Case Report: A novel EPAS1 mutation in a case of paraganglioma complicated with polycythemia and atrial septal defect.

Background: Paraganglioma is a rare neuroendocrine tumor and is highly associated with hereditary susceptibility genes, often occurring as part of a genetic syndrome. The genetic heterogeneity of paraganglioma poses challenges in diagnosis, counseling, and clinical management. Case summary: We present the case of a 60-year-old woman with hypertension, atrial septal defect, and polycythemia, who experienced paroxysmal palpitations, sweating, headache, abdominal pain, nausea, and vomiting. Her blood pressure was severely unstable. Blood laboratory tests revealed elevated catecholamine levels, contrast-enhanced CT of her whole abdomen showed a round retroperitoneal mass with soft tissue density, and somatostatin receptor imaging (68Ga PET-CT) indicated a retroperitoneal mass with abnormally increased expression of somatostatin receptor. It is interesting to note that whole exome sequencing (WES) analyses on both blood and tumor samples revealed a novel EPAS1 mutation, specifically the c.2501A > G; p.Tyr834Cys variant, which has never been reported. The patient was diagnosed with paraganglioma and underwent successful Da Vinci robot-assisted laparoscopic resection of the retroperitoneal tumor. During a 3-month follow-up period, her blood pressure stabilized, and her symptoms significantly improved. Conclusion: This case reveals that the EPSA1 mutation may be the primary driver of paraganglioma complicated by atrial septal defect and polycythemia. Additionally, the utilization of Da Vinci robot-assisted laparoscopic surgery contributed to a favorable prognosis for the patient.

Identification of a novel TBX5 c.755 + 1 G > A variant and related pathogenesis in a family with Holt-Oram syndrome.

The proband with congenital heart disease and abnormal thumb was clinically diagnosed as Holt-Oram syndrome (HOS). A novel variant, T-box transcription factor 5 (TBX5) c.755 + 1 G > A, was identified in the proband via whole exome sequencing and validated using Sanger sequencing. Pedigree analysis and clinical examinations revealed three/seven individuals over three generations within the family, with features suggestive of HOS. Deep amplicon sequencing confirmed that the allele frequencies of the novel variant in the proband (III-1), her brother (III-2), and her mother (II-2) were 50%, 48.3%, and 38.1%, respectively, indicating that III-1 and III-2 harbored heterozygous variants, while II-2 harbored mosaic heterozygous variants. The minigene splicing assay showed that the novel variant affected the normal splicing of exon 7, resulting in the production of abnormal TBX5 transcripts. Reverse transcription-quantitative polymerase chain reaction and western blot analyses revealed that the novel variant upregulated TBX5 expression at the transcriptional and translational levels. Nuclear localization assay demonstrated impaired nuclear localization of the mutant TBX5. Cell viability assay revealed the inhibition of cell activity by the mutant TBX5. Our findings indicate that the novel variant was potentially induced HOS, probably by causing aberrant splicing, reducing the enrichment of nuclear TBX5 protein, and inhibiting cellular proliferation.

CArdiac MagnEtic resonance assessment of bi-Atrial fibrosis in secundum atrial septal defects patients: CAMERA-ASD study.

AIMS: Atrial septal defects (ASD) are associated with atrial arrhythmias, but the arrhythmia substrate in these patients is poorly defined. We hypothesized that bi-atrial fibrosis is present and that right atrial fibrosis is associated with atrial arrhythmias in ASD patients. We aimed to evaluate the extent of bi-atrial fibrosis in ASD patients and to investigate the relationships between bi-atrial fibrosis, atrial arrhythmias, shunt fraction, and age. METHODS AND RESULTS: Patients with uncorrected secundum ASDs (n = 36; 50.4 ± 13.6 years) underwent cardiac magnetic resonance imaging with atrial late gadolinium enhancement. Comparison was made to non-congenital heart disease patients (n = 36; 60.3 ± 10.5 years) with paroxysmal atrial fibrillation (AF). Cardiac magnetic resonance parameters associated with atrial arrhythmias were identified and the relationship between bi-atrial structure, age, and shunt fraction studied. Bi-atrial fibrosis burden was greater in ASD patients than paroxysmal AF patients (20.7 ± 14% vs. 10.1 ± 8.6% and 14.8 ± 8.5% vs. 8.6 ± 6.1% for right and left atria respectively, P = 0.001 for both). In ASD patients, right atrial fibrosis burden was greater in those with than without atrial arrhythmias (33.4 ± 18.7% vs. 16.8 ± 10.3%, P = 0.034). On receiver operating characteristic analysis, a right atrial fibrosis burden of 32% had a 92% specificity and 71% sensitivity for predicting the presence of atrial arrhythmias. Neither age nor shunt fraction was associated with bi-atrial fibrosis burden. CONCLUSION: Bi-atrial fibrosis burden is greater in ASD patients than non-congenital heart disease patients with paroxysmal AF. Right atrial fibrosis is associated with the presence of atrial arrhythmias in ASD patients. These findings highlight the importance of right atrial fibrosis to atrial arrhythmogenesis in ASD patients.

Immunometabolic factors in adolescent chronic disease are associated with Th1 skewing of invariant Natural Killer T cells.

Invariant Natural Killer T (iNKT) cells respond to the ligation of lipid antigen-CD1d complexes via their T-cell receptor and are implicated in various immunometabolic diseases. We considered that immunometabolic factors might affect iNKT cell function. To this end, we investigated iNKT cell phenotype and function in a cohort of adolescents with chronic disease and immunometabolic abnormalities. We analyzed peripheral blood iNKT cells of adolescents with cystic fibrosis (CF, n = 24), corrected coarctation of the aorta (CoA, n = 25), juvenile idiopathic arthritis (JIA, n = 20), obesity (OB, n = 20), and corrected atrial septal defect (ASD, n = 25) as controls. To study transcriptional differences, we performed RNA sequencing on a subset of obese patients and controls. Finally, we performed standardized co-culture experiments using patient plasma, to investigate the effect of plasma factors on iNKT cell function. We found comparable iNKT cell numbers across patient groups, except for reduced iNKT cell numbers in JIA patients. Upon ex-vivo activation, we observed enhanced IFN-γ/IL-4 cytokine ratios in iNKT cells of obese adolescents versus controls. The Th1-skewed iNKT cell cytokine profile of obese adolescents was not explained by a distinct transcriptional profile of the iNKT cells. Co-culture experiments with patient plasma revealed that across all patient groups, obesity-associated plasma factors including LDL-cholesterol, leptin, and fatty-acid binding protein 4 (FABP4) coincided with higher IFN-γ production, whereas high HDL-cholesterol and insulin sensitivity (QUICKI) coincided with higher IL-4 production. LDL and HDL supplementation in co-culture studies confirmed the effects of lipoproteins on iNKT cell cytokine production. These results suggest that circulating immunometabolic factors such as lipoproteins may be involved in Th1 skewing of the iNKT cell cytokine response in immunometabolic disease.

Novel insertion mutation (Arg1822_Glu1823dup) in MYH6 coiled-coil domain causing familial atrial septal defect.

OBJECTIVE: Atrial septal defect, secundum (ASD Ⅱ, OMIM: 603642) is the second common congenital heart defect (CHD) in China. However, the genetic etiology of familial ASD II remains elusive. METHODS AND RESULTS: Using whole-exome sequencing (WES) and Sanger sequencing, we identified a novel myosin heavy chain 6 (MYH6) gene insertion variation, NM_002471.3: c.5465_5470dup (Arg1822_Glu1823dup), in a large Chinese Han family with ASD II. The variant Arg1822_Glu1823dup co-segregated with the disease in this family with autosomal dominant inheritance. The insertion variant located in the coiled-coil domain of the MYH6 protein, which is highly conserved across homologous myosin proteins and species. In transfected myoblast C2C12 cell lines, the MYH6 Arg1822_Glu1823dup variant significantly impaired myofibril formation and increased apoptosis but did not significantly reduce cell viability. Furthermore, molecular simulations revealed that the Arg1822_Glu1823dup variant impaired the myosin α-helix, increasing the stability of the coiled-coil myosin dimer, suggesting that this variant has an effect on the coiled-coil domain self-aggregation. These findings indicate that Arg1822_Glu1823dup variant plays a crucial role in the pathogenesis of ASD II. CONCLUSION: Our findings expand the spectrum of MYH6 variations associated with familial ASD II and may provide a molecular basis in genetic counseling and prenatal diagnosis for this Chinses family.

Functional analysis of two novel TBX5 variants present in individuals with Holt-Oram syndrome with different clinical manifestations.

Holt-Oram syndrome (HOS) is a rare disorder characterized by cardiac and upper-limb defects. Pathogenic variants in TBX5-a gene encoding a transcription factor important for heart and skeletal development-are the only known cause of HOS. Here, we present the identification and functional analysis of two novel TBX5 pathogenic variants found in two individuals with HOS presenting distinct phenotypes. The individual with the c.905delA variant has a severe cardiac phenotype but mild skeletal defects, unlike the individual with the c.246_249delGATG variant who has no cardiac problems but severe upper limbs malformations, including phocomelia. Both frameshift variants, c.246_249delGATG and c.905delA, generate mRNAs harbouring premature stop codons which, if not degraded by nonsense mediated decay, will lead to the production of shorter TBX5 proteins, p.Gln302Argfs*92 and p.Met83Phefs*6, respectively. Immunocytochemistry results suggest that both mutated proteins are produced and furthermore, like the wild-type protein, p.Gln302Argfs*92 mutant appears to be mainly localized in the nucleus, in contrast with p.Met83Phefs*6 mutant that displays a higher level of cytoplasmic localization. In addition, luciferase activity analysis revealed that none of the TBX5 mutants are capable of transactivating the NPPA promoter. In conclusion, our results provide evidence that both pathogenic variants cause a severe TBX5 loss-of-function, dramatically reducing its biological activity. The absence of cardiac problems in the individual with the p.Met83Phefs*6 variant supports the existence of other mechanisms/genes underlying the pathogenesis of HOS and/or the existence of an age-related delay in the development of a more serious cardiac phenotype. Further studies are required to understand the differential effects observed in the phenotypes of both individuals.

Deployed Dimensions of the GORE® CARDIOFORM ASD Occluder as Function of Defect Size.

The GORE® CARDIOFORM ASD occluder (ASDO) is approved for closure of ASDs up to 35 mm diameter. With an adaptable central waist, each device size is suitable over a range of defect diameters. Understanding deployed dimensions across various defect sizes will assist operators. Therefore, this study investigates the deployed dimensions of the ASDO as a function of defect size. A 2-mm-thick ASD model with circular defects ranging from 5 to 35 mm was 3D printed. Diameter, width, and left-right disc diameter were measured by fluoroscopy after ASDO devices were deployed in applicable defects. Linear regression evaluated relationships between device size, defect size, and deployed dimensions. Six ASDOs of each size (27, 32, 37, 44, and 48 mm) were deployed in all applicable defects. There was significant ASDO size-defect size interaction in determining deployed ASDO diameter. Diameter was positively associated with defect size for 48-mm (B = 0.13, p < 0.001) and 44-mm (B = 0.11, p < 0.001) ASDOs, while no association was seen for 27-mm, 32-mm, or 37-mm ASDOs. No such interaction existed for deployed width or left-right disc difference. Controlling for ASDO size, width (B = - 0.12, p < 0.001) and left-right disc difference (B = - 0.06, p < 0.001) were negatively associated with defect size. In smaller defects, the 44-mm and 48-mm ASDOs display progressive diameter foreshortening, and all devices display progressive increase in width and left-right disc asymmetry. Anticipating the degree of diameter foreshortening may be critical when attempting closure of fenestrated lesions and/or in patients with limited total atrial septal length.

TBX3 and TBX5 duplication: A family with an atypical overlapping Holt-Oram/ulnar-mammary syndrome phenotype.

Holt-Oram syndrome (HOS) is a rare, autosomal dominant heart-hand syndrome caused by mutations in the TBX5 gene. A wide spectrum of TBX5 mutations have been reported previously, most resulting in a null allele leading to haploinsufficiency. TBX5 gene duplications have been previously reported in association with typical and atypical HOS phenotypes. Ulnar-Mammary syndrome (UMS) is a distinct rare, autosomal dominant condition caused by mutations in the TBX3 gene. TBX5 and TBX3 are physically linked in cis on human chromosome 12 and contiguous chromosome 12q24 deletions comprising both TBX5 and TBX3 genes have been previously reported but to our knowledge, duplications have never been described. We report on a large German family with at least 17 affected individuals over 6 generations bearing a duplication at 12q24.21 identified on array-CGH comprising both TBX5 and TBX3 genes. Affected patients are presenting with HOS and UMS symptoms, consisting of variable limb anomalies involving the radial and the ulnar rays and cardiac findings such as congenital heart defects, persistent arterial duct or aortic stenosis, and non-classical symptoms, such as supernumerary nipples and cardiomyopathy. Fluorescence in situ hybridisation confirmed a tandem duplication at the 12q24.21 locus. This is the first report of a contiguous TBX3/TBX5 duplication associated with HOS/UMS phenotype.

Cardiac Anomalies in Two White-Tailed Eagles (Haliaeetus albicilla) in Sweden.

Severe cardiomegaly with an atrial septal defect was discovered during necropsy of a subadult White-tailed Eagle (Haliaeetus albicilla) found dead in the wild. A thin membrane composed of fibromuscular tissue separated the left atrium into two chambers, most consistent with that described for cor triatriatum sinister (CTS) in other species. Seventeen months later, necropsy of an adult White-tailed Eagle again revealed CTS. This lesion has not been reported previously in raptors.

Giant Eustachian valve and Thebesian valve-A highly deceptive structures as atrial septal defect rims.

The Eustachian valve (EV) of the inferior vena cava and the Thebesian valve (TV) of the coronary sinus are incompletely regressed structures of embryonic sinus venosus. In the majority of cases, the EV and TV disappear completely after birth or are represented only by a thin crescentic fold. On echocardiography, these vestiges may mimic abnormal structures. We report a case with giant EV and TV which were initially misinterpreted as rims of an atrial septal defect (ASD) leading to the false diagnosis of ostium secundum ASD.

Familial dilated cardiomyopathy associated with pathogenic TBX5 variants: Expanding the cardiac phenotype associated with Holt-Oram syndrome.

Holt-Oram syndrome (HOS) is a rare, autosomal dominant disorder caused by heterozygous pathogenic variants in cardiac T-box transcription factor, TBX5. Classically, it is associated with upper limb malformations and variable cardiac abnormalities. Limb manifestations are considered to be invariably present, ranging in severity from limitation in movement, to triphalangeal thumbs, absent thumbs, shortened forearms, or phocomelia. Cardiac involvement is characterized by congenital heart defects, most commonly septal structural malformations, and conduction system disease. Recently, novel TBX5 variants have also been reported in association with dilated cardiomyopathy (DCM). In this context, we report eight individuals from four unrelated families, in whom pathogenic variants in TBX5 segregated with an atypical HOS phenotype. Affected individuals exhibit relatively mild skeletal features of HOS, with a predominant cardiac phenotype, which includes several individuals affected by non-ischaemic DCM. To our knowledge, these represent the first reported cases of DCM in families with skeletal features of HOS, some of whom also harbored variants previously linked to a classical HOS phenotype (p. Arg279* and p.Arg237Gln). This finding supports diverse roles of TBX5 in cardiovascular development and function, and confirms the importance of long-term cardiac surveillance for individuals affected by HOS. Furthermore, these families highlight the wide phenotypic variability of HOS, which may include comparatively mild upper limb findings in respect to cardiac manifestations.

Surgical Management of Congenital Bilateral Multiple Atrial Aneurysms.

Congenital atrial aneurysms are a rare malformation, often associated with supraventricular arrhythmias. Here, we present the case of a child with biatrial aneurysms and a type 2 atrioseptal defect. Directly after birth the girl became symptomatic with incessant ectopic atrial tachyarrhythmia. On echocardiography, multiple biatrial aneurysms and septations were observed. The diagnosis was confirmed with computed tomography. After 7 months of antiarrhythmic therapy, the child underwent surgical intervention by aneurysm resection, atrioseptal defect closure, and ablation. Since then the patient has been in stable sinus rhythm.

Differences in morbidity and mortality in Down syndrome are related to the type of congenital heart defect.

Morbidity and mortality in Down syndrome (DS) are mainly related to congenital heart defects (CHDs). While CHDs with high prevalence in DS (typical CHDs), such as endocardial cushion defects, have been extensively described, little is known about the impact of less common CHDs (atypical CHDs), such as aortic coarctation and univentricular hearts. In our single-center study, we analyzed, in observational, retrospective manner, data regarding cardiac features, surgical management, and outcomes of a cohort of DS patients. Literature review was performed to investigate previously reported studies on atypical CHDs in DS. Patients with CHDs were subclassified as having typical or atypical CHDs. Statistical analysis was performed for comparison between the groups. The study population encompassed 859 DS patients, 72.2% with CHDs, of which 4.7% were atypical. Statistical analysis showed a significant excess in multiple surgeries, all-cause mortality and cardiac mortality in patients with atypical CHDs (p = .0067, p = .0038, p = .0001, respectively). According to the Kaplan-Meier method, survival at 10 and 40 years was significantly higher in typical CHDs (99 and 98% vs. 91 and 84%, log rank <0.05). Among atypical CHDs, it seems that particularly multiple complex defects in univentricular physiology associate with a worse outcome. This may be due to the surgical difficulty in managing univentricular hearts with multiple defects concurring to the clinical picture or to the severity of associated defects themselves. Further studies need to address this specific issue, also considering the higher pulmonary pressures, infective complications, and potential comorbidities in DS patients.

Long-Term Follow-Up of Transthoracic Echocardiography-Guided Transcatheter Closure of Large Atrial Septal Defects (≥ 30 mm) Using the SHSMA Occluder.

Transcatheter closure of large atrial septal defects (ASDs) remains controversial. The aim of this study was to evaluate the feasibility and safety of transthoracic echocardiography (TTE)-guided transcatheter closure of large ASDs. Patients with large secundum ASDs (≥ 30 mm) who underwent device closure were retrospectively reviewed. TTE was performed to guide ASD occluder positioning and assess the immediate and long-term outcomes. A total of 60 patients (median age 43.5 years, range 15-78 years) were enrolled in the study. The median ASD size was 35 mm (range 30-42 mm). Mild to moderate pulmonary hypertension was observed in 36 patients (60%). Thirty-one patients (51.7%) had one short rim, and 18 patients (30.0%) had two deficient rims. Placement of the device was successful in 57 patients (95%), and the median device size was 42 mm (range 40-50 mm). Dislodgement of the device occurred in three patients with two deficient rims: a larger device was redeployed in one case, and two patients required surgical repair. During a median follow-up of 37 months (range 6-83 months), no residual shunts, erosion, or embolization were noted, and pulmonary hypertension resolved in 75% of the patients. Thus t vast majority (95%) of large ASDs can be successfully closed percutaneously using the Chinese-made Shanghai Shape Memory Alloy (SHSMA) occluder under TTE guidance. Long-term follow-up showed that transcatheter closure could become a safe and effective alternative to surgery in select large ASDs.

Tbx5 inhibits hedgehog signaling in determination of digit identity.

Dominant TBX5 mutation causes Holt-Oram syndrome (HOS), which is characterized by limb defects in humans, but the underlying mechanistic basis is unclear. We used a mouse model with Tbx5 conditional knockdown in Hh-receiving cells (marked by Gli1+) during E8 to E10.5, a previously established model to study atrial septum defects, which displayed polydactyly or hypodactyly. The results suggested that Tbx5 is required for digit identity in a subset of limb mesenchymal cells. Specifically, Tbx5 deletion in this cell population decreased cell apoptosis and increased the proliferation of handplate mesenchymal cells. Furthermore, Tbx5 was found to negatively regulate the Hh-signaling activity through transcriptional regulation of Ptch1, a known Hh-signaling repressor. Repression of Hh-signaling through Smo co-mutation in Tbx5 heterozygotes rescued the limb defects, thus placing Tbx5 upstream of Hh-signaling in limb defects. This work reveals an important missing component necessary for understanding not only limb development but also the molecular and genetic mechanisms underlying HOS.

Myocardial overexpression of ANKRD1 causes sinus venosus defects and progressive diastolic dysfunction.

AIMS: Increased Ankyrin Repeat Domain 1 (ANKRD1) levels linked to gain of function mutations have been associated to total anomalous pulmonary venous return and adult cardiomyopathy occurrence in humans. The link between increased ANKRD1 level and cardiac structural and functional disease is not understood. To get insight into this problem, we have generated a gain of function ANKRD1 mouse model by overexpressing ANKRD1 in the myocardium. METHODS AND RESULTS: Ankrd1 is expressed non-homogeneously in the embryonic myocardium, with a dynamic nucleo-sarcomeric localization in developing cardiomyocytes. ANKRD1 transgenic mice present sinus venosus defect, which originates during development by impaired remodelling of early embryonic heart. Adult transgenic hearts develop diastolic dysfunction with preserved ejection fraction, which progressively evolves into heart failure, as shown histologically and haemodynamically. Transgenic cardiomyocyte structure, sarcomeric assembly, and stability are progressively impaired from embryonic to adult life. Postnatal transgenic myofibrils also present characteristic functional alterations: impaired compliance at neonatal stage and impaired lusitropism in adult hearts. Altogether, our combined analyses suggest that impaired embryonic remodelling and adult heart dysfunction in ANKRD1 transgenic mice present a common ground of initial cardiomyocyte defects, which are exacerbated postnatally. Molecular analysis showed transient activation of GATA4-Nkx2.5 transcription in early transgenic embryos and subsequent dynamic transcriptional modulation within titin gene. CONCLUSIONS: ANKRD1 is a fine mediator of cardiomyocyte response to haemodynamic load in the developing and adult heart. Increased ANKRD1 levels are sufficient to initiate an altered cellular phenotype, which is progressively exacerbated into a pathological organ response by the high ventricular workload during postnatal life. Our study defines for the first time a unifying picture for ANKRD1 role in heart development and disease and provides the first mechanistic link between ANKRD1 overexpression and cardiac disease onset.

Biallelic VPS35L pathogenic variants cause 3C/Ritscher-Schinzel-like syndrome through dysfunction of retriever complex.

BACKGROUND: 3C/Ritscher-Schinzel syndrome is characterised by congenital cranio-cerebello-cardiac dysplasia, where CCDC22 and WASHC5 are accepted as the causative genes. In combination with the retromer or retriever complex, these genes play a role in endosomal membrane protein recycling. We aimed to identify the gene abnormality responsible for the pathogenicity in siblings with a 3C/Ritscher-Schinzel-like syndrome, displaying cranio-cerebello-cardiac dysplasia, coloboma, microphthalmia, chondrodysplasia punctata and complicated skeletal malformation. METHODS: Exome sequencing was performed to identify pathogenic variants. Cellular biological analyses and generation of knockout mice were carried out to elucidate the gene function and pathophysiological significance of the identified variants. RESULTS: We identified compound heterozygous pathogenic variants (c.1097dup; p.Cys366Trpfs*28 and c.2755G>A; p.Ala919Thr) in the VPS35L gene, which encodes a core protein of the retriever complex. The identified missense variant lacked the ability to form the retriever complex, and the frameshift variant induced non-sense-mediated mRNA decay, thereby confirming biallelic loss of function of VPS35L. In addition, VPS35L knockout cells showed decreased autophagic function in nutrient-rich and starvation conditions, as well as following treatment with Torin 1. We also generated Vps35l-/- mice and demonstrated that they were embryonic lethal at an early stage, between E7.5 and E10.5. CONCLUSIONS: Our results suggest that biallelic loss-of-function variants in VPS35L underlies 3C/Ritscher-Schinzel-like syndrome. Furthermore, VPS35L is necessary for autophagic function and essential for early embryonic development. The data presented here provide a new insight into the critical role of the retriever complex in fetal development.