Identification of pathogenic variants in the brazilian cohort with familial hypercholesterolemia using exon-targeted gene sequencing

Familial hypercholesterolemia (FH) is a monogenic disease characterized by high plasma low-density lipoprotein cholesterol (LDL-c) levels and increased risk of premature atherosclerotic cardiovascular disease. Mutations in FH-related genes account for 40% of FH cases worldwide. In this study, we aimed to assess the pathogenic variants in FH-related genes in the Brazilian FH cohort FHBGEP using exon-targeted gene sequencing (ETGS) strategy. FH patients (n = 210) were enrolled at five clinical sites and peripheral blood samples were obtained for laboratory testing and genomic DNA extraction. ETGS was performed using MiSeq platform (Illumina). To identify deleterious variants in LDLR, APOB, PCSK9, and LDLRAP1, the long-reads were subjected to Burrows-Wheeler Aligner (BWA) for alignment and mapping, followed by variant calling using Genome Analysis Toolkit (GATK) and ANNOVAR for variant annotation. The variants were further filtered using in-house custom scripts and classified according to the American College Medical Genetics and Genomics (ACMG) guidelines. A total of 174 variants were identified including 85 missense, 3 stop-gain, 9 splice-site, 6 InDel, and 71 in regulatory regions (3'UTR and 5'UTR). Fifty-two patients (24.7%) had 30 known pathogenic or likely pathogenic variants in FH-related genes according to the American College Medical and Genetics and Genomics guidelines. Fifty-three known variants were classified as benign, or likely benign and 87 known variants have shown uncertain significance. Four novel variants were discovered and classified as such due to their absence in existing databases. In conclusion, ETGS and in silico prediction studies are useful tools for screening deleterious variants and identification of novel variants in FH-related genes, they also contribute to the molecular diagnosis in the FHBGEP cohort.

Identification of pathogenic variants in the Brazilian cohort with Familial hypercholesterolemia using exon-targeted gene sequencing.

Familial hypercholesterolemia (FH) is a monogenic disease characterized by high plasma low-density lipoprotein cholesterol (LDL-c) levels and increased risk of premature atherosclerotic cardiovascular disease. Mutations in FH-related genes account for 40% of FH cases worldwide. In this study, we aimed to assess the pathogenic variants in FH-related genes in the Brazilian FH cohort FHBGEP using exon-targeted gene sequencing (ETGS) strategy. FH patients (n = 210) were enrolled at five clinical sites and peripheral blood samples were obtained for laboratory testing and genomic DNA extraction. ETGS was performed using MiSeq platform (Illumina). To identify deleterious variants in LDLR, APOB, PCSK9, and LDLRAP1, the long-reads were subjected to Burrows-Wheeler Aligner (BWA) for alignment and mapping, followed by variant calling using Genome Analysis Toolkit (GATK) and ANNOVAR for variant annotation. The variants were further filtered using in-house custom scripts and classified according to the American College Medical Genetics and Genomics (ACMG) guidelines. A total of 174 variants were identified including 85 missense, 3 stop-gain, 9 splice-site, 6 InDel, and 71 in regulatory regions (3'UTR and 5'UTR). Fifty-two patients (24.7%) had 30 known pathogenic or likely pathogenic variants in FH-related genes according to the American College Medical and Genetics and Genomics guidelines. Fifty-three known variants were classified as benign, or likely benign and 87 known variants have shown uncertain significance. Four novel variants were discovered and classified as such due to their absence in existing databases. In conclusion, ETGS and in silico prediction studies are useful tools for screening deleterious variants and identification of novel variants in FH-related genes, they also contribute to the molecular diagnosis in the FHBGEP cohort.

Clinical Features, Genetic Findings, and Risk Stratification in Arrhythmogenic Right Ventricular Cardiomyopathy: Data From a Brazilian Cohort.

BACKGROUND: Arrhythmogenic right ventricular cardiomyopathy (ARVC), a rare inherited disease, causes ventricular tachycardia, sudden cardiac death, and heart failure (HF). We investigated ARVC clinical features, genetic findings, natural history, and the occurrence of life-threatening arrhythmic events (LTAEs), HF death, or heart transplantation (HF-death/HTx) to identify risk factors. METHODS: The clinical course of 111 consecutive patients with definite ARVC, predictors of LTAE, HF-death/HTx, and combined events were analyzed in the entire cohort and in a subgroup of 40 patients without sustained ventricular arrhythmia before diagnosis. RESULTS: The 5-year cumulative probability of LTAE was 30% and HF-death/HTx was 10%. Predictors of HF-death/HTx were reduced right ventricle ejection fraction (HR: 0.93; P=0.010), HF symptoms (HR: 4.37; P=0.010), epsilon wave (HR: 4.99; P=0.015), and number of leads with low QRS voltage (HR: 1.28; P=0.001). Each additional lead with low QRS voltage increased the risk of HF-death/HTx by 28%. Predictors of LTAE were prior syncope (HR: 1.81; P=0.040), number of leads with T wave inversion (HR: 1.17; P=0.039), low QRS voltage (HR: 1.12; P=0.021), younger age (HR: 0.97; P=0.006), and prior ventricular arrhythmia/ventricular fibrillation (HR: 2.45; P=0.012). Each additional lead with low QRS voltage increased the risk of LTAE by 17%. In patients without ventricular arrhythmia before clinical diagnosis of ARVC, the number of leads with low QRS voltage (HR: 1.68; P=0.023) was independently associated with HF-death/HTx. CONCLUSIONS: Our study demonstrated the characteristics of a specific cohort with a high prevalence of arrhythmic burden at presentation, male predominance, younger age and HF severe outcomes. Our main results suggest that the presence and extension of low QRS voltage can be a risk predictor for HF-death/HTx in ARVC patients, regardless of the arrhythmic risk. This study can contribute to the global ARVC risk stratification, adding new insights to the international current scientific knowledge.

Epigenetic regulation of transcription factor binding motifs promotes Th1 response in Chagas disease cardiomyopathy.

Chagas disease, caused by the protozoan Trypanosoma cruzi, is an endemic parasitic disease of Latin America, affecting 7 million people. Although most patients are asymptomatic, 30% develop complications, including the often-fatal Chronic Chagasic Cardiomyopathy (CCC). Although previous studies have demonstrated some genetic deregulations associated with CCCs, the causes of their deregulations remain poorly described. Based on bulk RNA-seq and whole genome DNA methylation data, we investigated the genetic and epigenetic deregulations present in the moderate and severe stages of CCC. Analysis of heart tissue gene expression profile allowed us to identify 1407 differentially expressed transcripts (DEGs) specific from CCC patients. A tissue DNA methylation analysis done on the same tissue has permitted the identification of 92 regulatory Differentially Methylated Regions (DMR) localized in the promoter of DEGs. An in-depth study of the transcription factors binding sites (TFBS) in the DMRs corroborated the importance of TFBS's DNA methylation for gene expression in CCC myocardium. TBX21, RUNX3 and EBF1 are the transcription factors whose binding motif appears to be affected by DNA methylation in the largest number of genes. By combining both transcriptomic and methylomic analysis on heart tissue, and methylomic analysis on blood, 4 biological processes affected by severe CCC have been identified, including immune response, ion transport, cardiac muscle processes and nervous system. An additional study on blood methylation of moderate CCC samples put forward the importance of ion transport and nervous system in the development of the disease.

Racial inequalities in multimorbidity: baseline of the Brazilian Longitudinal Study of Adult Health (ELSA-Brasil).

BACKGROUND: Evidence of multimorbidity has come mainly from high-income regions, while disparities among racial groups have been less explored. This study examined racial differences in multimorbidity in the multiracial cohort of the Longitudinal Study of Adult Health (Estudo Longitudinal de Saúde do Adulto), ELSA-Brasil. METHODS: The study examined baseline (2008-2010) data for 14 099 ELSA-Brasil participants who self-reported being white, mixed-race, or black. A list of 16 morbidities was used to evaluate multimorbidity, operationalised by simple count into ≥ 2, ≥ 3, ≥ 4, ≥ 5 and ≥ 6 morbidities, in addition to evaluating the number of coexisting conditions. Prevalence ratios (PR) were estimated from logistic models and a quantile model was used to examine racial differences graphically in the distribution quantiles for the number of morbidities. RESULTS: Overall prevalence of multimorbidity (≥ 2 morbidities) was 70% and, after controlling for age and sex, was greater among mixed-race and black participants - by 6% (PR: 1.06; 95% CI: 1.03-1.08) and 9% (PR: 1.09; 95% CI: 1.06-1.12), respectively - than among white participants. As the cutoff value for defining multimorbidity was raised, so the strength of the association increased, especially among blacks: if set at ≥ 6 morbidities, the prevalence was 27% greater for those of mixed-race (PR: 1.27; 95% CI: 1.07-1.50) and 47% greater for blacks (PR: 1.47; 95% CI: 1.22-1.76) than for whites. The disparities were smaller in the lower morbidity distribution quantiles and larger in the upper quantiles, indicating a heavier burden of disease, particularly on blacks. CONCLUSIONS: Multimorbidity was common among adults and older adults in a Brazilian cohort, but important racial inequalities were found. Raising the cutoff point for defining multimorbidity revealed stronger associations between race/skin colour and multimorbidity, indicating a higher prevalence of multimorbidity among mixed-race and black individuals than among whites and that the former groups coexisted more often with more complex health situations (with more coexisting morbidities). Interventions to prevent and manage the condition of multimorbidity that consider the social determinants of health and historically discriminated populations in low- and middle-income regions are necessary.

Body mass index is superior to other body adiposity indexes in predicting incident hypertension in a highly admixed sample after 10-year follow-up: The Baependi Heart Study.

Hypertension is the leading cause of overall mortality in low- and middle-income countries. In Brazil, there is paucity of data on the determinants of incident hypertension and related risk factors. We aimed to determine the incidence of hypertension in a sample from the Brazilian population and investigate possible relationships with body adiposity indexes. We assessed risk factors associated with cardiovascular disease, including adiposity body indexes and biochemical analysis, in a sample from the Baependi Heart Study before and after a 10-year follow-up. Hypertension was defined by the presence of systolic blood pressure (SBP) ≥140 mmHg and/or diastolic blood pressure ≥90 mmHg or the use of antihypertensive drugs. From an initial sample of 1693 participants, 498 (56% women; mean age 38 ± 13 years) were eligible to be included. The overall hypertension incidence was 24.3% (22.3% in men and 25.6% in women). Persons who developed hypertension had higher prevalence of obesity, higher levels for blood pressure, higher frequency of dyslipidemia, and higher body adiposity indexes at baseline. The best prediction model for incident hypertension includes age, sex, HDL-c, SBP, and Body Mass Index (BMI) [AUC = 0.823, OR = 1.58 (95% CI 1.23-2.04)]. BMI was superior in its predictive capacity when compared to Body Adiposity Index (BAI), Body Roundness Index (BRI), and Visceral Adiposity Index (VAI). Incident hypertension in a sample from the Brazilian population was 24.3% after 10-year follow-up and BMI, albeit the simpler index to be calculated, is the best anthropometric index to predict incident hypertension.

The Clinical Genome Resource (ClinGen) Familial Hypercholesterolemia Variant Curation Expert Panel consensus guidelines for LDLR variant classification.

PURPOSE: In 2015, the American College of Medical Genetics and Genomics (ACMG) and the Association for Molecular Pathology (AMP) published consensus standardized guidelines for sequence-level variant classification in Mendelian disorders. To increase accuracy and consistency, the Clinical Genome Resource Familial Hypercholesterolemia (FH) Variant Curation Expert Panel was tasked with optimizing the existing ACMG/AMP framework for disease-specific classification in FH. In this study, we provide consensus recommendations for the most common FH-associated gene, LDLR, where >2300 unique FH-associated variants have been identified. METHODS: The multidisciplinary FH Variant Curation Expert Panel met in person and through frequent emails and conference calls to develop LDLR-specific modifications of ACMG/AMP guidelines. Through iteration, pilot testing, debate, and commentary, consensus among experts was reached. RESULTS: The consensus LDLR variant modifications to existing ACMG/AMP guidelines include (1) alteration of population frequency thresholds, (2) delineation of loss-of-function variant types, (3) functional study criteria specifications, (4) cosegregation criteria specifications, and (5) specific use and thresholds for in silico prediction tools, among others. CONCLUSION: Establishment of these guidelines as the new standard in the clinical laboratory setting will result in a more evidence-based, harmonized method for LDLR variant classification worldwide, thereby improving the care of patients with FH.

Kefir metabolites in a fly model for Alzheimer's disease.

Alzheimer's Disease (AD) is the most common cause of dementia among elderly individuals worldwide, leading to a strong motor-cognitive decline and consequent emotional distress and codependence. It is traditionally characterized by amyloidogenic pathway formation of senile plaques, and recent studies indicate that dysbiosis is also an important factor in AD's pathology. To overcome dysbiosis, probiotics-as kefir-have shown to be a great therapeutic alternative for Alzheimer's disease. In this present work, we explored kefir as a probiotic and a metabolite source as a modulator of microbiome and amyloidogenic pathway, using a Drosophila melanogaster model for AD (AD-like flies). Kefir microbiota composition was determined through 16S rRNA sequencing, and the metabolome of each fraction (hexane, dichloromethane, ethyl acetate, and n-butanol) was investigated. After treatment, flies had their survival, climbing ability, and vacuolar lesions accessed. Kefir and fraction treated flies improved their climbing ability survival rate and neurodegeneration index. In conclusion, we show that kefir in natura, as well as its fractions may be promising therapeutic source against AD, modulating amyloidogenic related pathways.

Triglyceride glucose index as a tool to motivate early lifestyle modification in young adults at diabetes risk: The Baependi Heart Study.

Considering that the incidence of type 2 diabetes mellitus (T2DM) has been increasing especially in developing countries and becoming a global public health problem, this study aims to evaluate the association between triglyceride glucose index (TyG) - which is a mathematical product of the fasting blood glucose and triglyceride levels - and incident T2DM in an adult sample in the Baependi Heart Study (BHS). The data were from the BHS cohort consisting of two periods: cycle 1 (2005-2006; n = 1712; 119 families) and cycle 2 (2010-2013; n = 3017; 127 families). A total of 1121 individuals (both sexes, 18-100 years) were selected if they were assessed in both cycles and not diagnosed with T2DM at baseline (cycle 1). Our findings showed that a participant's risk of developing T2DM increased almost 10 times for a one-unit increase in the TyG (odds ratio OR = 10.17, 95% CI, 7.51-13.93). The association when stratified by age was OR = 28.13 [95% CI, 14.03-56.41] for young adults, meaning that the risk of developing T2DM increased more than 28 times for a one-unit increase in the TyG. For the other groups, young middle-aged adults, old middle-aged adults, and seniors, we found OR = 4.84 [95% CI, 2.91-8.06], OR = 28.73 [95% CI, 10.63-77.65, and OR = 9.88 [95% CI, 3.16-30.90], respectively. A higher TyG implies a significant increase in the risk of developing T2DM, which could be an important screening tool to target early lifestyle intervention in Brazil.

Body adiposity index in assessing the risk of type 2 diabetes mellitus development: the Baependi Heart Study.

BACKGROUND: The association between diabetes and obesity is very well established. Faced with this, several anthropometric indices of adiposity are often involved in studies on diabetes. Our main goal in this paper is to evaluate the association between body adiposity index (BAI) and type 2 diabetes mellitus (T2DM) in a sample of the Brazilian population after 5-year follow-up. METHODS: The data used come from the Baependi Heart Study cohort, which consists of two periods: cycle 1 (2005-2006) and cycle 2 (2010-2013). Individuals of both sexes (n = 1121) were selected by excluding participants with type 2 diabetes mellitus at baseline or those that were lost to follow-up. RESULTS: The diabetic subjects showed higher systolic blood pressure, BAI, body mass index, waist circumference and fasting glucose levels. In addition, using mixed-effects logistic regression, we found that the elevation of a single unit of BAI represented an increase of 8.4% in the risk of a patient developing T2DM (OR = 1.084 [95% CI 1.045-1.124]). CONCLUSIONS: Obesity is recognised as one of the most important risk factors for T2DM and BAI has proven to be a useful tool in estimating the risk of a patient developing T2DM in a Brazilian population.