RESUMO
Chronic kidney disease (CKD) is represented by a diminished filtration capacity of the kidneys. End-stage renal disease patients need dialysis treatment to remove waste and toxins from the circulation. However, endogenously produced uremic toxins (UTs) cannot always be filtered during dialysis. UTs are among the CKD-related factors that have been linked to maladaptive and pathophysiological remodeling of the heart. Importantly, 50% of the deaths in dialysis patients are cardiovascular related, with sudden cardiac death predominating. However, the mechanisms responsible remain poorly understood. The current study aimed to assess the vulnerability of action potential repolarization caused by exposure to pre-identified UTs at clinically relevant concentrations. We exposed human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and HEK293 chronically (48 h) to the UTs indoxyl sulfate, kynurenine, or kynurenic acid. We used optical and manual electrophysiological techniques to assess action potential duration (APD) in the hiPSC-CMs and recorded IKr currents in stably transfected HEK293 cells (HEK-hERG). Molecular analysis of KV11.1, the ion channel responsible for IKr, was performed to further understand the potential mechanism underlying the effects of the UTs. Chronic exposure to the UTs resulted in significant APD prolongation. Subsequent assessment of the repolarization current IKr, often most sensitive and responsible for APD alterations, showed decreased current densities after chronic exposure to the UTs. This outcome was supported by lowered protein levels of KV11.1. Finally, treatment with an activator of the IKr current, LUF7244, could reverse the APD prolongation, indicating the potential modulation of electrophysiological effects caused by these UTs. This study highlights the pro-arrhythmogenic potential of UTs and reveals a mode of action by which they affect cardiac repolarization.
Assuntos
Células-Tronco Pluripotentes Induzidas , Insuficiência Renal Crônica , Humanos , Toxinas Urêmicas , Células HEK293 , Potenciais de Ação , Células-Tronco Pluripotentes Induzidas/metabolismo , Diálise Renal , Miócitos Cardíacos , Insuficiência Renal Crônica/metabolismoRESUMO
Inherited forms of arrhythmogenic and dilated cardiomyopathy (ACM and DCM) are characterized by variable disease expression and age-related penetrance. Calcium (Ca2+) is crucially important for proper cardiac function, and dysregulation of Ca2+ homeostasis seems to underly cardiomyopathy etiology. A polymorphism, c.286T>G p.(Ser96Ala), in the gene encoding the histidine-rich Ca2+ binding (HRC) protein, relevant for sarcoplasmic reticulum Ca2+ cycling, has previously been associated with a marked increased risk of life-threatening arrhythmias among idiopathic DCM patients. Following this finding, we investigated whether p.(Ser96Ala) affects major cardiac disease manifestations in carriers of the phospholamban (PLN) c.40_42delAGA; p.(Arg14del) pathogenic variant (cohort 1); patients diagnosed with, or predisposed to, ACM (cohort 2); and DCM patients (cohort 3). We found that the allele frequency of the p.(Ser96Ala) polymorphism was similar across the general European-American population (control cohort, 40.3-42.2%) and the different cardiomyopathy cohorts (cohorts 1-3, 40.9-43.9%). Furthermore, the p.(Ser96Ala) polymorphism was not associated with life-threatening arrhythmias or heart failure-related events across various patient cohorts. We therefore conclude that there is a lack of evidence supporting the important role of the HRC p.(Ser96Ala) polymorphism as a modifier in cardiomyopathy, refuting previous findings. Further research is required to identify bona fide genomic predictors for the stratification of cardiomyopathy patients and their risk for life-threatening outcomes.
Assuntos
Cardiomiopatias , Cardiomiopatia Dilatada , Humanos , Arritmias Cardíacas/genética , Arritmias Cardíacas/metabolismo , Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Cardiomiopatias/genética , Cardiomiopatia Dilatada/genética , Histidina/genética , Polimorfismo GenéticoRESUMO
In arrhythmogenic cardiomyopathy (ACM) pathogenic variants are found in genes encoding desmosomal proteins and in non-desmosomal genes, such as phospholamban (PLN, p.Arg14del variant). Previous research showed that plakoglobin protein levels and localization in the cardiac tissue of ACM patients, and PLN p.Arg14del patients diagnosed with an ACM phenotype, are disturbed. Moreover, the effects of pathogenic variants in desmosomal genes are reflected in non-cardiac tissues like buccal mucosa cells (BMC) which could serve as a promising new and non-invasive tool to support diagnosis. We collected the BMC of 33 ACM patients, 17 PLN p.Arg14del patients and 34 controls, labelled the BMC with anti-plakoglobin antibodies at different concentrations, and scored their membrane labelling. We found that plakoglobin protein levels were significantly reduced in BMC obtained from diagnosed ACM patients and preclinical variant carriers when compared to controls. This effect was independent from age and sex. Moderate to strong correlations were found with the revised 2010 Task Force Criteria score which is commonly used for ACM diagnosis (rs = -0.67, n = 64, p < 0.0001 and rs = -0.71, n = 64, p < 0.0001). In contrast, plakoglobin scores in PLN p.Arg14del patients were comparable to controls (p > 0.209), which suggests differences in underlying etiology. However, for the individual diagnosis of the 'classical' ACM patient, this method might not be discriminative enough to distinguish true patients from variant carriers and controls, because of the high interindividual variability.
Assuntos
Displasia Arritmogênica Ventricular Direita/diagnóstico , Displasia Arritmogênica Ventricular Direita/patologia , Mucosa Bucal/patologia , Adulto , Displasia Arritmogênica Ventricular Direita/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Desmossomos/metabolismo , Desmossomos/patologia , Progressão da Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mucosa Bucal/metabolismo , gama Catenina/metabolismoRESUMO
Arrhythmogenic cardiomyopathy (ACM) is a progressive inheritable disease which is characterized by a gradual fibro-(fatty) replacement of the myocardium. Visualization of diffuse and patchy fibrosis patterns is challenging using clinically applied cardiac imaging modalities (e.g., late gadolinium enhancement, LGE). During collagen synthesis and breakdown, carboxy-peptides are released into the bloodstream, specifically procollagen type-I carboxy-terminal propeptides (PICP) and collagen type-I carboxy-terminal telopeptides (ICTP). We collected the serum and EDTA blood samples and clinical data of 45 ACM patients (age 50.11 ± 15.53 years, 44% female), divided into 35 diagnosed ACM patients with a 2010 ARVC Task Force Criteria score (TFC) ≥ 4, and 10 preclinical variant carriers with a TFC < 4. PICP levels were measured using an enzyme-linked immune sorbent assay and ICTP levels with a radio immunoassay. Increased PICP/ICTP ratios suggest a higher collagen deposition. We found significantly higher PICP and PICP/ICTP levels in diagnosed patients compared to preclinical variant carriers (p < 0.036 and p < 0.027). A moderate negative correlation existed between right ventricular ejection fractions (RVEF) and the PICP/ICTP ratio (r = -0.46, p = 0.06). In addition, significant correlations with left ventricular function (LVEF r = -0.53, p = 0.03 and end-systolic volume r = 0.63, p = 0.02) were found. These findings indicate impaired contractile performance due to pro-fibrotic remodeling. Follow-up studies including a larger number of patients should be performed to substantiate our findings and the validity of those levels as potential promising biomarkers in ACM.
RESUMO
Ribosome biogenesis in eukaryotes requires the coordinated production and assembly of 80 ribosomal proteins and four ribosomal RNAs (rRNAs), and its rate must be synchronized with cellular growth. Here, we showed that the Microprocessor complex, which mediates the first step of microRNA processing, potentiated the transcription of ribosomal protein genes by eliminating DNA/RNA hybrids known as R-loops. Nutrient deprivation triggered the nuclear export of Drosha, a key component of the Microprocessor complex, and its subsequent degradation by the E3 ubiquitin ligase Nedd4, thereby reducing ribosomal protein production and protein synthesis. In mouse erythroid progenitors, conditional deletion of Drosha led to the reduced production of ribosomal proteins, translational inhibition of the mRNA encoding the erythroid transcription factor Gata1, and impaired erythropoiesis. This phenotype mirrored the clinical presentation of human "ribosomopathies." Thus, the Microprocessor complex plays a pivotal role in synchronizing protein synthesis capacity with cellular growth rate and is a potential drug target for anemias caused by ribosomal insufficiency.
Assuntos
Biossíntese de Proteínas , Proteínas Ribossômicas/biossíntese , Ribossomos , Animais , Eritropoese , Camundongos , RNA Ribossômico/metabolismo , Proteínas Ribossômicas/genética , Ribossomos/metabolismoRESUMO
Background: Pathogenic variants in phospholamban (PLN, like p. Arg14del), are found in patients diagnosed with arrhythmogenic (ACM) and dilated cardiomyopathy (DCM). Fibrosis formation in the heart is one of the hallmarks in PLN p.Arg14del carriers. During collagen synthesis and breakdown, propeptides are released into the circulation, such as procollagen type I carboxy-terminal propeptide (PICP) and C-terminal telopeptide collagen type I (ICTP). Aim: To investigate if PICP/ICTP levels in blood are correlative biomarkers for clinical disease severity and outcome in PLN p.Arg14del variant carriers. Methods: Serum and EDTA blood samples were collected from 72 PLN p.Arg14del carriers (age 50.5 years, 63% female) diagnosed with ACM (n = 12), DCM (n = 14), and preclinical variant carriers (n = 46). PICP levels were measured with an enzyme-linked immune sorbent assay and ICTP with a radio immuno-assay. Increased PICP/ICTP ratios suggest a higher collagen deposition. Clinical data including electrocardiographic, and imaging results were adjudicated from medical records. Results: No correlation between PICP/ICTP ratios and late gadolinium enhancement (LGE) was found. Moderate correlations were found between the PICP/ICTP ratio and end-diastolic/systolic volume (both r s = 0.40, n = 23, p = 0.06). PICP/ICTP ratio was significantly higher in patients with T wave inversion (TWI), especially in leads V4-V6, II, III, and aVF (p < 0.022) and in patients with premature ventricular contractions (PVCs) during an exercise tolerance test (p = 0.007). Conclusion: High PICP/ICTP ratios correlated with clinical parameters, such as TWI and PVCs. Given the limited size and heterogeneity of the patient group, additional studies are required to substantiate the incremental prognostic value of these fibrosis biomarkers in PLN p.Arg14del patients.
RESUMO
Arrhythmogenic cardiomyopathy (ACM) is a life-threatening cardiac disease caused by mutations in genes predominantly encoding for desmosomal proteins that lead to alterations in the molecular composition of the intercalated disc. ACM is characterized by progressive replacement of cardiomyocytes by fibrofatty tissue, ventricular dilatation, cardiac dysfunction, and heart failure but mostly dominated by the occurrence of life-threatening arrhythmias and sudden cardiac death (SCD). As SCD appears mostly in apparently healthy young individuals, there is a demand for better risk stratification of suspected ACM mutation carriers. Moreover, disease severity, progression, and outcome are highly variable in patients with ACM. In this review, we discuss the aetiology of ACM with a focus on pro-arrhythmic disease mechanisms in the early concealed phase of the disease. We summarize potential new biomarkers which might be useful for risk stratification and prediction of disease course. Finally, we explore novel therapeutic strategies to prevent arrhythmias and SCD in the early stages of ACM.
Assuntos
Potenciais de Ação , Displasia Arritmogênica Ventricular Direita/fisiopatologia , Frequência Cardíaca , Remodelação Ventricular , Animais , Displasia Arritmogênica Ventricular Direita/genética , Displasia Arritmogênica Ventricular Direita/mortalidade , Displasia Arritmogênica Ventricular Direita/terapia , Morte Súbita Cardíaca/etiologia , Morte Súbita Cardíaca/prevenção & controle , Predisposição Genética para Doença , Humanos , Mutação , Fenótipo , Prevalência , Prognóstico , Medição de Risco , Fatores de RiscoRESUMO
Sudden cardiac death is a leading cause of death worldwide, mainly caused by highly disturbed electrical activation patterns in the heart. Currently, murine models are the most popular model to study underlying molecular mechanisms of inherited or acquired cardiac electrical abnormalities, although the numerous electrophysiological discrepancies between mouse and human raise the question whether mice are the optimal model to study cardiac rhythm disorders. Recently it has been uncovered that the zebrafish cardiac electrophysiology seems surprisingly similar to the human heart, mainly because the zebrafish AP contains a clear plateau phase and ECG characteristics show alignment with the human ECG. Although, before using zebrafish as a model to study cardiac arrhythmogenesis, however, it is very important to gain a better insight into the electrophysiological characteristics of the zebrafish heart. In this review we outline the electrophysiological machinery of the zebrafish cardiomyocytes, with a special focus on the intracellular Ca2+ dynamics and excitation-contraction coupling. We debate the potential of zebrafish as a model to study human cardiovascular diseases and postulate steps to employ zebrafish into a more 'humanized' model.