Isolation and molecular characterization of circulating extracellular vesicles from blood of chronic Chagas disease patients.

Extracellular vesicles (EVs) are lipid bilayer envelopes that encase several types of molecules. Their contents mostly reflect their cell origin and possible targets at other locations in the organism and can be modified in pathological conditions to interfere with intercellular communication, thus promoting disease establishment and development. These characteristics, in addition to their presence in virtually all body fluids, make such vesicles ideal for biomarker discovery in human diseases. Here, we describe the effect of different anticoagulants and the combination of two purification methods for isolation and characterization of circulating EVs from blood of chronic Chagas disease (CCD) patients. We illustrated this procedure by studying a population of patients with Chagas disease at the indeterminate chronic stage, in which the Trypanosoma cruzi is very scarce in circulation. EVs were harvested from blood collected without or with different anticoagulants. Protein and nanoparticle tracking analysis was used to measure EVs size and concentration. The EVs were purified by ultracentrifugation, followed by size-exclusion chromatography and characterized by chemiluminescent enzyme-linked immunosorbent assay and dot blot using antibodies that recognized parasite-derived EVs, such as hyperimmune sera, polyclonal and monoclonal antibodies against trans-sialidase and mucins. In parallel, antibodies against classical human EV markers CD9, CD63, CD81, and CD82, were also analyzed. The results showed that anticoagulants did not interfere with the analyzed parameters and circulating EVs from CCD patients contain T. cruzi antigens and classical human exosomal markers. Overall, our protocol is adequate for the isolation of the total circulating EVs and can serve as an important basis for further studies on biomarker discovery in Chagas' disease.

Rare Pathogenic Variants in Mitochondrial and Inflammation-Associated Genes May Lead to Inflammatory Cardiomyopathy in Chagas Disease.

Cardiomyopathies are an important cause of heart failure and sudden cardiac death. Little is known about the role of rare genetic variants in inflammatory cardiomyopathy. Chronic Chagas disease cardiomyopathy (CCC) is an inflammatory cardiomyopathy prevalent in Latin America, developing in 30% of the 6 million patients chronically infected by the protozoan Trypanosoma cruzi, while 60% remain free of heart disease (asymptomatic (ASY)). The cytokine interferon-γ and mitochondrial dysfunction are known to play a major pathogenetic role. Chagas disease provides a unique model to probe for genetic variants involved in inflammatory cardiomyopathy. METHODS: We used whole exome sequencing to study nuclear families containing multiple cases of Chagas disease. We searched for rare pathogenic variants shared by all family members with CCC but absent in infected ASY siblings and in unrelated ASY. RESULTS: We identified heterozygous, pathogenic variants linked to CCC in all tested families on 22 distinct genes, from which 20 were mitochondrial or inflammation-related - most of the latter involved in proinflammatory cytokine production. Significantly, incubation with IFN-γ on a human cardiomyocyte line treated with an inhibitor of dihydroorotate dehydrogenase brequinar (enzyme showing a loss-of-function variant in one family) markedly reduced mitochondrial membrane potential (ΔψM), indicating mitochondrial dysfunction. CONCLUSION: Mitochondrial dysfunction and inflammation may be genetically determined in CCC, driven by rare genetic variants. We hypothesize that CCC-linked genetic variants increase mitochondrial susceptibility to IFN-γ-induced damage in the myocardium, leading to the cardiomyopathy phenotype in Chagas disease. This mechanism may also be operative in other inflammatory cardiomyopathies.

Effect of exercise training on cardiovascular autonomic and muscular function in subclinical Chagas cardiomyopathy: a randomized controlled trial.

PURPOSE: Patients with chronic chagasic cardiomyopathy with preserved ventricular function present with autonomic imbalance. This study evaluated the effects of exercise training (ET) in restoring peripheral and cardiac autonomic control and skeletal muscle phenotype in patients with subclinical chronic chagasic cardiomyopathy. METHODS: This controlled trial (NCT02295215) included 24 chronic chagasic cardiomyopathy patients who were randomized www.random.org/lists/ into two groups: those who underwent exercise training (n = 12) and those who continued their usual activities (n = 12). Eight patients completed the exercise training protocol, and 10 patients were clinically followed up for 4 months. Muscular sympathetic nerve activity was measured by microneurography and muscle blood flow (MBF) using venous occlusion plethysmography. The low-frequency component of heart rate variability in normalized units (LFnuHR) reflects sympathetic activity in the heart, and the low-frequency component of systolic blood pressure variability in normalized units reflects sympathetic activity in the vessels. The infusion of vasoactive drugs (phenylephrine and sodium nitroprusside) was used to evaluate cardiac baroreflex sensitivity, and a vastus lateralis muscle biopsy was performed to evaluate atrogin-1 and MuRF-1 gene expression. RESULTS: The baroreflex sensitivity for increases (p = 0.002) and decreases (p = 0.02) in systolic blood pressure increased in the ET group. Muscle blood flow also increased only in the ET group (p = 0.004). Only the ET group had reduced resting muscular sympathetic nerve activity levels (p = 0.008) and sympathetic activity in the heart (LFnu; p = 0.004) and vessels (p = 0.04) after 4 months. Regarding skeletal muscle, after 4 months, participants in the exercise training group presented with lower atrogin-1 gene expression than participants who continued their activities as usual (p = 0.001). The reduction in muscular sympathetic nerve activity was positively associated with reduced atrogin-1 (r = 0.86; p = 0.02) and MuRF-1 gene expression (r = 0.64; p = 0.06); it was negatively associated with improved baroreflex sensitivity both for increases (r = -0.72; p = 0.020) and decreases (r = -0.82; p = 0.001) in blood pressure. CONCLUSIONS: ET improved cardiac and peripheral autonomic function in patients with subclinical chagasic cardiomyopathy. ET reduced MSNA and sympathetic activity in the heart and vessels and increased cardiac parasympathetic tone and baroreflex sensitivity. Regarding peripheral muscle, after 4 months, patients who underwent exercise training had an increased cross-sectional area of type I fibers and oxidative metabolism of muscle fibers, and decreased atrogin-1 gene expression, compared to participants who continued their activities as usual. In addition, the reduction in MSNA was associated with improved cardiac baroreflex sensitivity, reduced sympathetic cardiovascular tone, and reduced atrogin-1 and MuRF-1 gene expression. TRIAL REGISTRATION: ID: NCT02295215. Registered in June 2013.

Impact of Aortic Valve Function and the Need for Aortic Valve Repair on Long-Term Outcomes of Valve-Sparing Aortic Root Replacement: 13-Year Experience of David Operation.

BACKGROUND: The best approach for aortic root disease remains controversial. Composite valve-graft conduit (CVG) replacement offers good results at short-term and long-term follow-up; on the other hand, valve-sparing aortic root replacement (VSARR) has proven to be an excellent treatment alternative. This study aimed to analyse the outcomes after VSARR and compare whether preoperative moderate or severe aortic regurgitation (AR) and or the need for aortic valve repair (AVR) during this procedure influenced survival and freedom from reoperation rates. METHODS: From September 2005 to June 2018, 104 patients underwent VSARR using the reimplantation technique: 64% presented with preoperative moderate or severe AR, concomitant AVR was performed in 43.3%, Marfan syndrome was present in 16.3%, and 12.5% had a bicuspid aortic valve. Complete follow-up was obtained in 91% of the sample, echocardiographic results were available for 86% and the mean follow-up time was 1,893 days. RESULTS: In-hospital mortality was 2.9% and one death occurred 42 days after hospital discharge. In the latest echocardiographic assessment, 88.3% presented with mild AR or better. Freedom from reoperation at 8 years was 95.4%. There was no case of endocarditis and one patient had a stroke 2 years after the operation. There were no between-group differences in morbidity, mortality and complications during the follow-up. CONCLUSION: VSARR can be performed with low mortality rates and reasonable durability of the aortic valve. Neither moderate or severe AR nor the need for aortic valve repair during the procedure altered survival and freedom from reoperation.

The amount of late gadolinium enhancement outperforms current guideline-recommended criteria in the identification of patients with hypertrophic cardiomyopathy at risk of sudden cardiac death.

BACKGROUND: Identifying the patients with hypertrophic cardiomyopathy (HCM) in whom the risk of sudden cardiac death (SCD) justifies the implantation of a cardioverter-defibrillator (ICD) in primary prevention remains challenging. Different risk stratification and criteria are used by the European and American guidelines in this setting. We sought to evaluate the role of cardiovascular magnetic resonance (CMR) late gadolinium enhancement (LGE) in improving these risk stratification strategies. METHODS: We conducted a multicentric retrospective analysis of HCM patients who underwent CMR for diagnostic confirmation and/or risk stratification. Eligibility for ICD was assessed according to the HCM Risk-SCD score and the American College of Cardiology Foundation/American Heart Association (ACCF/AHA) algorithm. The amount of LGE was quantified (LGE%) and categorized as 0%, 0.1-10%, 10.1-19.9% and ≥ 20%. The primary endpoint was a composite of SCD, aborted SCD, sustained ventricular tachycardia (VT), or appropriate ICD discharge. RESULTS: A total of 493 patients were available for analysis (58% male, median age 46 years). LGE was present in 79% of patients, with a median LGE% of 2.9% (IQR 0.4-8.4%). The concordance between risk assessment by the HCM Risk-SCD, ACCF/AHA and LGE was relatively weak. During a median follow-up of 3.4 years (IQR 1.5-6.8 years), 23 patients experienced an event (12 SCDs, 6 appropriate ICD discharges and 5 sustained VTs). The amount of LGE was the only independent predictor of outcome (adjusted HR: 1.08; 95% CI: 1.04-1.12; p <  0.001) after adjustment for the HCM Risk-SCD and ACCF/AHA criteria. The amount of LGE showed greater discriminative power (C-statistic 0.84; 95% CI: 0.76-0.91) than the ACCF/AHA (C-statistic 0.61; 95% CI: 0.49-0.72; p for comparison < 0.001) and the HCM Risk-SCD (C-statistic 0.68; 95% CI: 0.59-0.78; p for comparison = 0.006). LGE was able to increase the discriminative power of the ACCF/AHA and HCM Risk-SCD criteria, with net reclassification improvements of 0.36 (p = 0.021) and 0.43 (p = 0.011), respectively. CONCLUSIONS: The amount of LGE seems to outperform the HCM Risk-SCD score and the ACCF/AHA algorithm in the identification of HCM patients at increased risk of SCD and reclassifies a relevant proportion of patients.

Dysregulation of insulin levels in Chagas heart disease is associated with altered adipocytokine levels.

Metabolic, inflammatory, and autonomic nervous system (ANS) dysfunction are present in patients with heart failure. However, whether these changes are due to left ventricular dysfunction or heart failure etiology is unknown. We evaluated metabolism and inflammatory activity in patients with idiopathic dilated cardiomyopathy (IDC) and Chagas cardiomyopathy (CHG) and their correlation with the ANS. Forty-six patients were divided into 3 groups: IDC, CHG, and control. We evaluated adiponectin, leptin, insulin, interleukin-6, and tumor necrosis factor-alpha. ANS were analyzed by heart rate variability in time and frequency domains on a 24-hour Holter monitor. Levels of glucose, cholesterol, leptin, and adiponectin did not show differences between groups. Insulin levels were lower in CHG group (5.4 ± 3.3 µU/mL) when compared with control (8.0 ± 4.9 µU/mL) and IDC (9.9 ± 5.0 µU/mL) groups (p = 0.007). Insulin was positively associated with LFr/HFr ratio (r = 0.562; p = 0.029) and with the LFr component (r = 0.562; p = 0.029) and negatively associated with adiponectin (r = -0.603; p = 0.017) in CHG group. The addition of an adiponectin unit reduced average insulin by 0.332 µg/mL. Insulin levels were decreased in the CHG group when compared with the IDC group and were associated with ANS indexes and adiponectin levels.

The learning curve effect on outcomes with frozen elephant trunk technique for extensive thoracic aorta disease.

OBJECTIVE: The purpose of this study was to analyze the learning curve effect on hospital mortality, postoperative outcomes, freedom from reintervention in the aorta and long-term survival after frozen elephant trunk (FET) operation. METHODS: From July 2009 to June 2018, 79 patients underwent surgery with the FET technique. They had type A aortic dissection (acute 7.6%, chronic 33%), type B aortic dissection (acute 1.26%, chronic 34.2%), and complex thoracic aortic aneurysm (24%). 27.8% were reoperations and 43% received concomitant cardiac procedures. To compare the results, the sample was divided into group 1 (G1) (first half of the sample - operations from 2009 to 2014) and group 2 (G2) (first half of the sample - operations from 2015 to 2018). RESULTS: The in-hospital mortality was 20.25%, 30.7% for G1 and 10% for G2 (P = .02). The mean cardiopulmonary bypass time, myocardial ischemia time, and selective cerebral perfusion at 25°C time were 154 ± 31, 118 ± 32, and 59 ± 12 minutes, respectively, similar for both groups. Stroke and spinal cord injury occurred in four and two patients, with no difference between groups (P = .61 and P = .24). The necessity for secondary intervention on the downstream aorta for both groups was also similar (P = .136). Five of sixty-three surviving patients died during the follow-up period and the estimated survival rate was different between groups 49% vs 88% (P = .007). CONCLUSION: The learning curve with the FET procedure had a significant impact on hospital mortality and midterm survival over the follow-up period, albeit did not influence the freedom from reintervention on the downstream aorta.

Erythropoietin reduces collagen deposition after myocardial infarction but does not improve cardiac function.

Myocardial remodeling includes inappropriate collagen deposition in the interstitium. Erythropoietin (EPO) may have cardioprotective effects. We aimed to assess the role of EPO on myocardial remodeling during the chronic phase. We studied 60 Wistar rats divided into the following groups: control (CT), control + EPO (CT + EPO), myocardial infarction + EPO (MI + EPO), and myocardial infarction (MI). The interstitial collagen volume fraction (ICVF) was quantified and echocardiography was performed. We quantified asymmetric dimethylarginine and glutathione by ELISA, and used real-time PCR to assess apoptosis and inflammation. Western blotting was used to evaluate inflammatory proteins and tissue inhibitors of metalloproteinases (TIMPs), and TUNEL staining was used to detect apoptosis. For matrix metalloproteinases (MMPs), we performed zymography. Parametric and nonparametric analyses were performed according to normality testing. ICVF was greater in MI groups (p < 0.001) and was attenuated by EPO (p = 0.05). The MMP-2 did not show any difference between groups. The TIMP-1 and TIMP-2 did not have difference between groups. The MI groups had worse fraction shortening (p < 0.001), without EPO protection (p = 0.666). The MI groups had increased left ventricle diastolic dimension (p < 0.001) without EPO attenuation (p = 0.79). EPO did not act on oxidative stress. Apoptosis and inflammation were not modulated by EPO. We concluded that EPO attenuated interstitial collagen accumulation, but did not protect from heart dilation or dysfunction.

Complication of hybrid treatment in type B aortic dissection diagnosed by echocardiography.

This case illustrates an unusual and fatal complication after endovascular treatment of type B aortic dissection and highlights the role of echocardiography in the early diagnosis of complications. In this case, a patient with previous diagnosis of chronic type B aortic dissection and moderate aortic regurgitation underwent endovascular repair of the proximal descending aorta and conservative surgical correction of the aortic valve. On early postoperative, a transesophageal echocardiogram and aortic angiotomography demonstrated proximal endoleak by contrast extravasation around the proximal graft attachment site, causing compression of the stent in its middle portion, resulting in narrowing with reduced cross-sectional area.

Exercise-induced quantitative microvolt T-wave alternans in hypertrophic cardiomyopathy.

BACKGROUND/PURPOSE: Patients with hypertrophic cardiomyopathy (HCM) have elevated risk for sudden cardiac death (SCD). Our study aimed to quantitatively characterize microvolt T-wave alternans (TWA), a potential arrhythmia risk stratification tool, in this HCM patient population. METHODS: TWA was analyzed with the quantitative modified moving average (MMA) in 132 HCM patients undergoing treadmill exercise testing, grouped according to Maron score risk factors as high-risk (H-Risk, n=67,), or low-risk (L-Risk, n=65, without these risk factors). RESULTS: TWA levels were much higher for the H-Risk than for the L-Risk group (101.40±75.61 vs. 54.35±46.26µV; p<0.0001). A 53µV cut point, set by receiver operator characteristic (ROC), identified H-Risk patients (82% sensitivity, 69% specificity). CONCLUSIONS: High TWA levels were found for hypertrophic cardiomyopathy patients. Abnormal TWA associated with major risk factors for SCD: non-sustained ventricular tachycardia on Holter (p=0.001), family history of SCD (p=0.006), septal thickness ≥30mm (p<0.001); and inadequate blood pressure response to effort (p=0.04).

Chagas' heart disease: gender differences in myocardial damage assessed by cardiovascular magnetic resonance.

BACKGROUND: Since a male-related higher cardiovascular morbidity and mortality in patients with Chagas' heart disease has been reported, we aimed to investigate gender differences in myocardial damage assessed by cardiovascular magnetic resonance (CMR). METHODS AND RESULTS: Retrospectively, 62 seropositive Chagas' heart disease patients referred to CMR (1.5 T) and with low probability of having significant coronary artery disease were included in this analysis. Amongst both sexes, there was a strong negative correlation between LV ejection fraction and myocardial fibrosis (male r = 0.64, female r = 0.73, both P < 0.001), with males showing significantly greater myocardial fibrosis (P = 0.002) and lower LV ejection fraction (P < 0.001) than females. After adjustment for potential confounders, gender remained associated with myocardial dysfunction, and 53% of the effect was mediated by myocardial fibrosis (P for mediation = 0.004). Also, the transmural pattern was more prevalent among male patients (23.7 vs. 9.9%, P < 0.001) as well as the myocardial heterogeneity or gray zone (2.2 vs. 1.3 g, P = 0.003). CONCLUSIONS: We observed gender-related differences in myocardial damage assessed by CMR in patients with Chagas' heart disease. As myocardial fibrosis and myocardial dysfunction are associated to cardiovascular outcomes, our findings might help to understand the poorer prognosis observed in males in Chagas' disease.

Functional IL18 polymorphism and susceptibility to Chronic Chagas Disease.

BACKGROUND: Chronic Chagas Disease cardiomyopathy (CCC), a life-threatening inflammatory dilated cardiomyopathy, affects 30% of the approximately 8 million patients infected by Trypanosoma cruzi, the rest of the infected subjects remaining asymptomatic (ASY). The Th1 T cell-rich myocarditis plays a pivotal role in CCC pathogenesis. Local expression of IL-18 in CCC myocardial tissue has recently been described. IL-18 could potentially amplify the process by inducing increased expression of IFN-γ which in turn can increase the production of IL-18, thereby creating a positive feedback mechanism. In order to assess the contribution of the IL-18 to susceptibility to Chronic Chagas Disease, we investigated the association between a single nucleotide polymorphism (SNP) located in the IL-18 gene with the risk of developing Chagas cardiomyopathy. METHODS AND RESULTS: We analyzed the rs2043055 marker in the IL18 gene in a cohort of Chagas disease cardiomyopathy patients (n=849) and asymptomatic subjects (n=202). We found a significant difference in genotype frequencies among moderate and severe CCC patients with ventricular dysfunction. CONCLUSIONS: Our analysis suggests that the IL18 rs2043055 polymorphism- or a SNP in tight linkage disequilibrium with it- may contribute to modulating the Chagas cardiomyopathy outcome.

Myocardial tissue characterization in Chagas' heart disease by cardiovascular magnetic resonance.

BACKGROUND: Chagas' heart disease is an important public health problem in South America. Several aspects of the pathogenesis are not fully understood, especially in its subclinical phases. On pathology Chagas' heart disease is characterized by chronic myocardial inflammation and extensive myocardial fibrosis. The latter has also been demonstrated by late gadolinium enhancement (LGE) by cardiovascular magnetic resonance (CMR). In three clinical phases of this disease, we sought to investigate the presence of LGE, myocardial increase in signal intensity in T2-weighted images (T2W) and in T1-weighted myocardial early gadolinium enhancement (MEGE), previously described CMR surrogates for myocardial fibrosis, myocardial edema and hyperemia, respectively. METHODS: Fifty-four patients were analyzed. Sixteen patients with the indeterminate phase (IND), seventeen patients with the cardiac phase with no left ventricular systolic dysfunction (CPND), and twenty-one patients with the cardiac phase with left ventricular systolic dysfunction (CPD). All patients underwent 1.5 T CMR scan including LGE, T2W and MEGE image sequences to evaluate myocardial abnormalities. RESULTS: Late gadolinium enhancement was present in 72.2 % of all patients, in 12.5 % of IND, 94.1 % of the CPND and 100 % of the CPD patients (p < 0.0001). Myocardial increase in signal intensity in T2-weighted images (T2W) was present in 77.8 % of all patients, in 31.3 % of the IND, 94.1 % of the CPND and 100 % of the CPD patients (p < 0.0001). T1-weighted myocardial early gadolinium enhancement (MEGE) was present in 73.8 % of all patients, in 25.0 % of the IND, 92.3 % of the CPND and 94.1 % of the CPD (p < 0.0001). A good correlation between LGE and T2W was observed (r = 0.72, and p < 0.001). CONCLUSIONS: Increase in T2-weighted (T2W) myocardial signal intensity and T1-weighted myocardial early gadolinium enhancement (MEGE) can be detected by CMR in patients throughout all phases of Chagas' heart disease, including its subclinical presentation (IND). Moreover, those findings were parallel to myocardial fibrosis (LGE) in extent and location and also correlated with the degree of Chagas' heart disease clinical severity. These findings contribute to further the knowledge on pathophysiology of Chagas' heart disease, and might have therapeutic and prognostic usefulness in the future.

Noncompaction cardiomyopathy: a substrate for a thromboembolic event.

BACKGROUND: Noncompaction cardiomyopathy (NCC) is a rare genetic cardiomyopathy characterized by a thin, compacted epicardial layer and an extensive noncompacted endocardial layer. The clinical manifestations of this disease include ventricular arrhythmia, heart failure, and systemic thromboembolism. CASE PRESENTATION: A 43-year-old male was anticoagulated by pulmonary thromboembolism for 1 year when he developed progressive dyspnea. Cardiovascular magnetic resonance imaging showed severe biventricular trabeculation with an ejection fraction of 15%, ratio of maximum noncompacted/compacted diastolic myocardial thickness of 3.2 and the presence of exuberant biventricular apical thrombus. CONCLUSION: Still under discussion is the issue of which patients and when they should be anticoagulated. It is generally recommended to those presenting ventricular systolic dysfunction, antecedent of systemic embolism, presence of cardiac thrombus and atrial fibrillation. In clinical practice the patients with NCC and ventricular dysfunction have been given oral anticoagulation, although there are no clinical trials showing the real safety and benefit of this treatment.

Molecular basis for the improvement in muscle metaboreflex and mechanoreflex control in exercise-trained humans with chronic heart failure.

Previous studies have demonstrated that muscle mechanoreflex and metaboreflex controls are altered in heart failure (HF), which seems to be due to changes in cyclooxygenase (COX) pathway and changes in receptors on afferent neurons, including transient receptor potential vanilloid type-1 (TRPV1) and cannabinoid receptor type-1 (CB1). The purpose of the present study was to test the hypotheses: 1) exercise training (ET) alters the muscle metaboreflex and mechanoreflex control of muscle sympathetic nerve activity (MSNA) in HF patients. 2) The alteration in metaboreflex control is accompanied by increased expression of TRPV1 and CB1 receptors in skeletal muscle. 3) The alteration in mechanoreflex control is accompanied by COX-2 pathway in skeletal muscle. Thirty-four consecutive HF patients with ejection fractions <40% were randomized to untrained (n = 17; 54 ± 2 yr) or exercise-trained (n = 17; 56 ± 2 yr) groups. MSNA was recorded by microneurography. Mechanoreceptors were activated by passive exercise and metaboreceptors by postexercise circulatory arrest (PECA). COX-2 pathway, TRPV1, and CB1 receptors were measured in muscle biopsies. Following ET, resting MSNA was decreased compared with untrained group. During PECA (metaboreflex), MSNA responses were increased, which was accompanied by the expression of TRPV1 and CB1 receptors. During passive exercise (mechanoreflex), MSNA responses were decreased, which was accompanied by decreased expression of COX-2, prostaglandin-E2 receptor-4, and thromboxane-A2 receptor and by decreased in muscle inflammation, as indicated by increased miRNA-146 levels and the stable NF-κB/IκB-α ratio. In conclusion, ET alters muscle metaboreflex and mechanoreflex control of MSNA in HF patients. This alteration with ET is accompanied by alteration in TRPV1 and CB1 expression and COX-2 pathway and inflammation in skeletal muscle.

Clinical predictors of a positive genetic test in hypertrophic cardiomyopathy in the Brazilian population.

BACKGROUND: Hypertrophic cardiomyopathy is a genetic autosomal dominant disease characterized by left ventricular hypertrophy. The molecular diagnosis is important but still expensive. This work aimed to find clinical predictors of a positive genetic test in a Brazilian tertiary centre cohort of index cases with HCM. METHODS: In the study were included patients with HCM clinical diagnosis. For genotype x phenotype comparison we have evaluated echocardiographic, electrocardiographic, and nuclear magnetic resonance measures. All patients answered a questionnaire about familial history of HCM and/or sudden death. ß-myosin heavy chain, myosin binding protein C, and troponin T genes were sequenced for genetic diagnosis. RESULTS: The variables related to a higher probability of a positive genetic test were familial history of HCM, higher mean heart frequency, presence of NSVT and lower age. Probabilities of having a positive molecular genetic test were calculated from the final multivariate logistic regression model and were used to identify those with a higher probability of a positive molecular diagnosis. CONCLUSIONS: We developed an easy and fast screening method that takes into account only clinical data that can help to select the patients with a high probability of positive genetic results from molecular sequencing of Brazilian HCM patients.

Severe acute pancreatitis: a possible role of intramyocardial cytokine production.

CONTEXT: Several mechanisms are involved in the development of the local and systemic response in acute pancreatitis. Cardiovascular system may be affected throughout the clinical course of acute pancreatitis. The aim was to evaluate local myocardial cytokine production, as well as, functional and histological myocardial alterations in severe acute pancreatitis. METHODS: The animals were divided into three groups: Group 1: control; Group 2: sham; Group 3: severe acute pancreatitis. Echocardiographic assessment of cardiac function, serum levels of amylase and cytokines (TNF-α, IL-6 and IL-10), and mRNA expression of TNF-α, IL-6 and TGF-ß were measured. Myocardial tissue alterations were analysed by histological examination. RESULTS: The serum TNF-α and IL-10 levels were significant higher in AP 2h group. The mRNA IL-6 levels from group AP 2h were statistically higher. The mRNA TNF-α level from sham group and AP 2h were statistically lower. Significant changes in the left ventricular diameter were found in AP 2h and AP 12h groups. There were statistical changes for vacuolar degeneration, picnosis and loss of nucleus, and lymphocytes. CONCLUSION: We found cardiac and histological changes compatible with the inflammatory process triggered by SAP with the promotion of local myocardial cytokine production.

Screening of MYH7, MYBPC3, and TNNT2 genes in Brazilian patients with hypertrophic cardiomyopathy.

BACKGROUND: Hypertrophic cardiomyopathy (HC) is the most prevalent genetic cardiac disease caused by a mutation in sarcomeres, Z-disks, or calcium-handling genes and is characterized by unexplained left ventricular hypertrophy. The aim of this study was to determine the genetic profile of Brazilian patients with HC and correlate the genotype with the phenotype. METHODS: We included 268 index patients from São Paulo city and 3 other cities in Brazil and extracted their DNA from whole blood. We amplified the coding sequencing of MYH7, MYBPC3, and TNNT2 genes and sequenced them with an automatic sequencer. RESULTS: We identified causal mutations in 131 patients (48.8%). Seventy-eight (59.5%) were in the MYH7 gene, 50 (38.2%) in the MYBPC3 gene, and 3 (2.3%) in the TNNT2 gene. We identified 69 mutations, 24 not previously described. Patients with an identified mutation were younger at diagnosis and at current age, had a higher mean heart rate and higher nonsustained ventricular tachycardia frequency compared with those without a mutation. Patients with MYH7 gene mutations had a larger left atrium and higher frequency of atrial fibrillation than did patients with MYBPC3 gene mutations. CONCLUSION: The presence of a mutation in one of the genes suggests a worse prognosis. Mutations in the MYH7 gene, rather than in the MYBPC3 gene, were also related to a worse prognosis. This is the first work characterizing HC molecular epidemiology in the Brazilian population for the 3 most important genes.

Exercise-induced decrease in myocardial high-energy phosphate metabolites in patients with Chagas heart disease.

BACKGROUND: The influence of exercise on cardiac metabolic response in patients with Chagas disease is incompletely understood. METHODS AND RESULTS: Changes in cardiac energetic metabolism were investigated in Chagas disease patients before and during isometric handgrip exercise with (31)P magnetic resonance spectroscopy (MRS). Twenty-eight patients (10 with systolic dysfunction: group I; 10 with normal systolic function and electrocardiogram (ECG) abnormalities: group II; and 8 asymptomatic without ECG abnormalities: group III) and 8 healthy control subjects (group C) were evaluated by electrocardiogram, echocardiogram, functional tests for coronary artery disease, and image-selected localized cardiac (31)P-MRS. The myocardial phosphocreatine to [ß-phosphate]adenosine triphosphate ratio (PCr/ß-ATP) was measured at rest and during isometric handgrip exercise. Exercise testing or 99mTc-sestamibi scintigraphy were negative for myocardial ischemia in all individuals. At rest, cardiac PCr/ß-ATP was decreased in all Chagas groups (1.23 ± 0.37) versus group C (1.88 ± 0.08; P < .001) and was lower in group I (0.89 ± 0.24) versus groups II (1.44 ± 0.23) and III (1.40 ± 0.37; P < .001). There was no stress-induced change in cardiac PCr/ß-ATP (1.88 ± 0.08 at rest vs 1.89 ± 0.08 during exercise; P = NS) in group C. Mean cardiac PCr/ß-ATP was 0.89 ± 0.24 and 0.56 ± 0.21 at rest and during exercise, respectively, in group I (37% decrease; P < .001). In group II, PCr/ß-ATP was 1.44 ± 0.23 at rest and 0.97 ± 0.37 during exercise (33% decrease; P < .001). In group III, PCr/ß-ATP was 1.40 ± 0.37 at rest and 0.60 ± 0.19 during exercise (57% decrease; P < .001). CONCLUSIONS: Myocardial high-energy phosphates are reduced at rest in Chagas heart disease patients, and the reduction is greater in patients with left ventricular dysfunction. Regardless of left ventricular function, Chagas patients exhibit an exercise-induced decline in cardiac high-energy phosphates consistent with myocardial ischemia, suggesting the possibility that this metabolic approach may offer a tool to probe new interventions in Chagas disease patients.

Prognostic Evaluation of Microvolt T-Wave Alternans in Hypertrophic Cardiomyopathy: 9-year Clinical Follow-up. / Avaliação Prognóstica da Microalternância da onda T na Cardiomiopatia Hipertrófica em um Seguimento Clínico de 9 anos.

BACKGROUND: Sudden cardiac death (SCD) resulting from ventricular arrhythmia is the main complication of hypertrophic cardiomyopathy (HCM). Microvolt T-wave alternans (MTWA) is associated with the occurrence of ventricular arrhythmias in several heart diseases, but its role in HCM remains uncertain. OBJECTIVE: To evaluate the association of MTWA with the occurrence of SCD or potentially fatal ventricular arrhythmias in HCM patients in a long-term follow-up. METHODS: Patients diagnosed with HCM and NYHA functional class I-II were consecutively selected. At the beginning of the follow-up, the participants performed the MTWA evaluation using the modified moving average during the stress test. The results were classified as altered or normal. The composite endpoint of SCD, ventricular fibrillation, sustained ventricular tachycardia (SVT) or appropriate implantable cardiac defibrillation (ICD) therapy was assessed. The level of significance was set at 5%. RESULTS: A total of 132 patients (mean age of 39.5 ± 12.6 years) were recruited and followed for a mean of 9.5 years. The MTWA test was altered in 74 (56%) participants and normal in 58 (44%). Nine events (6.8%) occurred during the follow-up, with a prevalence of 1.0%/year - six SCDs, two appropriate ICD shocks and one episode of (SVT). Altered MTWA was associated with non-sustained ventricular tachycardia on Holter (p = 0.016), septal thickness ≥30 mm (p < 0.001) and inadequate blood pressure response to effort (p = 0.046). Five patients with altered MTWA (7%) and four patients with normal MTWA (7%) had the primary outcome [OR = 0.85 (95% CI: 0.21 - 3.35, p=0.83)]. Kaplan-Meir event curves showed no differences between normal and altered MTWA. CONCLUSION: Altered MTWA was not associated with the occurrence of SCD or potentially fatal ventricular arrhythmias in HCM patients, and the low rate of these events during long-term follow-up suggests the good prognosis of this heart disease.

FUNDAMENTO: A morte súbita cardíaca (MSC), decorrente de arritmias ventriculares, é a principal complicação da cardiomiopatia hipertrófica (CMH). A microalternância da onda T (MAOT) está associada à ocorrência de arritmias ventriculares em diversas cardiopatias, mas seu papel na CMH permanece incerto. OBJETIVO: Avaliar associação da MAOT com a ocorrência de MSC ou arritmias ventriculares malignas em pacientes com CMH. MÉTODO: Pacientes com diagnóstico de CMH e classe funcional I-II (NYHA) foram selecionados de forma consecutiva. No início do seguimento os participantes realizaram a avaliação da MAOT pela metodologia da média móvel modificada no teste de esforço. Os resultados foram classificados em alterado ou normal. O desfecho foi composto por MSC, fibrilação ventricular, taquicardia ventricular sustentada (TVS) e terapia apropriada do cardioversor desfibrilador implantável (CDI). O nível de significância estatística foi de 5%. RESULTADOS: Um total de 132 pacientes (idade média de 39,5±12,6 anos) foram incluídos, com tempo de seguimento médio de 9,5 anos. A MAOT foi alterada em 74 (56%) participantes e normal em 58 (44%). Durante o seguimento, nove (6,8%) desfechos ocorreram, com prevalência de 1,0%/ano, sendo seis casos de MSC, dois choques apropriados do CDI e um episódio de TVS. MAOT alterada foi associada à taquicardia ventricular não sustentada no Holter (p=0,016), espessura septal≥30 mm (p<0,001) e resposta inadequada da pressão arterial ao esforço (p=0,046). Cinco pacientes (7%) e quatro pacientes (7%) com MAOT alterada e normal, respectivamente, apresentaram desfecho primário [OR=0,85(IC95%: 0,21­3,35, p=0,83)]. Curvas de eventos de Kaplan-Meir não apresentaram diferenças entre MAOT normal e alterada. CONCLUSÃO: A MAOT alterada não foi associada à ocorrência de MSC ou arritmias ventriculares potencialmente fatais em pacientes com CMH, e a baixa taxa desses eventos em um seguimento em longo prazo sugere o bom prognóstico dessa cardiopatia.