Supplementation with selenium and coenzyme Q10 in an elderly Swedish population low in selenium - positive effects on thyroid hormones, cardiovascular mortality, and quality of life.

BACKGROUND: Selenium-dependent deiodinases play a central role in thyroid hormone regulation and metabolism. In many European countries, insufficient selenium intake may consequently lead to adverse effects on thyroid function. In this randomised placebo-controlled double-blind study, we examined the effect of supplementation with selenium and coenzyme Q10 on thyroid hormonal status, cardiovascular (CV) mortality and health-related quality of life (Hr-QoL). METHODS: Free T3, free T4, reverse T3, and TSH were determined in 414 individuals at baseline, and the effect of selenium yeast (200 µg/day) and coenzyme Q10 (200 mg/day) supplementation on hormone concentrations, CV mortality and Hr-QoL was evaluated after 48 months using Short Form 36 (SF-36). Pre-intervention plasma selenium was low, mean 67 µg/L, corresponding to an estimated intake of 35 µg/day. Changes in concentrations of thyroid hormones following the intervention were assessed using T-tests, repeated measures of variance, and ANCOVA analyses. RESULTS: In the total population, the group with the lowest selenium concentration at baseline presented with significantly higher levels of TSH and lower levels of fT3 as compared to subjects with the highest selenium concentration. Supplementation with selenium and coenzyme Q10 for 4 years significantly increased fT3 and rT3, decreased fT4, and diminished the increase in TSH levels compared with placebo treatment (p = 0.03, all). In the placebo group, TSH and fT4 values above the median were associated with an increase in 10-year CV mortality, as compared with the mortality rate among those with TSH and fT4 below the median (p < 0.04, both), with no difference in mortality rate according to TSH and fT4 levels in the active intervention group. Similarly, TSH > median and fT3 < median were associated with a decline in mental Hr-QoL measures vs. TSH < and fT3 > median in the placebo group during 4 years of follow-up, but this was wiped out in the active group. CONCLUSIONS: Supplementation with selenium and coenzyme Q10 had a beneficial effect on thyroid hormones with respect to CV mortality and Hr-QoL outcomes. The initial deficient selenium status was associated with an impaired thyroid function and the changes in thyroid hormone levels can be explained by increased activity of deiodinases. We conclude that a substantial part of the elderly study population might suffer from suboptimal thyroidal function with adverse clinical implications due to selenium deficiency. TRIAL REGISTRATION: This study was registered at ClinicalTrials.gov and has the identifier NCT01443780. Since it was not mandatory to register at the time the study began, the study has been registered retrospectively.

Effects of an Intervention with Selenium and Coenzyme Q10 on Five Selected Age-Related Biomarkers in Elderly Swedes Low in Selenium: Results That Point to an Anti-Ageing Effect-A Sub-Analysis of a Previous Prospective Double-Blind Placebo-Controlled Randomised Clinical Trial.

Background: Ageing is associated with cardiovascular disease (CVD). As no single biomarker reflects the full ageing process, we aimed to investigate five CVD- and age-related markers and the effects of selenium and coenzyme Q10 intervention to elucidate the mechanisms that may influence the course of ageing. Methods: This is a sub-study of a previous prospective double-blind placebo-controlled randomized clinical trial that included 441 subjects low in selenium (mean age 77, 49% women). The active treatment group (n = 220) received 200 µg/day of selenium and 200 mg/day of coenzyme Q10, combined. Blood samples were collected at inclusion and after 48 months for measurements of the intercellular adhesion molecule (ICAM-1), adiponectin, leptin, stem cell factor (SCF) and osteoprotegerin (OPG), using ELISAs. Repeated measures of variance and ANCOVA evaluations were used to compare the two groups. In order to better understand and reduce the complexity of the relationship between the biomarkers and age, factor analyses and structural equation modelling (SEM) were performed, and a structural model is presented. Results: Correlation analyses of biomarker values at inclusion in relation to age, and relevant markers related to inflammation, endothelial dysfunction and fibrosis, demonstrated the biomarkers' association with these pathological processes; however, only ICAM1 and adiponectin were directly correlated with age. SEM analyses showed, however, that the biomarkers ICAM-1, adiponectin, SCF and OPG, but not leptin, all had significant associations with age and formed two independent structural factors, both significantly related to age. While no difference was observed at inclusion, the biomarkers were differently changed in the active treatment and placebo groups (decreasing and increasing levels, respectively) at 48 months (p ≤ 0.02 in all, adjusted), and in the SEM model, they showed an anti-ageing impact. Conclusions: Supplementation with selenium/Q10 influenced the analysed biomarkers in ways indicating an anti-ageing effect, and by applying SEM methodology, the interrelationships between two independent structural factors and age were validated.

The NLRP3 Genetic Variant (rs10754555) Reduces the Risk of Adverse Outcome in Middle-Aged Patients with Chronic Coronary Syndrome.

Background: Inflammation is central in development of cardiovascular disease (CVD). Aberrant function of the Nod-Like Receptor Protein 3 (NLRP3) inflammasome, a central mediator in the proinflammatory response, has been associated with atherosclerosis. The influence of genetic determinants on this inflammatory pathway and its downstream effects is less known. We aimed to investigate the frequency of a single NLRP3 gene variant according to clinical outcome in CVD and its influence on NLRP3-related markers. Methods: In this observational study, we included 1001 patients with chronic coronary syndrome. Blood samples were drawn at inclusion, including whole-blood and PAXgene tubes for DNA and RNA isolation, respectively. Allelic discrimination of the NLRP3 single nucleotide polymorphism rs10754555 was performed; and gene expression of NLRP3, Toll-Like Receptor 4, Interleukin- (IL-) 1ß, and IL-18 was relatively quantified, both methods by RT-PCR. Circulating IL-6, high-sensitivity (hs) C-reactive protein, IL-18, and IL-12 were measured by enzyme-like immunosorbent assays. Clinical endpoints during 2 years (n = 106) were a composite of unstable angina pectoris, myocardial infarction, nonhemorrhagic stroke, and death. Results: Minor allele frequency of the NLRP3 variant was 0.36. In all, no association of the NLRP3 variant with clinical subgroups or outcome was found, neither any significant influence on the genes' mRNA expression or circulating protein. However, in subjects < 56 years (25 percentile), the variant G-allele is associated with significant lower risk of suffering a composite event (OR = 0.43 (95% CI 0.19, 0.97), p = 0.043, adjusted). In the same age group, the NLRP3 gene was accordingly downregulated in G-allele carriers vs. noncarriers, and circulating IL12 was significantly reduced (p < 0.05, both). In subjects > 56 years, no significant effect of the variant was observed. Conclusion: The age-related reduced risk of composite endpoint in rs10754555 G-allele carriers accompanied by diminished NLRP3 mRNA expression is hypothesis generating and needs to be further explored. The study is registered at http://www.clinicaltrials.gov, with identification number NCT00222261.

Genetic Variation in ADAMTS13 is Related to VWF Levels, Atrial Fibrillation and Cerebral Ischemic Events.

INTRODUCTION: ADAMTS13 cleaves von Willebrand factor (VWF) multimers into less active fragments. Both markers have been related to cardiovascular disease (CVD). We aimed to investigate the influence of ADAMTS13 single nucleotide polymorphisms (SNPs) on levels of ADAMTS13 and VWF, and CVD. METHODS: The c.1342C>G, g.41635A>G and c.2699C>T polymorphisms were determined in patients with chronic coronary syndrome (n = 1000). VWF and ADAMTS13 were analyzed. Clinical endpoints after 2 years (n = 106) were unstable angina pectoris, myocardial infarction, non-hemorrhagic stroke and death. RESULTS: The SNPs did not affect ADAMTS13 levels. The 41635A-allele associated with higher VWF levels (P < .001). Patients with the 1342G-allele had significantly higher frequency of previous atrial fibrillation (n = 26, P = .016) and cerebral ischemic events (n = 47, P = .030). Heterozygous of the 1342CG variant experienced more clinical endpoints compared to homozygous (CC and GG) (P = .028). CONCLUSION: The association between the 41635A-allele and VWF indicates a role for this polymorphism in VWF regulation. ADAMTS13 has previously been linked to atrial fibrillation and ischemic stroke, and our findings suggest that the 1342G-allele may be of significance. The association between the 1342CG genotype and endpoints needs further investigations.Clinicaltrials.gov, ASCET, NCT00222261. https://clinicaltrials.gov/ct2/show/NCT00222261?term=NCT00222261&draw=2&rank=1.

Changes in circulating sirtuin 1 after bariatric surgery.

BACKGROUND AND AIMS: Obesity is associated with chronic inflammation and oxidative stress. Weight loss after bariatric surgery improves the inflammatory state and risk of cardiovascular disease. Improvement in metabolic dysfunction might be associated with changes in the activity of sirtuin 1 (SIRT1) and we aimed to investigate the effect of bariatric surgery on its circulating levels. METHODS AND RESULTS: This is a sub-study of a prospective cohort study, including 110 subjects with morbid obesity. The surgical procedure was either laparoscopic Roux-en-Y gastric bypass (RYGB) or sleeve gastrectomy (SG). Blood was sampled at inclusion and six and 12 months after surgery. SIRT1 was measured in EDTA plasma with an enzyme-linked immunosorbent assay. The mean age in the population was 43 years, 80% were women and mean body mass index (BMI) was 38.8 kg/m2. RYGB and SG were performed in 89 and 21 subjects, respectively. SIRT1 concentration was significantly reduced from baseline to six and 12 months after surgery, with mean values (SD) 156.8 (82.6), 119.5 (65.6) and 94.9 (45.6) ng/mL, respectively, (p ≤ 0.002, all), accompanied by significant reductions in C-reactive protein (CRP), BMI and triglycerides from inclusion (p < 0.001, all). Type of surgery did not differently modify SIRT1 levels (p = 0.09). CRP and triglycerides were both positively predictive of SIRT1 levels (p ≤ 0.001, both). CONCLUSION: SIRT1 concentration was significantly lower six and 12 months after bariatric surgery. CRP and triglycerides independently predicted SIRT1 levels, suggesting that reduction in SIRT1 levels might not intrinsically be related to weight reduction, but to improvement in metaflammation.

TERT and TET2 Genetic Variants Affect Leukocyte Telomere Length and Clinical Outcome in Coronary Artery Disease Patients-A Possible Link to Clonal Hematopoiesis.

Inherited and acquired mutations in hematopoietic stem cells can cause clonal expansion with increased risk of cardiovascular disease (CVD), a condition known for the clonal hematopoiesis of indeterminate potential (CHIP). Inherited genetic variants in two CHIP-associated genome loci, the telomerase gene telomerase enzyme reverse transcriptase (TERT) (rs7705526) and the epigenetic regulator ten−eleven translocation 2 (TET2) (rs2454206), were investigated in 1001 patients with stable coronary artery disease (CAD) (mean age 62 years, 22% women), with regards to cardiovascular outcome, comorbidities, and leukocyte telomere length. Over 2 years, mutated TERT increased the risk two-fold for major clinical events (MACEs) in all patients (p = 0.004), acute myocardial infarction (AMI) in male patients (p = 0.011), and stroke in female patients (p < 0.001). Mutated TET2 correlated with type 2 diabetes (p < 0.001), the metabolic syndrome (p = 0.002), as well as fasting glucose, HbA1c, and shorter telomeres (p = 0.032, p = 0.003, and p = 0.016, respectively). In conclusion, our results from stable CAD patients highlight TERTs' role in CVD, and underline TET2s' role in the epigenetic regulation of lifestyle-related diseases.

Abdominal Adipose Tissue Associates With Adiponectin and TNFα in Middle-Aged Healthy Men.

Introduction: Adipokines are highly active biopeptides involved in glucose metabolism, insulin regulation and the development and progression of obesity and its associated diseases. It includes, among others, adiponectin, visfatin and tumor necrosis factor alpha (TNFα). The sources of adipokines and their associations with glucometabolic variables are not completely understood. Aim: In this cross-sectional study, we aimed to investigate whether gene expression levels in subcutaneous adipose tissue (SAT) of selected adipokines and their corresponding circulating levels associate with the amount of AT in superficial (sSAT), deep (dSAT) and visceral AT (VAT), assessed by computed tomography (CT). Any association with glucometabolic variables were also explored. Methods: In 103 healthy Caucasian men, aged 39.5 years, fasting venous blood and SAT samples from the gluteal region were collected. Ninety-four of the participants underwent CT assessment of the abdominal AT, which was divided into VAT, sSAT and dSAT. Circulating levels of adipokines were measured by ELISA and AT gene-expression by PCR. Insulin sensitivity was determined by glucose clamp, assessing glucose disposal rate (GDR). Results: Circulating adiponectin and TNFα gene expression correlated inversely and positively to the amount of AT in all three compartments (r=-0.266 to -0.276, p<0.05 for all) and (r=0.323 - 0.368, p<0.05 for all), respectively, with strongest correlations to the amount in sSAT and dSAT. When dividing AT compartments into quartiles, a tendency was observed towards lower circulating adiponectin and higher TNFα gene expression levels, respectively, with increasing amount of sSAT and dSAT. Circulating adiponectin correlated inversely to insulin, C-peptide and waist circumference (r=-456 to -0.373, p<0.001) and positively to GDR (r=0.356, p<0.001). AT-expressed visfatin correlated inversely to insulin and C-peptide (r=-0.370 and r=-0.404, p<0.001). Conclusion: Increased amount of AT is associated with lower levels of adiponectin and increased levels of TNFα AT expression.

Complement activation in association with clinical outcomes in ST-elevation myocardial infarction.

Introduction: The complement system and neutrophil extracellular traps (NETs) might contribute to ischemia-reperfusion injury in ST-elevation myocardial infarction (STEMI). We aimed to estimate associations between complement activation and NETs in STEMI, and their prognostic value on clinical endpoints. Methods: In this cohort study, 864 patients admitted for PCI during STEMI were included. Complement activation was analyzed by the terminal complement complex (TCC), while NETs were analyzed by myeloperoxidase-DNA, citrullinated histone 3 (CitH3) and dsDNA. The composite endpoint was reinfarction, unscheduled revascularization, stroke, hospitalization due to heart failure, or death, and the secondary endpoint was total mortality. The association between TCC and clinical endpoints was assessed by Cox regression and ROC curve analysis. Results: TCC was weakly correlated to dsDNA (r = 0.127, p < 0.001) and CitH3 (r = 0.102, p = 0.003). After a median follow-up time of 4.6 years, 184 (21.3 %) patients had reached a clinical endpoint. TCC was not associated with the composite endpoint, but with total mortality (HR: 1.673, 95 % CI: [1.014, 2.761], p = 0.044). The significant association was lost when adjusting for CRP, NT-proBNP, LVEF and time from symptoms to PCI. In ROC curve analysis of total mortality, the AUC for TCC alone was 0.549 (95 % CI: [0.472, 0.625]), AUC for dsDNA alone was 0.653 (95 % CI: [0.579, 0.720]), while AUC for TCC and dsDNA combined was 0.660 (95 % CI: [0.590, 0.730]). Conclusions: In this STEMI cohort, TCC was not associated with the composite endpoint, but somewhat with total mortality. Combining TCC and dsDNA did not increase the prognostic value compared to dsDNA alone.

Adiponectin in relation to exercise and physical performance in patients with type 2 diabetes and coronary artery disease.

INTRODUCTION: Adipokines, expressed by adipose tissue (AT), have been associated with metabolic disturbances and coronary artery disease (CAD). The impact of exercise training on the AT in patients suffering from both diabetes and CAD is unknown. To gain knowledge on changes in ATs' inflammatory profile in such a population, we investigated the effects of long-term exercise on selected adipokines and their associations with physical performance and glucometabolic variables. Adiponectin was selected based on its anti-atherogenic and anti-diabetic properties and visfatin and tumour necrosis factor (TNF) for their association with atherosclerosis and metabolic disorders. Not many studies have focused on the effects of long-term exercise training on adipokines in patients with concomitant T2DM and CAD. METHODS: Patients with type 2 diabetes and CAD (n = 137), 41-81 years, 17.2% females, were randomized in a 1:1 manner to an exercise group, who underwent 1 year of 150 min weekly combined strength and endurance exercise, or a control group. AT from the gluteal region and blood samples were obtained at baseline and after 12 months, along with a physical performance test, assessed by the VO2 peak. Circulating protein levels were measured by ELISA. RNA was extracted from AT and expression levels were relatively quantified by PCR. RESULTS: After 1 year, no significant difference in the change in the investigated markers between the intervention group and the control group was observed. Changes in circulating adiponectin and VO2 peak correlated in the total population (r = 0.256, p = 0.008). At baseline, circulating adiponectin and TNF correlated inversely with insulin and with C-peptide and VO2peak, respectively (p < 0.001, all). CONCLUSION: In this population with concomitant diabetes and CAD, ATs' inflammatory profile remained unchanged apparently after 1 year of exercise intervention. Changes in the VO2peak were nevertheless, related to changes in circulating adiponectin levels. TRIAL REGISTRATION: http://www.clinicaltrials.gov NCT01232608.

Impact of Selenium on Biomarkers and Clinical Aspects Related to Ageing. A Review.

Selenium (Se) is an essential dietary trace element that plays an important role in the prevention of inflammation, cardiovascular diseases, infections, and cancer. Selenoproteins contain selenocysteine in the active center and include, i.a., the enzymes thioredoxin reductases (TXNRD1-3), glutathione peroxidases (GPX1-4 and GPX6) and methionine sulfoxide reductase, involved in immune functions, metabolic homeostasis, and antioxidant defense. Ageing is an inevitable process, which, i.a., involves an imbalance between antioxidative defense and reactive oxygen species (ROS), changes in protein and mitochondrial renewal, telomere attrition, cellular senescence, epigenetic alterations, and stem cell exhaustion. These conditions are associated with mild to moderate inflammation, which always accompanies the process of ageing and age-related diseases. In older individuals, Se, by being a component in protective enzymes, operates by decreasing ROS-mediated inflammation, removing misfolded proteins, decreasing DNA damage, and promoting telomere length. Se-dependent GPX1-4 and TXNRD1-3 directly suppress oxidative stress. Selenoprotein H in the cell nucleus protects DNA, and selenoproteins residing in the endoplasmic reticulum (ER) assist in the removal of misfolded proteins and protection against ER stress. In this review, we highlight the role of adequate Se status for human ageing and prevention of age-related diseases, and further its proposed role in preservation of telomere length in middle-aged and elderly individuals.

Effect of intermittent and continuous caloric restriction on Sirtuin1 concentration depends on sex and body mass index.

BACKGROUND & AIMS: The favorable effect of caloric restriction (CR) on health span is well known and partly mediated by the sirtuin system. Sirtuin1, a regulator of energy homeostasis in response to nutrient availability, is activated by CR. We therefore investigated effects of two different CR regimens on Sirtuin1 concentrations. METHODS & RESULTS: The study included 112 abdominally obese subjects, randomized to intermittent or continuous CR for 1 year. Blood samples and anthropometric measures were collected at baseline and after 12 months. Sirtuin1 concentrations were measured by ELISA. Sirtuin1 correlated significantly to BMI at baseline (r = .232, p = 0.019). Mean reduction in body-weight was 8.0 and 9.0 kg after intermittent and continuous CR, respectively. After 1 year, no significant between-group differences in Sirtuin1 levels were observed according to regimen (p = 0.98) and sex (p = 0.41). An increase in median Sirtuin1 concentrations (pg/mL) [25, 75 percentiles] from baseline was observed after intermittent CR in the total population (884 [624, 1285] vs.762 [530, 1135]; p = 0.041), most marked in men (820 [623, 1250] vs. 633 [524, 926]; p = 0.016). Improvement in BMI after 1 year correlated to Sirtuin1 changes, but varied according to sex. In women, Spearman's rho = .298, p = 0.034, with stronger correlation in the intermittent CR group (r = .424, p = 0.049). In men, there was an inverse relation to Sirtuin1 changes, only in the intermittent CR group (r = -.396, p = 0.045). CONCLUSIONS: Effects on Sirtuin1 concentrations after 1 year of CR are sex and BMI-related. Intermittent CR regimen affected Sirtuin1 to a stronger extent than continuous CR, suggesting individualized dietary intervention.

Biomarkers of ageing and cardiac remodeling are associated with atrial fibrillation.

Objectives. Ageing is one of the strongest risk factors for atrial fibrillation (AF), and additional risk factors are also closely related to ageing. Remodeling is part of the pathophysiology of AF, and a possible common denominator of ageing and other AF risk factors. The aim of this study was to investigate any association between the presence of AF and the ageing biomarkers, leukocyte telomere length (LTL) and sirtuin-1 (SIRT-1), and the cardiac remodeling biomarkers Galectin-3 and sST2 in elderly myocardial infarction (MI) patients. Design. Patients were included after admission for MI. Diagnosis of AF was retrieved from medical records and classified as either history of AF before MI or new onset from admission to study inclusion. SIRT-1, sST2 and Galectin-3 were analyzed by ELISAs and LTL by qPCR. Results. In total, 299 patients were included, median age 75 years, 70.2% male. A history of AF was recorded in 38 patients and 30 patients experienced new onset AF. Higher levels of SIRT-1 were associated with lower risk of having a history of AF (OR = 0.46 (95% CI 0.26, 0.81), p = 0.007), whereas higher sST2 levels were associated with higher risk of AF (OR = 4.13 (95% CI 1.69, 10.13), p = 0.002). Results remained significant after adjustment for other AF risk factors. No significant associations with AF were found for Galectin-3 or LTL. None of the biomarkers associated with new onset AF. Conclusion. In elderly patients with MI, higher ST2 and lower SIRT-2 levels were associated with higher prevalence of AF, possibly reflecting both ageing and the remodeling phenomena in AF. Clinical trials registration: ClinicalTrials.gov (NCT01841944).

NETosis in Long-Term Type 1 Diabetes Mellitus and Its Link to Coronary Artery Disease.

Background: Neutrophil extracellular traps NETs have been linked to glucose and the pathogenesis of type 1 diabetes mellitus (T1DM). NETs also play a role in vascular inflammation and the development of coronary artery disease (CAD). The role of NETs in CAD progression in patients with long-term T1DM is unclear. We aimed to 1) investigate whether levels of circulating NETs markers were elevated in long-term T1DM subjects compared to controls, and 2) explore whether levels of NETs were related to the presence of CAD. Material and Methods: 102 patients with > 45 years of T1DM and 75 age-matched controls were enrolled in a cross-sectional study. Median age was 62 years. Computed tomography coronary angiography (CTCA) was performed in 148 subjects without established coronary heart disease. For the current study, CAD was defined as a coronary artery stenosis >50%. Double-stranded deoxyribonucleic acid (dsDNA) was measured by a nucleic acid stain, myeloperoxidase-DNA (MPO-DNA), citrullinated histone 3 (H3Cit) and peptidylarginine deiminase 4 (PAD4) by ELISAs, while gene expression of PAD4 was measured in leukocytes from PAXgene tubes. Results: Circulating MPO-DNA levels were significantly lower in patients with T1DM than in controls (0.17 vs 0.29 OD, p<0.001), while dsDNA, H3Cit, PAD4 and gene expression of PAD4 did not differ with respect to the presence of T1DM. There were no significant associations between NETs markers and HbA1c in the T1DM group. None of the NETs markers differed according to the presence of CAD in patients with T1DM. While all circulating NETs markers correlated significantly with circulating neutrophils in the control group (r=0.292-393, p<0.014), only H3Cit and PAD4 correlated with neutrophils in the T1DM group (r= 0.330-0.449, p ≤ 0.001). Conclusions: In this cross-sectional study of patients with long-term T1DM and age-matched controls, circulating NETs levels were not consistently associated with the presence of T1DM or glycemic status, and did not differ according to the presence of CAD in patients with T1DM. Our results entail the possibility of altered neutrophil function and reduced NETosis in T1DM. This warrants further investigation.

Complement Activation in Association with Markers of Neutrophil Extracellular Traps and Acute Myocardial Infarction in Stable Coronary Artery Disease.

Complement activation and neutrophil extracellular traps (NETs) have both been suggested to drive atherosclerotic plaque progression. Although experimental studies suggest interplay between these two innate immunity components, the relevance in patients with coronary artery disease (CAD) is unclear. The aim of this study was to assess associations between complement activation and NETs in patients with stable CAD and examine the role of complement activation on clinical outcome. Blood samples from a cohort of patients with angiographically verified stable CAD (n = 1001) were analyzed by ELISA for the terminal complement complex (TCC) and by relative quantification for gene expression of the C5a receptor 1 (C5aR1) as markers of complement activation. As markers of NETs, dsDNA was analyzed by fluorescent nucleic acid stain and myeloperoxidase-DNA (MPO-DNA) by ELISA. Clinical outcome was defined as unstable angina, nonhemorrhagic stroke, acute myocardial infarction (MI), or death (n = 106, whereof 36 MI). Levels of TCC and C5aR1 were not significantly correlated to dsDNA (TCC: r = -0.045, p = 0.153; C5aR1: r = -0.060, p = 0.434) or MPO-DNA (TCC: r = 0.026, p = 0.414; C5aR1: r = 0.123, p = 0.107). When dividing TCC and C5aR1 levels into quartiles (Q), levels of MPO-DNA differed significantly across quartiles (TCC: p = 0.008, C5aR1: 0.049), while dsDNA did not (TCC: p = 0.181, C5aR1: p = 0.771). Patients with TCC levels in Q4 had significantly higher levels of MPO-DNA than Q1-3 (p = 0.019), and C5aR1 levels in Q3-4 had significantly higher levels of MPO-DNA than Q1-2 (p = 0.046). TCC levels did not differ between patients experiencing a clinical endpoint or not, but high levels were associated with increased risk of acute MI (OR. 1.97, 95% CI: 0.99-3.90, p = 0.053) during two-year follow up, also when adjusted for relevant covariates. In conclusion, TCC and C5aR1 were moderately associated with the NET marker MPO-DNA, and TCC levels were related to the risk of future MI in this cohort of patients with stable CAD.

Leukocyte telomere length and serum polyunsaturated fatty acids, dietary habits, cardiovascular risk factors and features of myocardial infarction in elderly patients.

BACKGROUND: Telomeres are non-coding sequences at the end of eukaryote chromosomes, which in complex with associated proteins serve to protect subtelomeric DNA. Telomeres shorten with each cell division, are regarded as a biomarker for aging and have also been suggested to play a role in atherosclerosis and cardiovascular disease (CVD). The aim of the present study was to explore the associations between leukocyte telomere length and serum polyunsaturated fatty acids, diet, cardiovascular risk factors and features of myocardial infarction (MI) in elderly patients. METHODS: The material is based upon the first 299 included patients in the OMEMI trial, where patients aged 70-82 years of age are randomized to receive omega-3 supplements or corn oil (placebo) after MI. Patients were included 2-8 weeks after the index MI. DNA was extracted from whole blood, and leukocyte telomere length (LTL) was analyzed by qPCR and reported as a number relative to a reference gene. Serum long chain polyunsaturated fatty acid (LCPUFA) content was analyzed by gas chromatography. Diet was evaluated with the validated SmartDiet food frequency questionnaire. Medical records, patient interviews and clinical examination provided previous medical history and anthropometric data. Non-parametric statistical tests were used. RESULTS: Median (25, 75 percentile) LTL was 0.55 (0.42, 0.72). Patients had a median age of 75 years, 70.2% were male and 45.2% used omega-3 supplements. There was a weak, but significant correlation between LTL and linoleic acid (r = 0.139, p = 0.017), but not with other LCPUFAs. There was a trend towards longer telomeres with a healthier diet, but this did not reach statistical significance (p = 0.073). No associations were found between LTL and CVD risk factors or features of MI. CONCLUSIONS: In our population of elderly with a recent myocardial infarction LTL was associated with linoleic acid concentrations, but not with other LCPUFAs. Patients with a healthy diet tended to have longer telomeres. The limited associations may be due to age and the narrow age-span in our population. Further studies, designed to detect longitudinal changes should be performed to explore the role of telomeres in cardiovascular aging. TRIAL REGISTRATION: Clinical trials no. NCT01841944, registration date April 29, 2013.

Neutrophil Extracellular Trap Components Associate with Infarct Size, Ventricular Function, and Clinical Outcome in STEMI.

BACKGROUND: The relevance of neutrophil extracellular traps (NETs) in acute ST-elevation myocardial infarction (STEMI) is unclear. We explored the temporal profile of circulating NET markers and their associations to myocardial injury and function and to adverse clinical events in STEMI patients. METHODS AND RESULTS: In 259 patients, blood samples were drawn before and after PCI, on day 1, and after 4 months. Double-stranded deoxyribonucleic acid (dsDNA) and myeloperoxidase-DNA (MPO-DNA) were measured in serum by a nucleic acid stain and ELISA. Cardiac magnetic resonance imaging assessed microvascular obstruction (MVO), area at risk, infarct size, myocardial salvage index, left ventricular ejection fraction (LVEF), and change in indexed left ventricular end-diastolic volume (LVEDVi). Clinical events were registered after 12 months. dsDNA and MPO-DNA levels were highest before PCI, with reduced levels thereafter (all p ≤ 0.02). Patients with high vs. low day 1 dsDNA levels (>median; 366 ng/ml) more frequently had MVO, larger area at risk, larger infarct size acutely and after 4 months, and lower myocardial salvage index (all p < 0.03). Moreover, they had lower LVEF acutely and after 4 months, and larger change in LVEDVi (all p ≤ 0.014). High day 1 dsDNA levels also associated with risk of having a large infarct size (>75th percentile) and low LVEF (≤49%) after 4 months when adjusted for gender, time from symptoms to PCI, and infarct localization (OR 2.3 and 3.0, both p < 0.021), and patients with high day 1 dsDNA levels were more likely to experience an adverse clinical event, also when adjusting for peak troponin T (hazard ratio 5.1, p = 0.012). No such observations were encountered for MPO-DNA. CONCLUSIONS: High day 1 dsDNA levels after STEMI were associated with myocardial infarct size, adverse left ventricular remodeling, and clinical outcome. Although the origin of dsDNA could be discussed, these observations indicate a potential role for dsDNA in acute myocardial ischemia. This trial is registered with S-08421d, 2008/10614 (Regional Committee for Medical Research Ethics in South-East Norway (2008)).

Effects of exercise training on inflammasome-related mediators and their associations to glucometabolic variables in patients with combined coronary artery disease and type 2 diabetes mellitus: Sub-study of a randomized control trial.

BACKGROUND: Adipose tissue produces pro-inflammatory mediators involved in the atherosclerotic process. We investigated whether 12-month exercise training in patients with type 2 diabetes mellitus and coronary artery disease would reduce circulating levels and genetic expression of mediators in the interleukin-18, Caspase-1 and NLR pyrin domain containing 3 pathways. Correlations to glucometabolic variables; fasting glucose, HbA1c, duration of diabetes, insulin, C-peptide, insulin resistance (measured by homeostatic model assessment indexes - insulin resistance) and body mass index at baseline were further assessed. METHODS: 137 patients (aged 41-81 years, 17.2% female participants) were included and randomized to a 12-month exercise programme or to a control group. Fasting blood and adipose tissue samples were taken at inclusion and after 12 months. RESULTS: No statistically significant difference in changes of any variable between the intervention and the control group was found. At baseline, a positive correlation between insulin and homeostatic model assessment indexes - insulin resistance, interleukin-18 expression in adipose tissue and an inverse correlation between some glucometabolic variables and leukocyte expression of NLR pyrin domain containing 3 and Caspase-1 were observed. CONCLUSION: No significant effects of long-term exercise training were observed on the inflammasome-related mediators in our patients with combined coronary artery disease and type 2 diabetes mellitus. The observed correlations may indicate a pro-inflammatory state in adipose tissue by overweight and a compensatory downregulation of these mediators in circulating leucocytes.

Novel biomolecules of ageing, sex differences and potential underlying mechanisms of telomere shortening in coronary artery disease.

Telomere length (TL), growth differentiate factor (GDF)11, insulin growth factor (IGF)1, sirtuin (SIRT)1 and inflammatory processes have been related to ageing and age-related diseases, like coronary artery disease (CAD). We aimed to investigate the associations between leukocyte TLs (LTLs), chronological age, sex and comorbidities in CAD patients. Any covariations between LTL, GDF11, IGF1, SIRT-1 and pro-inflammatory cytokines were further assessed. METHODS: In 300 patients with stable CAD (age 36-81 years, 20% females), DNA and RNA were isolated from whole blood for PCR analysis and relative quantification of LTLs and gene-expression of GDF11, IGF1,SIRT1, IL-12, IL-18 and IFNÆ´, respectively. Serum was prepared for the analyses of circulating IL-18, IL-12, IL-6 and TNFα. RESULTS: Patients with previous myocardial infarction (MI) presented with 20% shorter LTLs vs. patients without (p = 0.019) indicating LTLs to be of importance for CAD severity. The observation however, was only observed in men (p = 0.009, n = 115), in which the upper LTL quartile associated with 64% lower frequency of previous MI compared to quartile 1-3 (p = 0.005, adjusted). LTLs were not differently distributed according to sex or comorbidities such as hypertension, diabetes type 2 and metabolic syndrome. LTLs and GDF11 were inversely correlated to age (r = -0.17; p = 0.007 and r = -0.16; p = 0.010, respectively), however, separated in gender, LTL only in women (r = -0.37) and GDF11 only in men (r = -0.19) (p = 0.006, both). GDF11 and SIRT1 were strongly inter-correlated (r = 0.56, p ≤ 0.001), suggesting common upstream regulators. LTLs were moderately correlated to GDF11 and SIRT1 in overweight women (BMI ≥ 25 kg/m2) (r = 0.41; p = 0.027 and 0.43; p = 0.020, respectively), which may reflect common life-style influences on LTLs and these markers. In all women, we observed further that the highest LTL quartile associated with higher GDF11 and SIRT expression and lower circulating levels of IL-12, IL-18 and TNFα, as compared to quartile 1, which may indicate lifestyle influences on female LTLs. In men, the highest LTL quartile associated with lower IFNÆ´ expression and lower circulating TNFα. Overall, the results indicate an association between chronic low-grade inflammation and LTLs. CONCLUSIONS: Shorter LTLs in CAD patients with previously suffered MI may indicate telomere attrition as part of its pathophysiology in men. The inverse association between LTLs and age exclusively in women underpins the previously reported decline in attrition rate in men with increasing age. As elevated GDF11 and SIRT1 along with attenuated pro-inflammatory cytokines seem to positively affect LTL in women, we hypothesize a potential sex-dimorphism in LTL regulation, which may implicate sex- adjusted health-preventive therapies.

Effects of exercise training on markers of adipose tissue remodeling in patients with coronary artery disease and type 2 diabetes mellitus: sub study of the randomized controlled EXCADI trial.

BACKGROUND: Investigate effects of long-term exercise on the remodeling markers MMP-9, TIMP-1, EMMPRIN and Galectin-3 in combined type 2 diabetes mellitus (T2DM) and coronary artery disease (CAD) patients. Any associations between these biomarkers and glucometabolic variables were further assessed at baseline. METHODS: 137 patients (age 41-81 years, 17.2% females) were included and randomized to a 12-months exercise program or to a control group. Fasting blood samples and subcutaneous adipose tissue (AT) samples were taken at inclusion and after 12-months. The intervention was a combination of aerobic and strength training for a minimum of 150 min per week. Circulating protein levels were measured by ELISA methods and RNA was extracted from AT and circulating leukocytes. Expression levels were relatively quantified by PCR. RESULTS: After 12 months of intervention, both AT-expression and circulating levels of EMMPRIN were increased in the exercise group (p < 0.05, both) with significant difference in change between the two groups (p < 0.05 both). No significant effect was observed on MMP-9, TIMP-1 and Galectin-3. Levels of TIMP-1 (AT-expression and circulating) were significantly correlated to insulin, and HOMA2- after Bonferroni correction (p = 0.001, by 48 performed correlations). CONCLUSION: The increase in levels of EMMPRIN after long-term exercise training, might indicate some degree of AT remodeling in these patients after 12-months of exercise, whether beneficial or not. The remodeling markers were to some extent associated with glucometabolic variables in our population with the combined disease.Trial registration clinicaltrials.gov, NCT01232608. Registered 2 November 2010.

The MMP-9 -1562 C/T polymorphism in the presence of metabolic syndrome increases the risk of clinical events in patients with coronary artery disease.

BACKGROUND AND OBJECTIVES: Elevated levels of matrix metalloproteinase (MMP)-9 have been associated with the metabolic syndrome (MetS) and cardiovascular events. The MMP-9 -1562 C/T polymorphism has furthermore been shown as a risk factor for coronary artery disease (CAD). The non-favourable cardiometabolic state in MetS may increase the risk. We aimed to investigate the influence of MMP-9 -1562 C/T polymorphism in subjects with CAD and MetS. METHODS: Patients (n = 1000) with verified CAD stratified in Mets +/- (n = 244/756), were analyzed for the MMP-9 -1562 C/T polymorphism and related to clinical events after 2 years follow-up. Serum levels of total MMP-9 and tissue inhibitor of matrix metalloproteinases (TIMP)-1 were analyzed in all, whereas MMP-9 activity, extracellular matrix metalloproteinase inducer (EMMPRIN), and expression of the two genes were analyzed in a subset of 240 randomly selected patients. RESULTS: Totally, 106 clinical endpoints were recorded. In MetS; the T-allele associated with 5.5 fold increase in event rate (p<0.0001), increased with number of MetS components, a 117% increase in total MMP-9 levels (TT homozygous, p = 0.05), significantly higher total- and endogenous active MMP-9 and TIMP-1 levels (p<0.01 all), and EMMPRIN was inversely correlated with pro- and endogenous active MMP-9 (p<0.05, both). In non-MetS; the T-allele was not associated with new events, nor higher MMP-9 levels. EMMPRIN was significantly correlated with total MMP-9 and TIMP-1 (p<0.01, both) and the two genes were inter-correlated (p<0.001). CONCLUSION: In CAD patients with MetS, the MMP-9 T-allele increased the risk of clinical events, probably mediated through elevated MMP-9 levels and altered MMP-9 regulation.