagnosis of symptomatic and asymptomatic structural heart disease. BNP and NT-proBNP may be equally useful as an aid in the differential diagnosis of probable signs or symptoms of HF. In contrast, NT-proBNP might be a more discerning marker of early cardiac dysfunction than BNP.

A tool to identify vascular patients who receive antiplatelet therapy and to distinguish between responders and non-responders to antiplatelet therapy could be of clinical importance. The present observational study was designed to investigate whether the PFA-1 00± device (Dade Behring) is suitable to detect long-term therapy of aspirin (100 mg/d) and/or clopidogrel (75 mg/d) in a cohort of patients with peripheral arterial disease (PAD). A total of 150 consecutive patients with PAD were studied; 34 patients were excluded from the study due to irregular intake of antiplatelet therapy or due to method limitations. Of the remaining 1 16 patients, 42 had no antiplatelet therapy, 47 had daily aspirin (100 mg) intake, 19 were administered clopidogrel 75 mg daily, and 10 received a medication with 100 mg aspirin plus clopidogrel 75 mg daily, all for at least 10 days. Nonparametric Kruskal-Wallis test with post hoc comparisons showed that collagen plus epinephrine (CEPI) closure times of the patient group receiving aspirin and the group receiving aspirin plus clopidogrel were similar (p > 0.05). In contrast, both patient groups exhibited prolonged CEPI values compared to patients without antiplatelet therapy and patients taking clopidogrel (p 0.05). However, Kruskal-Wallis test results revealed that collagen plus adenosine-5'-diphosphate closure times were not significantly different in all four patient groups (p=0.257). In conclusion, the PFA-100' device may be a suitable tool for monitoring aspirin 100 mg therapy, but it is not appropriate for the detection of clopidogrel administration in its current setup. Although it appears plausible that patients with PAD could benefit from monitoring platelet inhibition, clear evidence for this concept is still lacking.

Elevated serum total homocysteine, an established risk factor for peripheral arterial disease, is influenced by the vitamin B12 and folate status. Since these vitamins are inversely correlated with erythrocyte mean corpuscular volume, an investigation of whether mean corpuscular volume is higher in patients with symptomatic peripheral arterial disease than in healthy subjects was performed. Furthermore, a determination of predictors of increased mean corpus cular volume levels in this population free of symptomatic coronary artery disease, cere brovascular disease, and diabetes mellitus was carried out. From 469 consecutive patients with symptomatic peripheral arterial disease, 100 fulfilled study inclusion criteria. Peripheral arterial disease was confirmed by angiography. One hundred age-matched subjects without peripheral arterial disease as verified by ankle-brachial index measurements > 0.9 served as control subjects. Patients with PAD displayed a significantly higher mean corpuscular volume level (94.5 fl) than control subjects (90.9 fl, p< 0.001). Logistic regression analysis showed that current smoking status (p

Objective The aim of this study was to investigate the long-term in vitro stability of soluble ST2 (sST2). Design and methods EDTA plasma samples were drawn from 15 individuals with various diseases. The PresageTM ST2 assay was used for measurement of sST2 concentrations directly after blood collection and after storing plasma samples for 18 months at −20 °C and −80 °C. The default criterion for analyte stability was set at 95%. Results sST2 concentrations in the 15 individuals ranged from 12 U/mL to 140 U/mL. Directly after blood collection, the mean (± SD) sST2 concentration was 51 ± 37 U/mL, and absolute analyte recoveries were 50 ± 35 U/mL and 51 ± 34 U/mL after storage of samples for 18 months at −20 °C and −80 °C, respectively. Relative analyte recoveries after 18 months of storage at −20 °C and −80 °C were 99 ± 5% and 101 ± 7%. Conclusion sST2 is stable for at least 1.5 years in plasma samples stored at −20 °C and −80 °C.

The aim of this study was to evaluate the prognostic value of chromogranin A (CgA) and C-terminal endothelin-1 precursor fragment (CT-proET-1) in patients with acute destabilized heart failure.137 consecutive patients with acute destabilized heart failure attending the emergency department of a tertiary care hospital were prospectively enrolled. Plasma concentrations of CgA, CT-proET-1, and amino-terminal proBNP (NT-proBNP) were measured at baseline. The endpoint was defined as all-cause mortality; the study participants were followed up for 365 days.Decedents (n=41) had higher median plasma concentrations of CgA (9.7 vs. 6.0 nmol/L; p=0.002), CT-proET-1 (120 vs. 72 pmol/L; p=0.006), and NT-proBNP (5112 vs. 2610 ng/L; p0.001) at baseline than survivors (n=96). Applying Cox proportional-hazards regression analyses, increased CgA (6.6 nmol/L), CT-proET-1 (79 pmol/L), and NT-proBNP (3275 ng/L) revealed significant risk ratios of 1.96 (95% CI, 1.04-3.70) for CgA, 2.56 (95% CI, 1.33-4.95) for CT-proET-1, and 2.05 (95% CI, 1.09-3.87) for NT-proBNP. When the cohort was stratified according to median CgA and NT-proBNP concentrations, and to median CT-proET-1 and NT-proBNP concentrations, respectively, Cox proportional-hazards regression analyses showed the highest risk for death in patients with both increased CgA and NT-proBNP (risk ratio, 3.65; 95% CI, 1.44-9.28), and increased CT-proET-1 and NT-proBNP (risk ratio, 4.03; 95% CI, 1.61-8.88).Our study demonstrates that increased CgA and CT-proET-1 plasma concentrations at the initial presentation of patients with acute destabilized heart failure in the emergency department add independent prognostic information in addition to NT-proBNP measurement.

We read with interest the article by Fries and Krishnan about equipoise, design bias and randomized controlled trials [1]. It is important to stress that equipoise is not the principle underlying company-driven clinical trials, which are doubtlessly necessary and useful for medical progress. As a rule, companies' clinical research departments cannot afford the risk that their hypotheses are invalid and, thus, that their trials will fail. Furthermore, equipoise in non-commercial trials is a very complex issue, which we do not intend to discuss here. So, "positive expected outcomes" seems to be an interesting and realistic alternative to equipoise. We cannot, however, agree with the authors' control group considerations. From our point of view, it is not acceptable for subjects assigned to a control group to be denied standard treatment, even if the mean expected outcome (expected outcome in the verum group plus expected outcome in the control group divided by two) is positive. One of the authors' arguments for the admissibility of control group treatments below clinical standard is that patients are autonomous and can make decisions on their own. In the case of most health problems, patients are under enormous mental pressure and are not necessarily able to weigh-up the pros and cons in an objective way. Furthermore, the feasibility of a placebo-controlled trial should not depend on a company's estimation of by how far the new treatment will exceed standard treatment. Finally, and this is the essential point, it is dangerous to undermine the patients' right to get the best possible treatment. In most European countries, social security systems enable patients to get the best possible treatment. It is hard to understand why this level of treatment should be neglected in clinical trials. In conclusion, we think that the principle of "positive expected outcomes" is an interesting and realistic approach but we should not ignore the ethical principle of "best possible treatment for every patient".

Objective Factor V G1691A (Leiden), prothrombin G20210A, and methylenetetrahydrofolate reductase (MTHFR) C677T mutations are considered risk factors for venous thromboembolism. It remains to be characterized whether the presence of these relatively common mutations poses a risk for peripheral arterial disease (PAD). Therefore, we intended to test, by conducting a case-control study, the hypothesis that PAD was associated with an increased prevalence of factor V G1691A, prothrombin G20210A, and MTHFR C677T mutations. Methods The study comprised 433 patients admitted for inpatient diagnostics and treatment of PAD in patients with chronic limb ischemia. Patients with acute ischemia or malignancy were excluded. A total of 433 control subjects matched to the patients with PAD in a 1:1 design by sex, age (±2 years), and diabetes mellitus status were recruited. Factor V G1691A, prothrombin G20210A, and MTHFR C677T genotypes were assessed by polymerase chain reaction. Results For the factor V G1691A polymorphism, the genotype frequencies in PAD patients were 92.8% GG (normal homozygotes=wild type) and 7.2% GA (mutant heterozygotes), and in control subjects they were 94.0% GG and 6.0% GA (χ 2 test; P = .493). The distribution of the prothrombin G20210A genotypes was 96.3% GG (normal homozygotes=wild type) and 3.7% GA (mutant heterozygotes) in PAD patients and was 97.2% GG and 2.8% GA in control subjects (χ 2 test; P = .442). Genotype frequencies for the MTHFR C677T polymorphism were 47.8% CC (normal homozygotes=wild type), 43.4% CT (mutant heterozygotes), and 8.8% TT (mutant homozygotes) in PAD patients, compared with 47.1% CC, 44.1% CT, and 8.8% TT in control subjects (χ 2 test; P = .977). Accordingly, as determined by logistic regression analysis, no significant odds ratios for heterozygous or homozygous genotypes of the three polymorphisms could be observed. Conclusions PAD was not associated with an increased prevalence of factor V G1691A, prothrombin G20210A, and MTHFR C677T mutations in the population studied. Thus, there is no indication that of one of these mutations may be a risk factor for chronic limb ischemia. However, the role of these mutations in acute limb ischemia remains to be clarified.