Update on Genes Associated with Arrhythmogenic Cardiomyopathy

Arrhythmogenic cardiomyopathy is a rare genetic entity characterized by progressive fibro-fatty replacement of myocardium leading to malignant arrhythmias, syncope, and sudden cardiac death. Mostly it affects the right ventricle, but cases have also been described with biventricular and even isolated left ventricular involvement. The disease affects mainly young males and arrhythmias are usually induced by exercise. Arrhythmogenic cardiomyopathy has a genetic origin and is basically caused by deleterious alterations in genes encoding desmosomal proteins, especially plakophilin-2. To date, more than 400 rare genetic alterations have been identified in 18 genes, mainly with autosomal dominant inheritance, but some recessive forms have also been reported (Naxos disease and Carvajal syndrome). A comprehensive genetic analysis identifies a rare variant as potential cause of the disease in around 60% of patients, suggesting the existence of unknown genes as well as other genome alterations not yet discovered. Genetic interpretation classifies some of these rare variants as ambiguous, playing an uncertain role in arrhythmogenic cardiomyopathy. This makes a proper translation of genetic data into clinical practice difficult. Moreover, incomplete penetrance and variable phenotypic expression makes it difficult to arrive at the correct diagnosis. In the present chapter, we focus on recent advances in the knowledge regarding the genetic basis of arrhythmogenic cardiomyopathy

Personalized Interpretation and Clinical Translation of Genetic Variants Associated With Cardiomyopathies

Cardiomyopathies are a heterogeneous group of inherited cardiac diseases characterized by progressive myocardium abnormalities associated with mechanical and/or electrical dysfunction. Massive genetic sequencing technologies allow a comprehensive genetic analysis to unravel the cause of disease. However, most identified genetic variants remain of unknown clinical significance due to incomplete penetrance and variable expressivity. Therefore, genetic interpretation of variants and translation into clinical practice remain a current challenge. We performed retrospective comprehensive clinical assessment and genetic analysis in six families, four diagnosed with arrhythmogenic cardiomyopathy, and two diagnosed with hypertrophic cardiomyopathy (HCM). Genetic testing identified three rare variants (two non-sense and one small indel inducing a frameshift), each present in two families. Although each variant is currently classified as pathogenic and the cause of the diagnosed cardiomyopathy, the onset and/or clinical course differed in each patient. New genetic technology allows comprehensive yet cost-effective genetic analysis, although genetic interpretation, and clinical translation of identified variants should be carefully done in each family in a personalized manner

Prevalence of Pathogenic Variants in Cardiomyopathy-Associated Genes in Myocarditis.

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Rare Variants Associated with Arrhythmogenic Cardiomyopathy: Reclassification Five Years Later.

Genetic interpretation of rare variants associated with arrhythmogenic cardiomyopathy (ACM) is essential due to their diagnostic implications. New data may relabel previous variant classifications, but how often reanalysis is necessary remains undefined. Five years ago, 39 rare ACM-related variants were identified in patients with features of cardiomyopathy. These variants were classified following the American College of Medical Genetics and Genomics' guidelines. In the present study, we reevaluated these rare variants including novel available data. All cases carried one rare variant classified as being of ambiguous significance (82.05%) or likely pathogenic (17.95%) in 2016. In our comprehensive reanalysis, the classification of 30.77% of these variants changed, mainly due to updated global frequencies. As in 2016, nowadays most variants were classified as having an uncertain role (64.1%), but the proportion of variants with an uncertain role was significantly decreased (17.95%). The percentage of rare variants classified as potentially deleterious increased from 17.95% to 23.07%. Moreover, 83.33% of reclassified variants gained certainty. We propose that periodic genetic reanalysis of all rare variants associated with arrhythmogenic cardiomyopathy should be undertaken at least once every five years. Defining the roles of rare variants may help clinicians obtain a definite diagnosis

Discerning the Ambiguous Role of Missense TTN Variants in Inherited Arrhythmogenic Syndromes

The titin gene (TTN) is associated with several diseases, including inherited arrhythmias. Most of these diagnoses are attributed to rare TTN variants encoding truncated forms, but missense variants represent a diagnostic challenge for clinical genetics. The proper interpretation of genetic data is critical for translation into the clinical setting. Notably, many TTN variants were classified before 2015, when the American College of Medical Genetics and Genomics (ACMG) published recommendations to accurately classify genetic variants. Our aim was to perform an exhaustive reanalysis of rare missense TTN variants that were classified before 2015, and that have ambiguous roles in inherited arrhythmogenic syndromes. Rare missense TTN variants classified before 2015 were updated following the ACMG recommendations and according to all the currently available data. Our cohort included 193 individuals definitively diagnosed with an inherited arrhythmogenic syndrome before 2015. Our analysis resulted in the reclassification of 36.8% of the missense variants from unknown to benign/likely benign. Of all the remaining variants, currently classified as of unknown significance, 38.3% showed a potential, but not confirmed, deleterious role. Most of these rare missense TTN variants with a suspected deleterious role were identified in patients diagnosed with hypertrophic cardiomyopathy. More than 35% of the rare missense TTN variants previously classified as ambiguous were reclassified as not deleterious, mainly because of improved population frequencies. Despite being inconclusive, almost 40% of the variants showed a potentially deleterious role in inherited arrhythmogenic syndromes. Our results highlight the importance of the periodical reclassification of rare missense TTN variants to improve genetic diagnoses and help increase the accuracy of personalized medicine

Discerning the Ambiguous Role of Missense TTN Variants in Inherited Arrhythmogenic Syndromes

The titin gene (TTN) is associated with several diseases, including inherited arrhythmias. Most of these diagnoses are attributed to rare TTN variants encoding truncated forms, but missense variants represent a diagnostic challenge for clinical genetics. The proper interpretation of genetic data is critical for translation into the clinical setting. Notably, many TTN variants were classified before 2015, when the American College of Medical Genetics and Genomics (ACMG) published recommendations to accurately classify genetic variants. Our aim was to perform an exhaustive reanalysis of rare missense TTN variants that were classified before 2015, and that have ambiguous roles in inherited arrhythmogenic syndromes. Rare missense TTN variants classified before 2015 were updated following the ACMG recommendations and according to all the currently available data. Our cohort included 193 individuals definitively diagnosed with an inherited arrhythmogenic syndrome before 2015. Our analysis resulted in the reclassification of 36.8% of the missense variants from unknown to benign/likely benign. Of all the remaining variants, currently classified as of unknown significance, 38.3% showed a potential, but not confirmed, deleterious role. Most of these rare missense TTN variants with a suspected deleterious role were identified in patients diagnosed with hypertrophic cardiomyopathy. More than 35% of the rare missense TTN variants previously classified as ambiguous were reclassified as not deleterious, mainly because of improved population frequencies. Despite being inconclusive, almost 40% of the variants showed a potentially deleterious role in inherited arrhythmogenic syndromes. Our results highlight the importance of the periodical reclassification of rare missense TTN variants to improve genetic diagnoses and help increase the accuracy of personalized medicine

Nuevas aportaciones para el estudio de la Majestad Batlló: identificación y caracterización de la policromía subyacente

triomfant a la Creu de mitjan segle xii, exposada a la Col·lecció d’Art Romànic del Museu Nacional d’Art de Catalunya (MNAC 15937), ha estat estudiada de manera íntegra per caracteritzar-ne els aspectes matèrics i tècnics, i comprovar l’existència d’una policromia subjacent. Per portar a terme aquest estudi, s’han emprat mètodes d’anàlisi no destructives com l’examen radiogràfic, la reflectografia infraroja, les imatges amb llum ultraviolada i l’observació detallada amb llum visible i llum rasant, a través d’ulleres d’augment i lupa binocular, que s’han comparat i verificat amb les anàlisis quimicofísiques com la microscòpia òptica (OM), la microscòpia electrònica (SEM), amb microanàlisi elemental incorporada (SEM/EDXA), i l’espectroscòpia infraroja (FT-IR). Aquesta combinació de mètodes d’anàlisi ha servit per identificar dues policromies existents i determinar com era la policromia original de la Majestat i de la creu. Presentem, així, una hipotètica reconstrucció virtual de la decoració pictòrica subjacent, substancialment diferent de la policromia actual, que estem segurs que aportarà dades significatives per enriquir el repertori decoratiu de les majestats catalanes del romànic.La Majestad Batlló, una talla de Cristo triumfante en la Cruz de mediados del siglo xii, expuesta en la Colección de Arte Románico del Museu Nacional d’Art de Catalunya (MNAC 15937), ha sido estudiada de manera íntegra para caracterizar sus aspectos matéricos y técnicos, y comprobar la existencia de una policromía subyacente. Para llevar a cabo este estudio, se han utilizado métodos de análisis no destructivos como el examen radiográfico, la reflectografía infrarroja, las imágenes con luz ultravioleta y la observación detallada con luz visible y luz rasante, a través de lentes de aumento y lupa binocular, que se han comparado y verificado con los análisis quimicofísicos como la microscopia óptica (OM), la microscopia electrónica (SEM), con microanálisis elemental incorporado (SEM/EDXA), y la espectroscopia infrarroja (FT-IR). Esta combinación de métodos de análisis ha servido para identificar dos policromías existentes y determinar cómo era la policromía original de la Majestad y de la cruz. Presentamos, de esta manera, una hipotética reconstrucción virtual de la decoración pictórica subyacente, sustancialmente diferente de la policromía actual, que estamos seguros que aportará datos significativos para enriquecer el repertorio decorativo de las majestades catalanas del románico.The Batlló Majesty, a carving of Christ on the Cross from the middle of the 12th century, exhibited in the Museu Nacional d’Art de Catalunya’s Romanesque Art Collection (MNAC 15937), has been comprehensively studied in order to typify its material and technical aspects, and to verify the existence of underlying polychromy. In order to perform this study, non-destructive methods of analysis have been carried out like X-ray examination, infrared reflectography, images with ultraviolet light and detailed observation with visible light and raking light, through hand lenses and the binocular microscope, which have been compared and checked with chemical and physical analyses like the optical microscope (OM), electron microscope (SEM), with microanalysis of the elements incorporated (SEM/EDXA), and infrared spectroscopy (FT-IR). This combination of methods of analysis has been useful for identifying two existing polychromies and determining what the original polychromy of the Majesty and the cross was like. We thus present a hypothetical virtual reconstruction of the underlying pictorial decoration, substantially different from the present polychromy, which we are sure will contribute significant data to enrich the decorative repertoire of Catalan Romanesque Majesties

Noves aportacions per a l'estudi de la Majestat Batlló: identificació i caracterització de la policromia subjacent

La Majestat Batlló, una talla de Crist triomfant a la Creu de mitjan segle XII, exposada a la Col·lecció d'Art Romànic del Museu Nacional d'Art de Catalunya (MNAC15937), ha estat estudiada de manera integra per caracteritzar-ne els aspectes matèrics i tècnics, i comprovar l'existència d'una policromia subjacent. Per portar a terme aquest estudi, s'han emprat mètodes d'anàlisi no destructives com l'examen radiogràfic, la reflectografia infraroja, les imatges amb llum ultraviolada i l'observació detallada amb llum visible i llum rasant, a través d'ulleres d'augment i lupa binocular, que s'han comparat i verificat amb les anàlisis quimicofísiques com la microscòpia òptica (OM), la microscòpia electrònica (SEM), amb microanàlisi elemental incorporada (SEM/EDXA) i l'espectroscòpia infraroja (FT-IR). Aquesta combinació de mètodes d'anàlisi ha servit per identificar dues policromies existents i determinar com era la policromia original de la Majestat i de la creu. Presentem, així, una hipotètica reconstrucció virtual de la decoració pictòrica subjacent, substancialment diferent de la policromia actual, que estem segurs que aportarà dades significatives per enriquir el repertori decoratiu de les majestats catalanes del romànic

Alterations in Calcium Handling Are a Common Feature in an Arrhythmogenic Cardiomyopathy Cell Model Triggered by Desmosome Genes Loss

Arrhythmogenic cardiomyopathy (ACM) is an inherited cardiac disease characterized by fibrofatty replacement of the myocardium. Deleterious variants in desmosomal genes are the main cause of ACM and lead to common and gene-specific molecular alterations, which are not yet fully understood. This article presents the first systematic in vitro study describing gene and protein expression alterations in desmosomes, electrical conduction-related genes, and genes involved in fibrosis and adipogenesis. Moreover, molecular and functional alterations in calcium handling were also characterized. This study was performed d with HL1 cells with homozygous knockouts of three of the most frequently mutated desmosomal genes in ACM: PKP2, DSG2, and DSC2 (generated by CRISPR/Cas9). Moreover, knockout and N-truncated clones of DSP were also included. Our results showed functional alterations in calcium handling, a slower calcium re-uptake was observed in the absence of PKP2, DSG2, and DSC2, and the DSP knockout clone showed a more rapid re-uptake. We propose that the described functional alterations of the calcium handling genes may be explained by mRNA expression levels of ANK2, CASQ2, ATP2A2, RYR2, and PLN. In conclusion, the loss of desmosomal genes provokes alterations in calcium handling, potentially contributing to the development of arrhythmogenic events in ACM

Alterations in Calcium Handling Are a Common Feature in an Arrhythmogenic Cardiomyopathy Cell Model Triggered by Desmosome Genes Loss

Arrhythmogenic cardiomyopathy (ACM) is an inherited cardiac disease characterized by fibrofatty replacement of the myocardium. Deleterious variants in desmosomal genes are the main cause of ACM and lead to common and gene-specific molecular alterations, which are not yet fully understood. This article presents the first systematic in vitro study describing gene and protein expression alterations in desmosomes, electrical conduction-related genes, and genes involved in fibrosis and adipogenesis. Moreover, molecular and functional alterations in calcium handling were also characterized. This study was performed d with HL1 cells with homozygous knockouts of three of the most frequently mutated desmosomal genes in ACM: PKP2, DSG2, and DSC2 (generated by CRISPR/Cas9). Moreover, knockout and N-truncated clones of DSP were also included. Our results showed functional alterations in calcium handling, a slower calcium re-uptake was observed in the absence of PKP2, DSG2, and DSC2, and the DSP knockout clone showed a more rapid re-uptake. We propose that the described functional alterations of the calcium handling genes may be explained by mRNA expression levels of ANK2, CASQ2, ATP2A2, RYR2, and PLN. In conclusion, the loss of desmosomal genes provokes alterations in calcium handling, potentially contributing to the development of arrhythmogenic events in ACM