Search

Background High white blood cells (WBC) count is a well-recognized indicator of inflammation. Previous research has considered it a risk factor for coronary artery disease (CAD). Other new inflammatory risk markers are expensive to test, are not readily available, lack standardization and have not been confirmed as risk markers by multiple prospective studies. We aim to investigate whether elevated WBC count representsan independent risk factor for CAD in our Portuguese population. Material and methods A case-control study with 3,160 individuals (mean age 53.1 ± 7.8; 77.6% male), namely 1,723 coronary patients and 1,437 controls, was performed. Together with WBC, the following CAD risk factors were analyzed: smoking, hypertension, diabetes, dyslipidemia and obesity through bivariate analysis. Multivariate logistic regression evaluated the independent risk factors for CAD. Results and conclusions Comparing cases with controls, 47.2% were smokers vs. 23.8% (P < 0.0001); 71.0% were hypertensive individuals vs. 52.3% (P < 0.0001); 89.0% had dyslipidemiavs 70.4% (P < 0.0001); 33.8% had diabetes vs. 13.2% (P < 0.0001) and 34.2% were obese patients vs. 28.9% (P = 0.001). After multivariate regression, smoking (OR = 3.14, P < 0.0001), diabetes (OR = 2.89, P < 0.0001), dyslipidemia (OR = 2.72, P < 0.0001), hypertension (OR = 1.95, P < 0.0001) and WBC (OR = 1.05, P = 0.035) remained as independent risk factors for CAD. Elevated white blood counts are associated with CAD risk in our population. This marker is inexpensive, widely available and may further predict inflammation and coronaryartery disease risk.

Background White blood count (WBC) and C-reactive protein (CRP) elevation are well-recognized inflammation indicators and considered risk factors for coronary heart disease (CHD). Most of the other newly introduced inflammatory risk markers are expensive to test, are not readily available, lack standardization, and have not been confirmed by multiple prospective studies. Whether these classic inflammatory markers may predict majoradverse coronary events (MACE) in CHD patients is controversial. We aimed to evaluate whether high WBC and CRP are independent risk factors for MACE occurrence in CHD patients with an extended follow-up. Material and methods A cohort of 1,713 CHD patients (mean age 54.0 ± 7.5 years, 78.6% male) was prospectively followed during an extended time of 4.9 ± 3.4 years (range 1 to 17 years). Bivariate and multivariate Cox regression with WBC, CRP and traditional risk factors (age,gender, smoking, family history, dyslipidemia, diabetesand BMI) analyzed those significantlyassociated with MACE. Results and conclusions MACE group presented a higher percentage of non-smokers (P = 0.006); hypertension (P = 0.003); dyslipidemia (P = 0.025) and diabetes (P < 0.0001). WBC (P = 0.048) and CRP (P < 0.0001) were also significantly higher in patients with MACE relatively to non-MACE patients. After multivariate Cox analysis, WBC and CRP remained significant risk factors for MACE, with diabetes and hypertension presenting as strong statistical value. As a conclution, the inflammatory CRP and WBC factors are accurate to predict MACE in secondary prevention. These biomarkersare easily accessed, providing helpful information in the prognosis of cardiovascular patients.

ObjectiveTo assess the influence of presbylarynx and presbycusis on Voice Handicap Index and emotional status.MethodsA case–control, prospective, observational, cross-sectional study was conducted of patients aged 65 years or older referred to an otorhinolaryngology department from January to September 2020. Presbycusis was assessed by pure tone and vocal audiometry. Each subject underwent fibre-optic videolaryngoscopy with stroboscopy, and presbylarynx was considered when two or more of the following endoscopic findings were identified: vocal fold bowing, prominence of vocal processes in abduction, and a spindle-shaped glottal gap. Each subject completed two questionnaires: Voice Handicap Index and Geriatric Depression Scale (short-form).ResultsThe studied population included 174 White European subjects, with a mean age of 73.99 years, of whom 22.8 per cent presented both presbylarynx and presbycusis. Multivariate linear regression revealed that only presence and severity of presbylarynx had an influence on Voice Handicap Index-30 scores. However, both spindle-shaped glottal gap and presbycusis influenced Geriatric Depression Scale scores.ConclusionPresbylarynx has a strong association with the impact of voice on quality of life. Presbylarynx and presbycusis seem to have a cumulative effect on emotional status.

Introduction Coronary Heart Disease (CAD) is a multifactorial disease, including environmental and genetic risk factors. Current smoking, dyslipidemia and diabetes have a significant impact in long- term mortality and morbidity. However, several genetic variants associated with CAD but not with traditional risk factors (TRFs) has been reported to improve prediction of events and extended mortality, in younger CAD people. Aim To evaluate the clinical utility of a GRS composed by variants from GWAS associated to CAD but not with TRF to predict life-long residual risk in patients under 55 years old and a low level of TRFs. Methods We conducted a prospective study with 573 consecutive patients aged 150 mg/ml). We analysed several biochemical markers and performed a GRS with variants not associated with TRFs (TCF21 rs12190287, CDKN2B-AS1 rs1333049, CDKN2B rs4977574, PHACTR1 rs1332844, MIA3 rs17465637, ADAMTS7 rs3825807, ZC3HC1 rs11556924, SMAD3 rs17228212 and GJA4 rs618675). We studied the GRS association with a primary composite endpoint of all-cause vascular morbidity and mortality including recurrent acute coronary syndrome (myocardial infarct and unstable angina), coronary revascularization (coronary artery bypass grafting (CABG) and percutaneous coronary intervention (PCI), re-hospitalization for heart failure, ischemic stroke and cardiovascular dead. Results A total of 573 patients were studied and followed up for a mean of 4.7±4.0 years. There were 169 recurrent cardiovascular events. The GRS was sub-divided into terciles, verifying that patients in the third tercile (high risk) had a higher number of risk alleles. Compared with the low-risk GRS tercile, the multivariate-adjusted HR for recurrences was 1.520 (95% CI 1.011–2.286); p=0.044 for the intermediate-risk group and was 2.051 (95% CI 1.382–3.044); p Conclusions A multilocus GRS may identify young coronary disease patients with a low level of TRFs but at significant risk of long-term events recurrence. The genetic information may improve prediction discrimination, and reclassification over the conventional risk factors alone, providing better cost-effective therapeutic strategies. Funding Acknowledgement Type of funding sources: None. Figure 1

Introduction Epicardial adipose tissue (EAT) volume can be noninvasively detected by CT and has been suggested to predict major adverse cardiovascular events (MACE). Framingham Risk Score is one of a number of scoring systems used to determine an individual's chances of developing cardiovascular disease, hence identifying who is most likely to benefit from prevention. Objectives The purpose of this study was to determine net reclassification improvement (NRI) and improved risk prediction based on EAT volume, in comparison to a traditionally known cardiovascular risk score, such as the Framingham. Methods 895 asymptomatic volunteers were prospectively enrolled in a single Portuguese center (mean age 51.9±7.7, 78.5% male) and underwent a median follow-up time of 3.7 years (IQR 5.0). EAT volume was measured by Cardiac Computed Tomography (CCT) using a modified simplified method. For NRI assessment, EAT volume as a continuous variable was added to the Framingham Risk Score. Results After 3.7 median years of follow-up, 27 patients developed a MACE. Using NRI, the net proportion of events (netNRIe) that assigned a higher risk was 33.3% (better reclassified), and the net ratio of non-events (netNRIne) was 24.7%, resulting in a net reclassification index (netNRI) of 58.0%. When the new marker was included in the model, 58.0% of patients were better reclassified. In our work, a total of 33.3% of patients who suffered events (n=27) were correctly reclassified and assigned a higher risk. Conclusion EAT volume results in a high reclassification rate in an asymptomatic, low-risk population, demonstrating the benefit of this marker beyond traditional risk assessment models. Our study supports its application, especially in carefully selected individuals. Funding Acknowledgement Type of funding sources: None. Figure 1

Introduction Epicardial adipose tissue (EAT) is an emerging cardiovascular risk marker. It has been suggested to be an inflammatory mediator with a role in subclinical atherosclerosis and coronary artery disease. However, its prognostic relevance in hard clinical outcomes remains thoroughly unexplored in the literature. Purpose Evaluate the prognostic relevance of EAT, regarding the occurrence of major adverse cardiovascular events (MACE) in an asymptomatic population. Methods 895 asymptomatic volunteers were prospectively enrolled in a single Portuguese center (mean age 51.9±7.7, 78.5% male) and underwent a median follow-up time of 3.7 years (IQR 5.0). EAT volume was measured by Cardiac Computed Tomography (CCT) using a modified simplified method. Participants were distributed into two groups, above and below the EAT-volume median. We compared both groups regarding the occurrence of MACE through univariate analysis, Kaplan-Meier Survival curves and log-rank test. Association to subclinical atherosclerosis was addressed using correlation between EAT volume and calcium score (Agatson). Results There is a strong correlation between EAT volume and calcium score (r=0.205, p Conclusion In an asymptomatic population, EAT volume seems to be related to subclinical atherosclerosis and to the occurrence of adverse cardiovascular events on long-term follow-up. Our study addresses some unanswered questions, such as the prognostic relevance of EAT as an emerging cardiovascular risk marker. Funding Acknowledgement Type of funding sources: None.

Background Hepatocyte nuclear factor4 A (HNF4A) gene was considered by GWAS associated with atherosclerosis and CAD susceptibility. Loss-of-function mutations in human hepatocyte nuclear factor 4α (HNF4α), a transcriptor factor encoded by the HNF4A gene, are associated with maturity-onset diabetes of the young and lipid disorders. However, the mechanisms underlying the lipid disorders are poorly understood. Aim We propose identifying the genetic predisposition to atherosclerosis progression and events occurrence or regression and better prognosis, through a cohort study from GENEMACOR population. Methods We investigated a cohort of 1,712 patients who underwent coronary angiography with more than 70% stenosis of at least one main coronary vessel. 33 SNPs associated with the risk of CAD in previous GWAS were genotyped by TaqMan assays methodology. We evaluated the best genetic model associated with CAD prognosis (events) with a 95% CI in bivariate analysis. The hazard function was performed by a Cox survival regression model adjusted for age, sex, type 2 diabetes, hypertension, and hypercholesterolemia, to evaluate their relationship with the event's incidence. Finally, we constructed Kaplan–Meier cumulative-event curves for the significant genetic variants. Results Our evaluation revealed a SNP paradoxically associated with protection from atherosclerosis progression and events occurrence: rs1884613 C>G in the HNF4A gene on chromosome 20 dominant model [OR=0.653; 95% CI (0.522–0.817); p=0.0002]. Cox survival regression model showed a CAD protective effect of HNF4A with a Hazard ratio (HR) of 0.771; p=0.007. The Kaplan-Meier cumulative event analysis disclosed that the CG+GG vs CC genotype of rs1884613 HNF4α was associated with a better prognosis (Breslow test, p=0.004) at the end of the follow-up. Conclusion We identified, in this study, one SNPs paradoxically associated with a better CAD prognosis rs1884613 in HNF4A. The HNF4A gene variants could induce loss of HNF4α function, modifying and modulating hepatic lipase and lipid metabolism conferring a beneficial effect on atherosclerosis progression and events occurrence. Funding Acknowledgement Type of funding sources: None.

Background Metabolic syndrome (MetS) is a clinical condition composed of metabolic and cardiovascular risk factors, such as abdominal obesity, hyperglycemia, dyslipidemia and hypertension. Many patients with MetS suffer major adverse cardiovascular events (MACE) that are not adequately identified by traditional risk assessment, suggesting the need for early detection of subclinical coronary heart disease to identify those at high-risk. Coronary artery calcification (CAC) screening has added utility in categorizing patients with low, intermediate and high cardiovascular risk. Purpose Evaluate the prognostic role of CAC score in asymptomatic population patients with metabolic syndrome in cardiovascular events risk prediction. Methods A total of 1,122 asymptomatic individuals without known coronary heart disease, enrolled from GENEMACOR study, were followed for a mean of 5.3±3.4 years for the primary endpoint of all-cause of cardiovascular events. All were referred for computed tomography for the CAC scoring assessment. According to the Hoff's nomogram, 3 categories were created: low CAC (0≤CAC75). In a subgroup of 507 individuals with MetS and 615 controls, CAC values were compared by T-student and association of CAC severity with events occurrence was evaluated. Finally, a logistic regression model adjusted for CAC severity was performed in patients with MetS. Results Among our population, the extent of CAC differs significantly between men and women in the same age group. Patients with Mets (23.2%, n=115) had higher CAC scores than controls (219.0±486.0 vs 115.8±370.8, p Conclusion Our results point to the importance of the inclusion of CAC screening in patients with MetS to further stratify those patients that, despite tight control of cardiovascular risk factors, may benefit from more intensive therapies. This tool is a useful and straightforward method that could have a significant impact on the prognosis of future cardiovascular disease in patients with MetS. Funding Acknowledgement Type of funding sources: None.

Introduction TCF21 is expressed in cells that migrate into the developing plaque facilitating the repair of the vessel wall. However, the rs12190287 risk allele (C) of TCF21 can lead to reduced TCF21 expression being a risk factor for CAD. Purpose Investigate whether the variant rs12190287 G>C of TCF21 gene represents a risk factor for CAD in a Southern European population. Methods Case-control with 3139 individuals, 1723 CAD patients and 1416 controls, adjusted for age and gender. Genotyping of TCF21 rs12190287 G>C was performed by TaqMan Real-Time PCR. CAD association of each genetic model was evaluated. Multivariate logistic regression analysis adjusted for confound variables: smoking status, dyslipidemia, diabetes, physical inactivity, and hypertension, was made. Results TCF21 rs12190287 G>C has shown significant genotypic differences between cases and controls: GG 9.5% vs 11.9%; GC 43.2% vs 46.5% and CC 47.3% vs 41.6%. CAD risk was significant in all models: dominant (OR 1.28; 95% CI: 1.02–1.61; p=0.033); recessive (OR 1.26; 95% CI: 1.09–1.45; p=0.001); additive (OR 1.20; 95% CI: 1.08–1.34; p=0.001). After multivariate analysis, TCF21 variant was independently associated with CAD. Conclusion TCF21 variant rs12190287 G>C may be a risk factor for CAD. It is plausible that TCF21 loci exert its protective effect by promoting infiltration of fibromyocytes in the coronary wall lesion and fibrous layer and loss of TCF21 expression can result in fewer fibromyocytes to fibrous cap increasing vulnerability of the plaque. Funding Acknowledgement Type of funding sources: None. Variables associated with CAD risk

Journal of Statistics on Health Decision, vol. 3 n.º 1 (2021): Special Issue - Statistics on Health Decision Making: Public Health

Evidence points epicardial adipose tissue (EAT) as an emerging cardiovascular risk marker. Whether genetic polymorphisms linked with atherosclerosis are associated with higher EAT is still unknown. We aim to assess the role of genetic burden of atherosclerosis and its association to EAT in a cohort of asymptomatic individuals without coronary disease. A total of 996 participants were prospectively enrolled in a single Portuguese center. EAT volume was measured by Cardiac Computed Tomography and participants were distributed into 2 groups, above and below median EAT. SNPs were genotyped and linked to their respective pathophysiological axes. A multiplicative genetic risk score (mGRS) was constructed, representing the genetic burden of the studied SNPs. To evaluate the association between genetics and EAT, we compared both groups by global mGRS, mGRS by functional axes, and SNPs individually. Individuals above-median EAT were older, had a higher body mass index (BMI) and higher prevalence of hypertension, metabolic syndrome, diabetes, and dyslipidemia. They presented higher GRS, that remained an independent predictor of higher EAT volumes. The group with more EAT consistently presented higher polymorphic burden across numerous pathways. After adjustment, age, BMI, and mGRS of each functional axis emerged as independently related to higher EAT volumes. Amongst the 33 SNPs, MTHFR677 polymorphism emerged as the only significant and independent predictor of higher EAT volumes. Patients with higher polymorphism burden for atherosclerosis present higher EAT volumes. We present the first study in a Portuguese population, evaluating the genetic profile of EAT through GWAS and GRS, casting further insight into this complicated matter. info:eu-repo/semantics/publishedVersion

The inclusion of a genetic risk score (GRS) can modify the risk prediction of coronary artery disease (CAD), providing an advantage over the use of traditional models. The predictive value of the genetic information on the recurrence of major adverse cardiovascular events (MACE) remains controversial. A total of 33 genetic variants previously associated with CAD were genotyped in 1587 CAD patients from the GENEMACOR study. Of these, 18 variants presented an hazard ratio >1, so they were selected to construct a weighted GRS (wGRS). MACE discrimination and reclassification were evaluated by C-Statistic, Net Reclassification Index and Integrated Discrimination Improvement methodologies. After the addition of wGRS to traditional predictors, the C-index increased from 0.566 to 0.572 (p=0.0003). Subsequently, adding wGRS to traditional plus clinical risk factors, this model slightly improved from 0.620 to 0.622 but with statistical significance (p=0.004). NRI showed that 17.9% of the cohort was better reclassified when the primary model was associated with wGRS. The Kaplan-Meier estimator showed that, at 15-year follow-up, the group with a higher number of risk alleles had a significantly higher MACE occurrence (p=0.011). In CAD patients, wGRS improved MACE risk prediction, discrimination and reclassification over the conventional factors, providing better cost-effective therapeutic strategies. info:eu-repo/semantics/publishedVersion

Funding Acknowledgements Type of funding sources: None. OnBehalf GENEMACOR Introduction After the diagnosis of coronary artery disease (CAD), traditional risk factors such as diabetes mellitus, dyslipidemia, hypertension and smoking have been used to assess the risk of major cardiovascular adverse events (MACE). However, despite reduction of these factors, presence of MACE remains high. It is necessary to identify other causal risk factors for MACE in coronary patients and increased plasma Homocysteine (Hcy) level seems to be a likely candidate. However, the influence of Hcy levels in the prognosis of coronary patients presents a limited knowledge. Objective To evaluate the influence of high level of Hcy in MACE (defined as a composite endpoint of cardiovascular death, acute myocardial infarction, stroke, admission for heart failure and need to revascularization) of coronary artery patients. Materials and Methods Study analyses of 1687 patients selected from GENEMACOR study population, with at least one > 75% coronary stenosis by angiography. That population was divided in three terciles according to the Hcy level and the population of the 2nd tercil (Hcy 11.1-13.6mmol/L) was excluded. The end population of 1118 patients was a median age of 53.1 ± 7.9 years and 77.6% were men. We compared patients in the 1st (Hcy < 11.1mmol/L) and 3rd tercil (Hcy > 13.6mmol/L) during a mean follow up of 5.0 ± 4.8 years. Results 560 (50.1%) patients were included in the 1st tercil group (median age 51.6 ± 3 years, 72.0% men) and 558 (49.9%) patients were in the 3rd tercil group (median age 54.6 ± 3 years, 83.3% men). In our population, high levels of Hcy were associated with MACE (OR 1.43, 95% CI: 1.12-1.83, p 0.004). Conclusion In our population a higher level of Hcy was associated with adverse prognosis and increased occurrence of MACE. Knowing that elevated homocysteine levels are associated with increased risk of MACE, in these patients is essential to have a more intensive therapeutic strategy.

Funding Acknowledgements Type of funding sources: None. Introduction Coronary artery disease (CAD) remains a leading cause of morbidity and mortality worldwide. We know that plasma level of LDL cholesterol (LDL-C) is strongly associated with atherosclerosis, and its reduction with statins has led to a decrease in the incidence and complications of CAD. According to the 2019 ESC guidelines, in high-risk patient the aim is to achieve an absolute LDL-C treatment goal of Objective To evaluate the degree of LDL-C control in coronary artery disease patients according to ESC guidelines. Materials and Methods Study analyses of 1687 patients selected from GENEMACOR study population, with at least one > 75% coronary stenosis by angiography (median age 53.3 ± 3 years and 54.8% men). LDL-C was determined by chemical methods and all patients were statin treated. The population was divided in four groups according to LDL-C levels: inferior to 55mg/dL, inferior to 70mg/dL, inferior to 115mg/dL and superior to 115mg/dL. Results LDL-C mean value was 108.7mg/dL, median 105.1mg/dL (P25 83.0 and P75 127.4mg/dL). 150 (8.9%) patients had LDL-C < 55mg/dL vs 1537 (91.1%) with LDL-C ≥ 55 mg/dl. 275 (16.3%) patients had LDL-C < 70 mg/dL vs 1412 (83.7%) with LDL-C ≥ 70 mg/dL. 1084 (64.3%%) patients had LDL < 115 mg/dL vs 603 (35.7%) with LDL-C ≥ 115 mg/dL. Conclusion In our population LDL-C control levels was low, with 91.1% patients with LDL-C ≥ 55mg/dL and 83.7% patients with LDL ≥ 70 mg/dL. It is interesting to note that most of our patients have LDL-C levels above the recommend by the newest and, surprisingly, the 2016 dyslipidemia guidelines. It is therefore important to implement a more intensive treatment strategy of dyslipidemia in coronary patients.

Introduction Our group has recently validated and published a new score - KAsH score. KAsH consists of a continuous, multiplicative score based on 4 simple clinical variables available at first medical contact, proven to be a robust predictor of in-hospital mortality and all-cause mortality at 1 year follow-up in patients with myocardial infarction, putting it next to other well established risk scores. However, the role of KAsH in patients with myocardial injury (Mi), a largely uncharacterized group in the literature, remains unknown. Purpose We aim to assess the predictive power of KAsH in patients with myocardial injury (Mi), regarding in-hospital mortality and at 1 year follow-up. Methods Prospective registry of 250 patients admitted consecutively through the emergency department from January 2018 onward, with higher than P99th high-sensitive troponin assay. The kit used was Roche's Elecsys hsSTAT, and the P99th appointed by the manufacturer was 14 ng/L. All patients with chronic kidney disease ClCr KAsH = (Killip Kimbal × Age × Heart Rate) / Systolic BP We used a simplified Killip classification: without heart failure (1 point), with heart failure (2 points) and in shock (3 points). We assessed the score's association to mortality and its predictive value through ROC curves and their respective area under the curve (AUC). Results Both Killip and KAsH had a significant and positive association with in-hospital mortality (KK: p=0.02; KAsH: p0.1). Conclusions KAsH seems to maintain its in-hospital predictive value even in patients with Mi. To our knowledge, this is the first study that tries to apply risk scores and stratification tools to such a heterogeneous group of patients. By comprising hemodynamic variables, KAsH may actually be a better risk stratification tool than just the severity of heart failure on admission. However, unlike previously proven in myocardial infarction (MI), KAsH score and its hemodynamic variables do not seem to justify the high mortality on the long run behind these patients. More studies will be needed to address the complex causes behind long-term mortality of Mi patients. KASH table graph Funding Acknowledgement Type of funding source: None

Introduction Coronary atherosclerosis is an important pathophysiological mechanism in the development of coronary artery disease (CAD). While it has been proven via multiple studies that elevated levels of low-density lipoprotein (LDL) contribute to the development of atherosclerosis, high-density lipoprotein (HDL) is widely thought to have atheroprotective effects. Multiple epidemiologic studies have given the idea that high HDL levels protect against CAD, however, other trials also shown that its benefit can be paradoxical. Objective To evaluate the relationship between HDL levels and the extent of CAD (one vessel vs multivessel disease) in coronary artery patients. Materials and methods Study analyses of 1676 patients selected from GENEMACOR study population, with at least one >75% coronary stenosis by angiography (median age 53.3±7.9 years, 78.6% male, median HDL 43.0±11.1 mg/dL). Population was divided according to the HDL level quartiles (1st quartile HDL 49 mg/dL). Population of the 1st and 4th quartiles (825 patients, median age of 53.3±7.9 years and 78.7% male) were adjudicated and prospectively followed-up by 5.0±4.8 years. χ2 and T student tests were used to analyze the demographic, laboratorial, angiographic and anthropometric characteristics of the population according to HDL level. Results 420 (50.9%) patients were included in the 1st quartile group (median age 53.2±7.9 years, 85.7% men) and 405 (49.1%) patients were included in the 4th quartile group (median age 53.4±8.0 years, 71.4% men). In our population, lower levels of HDL were associated with increased risk of multivessel coronary disease (OR 1.63, 95% CI 1.23–2.14, p 0.001). Conclusion Despite uncertainties about HDL benefit, in our population a higer level of HDL was associated with a shortest extent of coronary artery disease. We conclude that higher levels of HDL can be considered protective in coronary patients, and strategies to increase HDL levels may indeed translate in improved outcomes in CAD disease. Funding Acknowledgement Type of funding source: None

Background Despite being a controversial subject, multiple guidelines mention the use of Coronary Artery Calcification (CAC) scoring in the cardiovascular risk prediction, in asymptomatic population. The inclusion of CAC scoring in traditional risk models may help in decision-make providing better cardiovascular risk stratification. Purpose The aim of our study is to estimate the impact of CAC scoring in cardiovascular events risk prediction in a model based on traditional risk factors (TRFs). Methods and results The study consisted of 1052 asymptomatic individuals free of known coronary heart disease, enrolled from GENEMACOR study and referred for computed tomography for the CAC scoring assessment. A cohort of 952 was followed for a mean of 5.2±3.2 years for the primary endpoint of all-cause of cardiovascular events. The following traditional risk factors were considered: (1) current cigarette smoking, (2) dyslipidemia, (3) diabetes mellitus, (4) hypertension and (5) family history of coronary heart disease. Among this population, the extent of CAC differs significantly between men and women in the same age group. Therefore, the distribution of CAC score by age and gender was done by using the Hoff's nomogram (a). According to this nomogram, 3 categories were created: low CAC (0≤CAC75). Two Cox regression models were created, the first only with TRFs and the second adding the CAC severity categories. When including CAC categories to the TRFs, the higher severity level presented a significant risk of MACE occurrence with an HR of 4.39 (95% CI 1.83–10.52; p=0.001). Conclusion Our results point to the importance of the inclusion of CAC in both primary and secondary prevention to an improved risk stratification. Larger prospective multicentre cohorts with longer follow-up should reproduce and validate these findings. Funding Acknowledgement Type of funding source: None

Background TCF21 gene, encodes a basic-helix- loop- helix transcription factor, playing a critical action in the development of epicardial progenitor cells that give rise to coronary artery smooth muscle cells (SMC) and cardiac fibroblasts. Recent data suggest that TCF21 may play a role in the state of differentiation of SMC precursor cells that migrate to vascular lesions and contribute to fibrous cap. Purpose Investigate the association of TCF21 rs12190287G>C variant with coronary artery disease (CAD) in a Portuguese population and its role on the prognosis. Methods Case-control study with 3120 participants, 1687 coronary patients with at least 75% obstruction of a major coronary artery and 1433 controls. Genotyping used the TaqMan technique (Applied Biosystems) and then a univariate and multivariate logistic regression analysis were performed. After a mean follow-up of 5.01±4.2 years (interquartile range 1.96–7.57), the occurrence of the combined Major Adverse Cardiovascular Events (MACE) (Cardiovascular Mortality, non-fatal Myocardial Infarction, new Revascularization, Cerebrovascular Disease and Peripheric Vascular Disease) were registered and analysed by Cox regression. Finally, Kaplan-Meier survival estimate was performed. Results In the total population, GC+CC genotype was found to be associated with CAD with an OR of 1.285; CI: 1.022–1.614; p=0.031. After multivariate logistic regression, adjusted to traditional risk factors, the association with CAD remained significant for this genotype (OR=1.340; CI: 1.042–1.723; p=0.022).After Cox regression adjusted for confounding variables (age and sex, hypertension, diabetes, smoking, dyslipidemia, eGFR, Ejection fraction Conclusion The mutated TCF21 variant can provide a new marker to identify patients at high cardiovascular risk and may representa potential target for gene therapy in future. Figure 1 Funding Acknowledgement Type of funding source: None

Background: Genetic risk score can quantify individual’s predisposition to coronary artery disease; however, its usefulness as an independent risk predictor remains inconclusive. Objective: To evaluate the incremental predictive value of a genetic risk score to traditional risk factors associated with coronary disease. Methods: Thirty-three genetic variants previously associated with coronary disease were analyzed in a case-control population with 2,888 individuals. A multiplicative genetic risk score was calculated and then divided into quartiles, with the 1st quartile as the reference class. Coronary risk was determined by logistic regression analysis. Then, a second logistic regression was performed with traditional risk factors and the last quartile of the genetic risk score. Based on this model, two ROC curves were constructed with and without the genetic score and compared by the Delong test. Statistical significance was considered when p values were less than 0.05. Results: The last quartile of the multiplicative genetic risk score revealed a significant increase in coronary artery disease risk (OR = 2.588; 95% CI: 2.090-3.204; p < 0.0001). The ROC curve based on traditional risk factors estimated an AUC of 0.72, which increased to 0.74 when the genetic risk score was added, revealing a better fit of the model (p < 0.0001). Conclusions: In conclusion, a multilocus genetic risk score was associated with an increased risk for coronary disease in our population. The usual model of traditional risk factors can be improved by incorporating genetic data. Resumo Fundamento: O escore de risco genético pode quantificar a predisposição do indivíduo em desenvolver doença arterial coronariana; no entanto, sua utilidade como preditor de risco independente permanece inconclusiva. Objetivo: Avaliar o incremento no valor preditivo de um escore de risco genético aos fatores de risco tradicionais associados à doença arterial coronariana. Métodos: Trinta e três variantes genéticas previamente associadas à doença arterial coronariana foram analisadas em uma população caso-controle com 2888 indivíduos. Um escore de risco genético multiplicativo foi calculado e dividido em quartis, com o 1º quartil como a classe de referência. O risco coronário foi determinado por análise de regressão logística. Uma segunda regressão logística foi realizada com fatores de risco tradicionais e o último quartil do escore de risco genético. Com base nesse modelo, duas curvas ROC foram construídas com e sem o escore de risco e comparadas pelo teste de DeLong. A significância estatística foi considerada quando os valores de p eram inferiores a 0,05. Resultados: O último quartil do score de risco genético multiplicativo revelou um aumento significativo no risco de doença arterial coronariana (OR = 2,588; IC 95%: 2,090-3,204; p < 0,0001). A curva ROC baseada nos fatores de risco tradicionais estimou uma AUC de 0,72, que aumentou para 0,74 quando o score de risco genético foi adicionado, revelando um ajuste melhor do modelo (p < 0,0001). Conclusões: Em conclusão, um escore de risco genético com múltiplos loci foi associado a um risco aumentado de doença coronariana na nossa população. O modelo usual de fatores de risco tradicionais pode ser melhorado pela incorporação de dados genéticos.

Introduction Coronary artery disease (CAD) is the leading cause of death worldwide, placing a major economic and resource burden on health and public health systems, so efforts are being made to accurately predict risk for major adverse cardiac events (MACE). The field of risk prediction and CAD prevention continues to evolve with the identification of novel risk factors and biomarkers, such as lipoprotein a [Lp)a]. Almost 20% of the population has elevated circulating levels of Lp(a), which is recognized as an independent risk factor for CAD, stroke, peripheral arterial disease, and aortic stenosis. Importantly, studies showed that this was particularly true for women. Objective To evaluate if the elevation of Lp(a) is associated with MACE in female, male or both. Materials and methods Case control study of 3050 subjects from the GENEMACOR study population. In female population (n=676): cases were 341 patients with at least one >75% coronary stenosis (median age 55.7±7.2) and 335 normal controls (median age 55.8±6) adjusted by age with cases. In male population (n=2374): 1278 patients with at least one >75% coronary stenosis (median age 52.7±8) and 1096 controls (median age 51.9±8) also adjusted by age. χ2 and T student tests were used to analyze the demographic, laboratorial, angiographic and anthropometric characteristics of the population. Lipoprotein (a) was determined by immunoturbidimetry. High Lp(a) level was considered if superior to 30 mg/dl. Logistic regression was used to evaluate Lp(a) as a risk factor for CAD in total, female and male populations. Results In female population 44.0% patients vs 21.2% controls (p30mg/dl. In male population 39.4% patients vs 23.8% controls (p30mg/dl. In total population Lp(a)>30mg/dl was a predictor for CAD (OR 2.24, 95% CI: 1.91–2.62, p30mg/dl was also a predictor for CAD either in male (OR 2.08, 95% CI: 1.74–2.5, p Conclusions As opposed to other studies, in our population elevated Lp(a) levels (>30mg/dl) were associated with elevated CAD risk, in both men and women. We conclude that Lp(a) can be considered an independent risk factor for CAD disease in our population, and further strategies for Lp(a) reduction may indeed translate in improved outcomes in CAD disease.

Introduction The complex interaction between genes and environmental factors contribute to individual-level risk of coronary artery disease (CAD), often resulting in premature CAD. The role for genetic risk scores in premature CAD is still controversial. Objective To evaluate the importance of conventional risk factors and of a genetic risk score in younger and older patients with coronary artery disease Methods From a group of 1619 pts with angiographic documented CAD from the GENEMACOR study, we selected 1276 pts admitted for ACS and analysed them in 2 groups (group A: ≤50 years, n=491 pts, 87.2% male, mean age 44±4.9 and group B: >50 years, n=785 pts, 75.2% male, mean age 57±4.2). Univariate analysis was used to characterize the traits of each group and we used ROC curves and respective AUCs to evaluate the power of genetics in the prediction of CAD, through a Genetic Risk Score (GRS). Results 99.3% of the young patients had at least one modifiable risk factor, 18.4% had 2 modifiable risk factors and 75.2% had 3 or more modifiable risk factors. The pattern of risk factors contributing to CAD were different among groups: family history (A: 27.5%, B: 21.4%, p=0.015) and smoking habits (A: 64.8%, B: 42.9%, p0.805, p=0.0178 and 0.748->0.761, p=0.0007 in patients under and over 50, respectively). Conclusion Coronary artery disease is not all the same, as premature CAD shares a unique and specific pattern of risk factors, clinical presentation, angiographic severity and prognosis. Genetics should not be used as an excuse to justify premature CAD, as there is frequently more than one potentially reversible risk factor present even in young patients and the additive predictive value of GRS is modest.

Type 2 Diabetes (T2D) is a risk factor for dysregulation of glomerular filtration rate (GFR) and albuminuria. However, it remains unclear whether this association is only causal. Genetic variants are inherited independent of potential confounding factors and represent a lifetime exposure. Aim Investigate whether the reduction of GFR is a direct consequence of T2D or there are other genetic mechanisms involved in the pathophysiology of the evolution to chronic kidney disease. Methods Cross-sectional study with a total of 2579 individuals was performed, of which 735 patients had T2D. Subjects were classified as `'diabetic” if they were taking oral anti-diabetic medication or insulin or if their fasting plasma glucose was higher than 7.0 mmol/l or 126 mg/dl. Within the diabetic group, we considered those with (n=63) and without (n=627) decreased GFR. GFR was calculated through the Cockcroft and Gault formula and decreased GFR was defined as GFR120 mg/ml, dyslipidemia, alcohol consumption, CAD diagnosis). A logistic regression was performed firstly with the risk factors solely; and secondly adding the genetic variants in order to evaluate the independent predictors of progression to renal failure in T2D. Results After the first multivariate logistic regression with all the risk factors for decreased GFR, only CAD remained in the equation, showing to be an independent risk factor for progression to renal failure, in T2D (OR=4.17; 95% CI: 1.64–10.59; p=0.003). In the second logistic regression, including risk factors and the genetic variants, only ZNF259 rs964184 showed an independent and significant association with the risk of decreased GFR (OR=3.03; 95% CI: 1.06–8.70; p=0.039). Conclusion This study shows that the variant ZNF259 rs964184 is associated with decreased kidney function, independently of other risk factors. This finding needs further investigation to clarify the genetic mechanism behind the association of rs964184 with decreased GFR, in Type 2 diabetes.

TCF21 is a member of the basic-helix-loop-helix (bHLH) transcriptor factor family, being critical for embryogenesis of the heart, kidney and spleen. TCF21 also regulates epicardium-derived cells differentiation into smooth muscle and fibroblast lineages. Aim Investigate the impact of TCF21 rs12190287 in the prediction and discrimination of CAD risk, individually or into a genetic risk score (GRS) formed by a set of 13 genetic variants. Methods We performed a case-control study with 3050 subjects (1619 coronary patients with 53.3±8 years; 78.9% male and 1431 controls with 52.8±8 years; 76.6% male) from GENEMACOR study. We investigated all traditional risk factors (TRF), as well as 13 genetic variants from GWAS with unknown pathophysiological pathway so far, including TCF21 (rs12190287), ZC3HC1 (rs11556924), PSRC1/SORTI (rs599839), PHACTR1 (rs1332844), MIA3 (rs17465637), SMAD3 (rs17228212), ZNF259 (rs964184), ADAMTS7 (rs3825807), CDKN2B (rs4977574), 9p21.3 (rs1333049), KIF6 (rs20455), PCSK9 (rs2114580) and GJA4 (rs618675). A multiplicative genetic risk score with these 13 genetic variants (m13GRS), was calculated. Subsequently, two logistic regressions were performed; primarily with all the TRF and all the genes individually and the second with TRF and m13GRS. Results The first multivariate analysis shows that, besides the strong association of the TRF with CAD risk (with smoking status on the top of the list, with an OR of 3.2; p Conclusion TCF21 rs12190287 is a risk factor for CAD in the Portuguese population, either individually or incorporated in a m13GRS. TCF21 risk is independent from TRF. In the future, TCF21 can provide a new clues to identify patients at high cardiovascular risk and become a potential target for gene therapy.

Essential hypertension (EH) is a disease in which both environment and genes have an important role. This study was designed to identify the interaction model between genetic variants and environmental risk factors that most highly potentiates EH development.We performed a case-control study with 1641 participants (mean age 50.6 ± 8.1 years), specifically 848 patients with EH and 793 controls, adjusted for gender and age. Traditional risk factors, biochemical and genetic parameters, including the genotypic discrimination of 14 genetic variants previously associated with EH, were investigated. Multifactorial dimensionality reduction (MDR) software was used to analyze gene-environment interactions. Validation was performed using logistic regression analysis with environmental risk factors, significant genetic variants, and the best MDR model.The best model indicates that the interactions among the ADD1 rs4961 640T allele, diabetes, and obesity (body mass index ≥30) increase approximately four-fold the risk of EH (odds ratio = 3.725; 95% confidence interval: 2.945-4.711; p 0.0001).This work showed that the interaction between the ADD1 rs4961 variant, obesity, and the presence of diabetes increased the susceptibility to EH four-fold. In these circumstances, lifestyle adjustment and diabetes control should be intensified in patients who carry the ADD1 variant.

Introdução: A hipertensão arterial é uma doença complexa, multifatorial, controlada por fatores genéticos e ambientais. Objetivo: Avaliar a susceptibilidade genética no aparecimento de hipertensão arterial e sua associação com os fatores de risco tradicionais na eclosão desta patologia. Material e Métodos: Estudo caso-controlo com 1712 indivíduos, idade média de 51,0 ± 7,9 anos (860 hipertensos e 852 controlos). Avaliaram-se os fatores tradicionais, bioquímicos e as variantes genéticas: ACE I/D rs4340, ACE A2350G rs4343, AGT T174M rs4762, AGT M235T rs699 AGTR1 A1166C rs5186, CYP11B2 -344 C/T rs1799998, ADRB1 R389G rs1801253, ADRB2 R16G rs1042713, ADD1 G460W rs4961, SCNN1G G173A rs5718, GNB3 C825T rs5443, ATP2B1 A/G rs2681472, CYP17A1 T/C rs11191548, SLC4A2 C/T rs2303934. Calculámos o risco de cada gene para a hipertensão, pelos modelos dominante, recessivo, co-dominante e multiplicativo. Através da regressão logística, avaliámos as variáveis associadas à hipertensão. Elaboraram-se curvas ROC com os fatores tradicionais e posteriormente adicionando as variantes genéticas associadas com hipertensão. Analisámos os dados através do SPSS for Windows 19.0 e MedCalc v. 13.3.3.0. Resultados: As variantes genéticas ADD1 G460W, GNB3 C825T, ACE I/D e ACE A2350G associaram-se à hipertensão. A curva ROC com os factores de risco tradicionais e estas variantes mostrou um incremento na capacidade preditiva de hipertensão (p = 0,018). Discussão: Segundo os resultados do nosso estudo as variantes genéticas que após análise univariada se associaram à hipertensão arterial foram a ACE I/D rs4340, ACE A2350G rs4343, ADD1 G460W rs4961, GNB3 C825T rs5443. As duas primeiras variantes relacionam-se com a hipertensão arterial por interferirem no sistema renina-angiotensina-aldosterona, que tem um importante papel na regulação da pressão arterial. Salienta-se o facto dos genes que codificam os componentes do sistema renina-angiotensinaaldosterona serem candidatos naturais ao desenvolvimento e progressão da hipertensão arterial. Também na nossa população os polimorfismos da alfa-aducina (ADD1 G460W rs4961), associaram-se à hipertensão arterial. Nesta população portuguesa, conhecida por ter elevado consumo de sal, faz sentido que estes polimorfismos, sejam relevantes na gestão do sal e da água e consequentemente, no aparecimento de hipertensão arterial. A variante genética GNB3 C825T rs5443 que interfere na sinalização intracelular também constituiu uma forte candidata à hipertensão arterial. Com a elaboração da curva ROC e cálculo das AUC inicialmente só com os fatores de risco tradicionais e posteriormente adicionando as variantes ADD1 G460W, GNB3 C825T, ACE I/D e ACE A2350G aos fatores de risco tradicionais, verificámos ter havido um incremento no risco preditivo de hipertensão arterial, relativamente ao existente só com os fatores de risco tradicionais, com significado estatístico (p = 0,018). Isto sugere que a hipertensão arterial é uma doença multifatorial, que resulta da interação de fatores ambientais, genéticos e estilos de vida que interagem entre si e levam ao aparecimento desta importante patologia. Conclusão: No nosso estudo os polimorfismos associados à hipertensão, estão ligados ao eixo renina-angiotensina-aldosterona (ACE I/D, ACE A2350G), bem como à gestão de sal e água (ADD1 G460W, GNB3 C825T). Através de uma análise multivariada, concluiuse que estas duas últimas variantes genéticas conjuntamente com quatro dos fatores tradicionais (tabagismo, hábitos alcoólicos, obesidade e diabetes) se associam de forma significativa e independente à hipertensão arterial essencial. Num modelo preditivo de hipertensão arterial, a introdução das variantes genéticas aumenta ligeiramente o valor preditivo do modelo. Introduction: Arterial hypertension is a complex, multifactorial disease, controlled by genetic and environmental factors. Objective: Evaluate the genetic susceptibility for developing arterial hypertension and its association with the traditional risk factors in the outbreak of this pathology. Material and Methods: Case-control study with 1712 individuals, mean age of 51.0 ± 7.9 years (860 hypertensive patients and 852 controls). Biochemical and traditional risk factors, and genetic variants were evaluated: ACE I/D rs4340, ACE A2350G rs4343, AGT T174M rs4762, AGT M235T rs699 AGTR1 A1166C rs5186, CYP11B2 -344 C/T rs1799998, ADRB1 R389G rs1801253, ADRB2 R16G rs1042713, ADD1 G460W rs4961, SCNN1G G173A rs5718, GNB3 C825T rs5443, ATP2B1 A/G rs2681472, CYP17A1 T/C rs11191548, SLC4A2 C/T rs2303934. The risk of each gene for hypertension was estimated by the dominant, recessive, co-dominant and multiplicative models. By logistic regression, variables associated with hypertension were evaluated. ROC curves were first performed with traditional risk factors and then adding the genetic variants associated with hypertension. Data were analyzed by SPSS for Windows 19.0 and MedCalc v. 13.3.3.0. Results: The genetic variants ADD1 G460W, GNB3 C825T, ACE I/D, ACE A2350G were associated with hypertension. ROC curve with traditional risk factors and these variants showed an increase in the predictive capacity of hypertension (p = 0.018). Discussion: According to the results of our study, the genetic variants found to be associated with hypertension were: ACE I/D rs4340, ACE A2350G rs4343, ADD1 G460W rs4961 and GNB3 C825T rs5443. The first two variants are associated with hypertension by interfering with the renin-angiotensin-aldosterone system, which plays an important role in regulating blood pressure. It should be noted that genes encoding the components of renin-angiotensin-aldosterone system are natural candidates for the development and progression of hypertension. In our population alpha-aducin polymorphism (ADD1 G460W rs4961) was also associated with hypertension. In a Portuguese population, known to have high salt intake, it makes sense that this polymorphism which is relevant in salt and water management may consequently be relevant in the onset of hypertension. The genetic variant GNB3 C825T rs5443 that affects intracellular signalling was also found to be a strong risk candidate for hypertension. Initially, with the elaboration of the ROC curve and calculation of the AUC using only with traditional risk factors and later by adding the variants ADD1 G460W, GNB3 C825T, ACE I/D and ACE A2350G to the traditional risk factors, we verified that genetic polymorphisms increased the predictive risk of hypertension, when compared to the risk given only by traditional risk factors, with statistical significance (p = 0.018). This suggests that hypertension is a multifactorial disease that results from the interaction of environmental, genetic and lifestyle factors that interact with each other and lead to the advent of this important pathology. Conclusion: In our study, the hypertension-associated polymorphisms are linked to the renin-angiotensin-aldosterone axis (ACE I/D, ACE A2350G), as well as to salt and water management (ADD1 G460W, GNB3 C825T). Through a multivariate analysis, it was concluded that these two last genetic variants together with four of the traditional risk factors (smoking, alcohol consumption, obesity and diabetes) are associated in a significant and independent way with essential hypertension. In a predictive model of hypertension, the introduction of genetic variants slightly increases the predictive value of the model. info:eu-repo/semantics/publishedVersion

The utility of genetic risk scores (GRS) as independent risk predictors remains inconclusive. Here, we evaluate the additive value of a multi-locus GRS to the Framingham risk score (FRS) in coronary artery disease (CAD) risk prediction. A total of 2888 individuals (1566 coronary patients and 1322 controls) were divided into three subgroups according to FRS. Multiplicative GRS was determined for 32 genetic variants associated to CAD. Logistic Regression and Area Under the Curve (AUC) were determined first, using the TRF for each FRS subgroup, and secondly, adding GRS. Different models (TRF, TRF+GRS) were used to classify the subjects into risk categories for the FRS 10-year predicted risk. The improvement offered by GRS was expressed as Net Reclassification Index and Integrated Discrimination Improvement. Multivariate analysis showed that GRS was an independent predictor for CAD (OR = 1.87; p

Essential hypertension (EH) is a complex disease in which physiological, environmental, and genetic factors are involved in its genesis. The genetic variant of the alpha-adducin gene (ADD1) has been described as a risk factor for EH, but with controversial results.The objective of this study was to evaluate the association of ADD1 (Gly460Trp) gene polymorphism with the EH risk in a population from Madeira Island.A case-control study with 1614 individuals of Caucasian origin was performed, including 817 individuals with EH and 797 controls. Cases and controls were matched for sex and age, by frequency-matching method. All participants collected blood for biochemical and genotypic analysis for the Gly460Trp polymorphism. We further investigated which variables were independently associated to EH, and, consequently, analyzed their interactions.In our study, we found a significant association between the ADD1 gene polymorphism and EH (odds ratio 2.484, P = .01). This association remained statistically significant after the multivariate analysis (odds ratio 2.548, P = .02).The ADD1 Gly460Trp gene polymorphism is significantly and independently associated with EH risk in our population. The knowledge of genetic polymorphisms associated with EH is of paramount importance because it leads to a better understanding of the etiology and pathophysiology of this pathology. info:eu-repo/semantics/publishedVersion

Several genetic risk scores (GRS) have been associated with cardiovascular disease; their role, however, in survival from proven coronary artery disease (CAD) have yielded conflicting results. Objective: The objective of this study was to evaluate long- term cardiovascular mortality according to the genetic risk score in a Southern European population with CAD. Methods: A cohort of 1464 CAD patients with angiographic proven CAD were followed up prospectively for up to 58.3 (interquartile range: 25.8- 88.1) months. Genotyping of 32 single- nucleotide polymorphisms previously associated with CAD was performed using oligonucleotides probes marked with fluorescence for each allele. GRS was constructed according to the additive model assuming codominance and categorised using the median (=26). Cox Regression analysis was performed to determine independent multivariate predictors of cardiovascular mortality. Kaplan- Meier survival curves compared high vs low GRS using log- rank test. C- index was done for our population, as a measure of discrimination in survival analysis model. Results: During a mean follow- up of 58.3 months, 156 patients (10.7%) died, 107 (7.3%) of CV causes. High GRS (≥26) was associated with reduced cardiovascular survival. Survival analysis with Cox regression model adjusted for 8 variables showed that high GRS, dyslipidemia, diabetes and 3- vessel disease were independent risk factors for cardiovascular mortality (HR=1.53, P=.037; HR=3.64, P=.012; HR=1.75, P=.004; HR=2.97, P

Funding: This study was supported by the European Regional Development Fund’s Operational Programme for the Enhancement of Economic Potential and Territorial Cohesion for the Autonomous Region of Madeira (INTERVIR ). Recent genetic studies have revealed an association between polymorphisms at the ADAMTS7 gene locus and coronary artery disease (CAD) risk. Functional studies have shown that a CAD-associated polymorphism (rs3825807) affects ADAMTS7 maturation and vascular smooth muscular cell (VSMC) migration. Here, we tested whether ADAMTS7 (A/G) SNP is associated with cardiovascular (CV) survival in patients with established CAD. A cohort of 1,128 patients with angiographic proven CAD, who were followed up prospectively for a mean follow- up period of 63 (range 6-182) mo, were genotyped for rs3825807 A/G. Survival statistics (Cox regression) compared heterozygous (AG) and wild-type (AA) with the reference homozygous GG. Kaplan- Meier (K-M) survival curves were performed according to ADAMTS7 genotypes for CV mortality. Results showed that 47.3% of patients were heterozygous (AG), 36.5% were homozygous for the wild-type allele (AA) and only 16.2% were homozygous for the GG genotype. During the follow-up period, 109 (9.7%) patients died, 77 (6.8%) of CV causes. Survival analysis showed that AA genotype was an independent risk factor for CV mortality compared with reference genotype GG (HR = 2.7, P = 0.025). At the end of follow-up, the estimated survival probability (K-M) was 89.8% for GG genotype, 82.2% for AG and 72.3% for AA genotype (P = 0.039). Carriage of the mutant G allele of the ADAMTS7 gene was associated with improved CV survival in patients with documented CAD. The native overfunctional ADAMTS7 allele (A) may accelerate VSMC migration and lead to neointimal thickening, atherosclerosis progression and acute plaque events. ADAMTS7 gene should be further explored in CAD for risk prediction, mechanistic and therapeutic goals. publishersversion published

Coronary artery disease (CAD) is the main cause of mortality in developed countries. Increased lipid peroxidation is associated with accelerated progression of atherosclerosis. Paraoxonase (PON1) is an antioxidant enzyme bound to high-density lipoprotein (HDL), which protects against lipid peroxidation and coronary artery disease. PON1 activity is under genetic control and its molecular basis is a polymorphism in the PON1 gene that shows two common isoforms: the wild Q form (192 Gln) with high ability to protect LDL from lipid peroxidation in vitro, and the mutated R (Arg) form with lower ability.To explore the interaction of the R allele of the paraoxonase gene and low HDL-cholesterol concentrations in CAD risk.The study population consisted of 818 individuals, 298 coronary patients, aged 55.0 +/- 10.3 years, 78.9% male, and 520 age and gender matched healthy controls, aged 53.3 +/- 11.7 years, 72.5% male. Low HDL-cholesterol was defined as0.90 mmol/l in men and1.11 mmol/l in women. Comparisons of genotypes between cases and controls were performed by a chi-square test. Statistical significance was accepted at p0.05. Odds ratios and 95% confidence intervals for the RR genotypes and HDL-deficient subjects were computed using univariate analysis (2 x 2 tables). To determine the interaction between the RR paraoxonase genotype and HDL-deficient subjects, we used 4 x 2 epidemiologic tables and synergy measures: the additive model (Rothman's synergy index, SI) and multiplicative model (Khoury's synergy index, SIM). The relative excess risk due to interaction (RERI) and the attributable proportion (AP) due to interaction (Rothman) were calculated.The PON1 RR192 polymorphism was associated with coronary heart disease (OR = 1.61; p = 0.043) in the whole population. HDL-deficient subjects with the RR192 genotype showed increased risk for CAD (OR = 17.38; p0.0001) compared to those with normal HDL and RR192 (OR = 1.39; p = 0.348) and HDL-deficient subjects not carrying the RR genotype (OR = 7.79; p0.0001). Synergy measures were SI = 2.3, SIM = 1.6; RERI = 9.2.These data suggest the existence of a synergistic effect of the PON1 RR192 genotype (with lower antioxidant ability) and HDL-deficient subjects in risk for development of CAD. The AP due to this interaction was 0.53, meaning that 53% of CAD was explained by this interaction.

Dissertação de Mestrado em Gestão de Ecossistemas A presente tese teve como objectivo o estudo da biologia e comportamento da Gralha-de-bico-vermelho (Pyrrhocorax pyrrhocorax) no Noroeste de Portugal, mais propriamente na serra do Barroso e do Alvão e em Pitões das Júnias. Foi dado ênfase à conservação da espécie recorrendo à análise do comportamento e sucesso reprodutor, à determinação de habitat potencial para a espécie, à sua dieta alimentar e à disponibilidade das presas avaliada pela colocação de pitfalls em habitats favoráveis e não favoráveis e à recuperação e monitorização de potenciais novos abrigos. Em simultâneo foi desenvolvida uma linha de interligação entre o estudo da biologia da espécie e a aplicação de novas tecnologias, através da realização de captura e transmissão de imagens em tempo real, no dormitório principal na serra do Barroso. Neste trabalho foi também construído um modelo estocástico-dinâmico, relacionado com a gestão de habitat da espécie, utilizando variáveis ambientais seleccionadas através de tratamento estatístico, com influência sobre a observação de indivíduos nos vários locais amostrados. Ao longo de um ano (de Setembro de 2007 a Setembro de 2008), foi avaliada a constituição da população, nos três núcleos em estudo, tendo-se registado uma elevada variação anual, no Barroso e em Pitões das Júnias e não tendo sido observado nenhum indivíduo, no Alvão. O habitat preferencial da espécie é composto por afloramentos rochosos graníticos associados a vegetação rasteira, áreas queimadas e lameiros. A análise às presas recolhidas nos locais de alimentação utilizados revelou maior abundância dos artrópodes: dominados pelos Heminoptera (Barroso) e pelos Araneae e Coleoptera (Alvão). A aplicação do sistema de recolha de imagens permitiu adquirir um maior conhecimento da etologia da espécie e da utilização sazonal de dormitórios. Os resultados da simulação do modelo estocástico-dinâmico são encorajadores, uma vez que parecem demonstrar a estabilidade do modelo em capturar a dinâmica da ocorrência da espécie, prevendo o padrão comportamental da sua abundância e das características do habitat principal. Em geral, este estudo representa uma contribuição útil para a avaliação do estado da espécie, fornecendo um ponto de partida interessante para desenvolver estudos mais globais, no âmbito da sua conservação, que permitam obter uma noção mais realista da distribuição, do sucesso reprodutor e do comportamento desta espécie, nesta região. The present thesis aims to study biology and behavior of the red-billed Chough (Pyrrhocorax pyrrhocorax), developed in the northwest of Portugal, more specifically in the Serra do Alvão and Barroso and Pitões das Júnias. An emphasis was given the scope of the species conservation by using the analysis of behavior and reproductive success, determination of potential habitat for the species, the respective diet and prey availability, assessed by placing pitfalls in favorable and non-favorable habitats and recovery and monitoring of potential new shelters. At the same time a line linking was developed between the biological study of the species and the application of new technologies, by digital capture and transmits images in real time, in the principal roost placed in Barroso region. In this work there was also built a dynamic-stochastic model, related to the management of specie habitat, using environmental variables selected by statistic treatment with influence on the observation of individuals in different sample places. Over a year (September 2007 to September 2008), the constitution of the population was evaluated, in the three different groups that varied throughout the year in the Barroso and in Pitões das Júnias and there no individuals observed in Alvão. The habitat used by species is mainly characterized by the presence of granitic rock outcrops associated with low vegetation, burned areas and grassland. The analysis of prey samples collected in feeding sites used by the red-billed Chough, revealed the abundance of the active arthropods, dominated by the Hymenoptera (Barroso) and by the Araneae and Coleoptera (Alvão). The application of image collection system was a powerful instrument for recording real-time processing of images captured, which allowed gain greater knowledge of ethology of specie and seasonal use of roosts. The StDM simulation results are encouraging since they seem to demonstrate the model reliability in capturing the dynamics of the species occurrence by predicting the behavioural pattern of their abundance and habitat main characteristics. Overall, this study seems to represent a useful contribution for the species status assessment, providing a useful starting point to develop more global studies, in the scope of their conservation wich order to be able to obtain a more realistic notion of the distribution, reproductive success and behavior of the red-billed Chough in the northwest of Portugal.

Gender is a well-known risk factor for mortality and morbidity after coronary artery bypass grafting and various reasons have been proposed to explain the poorer results observed in women. The study objective was to determine whether female gender was still an operative risk factor with the adoption of off-pump coronary artery bypass surgery.Of 2123 consecutive patients who underwent isolated CABG between November 2002 and December 2007, 1966 (92.6%) (481 women and 1485 men) were operated without cardiopulmonary bypass and form the study population. Women were older (69.0 vs. 64.7 years; p = 0.001) and had more severe angina (CCS class 3.1 vs. 2.7; p = 0.001), smaller body surface area (1.6 vs. 1.8 m2; p = 0.001), higher body mass index and greater incidence of diabetes (48.3% vs. 34.2%; p = 0.005) than men. There were no statistically significant differences in the incidence of three-vessel disease (69.5% vs. 71.9%) or left main disease (22.2% vs. 27.0%) but ejection fraction was higher in women (61.3% vs. 59.0%; p = 0.01). Risk factors for in-hospital mortality were identified by univariate analysis and logistic regression.The number of distal anastomoses was lower in women than in men (2.8 vs. 3.0; p = 0.001). In-hospital mortality was 2.0% in women and 0.8% in men (p = 0.01) and female gender was one of the eight risk factors for in-hospital mortality identified by univariate analysis. Using a logistic regression model, only age (OR: 1.7; 95% CI 1.01-1.14; p = 0.02) and logistic EuroSCORE (OR: 1.07; 95% CI 1.03-1.10; p0.001) were independently associated with in-hospital mortality. Logistic EuroSCORE was the only independent risk factor for major morbidity (OR: 1.07 95% CI 1.04-1.10; p0.001).Women who undergo coronary artery bypass grafting have an unfavorable clinical profile compared to men, resulting in higher unadjusted in-hospital mortality. However, in an unselected patient population undergoing off-pump CABG, female gender was not found to be an independent risk factor for mortality or major morbidity.

Various studies have compared coronary artery disease (CAD) patients with controls in order to determine which polymorphisms are associated with a higher risk of disease. The results have often been contradictory. Moreover, these studies evaluated polymorphisms in isolation and not in association, which is the way they occur in nature.Our purpose was to evaluate the risk of CAD in patients with associated polymorphisms in the same gene or in differen genes.We evaluated the risk associated with ACE DD, ACE 8 CC, ACT 174MM, AGT 235TT, MTHFR 677TT, MTHFR 1298AA, PON1 192RR and PON1 55MM in 298 CAD patients and 298 healthy individuals. We then evaluated the risk of associated polymorphisms in the same gene (ACE DD + ACE 8GG; AGT 174MM + AGT 235TT; MTHFR 677TT + MTHFR 1298AA). Finally, for the isolated polymorphisms which were significant, we evaluated the risk of polymorphism associations at different functional levels (ACE + AGT; ACE + MTHFR; ACE + PON1). Multiple logistic regression was used to identify independent risk factors for CAD.Isolated polymorphisms including ACE DD(p0.0001), ACE 8 gg (p=0.023), and MTHFR 1298AA (p = 0.049) presented with a significantly higher frequency in the CAD group. An association of polymorphisms in the same gene did not have an additive or synergistic effect, nor did it increase the risk of CAD. Polymorphic associations in different genes increased the risk of CAD, compared with the isolated polymorphisms. The association of ACE DD or ACE 8 GG with PON1 192RR increased the risk of CA fourfold (1.8 to 4.2). After logistic regression analysis, current smoking, family history, fibrinogen, diabetes, and the ACE DD or ACE 8 GG + MTHFR 1298AA and ACE DD or ACE 8 GG + PON1 192RR associations remained in the, model and proved to be independent predictors of CAD.The association of polymorphisms in the same gene did not increase the risk of the isolated polymorphism. The association of polymorphisms in genes belonging to different enzyme systems was always linked to increased risk compared to the isolated polymorphisms. This study may contribute to a better understanding of overall genetic risk for CAD rather than that associated with each polymorphism in isolation.

Complex diseases such as coronary artery disease (CAD), hypertension and diabetes are usually caused by individual susceptibility to multiple genes, environmental factors, and the interaction between them. The paraoxonase 1 (PON1) enzyme has been implicated in the pathogenesis of atherosclerosis and CAD. Two common polymorphisms in the coding region of the PON1 gene, which lead to a glutamine (Q)/arginine (R) substitution at position 192 and a leucine (L)/methionine (M) substitution at position 55, influence PON1 activity. Studies have investigated the association between these polymorphisms and CAD, but with conflicting results.1) To evaluate the association between PON1 polymorphisms and CAD risk; and 2) to study the interaction between PON1 polymorphisms and others in different candidate genes.We evaluated the risk of CAD associated with PON1 Q192R and L55M polymorphisms in 298 CAD patients and 298 healthy individuals. We then evaluated the risk associated with the interaction of the PON1 polymorphisms with ACE DD, ACE 8 GG and MTHFR 1298AA. Finally, using a logistic regression model, we evaluated which variables (genetic, biochemical and environmental) were linked significantly and independently with CAD.We found that the PON1 55MM genotype was more common in the CAD population, but this did not reach statistical significance as a risk factor for CAD, while PON1 192RR presented an 80% higher relative risk compared to the population without this polymorphism. The interaction between PON1 192RR and MTHFR 1298AA, sited in different genes, increased the risk for CAD, compared with the polymorphisms in isolation (OR=2.76; 95% CI=1.20-6.47; p=0.009), as did the association of PON1 192RR with ACE DD, which presented a 337% higher risk compared to the population without this polymorphic association (OR=4.37; 95% CI=1.47-13.87; p=0.002). Similarly, the association between PON1 192RR and ACE 8 GG was linked to an even higher risk (OR=6.23; 95% CI=1.67-27.37; p0.001). After logistic regression, smoking, family history, fibrinogen, diabetes, Lp(a) and the association of PON1 192RR + ACE 8 GG remained in the regression model and proved to be significant and independent risk factors for CAD. In the regression model the latter association had OR=14.113; p=0.018.When analyzed separately, the PON1 192RR genotype presented a relative risk for CAD 80% higher than in the population without this genotype. Its association with other genetic polymorphisms sited in different genes, coding for different enzymes and belonging to different physiological systems, always increased the risk for CAD. After correction for other conventional and biochemical risk factors, the PON1 192RR + ACE 8 GG association remained a significant and independent risk factor for CAD.