Grazina Urbonaviciene

Higher cystatin C level predicts long-term mortality in patients with peripheral arterial disease

AIMS Cystatin C and cathepsins could play a role in different processes and stages of the atherosclerotic disease. We aimed to investigate the relationship of cystatin C, and cathepsins L, and S, to lethal outcome in patients with peripheral arterial disease (PAD). METHODS AND RESULTS We studied 378 patients with established PAD. Cox regression was used to assess relationships between serum cystatin C or cathepsins L and S, and time to lethal outcome. The role of cystatin for prognosis of cardiovascular death was assessed with c-statistic, and net reclassification improvement (NRI). Patients with cystatin C levels above 1 mg/l (fifth quintile) had a significantly increased adjusted risk for all-cause and cardiovascular mortality compared to patients with cystatin C levels below or equal to 1 mg/l (hazard ratios (HR) 2.2, 95% CI 1.22-4.12, and HR 3.2, 95% CI 1.39-7.59, respectively). Furthermore, high cystatin C levels were related with higher all-cause (adjusted HR 2.99, 95% CI 1.31-6.85) and cardiovascular mortality (adjusted HR 4.36, 95% CI 1.07-18.8) among PAD patients without renal impairment. Although the addition of cystatin C to conventional risk factors improved the accuracy of risk prediction model for cardiovascular mortality (0.72-0.79; p=0.03), it did not reclassify a substantial proportion of patients to risk categories (NRI=0.12, p=0.128). CONCLUSIONS Higher cystatin C levels independently predicted 5 years all-cause, and cardiovascular death in PAD patients. However, a small improvement in discrimination with the addition of cystatin C to conventional risk factors, and no improvement in reclassification of risk categories suggest that clinical usefulness of cystatin C for predicting cardiovascular mortality in PAD population might be modest.

Impact of soluble TWEAK and CD163/TWEAK ratio on long-term cardiovascular mortality in patients with peripheral arterial disease

AIM Soluble tumor necrosis factor-like weak inducer of apoptosis (sTWEAK) has recently been introduced as a potential mediator of cardiovascular disease. We examined the associations between sTWEAK, its scavenger receptor sCD163, sCD163/sTWEAK ratio and risk for long-term all-cause and cardiovascular mortality in patients with lower-extremity peripheral arterial disease (PAD). METHODS sTWEAK and sCD163 serum levels were measured retrospectively in a cohort of 295 patients with symptomatic PAD followed for 6.1±2.1 years. The endpoints were defined as all-cause or cardiovascular death. The relationship between sTWEAK, sCD163 levels, sCD163/sTWEAK ratio, and times to fatal outcome was examined by Cox proportional hazards analysis. RESULTS sTWEAK levels were significantly lower (672 (IQR 515; 872)pg/ml vs. 814 (IQR 673; 957)pg/ml, p < 0.0001), and sCD163/sTWEAK ratio significantly higher (0.91 (IQR 0.63; 1.37) vs. 0.77 (IQR 0.55; 1.12), p = 0.008) in patients with critical limb ischemia (CLI) on admission as compared with those with intermittent claudication (IC). During follow-up, 80 (27%) patients died, hereof 33 (11.5%) of cardiovascular causes. Cox regression analysis revealed that an increase of 100 pg/ml of baseline sTWEAK were associated with a decreased risk for all cause [adjusted hazard ratio (HR) 0.89 (95%CI (0.80-0.99)), p = 0.043] and cardiovascular mortality [adjusted HR 0.83 (95% CI (0.69-0.99)), p = 0.038]. The patients with lower sTWEAK concentrations had a higher risk for cardiovascular death being more than two times as great as patients in the two upper tertiles (adjusted HR 2.2, 95% CI (1.06-4.87), p = 0.035). Similarly, the risk of cardiovascular death was 3-fold increased for patients in the upper tertile of sCD163/sTWEAK ratio as comparing with the patients in two lower tertiles (adjusted HR 3.04, 95% CI (1.44-6.43), p = 0.004). The model including sCD163/sTWEAK ratio have shown a significant improvement in accuracy of cardiovascular death prediction (the area under ROC curve 0.79 (0.72-0.86) vs. 0.84 (0.78-0.90), p = 0.019). CONCLUSIONS Decreased sTWEAK concentration, and increased sCD163/sTWEAK ratio were significantly and independently associated with long-term cardiovascular mortality in patients with lower-extremity PAD.

Proteomic identification of differentially expressed proteins in aortic wall of patients with ruptured and nonruptured abdominal aortic aneurysms

OBJECTIVE To compare the basic proteomic composition of abdominal aortic aneurysm (AAA) wall tissue in patients with nonruptured and ruptured aneurysms. METHODS A proteomic approach with two-dimensional gel electrophoresis (2D-PAGE) and mass spectrometry (MS) was used to identify differentially expressed proteins in AAA tissue from nine patients with nonruptured and eight patients with ruptured AAA. Computerized image analysis was used to detect protein spots. Differentially expressed protein spots were in-gel digested and identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Western blot analysis was used to confirm differential expression. RESULTS Seven differentially expressed proteins were detected among 745 protein spots, selecting spots whose average relative volumes differed more than twofold between the nonruptured and the ruptured group. Four protein spots were up-regulated in the ruptured group, and three were down-regulated. Five of the spots were identified. Among the upregulated spots, No. 605 was identified as peroxiredoxin-2. The up-regulation was confirmed by Western blotting. No. 381 was identified as an actin fragment. Two spots, Nos. 719 and 499, could not be identified. Among the down-regulated protein spots, No. 130 contained two peptides; one reliably determined peptide, FEDGVLDPDYPR, is found in vitronectin. Another peptide, QIDNPDYK, was borderline significant and found in calreticulin. The down-regulation of vitronectin was confirmed by Western blotting. Spot Nos. 193 and 199 both contained peptides from albumin with actin also present in No. 199. CONCLUSION The identified proteins suggest that the aortic wall of ruptured aneurysms responds to a stressful condition and that proteolytic degradation of the cytoskeleton and connective tissue may be part of the response.

Markers of inflammation in relation to long-term cardiovascular mortality in patients with lower-extremity peripheral arterial disease

BACKGROUND Elevated levels of inflammatory mediators reflect vascular inflammation, and play a significant role in the genesis of atherosclerosis, plaque instability and rupture. METHODS AND MATERIAL Plasma α-defensin and serum high sensitivity C reactive protein (hs-CRP) levels were examined in 463 patients with lower-extremity peripheral arterial disease (PAD). The relationships between inflammatory markers and lethal outcome were examined by Cox regression, and receiver operating characteristic (ROC) analysis. RESULTS Overall, 126 patients died, hereof 59 of cardiovascular causes. The patients with chronic critical limb ischemia (CLI) at baseline had significantly higher α-defensin and hs-CRP levels compared with patients with intermittent claudication (IC). For patients with IC, the relative risk for cardiovascular mortality was three times higher in patients within the upper tertile of α-defensin concentration (>162 μg/l), when compared with those in the two lower tertiles (HR 3.04 95% CI 1.26-7.32). The multivariable model revealed that IC-patients with high α-defensin and high hs-CRP concentration had more than 5 times higher risk for cardiovascular mortality than those with either high α-defensin or high hs-CRP alone, and low α-defensin or low hs-CRP concentrations (HR 5.16, 95% CI 1.78-14.8). Area under the ROC curve for combined use of high values of α-defensin and hs-CRP was 0.71 (95% CI 0.57-0.85). The addition of α-defensin or hs-CRP to conventional risk factors significantly improved the accuracy of risk prediction model for cardiovascular mortality. No associations were found among α-defensin, hs-CRP, and lethal outcome for patients with CLI. CONCLUSIONS Combined analysis of α-defensin and hs-CRP, adds prognostic information with regard to the long-term cardiovascular prognosis among patients with IC.

Association of serum adiponectin with risk for cardiovascular events in patients with peripheral arterial disease.

OBJECTS Adiponectin exerts anti-atherogenic and anti-inflammatory properties and may be important as a biomarker for cardiovascular disease (CVD). We examined whether serum adiponectin was linked with future cardiovascular events or all-cause death in patients with peripheral arterial disease (PAD). METHODS The study prospectively included 468 patients (58% male) with symptomatic PAD. Serum total adiponectin was determined by an in-house immunoassay. We used Cox regression, adjusted for age, gender, BMI, systemic hypertension, smoking status, diabetes mellitus, previous myocardial infarction (MI), ankle-brachial index (ABI), symptoms of leg ischemia, total cholesterol, and use of beta-blockers (BAB) and angiotensin-converting enzyme (ACE) inhibitors to assess possible relationship between serum adiponectin and time to first non-fatal cardiovascular event, and all-cause death. RESULTS During the median follow-up of 3.5 years, 215 new cases of non-fatal cardiovascular events and 97 all-cause deaths were detected. Adjusted Cox-regression analysis showed that a 1mg/l increase in serum adiponectin was associated with a decrease in the risk of non-fatal cardiovascular events to 0.68, (95% CI 0.47-0.99) in men, but not in women (HR 0.96 95% CI 0.55-1.70). The relative risk of adverse non-fatal cardiovascular events was 77% higher in male patients within the lower adiponectin tertile, when compared with those in the higher tertile (95% CI 1.05-2.97). Moreover, serum adiponectin was the only significant independent predictor of non-fatal cardiovascular events for men with severe PAD (HR=0.37, 95% CI, 0.16-0.89; p=0.026), whereas previous MI (p=0.92) and ABI (p=0.08) failed to reach statistical significance in the multivariable model. We did not obtain any significant associations between serum adiponectin and all-cause mortality. Multivariable model revealed that age and previous MI were independently associated with risk for all-cause death. CONCLUSIONS Lower levels of serum adiponectin were significantly associated with an increased risk for future non-fatal cardiovascular events in men with symptomatic PAD, but not in women.

Proteins associated with the size and expansion rate of the abdominal aortic aneurysm wall as identified by proteomic analysis

OBJECTIVES Identification of biomarkers for the natural history of abdominal aortic aneurysms (AAA) holds the key to non-surgical intervention and improved selection for AAA repair. We aimed to associate the basic proteomic composition of AAA wall tissue with the expansion rate and size in patients with AAA. METHODS A proteomic approach was used, consisting of two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and mass spectrometry (MS) to identify differentially expressed proteins in AAA tissue. Relevant protein spots were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). RESULTS Spearmans correlation analysis revealed 16 protein spots were strongly correlated with AAA expansion rate (ρ>±0.75). Nine protein spots were identified. Six protein spots showed correlation with AAA size (ρ>±0.5). Three protein spots were identified: vitronectin with traces of calreticulin, albumin and a spot containing two proteins: collagen α-3(VI) chain and vitamin D binding protein. Interestingly, in our previous study vitronectin was shown to be down-regulated in a ruptured AAA group compared with non-ruptured AAA. Western blot analysis in the present study confirmed a correlation of vitronectin bands with AAA size in aortic aneurysm tissue. CONCLUSION A proteomic approach seems valuable, and identified several candidates not previously associated with AAA. Larger studies are required to confirm the potential and clinical role of the identified proteins.

Predictors of post-operative mortality following treatment for non-ruptured abdominal aortic aneurysm

The aim of this prospective study of patients undergoing repair of non-ruptured abdominal aortic aneurysm between 1999 and 2003 was to evaluate and compare risk factors for mortality after surgery, to determine a complex of informative factors for lethal outcome, and to define patient risk groups. Logistic regression analysis revealed a complex of informative factors, including female gender, previous myocardial infarction, age greater than 75 years, and clinical course of abdominal aortic aneurysm as important indicators for lethal outcome. A risk score model identified low-, moderate- and high-risk groups with mortality rates of 2.9%, 8.0% and 44.4%, respectively.

IGF-I and IGFBP2 in peripheral artery disease: results of a prospective study

Abstract Background and objectives. The search for novel risk factors of cardiovascular disease (CVD) has provided valuable clinical data concerning underlying mechanism of disease. Increasing evidence indicates a possible involvement of insulin-like growth factor-I (IGF-I) and its binding protein 2 (IGFBP-2) in the pathogenesis of CVD disorders. The aim of this study was to examine the relationship between levels of IGF-I and IGFBP-2 with all-cause and CVD mortality in a prospective study of patients with lower-extremity peripheral artery disease (PAD). Methods and material. Serum IGF-I and IGFBP-2 levels were obtained in 440 patients (257 males) with symptomatic PAD. Patients were followed for a median of 6.1 (IQ 5.1–7.2) years. The relationship between times to lethal outcome and baseline serum IGF-I and IFGBP-2 levels were examined by Cox proportional hazard analysis. The role of IFGBP-2 for prognosis of CVD death was assessed with c-statistic. Results. During follow-up 115 (26%) patients (48 females and 67 males) died, and 53 (12%) died from CVD-related causes. Cox regression analysis revealed that an increase of 100 μg/l of baseline IFGBP-2 were significantly associated with an increased risk for CVD mortality [crude hazard ratio (HR) 1.14 (95% CI (1.05–1.23)), and adjusted HR 1.12 (95% CI (1.01–1.24))]. The receiver operating characteristic (ROC) analysis yielded area under curve of 0.61 (95% CI: 0.51–0.67, p = 0.022). However, the model including IFGBP-2 did not show a significant improvement in accuracy of CVD death prediction [the area under ROC curve 0.73 (0.66–0.80) vs. 0.75 (0.69–0.82), p = 0.696], and net reclassification improvement was 10.3% (p = 0.23). Conclusions. Increased IFGBP-2 concentration was significantly and independently associated with long-term CVD mortality in patients with lower-extremity PAD. However, risk prediction of CVD mortality did not improve by adding IFGBP-2 to a model containing conventional CVD risk factors.

Danish study of Non-Invasive testing in Coronary Artery Disease (Dan-NICAD): study protocol for a randomised controlled trial

BackgroundCoronary computed tomography angiography (CCTA) is an established method for ruling out coronary artery disease (CAD). Most patients referred for CCTA do not have CAD and only approximately 20–30 % of patients are subsequently referred to further testing by invasive coronary angiography (ICA) or non-invasive perfusion evaluation due to suspected obstructive CAD. In cases with severe calcifications, a discrepancy between CCTA and ICA often occurs, leading to the well-described, low-diagnostic specificity of CCTA. As ICA is cost consuming and involves a risk of complications, an optimized algorithm would be valuable and could decrease the number of ICAs that do not lead to revascularization.The primary objective of the Dan-NICAD study is to determine the diagnostic accuracy of cardiac magnetic resonance imaging (CMRI) and myocardial perfusion scintigraphy (MPS) as secondary tests after a primary CCTA where CAD could not be ruled out. The secondary objective includes an evaluation of the diagnostic precision of an acoustic technology that analyses the sound of coronary blood flow. It may potentially provide better stratification prior to CCTA than clinical risk stratification scores alone.Methods/designDan-NICAD is a multi-centre, randomised, cross-sectional trial, which will include approximately 2,000 patients without known CAD, who were referred to CCTA due to a history of symptoms suggestive of CAD and a low-risk to intermediate-risk profile, as evaluated by a cardiologist. Patient interview, sound recordings, and blood samples are obtained in connection with the CCTA. All patients with suspected obstructive CAD by CCTA are randomised to either stress CMRI or stress MPS, followed by ICA with fractional flow reserve (FFR) measurements. Obstructive CAD is defined as an FFR below 0.80 or as high-grade stenosis (>90 % diameter stenosis) by visual assessment.Diagnostic performance is evaluated as sensitivity, specificity, predictive values, likelihood ratios, and C statistics. Enrolment commenced in September 2014 and is expected to be complete in May 2016.DiscussionDan-NICAD is designed to assess whether a secondary perfusion examination after CCTA could safely reduce the number of ICAs where revascularization is not required. The results are expected to add knowledge about the optimal algorithm for diagnosing CAD.Trial registrationClinicaltrials.gov identifier, NCT02264717. Registered on 26 September 2014.

Diagnosing coronary artery disease after a positive coronary computed tomography angiography: the Dan-NICAD open label, parallel, head to head, randomized controlled diagnostic accuracy trial of cardiovascular magnetic resonance and myocardial perfusion scintigraphy

Aims Perfusion scans after coronary computed tomography angiography (CCTA) in patients with suspected coronary artery disease (CAD) may reduce unnecessary invasive coronary angiographies (ICAs). However, the diagnostic accuracy of perfusion scans after primary CCTA is unknown. The aim of this study was to determine the diagnostic accuracy of cardiac magnetic resonance (CMR) and myocardial perfusion scintigraphy (MPS) against ICA with fractional flow reserve (FFR) in patients suspected of CAD by CCTA. Methods and results Included were consecutive patients (1675) referred to CCTA with symptoms of CAD and low/intermediate risk profile. Patients with suspected CAD based on CCTA were randomized 1:1 to CMR or MPS followed by ICA with FFR. Obstructive CAD was defined as FFR ≤ 0.80 or > 90% diameter stenosis by visual assessment. After initial CCTA, 392 patients (23%) were randomized; 197 to CMR and 195 to MPS. Perfusion scans and ICA were completed in 292 patients (CMR 148, MPS 144). Based on the ICA, 117/292 (40%) patients were classified with CAD. Sensitivity, specificity, negative predictive value (NPV), and positive predictive value (PPV) for CMR were 41%, 95% CI [28-54], 84% [75-91], 62% [45-78], and 68% [58-76], respectively. For the MPS group 36% [24-50], 94% [87-98], 81% [61-93], and 68% [59-76], respectively. Conclusion Patients with low/intermediate CAD risk and a positive CCTA scan represent a challenge to perfusion techniques indicated by the low sensitivity of both CMR and MPS with FFR as a reference. The mechanisms underlying this discrepancy need further investigation.