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Congenital heart disease (CHD) is the most frequent kind of birth deformity in human beings and the leading cause of neonatal mortality worldwide. Although genetic etiologies encompassing aneuploidy, copy number variations, and mutations in over 100 genes have been uncovered to be involved in the pathogenesis of CHD, the genetic components predisposing to CHD in most cases remain unclear. We recruited a family with CHD from the Chinese Han population in the present investigation. Through whole-exome sequencing analysis of selected family members, a new

Pulmonary arterial hypertension (PAH) refers to a rare, progressive disorder that is characterized by occlusive pulmonary vascular remodeling, resulting in increased pulmonary arterial pressure, right-sided heart failure, and eventual death. Emerging evidence from genetic investigations of pediatric-onset PAH highlights the strong genetic basis underpinning PAH, and deleterious variants in multiple genes have been found to cause PAH. Nevertheless, PAH is of substantial genetic heterogeneity, and the genetic defects underlying PAH in the overwhelming majority of cases remain elusive. In this investigation, a consanguineous family suffering from PAH transmitted as an autosomal-dominant trait was identified. Through whole-exome sequencing and bioinformatic analyses as well as Sanger sequencing analyses of the PAH family, a novel heterozygous SOX17 mutation, NM_022454.4: c.379C>T; p. (Gln127*), was found to co-segregate with the disease in the family, with complete penetrance. The nonsense mutation was neither observed in 612 unrelated healthy volunteers nor retrieved in the population genetic databases encompassing the Genome Aggregation Database, the Exome Aggregation Consortium database, and the Single Nucleotide Polymorphism database. Biological analyses using a dual-luciferase reporter assay system revealed that the Gln127*-mutant SOX17 protein lost the ability to transcriptionally activate its target gene NOTCH1. Moreover, the Gln127*-mutant SOX17 protein exhibited no inhibitory effect on the function of CTNNB1-encode β-catenin, which is a key player in vascular morphogenesis. This research firstly links SOX17 loss-of-function mutation to familial PAH, which provides novel insight into the molecular pathogenesis of PAH, suggesting potential implications for genetic and prognostic risk evaluation as well as personalized prophylaxis of the family members affected with PAH.

Background Atrial fibrillation (AF) represents the most common clinical cardiac arrhythmia and substantially increases the risk of cerebral stroke, heart failure, and death. Although causative genes for AF have been identified, the genetic determinants for AF remain largely unclear. Objective This study aimed to investigate the molecular basis of AF in a Chinese kindred. Methods A 4-generation family with autosomal-dominant AF and other arrhythmias (atrioventricular block, sinus bradycardia, and premature ventricular contractions) was recruited. Genome-wide scan with microsatellite markers and linkage analysis as well as whole-exome sequencing analysis were performed. Electrophysiological characteristics and subcellular localization of the AF-linked mutant were analyzed using dual whole-cell patch clamps and confocal microscopy, respectively. Results A novel genetic locus for AF was mapped to chromosome 17q21.3, a 3.23-cM interval between markers D17S951 and D17S931, with a maximum 2-point logarithm of odds score of 4.2144 at marker D17S1868. Sequencing analysis revealed a heterozygous mutation in the mapping region, NM_005497.4:c.703A>T;p.(M235L), in the GJC1 gene encoding connexin45 (Cx45). The mutation cosegregated with AF in the family and was absent in 632 control individuals. The mutation decreased the coupling conductance in cell pairs (M235L/M235L, M235L/Cx45, M235L/Cx43, and M235L/Cx40), likely because of impaired subcellular localization. Conclusion This study defines a novel genetic locus for AF on chromosome 17q21.3 and reveals a loss-of-function mutation in GJC1 (Cx45) contributing to AF and other cardiac arrhythmias.

The number of confirmed COVID-19 cases has increased drastically; however, information regarding the impact of this disease on the occurrence of arrhythmias is scarce. The aim of this study was to determine the impact of COVID-19 on arrhythmia occurrence. This prospective study included patients with COVID-19 treated at the Leishenshan Temporary Hospital of Wuhan City, China, from February 24 to April 5, 2020. Demographic, comorbidity, and arrhythmias data were collected from patients with COVID-19 (n = 84) and compared with control data from patients with bacterial pneumonia (n = 84) infection. Furthermore, comparisons were made between patients with severe and nonsevere COVID-19 and between older and younger patients. Compared with patients with bacterial pneumonia, those with COVID-19 had higher total, mean, and minimum heart rates (all P < 0.01 ). Patients with severe COVID-19 (severe and critical type diseases) developed more atrial arrhythmias compared with those with nonsevere symptoms. Plasma creatine kinase isoenzyme (CKMB) levels ( P = 0.01 ) were higher in the severe group than in the nonsevere group, and there were more deaths in the severe group than in the nonsevere group (6 (15%) vs. 3 (2.30%); P = 0.05 ). Premature atrial contractions (PAC) and nonsustained atrial tachycardia (NSAT) were significantly positively correlated with plasma CKMB levels but not with high-sensitive cardiac troponin I or myoglobin levels. Our data demonstrate that COVID-19 patients have higher total, mean, and minimum heart rates compared with those with bacterial pneumonia. Patients with severe or critical disease had more frequent atrial arrhythmias (including PAC and AF) and higher CKMB levels and mortality than those with nonsevere symptoms.

Objectives Dilated cardiomyopathy (DCM) represents the most frequent form of cardiomyopathy, leading to heart failure, cardiac arrhythmias and death. Accumulating evidence convincingly demonstrates the crucial role of genetic defects in the pathogenesis of DCM, and over 100 culprit genes have been implicated with DCM. However, DCM is of substantial genetic heterogeneity, and the genetic determinants underpinning DCM remain largely elusive. Methods Whole-exome sequencing and bioinformatical analyses were implemented in a consanguineous Chinese family with DCM. A total of 380 clinically annotated control individuals and 166 more DCM index cases then underwent Sanger sequencing analysis for the identified genetic variation. The functional characteristics of the variant were delineated by utilizing a dual-luciferase assay system. Results A heterozygous variation in the MEF2A gene (encoding myocyte enhancer factor 2A, a transcription factor pivotal for embryonic cardiogenesis and postnatal cardiac adaptation), NM_001365204.1: c.718G>T; p. (Gly240*), was identified, and verified by Sanger sequencing to segregate with autosome-dominant DCM in the family with complete penetrance. The nonsense variation was neither detected in 760 control chromosomes nor found in 166 more DCM probands. Functional analyses revealed that the variant lost transactivation on the validated target genes MYH6 and FHL2, both causally linked to DCM. Furthermore, the variation nullified the synergistic activation between MEF2A and GATA4, another key transcription factor involved in DCM. Conclusions The findings firstly indicate that MEF2A loss-of-function variation predisposes to DCM in humans, providing novel insight into the molecular mechanisms of DCM and suggesting potential implications for genetic testing and prognostic evaluation of DCM patients.

Congenital heart disease (CHD) represents the most frequent developmental deformity in human beings and accounts for substantial morbidity and mortality worldwide. Accumulating investigations underscore the strong inherited basis of CHD, and pathogenic variations in100 genes have been related to CHD. Nevertheless, the heritable defects underpinning CHD remain elusive in most cases, mainly because of the pronounced genetic heterogeneity. In this investigation, a four-generation family with CHD was recruited and clinically investigated. Via whole-exome sequencing and Sanger sequencing assays in selected family members, a heterozygous variation in the SMAD4 gene (coding for a transcription factor essential for cardiovascular morphogenesis), NM_005359.6: c.285T A; p.(Tyr95*), was identified to be in co-segregation with autosomal-dominant CHD in the entire family. The truncating variation was not observed in 460 unrelated non-CHD volunteers employed as control subjects. Functional exploration by dual-reporter gene analysis demonstrated that Tyr95*-mutant SMAD4 lost transactivation of its two key downstream target genes NKX2.5 and ID2, which were both implicated with CHD. Additionally, the variation nullified the synergistic transcriptional activation between SMAD4 and GATA4, another transcription factor involved in CHD. These data strongly indicate SMAD4 may be associated with CHD and shed more light on the molecular pathogenesis underlying CHD, implying potential implications for antenatal precise prevention and prognostic risk stratification of the patients affected with CHD.

Atrial fibrillation (AF) represents the most common type of sustained cardiac arrhythmia in humans and confers a significantly increased risk for thromboembolic stroke, congestive heart failure and premature death. Aggregating evidence emphasizes the predominant genetic defects underpinning AF and an increasing number of deleterious variations in more than 50 genes have been involved in the pathogenesis of AF. Nevertheless, the genetic basis underlying AF remains incompletely understood. In the current research, by whole-exome sequencing and Sanger sequencing analysis in a family with autosomal-dominant AF and congenital patent ductus arteriosus (PDA), a novel heterozygous variation in the PRRX1 gene encoding a homeobox transcription factor critical for cardiovascular development, NM_022716.4:c.373G>T;p.(Glu125*), was identified to be in co-segregation with AF and PDA in the whole family. The truncating variation was not detected in 306 unrelated healthy individuals employed as controls. Quantitative biological measurements with a reporter gene analysis system revealed that the Glu125*-mutant PRRX1 protein failed to transactivate its downstream target genes SHOX2 and ISL1, two genes that have been causally linked to AF. Conclusively, the present study firstly links PRRX1 loss-of-function variation to AF and PDA, suggesting that AF and PDA share a common abnormal developmental basis in a proportion of cases.

Recently, mutations in the Kruppel-like factor 13 (

As the most common form of developmental malformation affecting the heart and endothoracic great vessels, congenital heart disease (CHD) confers substantial morbidity and mortality as well as socioeconomic burden on humans globally. Aggregating convincing evidence highlights the genetic origin of CHD, and damaging variations in over 100 genes have been implicated with CHD. Nevertheless, the genetic basis underpinning CHD remains largely elusive. In this study, via whole-exosome sequencing analysis of a four-generation family inflicted with autosomal-dominant CHD, a heterozygous SMAD1 variation, NM_005900.3: c.264C > A; p.(Tyr88 ∗ ), was detected and validated by Sanger sequencing analysis to be in cosegregation with CHD in the whole family. The truncating variation was not observed in 362 unrelated healthy volunteers employed as control persons. Dual-luciferase reporter gene assay in cultured COS7 cells demonstrated that Tyr88 ∗ -mutant SMAD1 failed to transactivate the genes TBX20 and NKX2.5, two already well-established CHD-causative genes. Additionally, the variation nullified the synergistic transcriptional activation between SMAD1 and MYOCD, another recognized CHD-causative gene. These data indicate SMAD1 as a new gene responsible for CHD, which provides new insight into the genetic mechanism underlying CHD, suggesting certain significance for genetic risk assessment and precise antenatal prevention of the family members inflicted with CHD.

Background Atrial fibrillation (AF) is the most common form of clinical cardiac dysrhythmia responsible for thromboembolic cerebral stroke, congestive heart failure, and death. Aggregating evidence highlights the strong genetic basis of AF. Nevertheless, AF is of pronounced genetic heterogeneity, and in an overwhelming majority of patients, the genetic determinants underpinning AF remain elusive. Methods and Results By genome‐wide screening with polymorphic microsatellite markers and linkage analysis in a 4‐generation Chinese family affected with autosomal‐dominant AF, a novel locus for AF was mapped to chromosome 1q24.2–q25.1, a 3.20‐cM (≈4.19 Mbp) interval between markers D1S2851 and D1S218, with the greatest 2‐point logarithm of odds score of 4.8165 for the marker D1S452 at recombination fraction=0.00. Whole‐exome sequencing and bioinformatics analyses showed that within the mapping region, only the mutation in the paired related homeobox 1 ( PRRX1 ) gene, NM_022716.4:c.319C>T;(p.Gln107*), cosegregated with AF in the family. In addition, sequencing analyses of PRRX1 in another cohort of 225 unrelated patients with AF revealed a new mutation, NM_022716.4:c.437G>T; (p.Arg146Ile), in a patient. The 2 mutations were absent in 908 control subjects. Biological analyses in HeLa cells demonstrated that the 2 mutants had significantly diminished transactivation on the target genes ISL1 and SHOX2 and markedly decreased ability to bind the promoters of ISL1 and SHOX2 (2 genes causally linked to AF), although with normal intracellular distribution. Conclusions This study first indicates that PRRX1 loss‐of‐function mutations predispose to AF, which provides novel insight into the molecular pathogenesis underpinning AF, implying potential implications for precisive prophylaxis and management of AF.

Congenital heart defect (CHD) represents the most common birth deformity, afflicting 1% of all births worldwide, and accounts for substantial morbidity and mortality. Increasing evidence highlights the pivotal roles of genetic etiologies in the pathogenesis of CHD, and pathogenic mutations in multiple genes, including TBX5 encoding a cardiac core transcription factor key to cardiovascular morphogenesis, have been involved in CHD. However, due to pronounced genetic heterogeneity of CHD, the genetic determinants underlying CHD in most cases remain obscure. In this investigation, by sequencing analysis of the coding exons and flanking introns of the TBX5 gene in 198 unrelated patients affected with CHD, a novel heterozygous mutation, NM_000192.3: c.692C>T; p. (Pro231Leu), was identified in an index patient with familial double outlet right ventricle (DORV), ventricular septal defect (VSD), and atrioventricular block (AVB). Genetic analysis of the proband's pedigree showed that the mutation co-segregated with the diseases. The missense mutation, which altered the amino acid conserved evolutionarily, was absent from 266 unrelated healthy subjects. Functional analyses with a dual-luciferase reporter assay system unveiled that the Pro231Leu-mutant TBX5 was associated with significantly reduced transcriptional activity on its target genes MYH6 and NPPA. Furthermore, the mutation disrupted the synergistic transactivation between TBX5 and NKX2-5 as well as GATA4, two other transcription factors causally linked to CHD. This study firstly links TBX5 loss-of-function mutation to familial DORV, VSD, and AVB, which provides novel insight into the mechanism underpinning CHD and AVB, suggesting potential implications for genetic evaluation and individualized treatment of patients affected by CHD and AVB.

Long noncoding RNAs (lncRNAs) are an emerging class of RNA species that could participate in some critical pathways and disease pathogenesis. However, the underlying molecular mechanism of lncRNAs in atrial fibrillation (AF) is still not fully understood. In the present study, we analyzed RNA-seq data of paired left and right atrial appendages from five patients with AF and other five patients without AF. Based on the gene expression profiles of 20 samples, we found that a majority of genes were aberrantly expressed in both left and right atrial appendages of patients with AF. Similarly, the dysregulated pathways in the left and right atrial appendages of patients with AF also bore a close resemblance. Moreover, we predicted regulatory lncRNAs that regulated the expression of adjacent protein-coding genes (PCGs) or interacted with proteins. We identified that NPPA and its antisense RNA NPPA-AS1 may participate in the pathogenesis of AF by regulating the muscle contraction. We also identified that RP11 - 99E15.2 and RP3 - 523K23.2 could interact with proteins ITGB3 and HSF2, respectively. RP11 - 99E15.2 and RP3 - 523K23.2 may participate in the pathogenesis of AF via regulating the extracellular matrix binding and the transcription of HSF2 target genes, respectively. The close association of the lncRNA-interacting proteins with AF further demonstrated that these two lncRNAs were also associated with AF. In conclusion, we have identified key regulatory lncRNAs implicated in AF, which not only improves our understanding of the lncRNA-related molecular mechanism underlying AF but also provides computationally predicted regulatory lncRNAs for AF researchers.

There are significant differences in clinical presentation and treatment of atrial fibrillation (AF) between women and men. The primary goal of AF management is to restore sinus rhythm and to prevent various complications, including stroke and heart failure. In many areas of AF, such as prevalence, clinical manifestations, morbidity, risk factors, pathophysiology, treatment strategies, and complications, gender-specific variability is observed and needs to be further addressed by large-scale population researches or randomized clinical trials, which help to promote the customization of AF treatment programs, hence to maximize the success rate of AF therapy in both sexes. This review highlights our current understanding of these gender differences in AF and how these differences affect treatment decisions on AF.

Introduction: As the most frequent type of birth defect in humans, congenital heart disease (CHD) leads to a large amount of morbidity and mortality as well as a tremendous socioeconomic burden. Accumulating studies have convincingly substantiated the pivotal roles of genetic defects in the occurrence of familial CHD, and deleterious variations in a great number of genes have been reported to cause various types of CHD. However, owing to pronounced genetic heterogeneity, the hereditary components underpinning CHD remain obscure in most cases. This investigation aimed to identify novel genetic determinants underlying CHD. Methods and results: A four-generation pedigree with high incidence of autosomal-dominant CHD was enrolled from the Chinese Han race population. Using whole-exome sequencing and Sanger sequencing assays of the family members available, a novel SOX7 variation in heterozygous status, NM_031439.4: c.310C>T; p.(Gln104*), was discovered to be in co-segregation with the CHD phenotype in the whole family. The truncating variant was absent in 500 unrelated healthy subjects utilized as control individuals. Functional measurements by dual-luciferase reporter analysis revealed that Gln104*-mutant SOX7 failed to transactivate its two important target genes, GATA4 and BMP2, which are both responsible for CHD. In addition, the nonsense variation invalidated the cooperative transactivation between SOX7 and NKX2.5, which is another recognized CHD-causative gene. Conclusion: The present study demonstrates for the first time that genetically defective SOX7 predisposes to CHD, which sheds light on the novel molecular mechanism underpinning CHD, and implies significance for precise prevention and personalized treatment in a subset of CHD patients.

Congenital heart defect (CHD) is the most common form of birth deformity and is responsible for substantial morbidity and mortality in humans. Increasing evidence has convincingly demonstrated that genetic defects play a pivotal role in the pathogenesis of CHD. However, CHD is a genetically heterogeneous disorder and the genetic basis underpinning CHD in the vast majority of cases remains elusive. This study was sought to identify the pathogenic mutation in the ISL1 gene contributing to CHD. A cohort of 210 unrelated patients with CHD and a total of 256 unrelated healthy individuals used as controls were registered. The coding exons and splicing boundaries of ISL1 were sequenced in all study subjects. The functional effect of an identified ISL1 mutation was evaluated using a dual-luciferase reporter assay system. A novel heterozygous ISL1 mutation, c.409G > T or p.E137X, was identified in an index patient with congenital patent ductus arteriosus and ventricular septal defect. Analysis of the proband’s pedigree revealed that the mutation co-segregated with CHD, which was transmitted in the family in an autosomal dominant pattern with complete penetrance. The nonsense mutation was absent in 512 control chromosomes. Functional analysis unveiled that the mutant ISL1 protein failed to transactivate the promoter of MEF2C, alone or in synergy with TBX20. This study firstly implicates ISL1 loss-of-function mutation with CHD in humans, which provides novel insight into the molecular mechanism of CHD, implying potential implications for genetic counseling and individually tailored treatment of CHD patients.

Atrial fibrillation (AF), as the most common sustained cardiac arrhythmia, is associated with substantially increased morbidity and mortality. Aggregating evidence demonstrates that genetic defects play a crucial role in the pathogenesis of AF, especially in familial AF. Nevertheless, AF is of pronounced genetic heterogeneity, and in an overwhelming majority of cases the genetic determinants underlying AF remain elusive. In the current study, 162 unrelated patients with familial AF and 238 unrelated healthy individuals served as controls were recruited. The coding exons and splicing junction sites of the SHOX2 gene, which encodes a homeobox-containing transcription factor essential for proper development and function of the cardiac conduction system, were sequenced in all study participants. The functional effect of the mutant SHOX2 protein was characterized with a dual-luciferase reporter assay system. As a result, a novel heterozygous SHOX2 mutation, c.580C>T or p.R194X, was identified in an index patient, which was absent from the 476 control chromosomes. Genetic analysis of the proband's pedigree revealed that the nonsense mutation co-segregated with AF in the family with complete penetrance. Functional assays demonstrated that the mutant SHOX2 protein had no transcriptional activity compared with its wild-type counterpart. In conclusion, this is the first report on the association of SHOX2 loss-of-function mutation with enhanced susceptibility to familial AF, which provides novel insight into the molecular mechanism underpinning AF, suggesting potential implications for genetic counseling and individualized management of AF patients.

As the most prevalent form of human birth defect, congenital heart disease (CHD) contributes to substantial morbidity, mortality and socioeconomic burden worldwide. Aggregating evidence has convincingly demonstrated that genetic defects exert a pivotal role in the pathogenesis of CHD, and causative mutations in multiple genes have been causally linked to CHD. Nevertheless, CHD is of pronounced genetic heterogeneity, and the genetic components underpinning CHD in the overwhelming majority of patients remain obscure. In this research, a four-generation consanguineous family suffering from CHD transmitted in an autosomal dominant mode was recruited. By whole-exome sequencing and bioinformatics analyses as well as Sanger sequencing analyses of the family members, a new heterozygous SOX17 variation, NM_022454.4: c.553G > T; p.(Glu185*), was identified to co-segregate with CHD in the family, with complete penetrance. The nonsense variation was neither detected in 310 unrelated healthy volunteers used as controls nor retrieved in such population genetics databases as the Exome Aggregation Consortium database, Genome Aggregation Database, and the Single Nucleotide Polymorphism database. Functional assays by utilizing a dual-luciferase reporter assay system unveiled that the Glu185*-mutant SOX17 protein had no transcriptional activity on its two target genes NOTCH1 and GATA4, which have been reported to cause CHD. Furthermore, the mutation abrogated the synergistic transactivation between SOX17 and NKX2.5, another established CHD-causing transcription factor. These findings firstly indicate SOX17 loss-of-function mutation predisposes to familial CHD, which adds novel insight to the molecular mechanism of CHD, implying potential implications for genetic risk appraisal and individualized prophylaxis of the family members affected with CHD.

TBX5 has been linked to Holt-Oram syndrome, with congenital heart defect (CHD) and atrial fibrillation (AF) being two major cardiac phenotypes. However, the prevalence of a TBX5 variation in patients with CHD and AF remains obscure. In this research, by sequencing analysis of TBX5 in 178 index patients with both CHD and AF, a novel heterozygous variation, NM_000192.3: c.577G>T; p.(Gly193*), was identified in one index patient with CHD and AF as well as bicuspid aortic valve (BAV), with an allele frequency of approximately 0.28%. Genetic analysis of the proband’s pedigree showed that the variation co-segregated with the diseases. The pathogenic variation was not detected in 292 unrelated healthy subjects. Functional analysis by using a dual-luciferase reporter assay system showed that the Gly193*-mutant TBX5 protein failed to transcriptionally activate its target genes MYH6 and NPPA. Moreover, the mutation nullified the synergistic transactivation between TBX5 and GATA4 as well as NKX2-5. Additionally, whole-exome sequencing analysis showed no other genes contributing to the diseases. This investigation firstly links a pathogenic variant in the TBX5 gene to familial CHD and AF as well as BAV, suggesting that CHD and AF as well as BAV share a common developmental basis in a subset of patients.

Dilated cardiomyopathy (DCM) represents the most frequent form of cardiomyopathy, leading to heart failure, cardiac arrhythmias and death. Accumulating evidence convincingly demonstrates the crucial role of genetic defects in the pathogenesis of DCM, and over 100 culprit genes have been implicated with DCM. However, DCM is of substantial genetic heterogeneity, and the genetic determinants underpinning DCM remain largely elusive.Whole-exome sequencing and bioinformatical analyses were implemented in a consanguineous Chinese family with DCM. A total of 380 clinically annotated control individuals and 166 more DCM index cases then underwent Sanger sequencing analysis for the identified genetic variation. The functional characteristics of the variant were delineated by utilizing a dual-luciferase assay system.A heterozygous variation in theThe findings firstly indicate that

Congenital heart defect (CHD) represents the most common birth deformity, afflicting 1% of all births worldwide, and accounts for substantial morbidity and mortality. Increasing evidence highlights the pivotal roles of genetic etiologies in the pathogenesis of CHD, and pathogenic mutations in multiple genes, including TBX5 encoding a cardiac core transcription factor key to cardiovascular morphogenesis, have been involved in CHD. However, due to pronounced genetic heterogeneity of CHD, the genetic determinants underlying CHD in most cases remain obscure. In this investigation, by sequencing analysis of the coding exons and flanking introns of the TBX5 gene in 198 unrelated patients affected with CHD, a novel heterozygous mutation, NM_000192.3: c.692CT; p. (Pro231Leu), was identified in an index patient with familial double outlet right ventricle (DORV), ventricular septal defect (VSD), and atrioventricular block (AVB). Genetic analysis of the proband's pedigree showed that the mutation co-segregated with the diseases. The missense mutation, which altered the amino acid conserved evolutionarily, was absent from 266 unrelated healthy subjects. Functional analyses with a dual-luciferase reporter assay system unveiled that the Pro231Leu-mutant TBX5 was associated with significantly reduced transcriptional activity on its target genes MYH6 and NPPA. Furthermore, the mutation disrupted the synergistic transactivation between TBX5 and NKX2-5 as well as GATA4, two other transcription factors causally linked to CHD. This study firstly links TBX5 loss-of-function mutation to familial DORV, VSD, and AVB, which provides novel insight into the mechanism underpinning CHD and AVB, suggesting potential implications for genetic evaluation and individualized treatment of patients affected by CHD and AVB.

Congenital bicuspid aortic valve (BAV), the most common form of birth defect in humans, is associated with substantial morbidity and mortality. Increasing evidence demonstrates that genetic risk factors play a key role in the pathogenesis of BAV. However, BAV is a genetically heterogeneous disease and the genetic determinants underpinning BAV in an overwhelming majority of patients remain unknown. In the present study, the coding exons and flanking introns of the GATA6 gene, which encodes a zinc-finger transcription factor essential for the normal development of the aortic valves, were sequenced in 152 unrelated patients with congenital BAV. The available relatives of a proband harboring an identified GATA6 mutation and 200 unrelated, ethnically matched healthy individuals used as controls were also genotyped for GATA6. The functional characteristics of the mutation were analyzed by using a dual-luciferase reporter assay system. As a result, a novel heterozygous GATA6 mutation, p.E386X, was identified in a family with BAV transmitted in an autosomal dominant mode. The nonsense mutation was absent in 400 control chromosomes. Biological assays revealed that the mutant GATA6 protein had no transcriptional activity compared with its wild-type counterpart. Furthermore, the mutation disrupted the synergistic transcriptional activation between GATA6 and GATA4, another transcription factor causally linked to BAV. In conclusion, this study firstly associates GATA6 loss-of-function mutation with enhanced susceptibility to familial BAV, which provides novel insight into the molecular mechanism of BAV, implying potential implications for genetic counseling and personalized management of BAV patients.

As a prevalent primary myocardial disease, dilated cardiomyopathy (DCM) represents the most common cause of heart failure in the young and the most frequent indication for cardiac transplantation. Aggregating evidence highlights the genetic basis of DCM. However, due to substantial genetic heterogeneity, the genetic defects of DCM in most cases remain elusive. In the current investigation, the entire coding exons and splicing junctions of the KLF5 gene, which encodes a key transcription factor required for cardiac structural and functional remodeling, were sequenced in 234 probands affected with DCM, and a heterozygous KLF5 mutation, NM_001730.5: c.1100T > A; p.(Leu367*), was identified in a proband. Genetic analysis of the proband's family members revealed that the identified KLF5 mutation co-segregated with DCM in the family with complete penetrance. The nonsense mutation was neither detected in 506 control individuals nor reported in such population-genetics databases as ExAC, dbSNP and gnomAD. Biological assays with a dual-luciferase reporter assay system demonstrated that the mutant KLF5 protein had no transcriptional activity when compared with its wild-type counterpart. Furthermore, the mutation abrogated the synergistic transactivation between KLF5 and NFKB1, another pivotal transcription factor that has been causally linked to DCM. The whole-exome sequencing analysis of the proband's family members revealed no other causative genes. The findings indicate KLF5 as a new gene contributing to DCM in humans, implying potential implications for the precision medicine of DCM.

Occurring in about 1% of all live births, congenital heart defects (CHDs) represent the most frequent type of developmental abnormality and account for remarkably increased infant morbidity and mortality. Aggregating studies demonstrate that genetic components have a key role in the occurrence of CHDs. Nevertheless, due to pronounced genetic heterogeneity, the genetic causes of CHDs remain unclear in most patients. In this research, 114 unrelated patients affected with CHDs and 218 unrelated individuals without CHDs served as controls were recruited. The coding regions and splicing donors/acceptors of the ISL1 gene, which codes for a transcription factor required for proper cardiovascular development, were screened for mutations by sequencing in all study participants. The functional characteristics of an identified ISL1 mutation were delineated with a dual-luciferase reporter assay system. As a result, a new heterozygous ISL1 mutation, NM_002202.2: c.225CG; p. (Tyr75*), was discovered in an index patient with double outlet right ventricle and ventricular septal defect. Analysis of the proband's family unveiled that the mutation co-segregated with the CHD phenotype. The nonsense mutation was absent in the 436 control chromosomes. Biological analysis showed that the mutant ISL1 protein had no transcriptional activity. Furthermore, the mutation nullified the synergistic activation between ISL1 and TBX20, another CHD-associated transcription factor. This research for the first time links an ISL1 loss-of-function mutation to double outlet right ventricle in humans, which adds insight to the molecular pathogenesis underpinning CHDs, suggesting potential implications for timely personalized management of CHD patients.

Congenital heart defect (CHD) is the most common type of birth defect in humans and a leading cause of infant morbidity and mortality. Previous studies have demonstrated that genetic defects play a pivotal role in the pathogenesis of CHD. However, the genetic basis of CHD remains poorly understood due to substantial genetic heterogeneity. In this study, the coding exons and splicing boundaries of the NR2F2 gene, which encodes a pleiotropic transcription factor required for normal cardiovascular development, were sequenced in 168 unrelated patients with CHD, and a novel mutation (c.247G > T, equivalent to p.G83X) was detected in a patient with double outlet right ventricle as well as ventricular septal defect. Genetic scanning of the mutation carrier's relatives available showed that the mutation was present in all affected family members but absent in unaffected family members. Analysis of the index patient's pedigree displayed that the mutation co-segregated with CHD, which was transmitted as an autosomal dominant trait with complete penetrance. The nonsense mutation was absent in 230 unrelated, ethnically-matched healthy individuals used as controls. Functional deciphers by using a dual-luciferase reporter assay system revealed that the mutant NR2F2 protein had no transcriptional activity as compared with its wild-type counterpart. Furthermore, the mutation abrogated the synergistic transcriptional activation between NR2F2 and GATA4, another core cardiac transcription factor associated with CHD. This study firstly associates NR2F2 loss-of-function mutation with an increased susceptibility to double outlet right ventricle in humans, which provides further significant insight into the molecular mechanisms underpinning CHD, suggesting potential implications for genetic counseling of CHD families and personalized treatment of CHD patients.

Aggregating evidence suggests that genetic determinants play a pivotal role in the pathogenesis of the congenitally bicuspid aortic valve (BAV). BAV is of pronounced genetic heterogeneity, and the genetic components underlying BAV in an overwhelming majority of patients remain elusive. In the current study, the whole coding exons and adjacent introns, as well as 5′ and 3′ untranslated regions of the GATA4 gene, which codes for a zinc-finger transcription factor crucial for the normal development of the aortic valve, were screened by direct sequencing in 150 index patients with congenital BAV. The available family members of an identified mutation carrier and 300 unrelated, ethnically matched healthy individuals used as controls were also genotyped for GATA4 . The functional effect of the mutation was characterized using a dual-luciferase reporter assay system. As a result, a novel heterozygous GATA4 mutation, p.E147X, was identified in a family with BAV transmitted in an autosomal dominant pattern. The nonsense mutation was absent in 600 control chromosomes. Functional deciphers revealed that the mutant GATA4 protein lost transcriptional activity compared with its wild-type counterpart. Furthermore, the mutation disrupted the synergistic transcriptional activation between GATA4 and NKX2.5, another transcription factor responsible for BAV. In conclusion, this study associates the GATA4 loss-of-function mutation with enhanced susceptibility to a BAV, thus providing novel insight into the molecular mechanism underpinning the BAV.

Atrial fibrillation (AF) represents the most common type of clinical cardiac arrhythmia and substantially increases the risks of cerebral stroke, heart failure and death. Accumulating evidence has convincingly demonstrated the strong genetic basis of AF, and an increasing number of pathogenic variations in over 50 genes have been causally linked to AF. Nevertheless, AF is of pronounced genetic heterogeneity, and the genetic determinants underpinning AF in most patients remain obscure. In the current investigation, a Chinese pedigree with AF as well as ventricular arrhythmias and hypertrophic cardiomyopathy was recruited. Whole exome sequencing and bioinformatic analysis of the available family members were conducted, and a novel heterozygous variation in the KLF15 gene (encoding Krüppel-like factor 15, a transcription factor critical for cardiac electrophysiology and structural remodeling), NM_014079.4: c.685A&gt, T, p.(Lys229*), was identified. The variation was verified by Sanger sequencing and segregated with autosomal dominant AF in the family with complete penetrance. The variation was absent from 300 unrelated healthy subjects used as controls. In functional assays using a dual-luciferase assay system, mutant KLF15 showed neither transcriptional activation of the KChIP2 promoter nor transcriptional inhibition of the CTGF promoter, alone or in the presence of TGFB1, a key player in the pathogenesis of arrhythmias and cardiomyopathies. The findings indicate KLF15 as a new causative gene responsible for AF as well as ventricular arrhythmias and hypertrophic cardiomyopathy, and they provide novel insight into the molecular mechanisms underlying cardiac arrhythmias and hypertrophic cardiomyopathy.

The Ca2+-sensitive cardiac troponin (cTn) is a hetero-trimer complex consisting of three subunits cTnC, cTnI, and cTnT, which has been recognized as an important biomarker and a potential target of cardiovascular diseases. Previously, several small-molecule agents such as levosimendan and pimobendan have been successfully developed to target this protein for the treatment of heart failure. Here, instead of small-molecule chemical drugs, we purposed rational derivation of self-inhibitory peptides as potential biologic disruptors of cTnC–cTnI interaction from the interaction complex interface. In the procedure, the crystal structure of cTn trimer was examined in detail using bioinformatics approach, from which a peptide-mediated interaction between the N-terminal domain of cTnC and the switch region of cTnI was identified. The switch is a 19-mer peptide segment Swt that contains a structured helical core capped by a short N-terminal tripeptide and a disordered C-terminal tail. Structural and energetic analysis revealed that the Swt peptide binds independently to cTnC N-terminal domain, which can be stripped from the intact cTnI subunit to interact effectively with cTnC. Further investigations found that truncation of two N-terminal residues and five C-terminal residues of the full-length Swt peptide, resulting in a shortened version namely Swt-ΔN2ΔC5 peptide, would not cause substantial loss in its binding potency to cTnC. The computational finding was then confirmed by using fluorescence-based affinity assays; the Swt and Swt-ΔN2ΔC5 peptides was experimentally measured to have a moderately high affinity to the recombinant protein of human cTnC N-terminal domain with K d values at micromolar level. The Swt and Swt-ΔN2ΔC5 are considered as inhibitory peptides that can be further optimized and modified to obtain high-affinity disruptors of cTnI–cTnC interaction.

Dilated cardiomyopathy (DCM), the most common form of primary myocardial disease, is a leading cause of congestive heart failure and the most common indication for heart transplantation. Recently, NKX2-5 mutations have been involved in the pathogenesis of familial DCM. However, the prevalence and spectrum of NKX2-5 mutations associated with sporadic DCM remain to be evaluated. In this study, the coding regions and flanking introns of the NKX2-5 gene, which encodes a cardiac transcription factor pivotal for cardiac development and structural remodeling, were sequenced in 210 unrelated patients with sporadic adult-onset DCM. A total of 300 unrelated healthy individuals used as controls were also genotyped for NKX2-5. The functional effect of the mutant NKX2-5 was investigated using a dual-luciferase reporter assay system. As a result, two novel heterozygous NKX2-5 mutations, p.R139W and p.E167X, were identified in 2 unrelated patients with sporadic adult-onset DCM, with a mutational prevalence of approximately 0.95%. The mutations were absent in 600 referential chromosomes and the altered amino acids were completely conserved evolutionarily across species. Functional assays revealed that the NKX2-5 mutants were associated with significantly reduced transcriptional activity. Furthermore, the mutations abrogated the synergistic activation between NKX2-5 and GATA4 as well as TBX20, two other cardiac key transcription factors that have been causally linked to adult-onset DCM. This study is the first to associate NKX2-5 loss-of-function mutations with enhanced susceptibility to sporadic DCM, which provides novel insight into the molecular etiology underpinning DCM, and suggests the potential implications for the genetic counseling and personalized treatment of the DCM patients.

As the most common form of birth defect in humans, congenital heart disease (CHD) is associated with substantial morbidity and mortality in both children and adults. Increasing evidence demonstrates that genetic defects play a pivotal role in the pathogenesis of CHD. However, CHD is of great heterogeneity, and in an overwhelming majority of cases, the genetic determinants underpinning CHD remain elusive. In the present investigation, the coding exons and flanking introns of the CASZ1 gene, which codes for a zinc finger transcription factor essential for the cardiovascular morphogenesis, were sequenced in 172 unrelated patients with CHD. As a result, a novel heterozygous CASZ1 mutation, p.L38P, was identified in an index patient with congenital ventricular septal defect (VSD). Genetic scanning of the mutation carrier's available family members revealed that the mutation was present in all affected patients but absent in unaffected individuals. Analysis of the proband's pedigree showed that the mutation co-segregated with VSD, which was transmitted as an autosomal dominant trait with complete penetrance. The missense mutation, which altered the amino acid that was highly conserved evolutionarily, was absent in 200 unrelated, ethnically-matched healthy subjects used as controls. Functional deciphers by using a dual-luciferase reporter assay system unveiled that the mutant CASZ1 had significantly reduced transcriptional activity as compared with its wild-type counterpart. To the best of our knowledge, the current study firstly identifies CASZ1 as a new gene predisposing to CHD in humans, which provides novel insight into the molecular mechanisms underlying CHD and a potential therapeutic target for CASZ1-associated CHD, suggesting potential implications for personalized prophylaxis and therapy of CHD.

Aim This study aimed to investigate the effect of 1,25-dihydroxy-vitamin D3 (1,25(OH)2D3) on primary chondrocytes cultured from patients with osteoarthritis (OA). Method Primary chondrocytes isolated from the tibial plateau of female OA patients were characterized by immunocytochemistry analysis. Using Cell Counting Kit-8 (CCK-8), cell viability was measured to select suitable 1,25(OH)2D3 concentrations for treating chondrocytes. RNA-sequencing was performed on primary chondrocytes treated with or without 1,25(OH)2D3. Differentially expressed genes (DEGs) as well as gene ontology (GO)-biological process (BP) and pathways affected by 1,25(OH)2D3 were identified. Protein-protein interaction (PPI) network was constructed, and the hub nodes in the PPI network were identified. qRT-PCR was conducted to confirm the expression levels of six DEGs. Results Positive collagen II staining confirmed the successful isolation of primary chondrocytes. CCK-8 assay showed maximal primary chondrocyte survival rate when treated with 10-5 μmol/L of 1,25(OH)2D3 for 72 h. RNA-sequencing results identified a total of 1036 DEGs, including 593 upregulated and 443 downregulated genes from 1,25(OH)2D3 treated and untreated cells. Further functional enrichment analyses showed the association of these DEGs with GO-BP terms such as response to the stimulus, cell proliferation, angiogenesis, and regulation of cell motility, and KEGG pathways, including TNF signaling pathway, IL-17 signaling pathway, cytokine-cytokine receptor interaction, and NF-kappa B signaling pathway. PPI network identified UBC, FOS, IFIT1, CDK1, and ISG15 as the hub nodes in the network. qRT-PCR results were in alignment with the results of RNA-sequencing. Conclusion Our study might provide a theoretical basis for the use of vitamin D in treating OA.

Congenital bicuspid aortic valve (BAV) represents the most common type of cardiac birth defect affecting 0.4‑2% of the general population, and accounts for a markedly increased incidence of life‑threatening complications, including valvulopathy and aortopathy. Accumulating evidence has demonstrated the genetic basis of BAV. However, the genetic basis for BAV in the majority of cases remains to be elucidated. In the present study, the coding regions and splicing donors/acceptors of the nuclear receptor subfamily 2 group F member 2 (NR2F2) gene, which encodes a transcription factor essential for proper cardiovascular development, were sequenced in 176 unrelated cases of congenital BAV. The available family members of the proband carrying an identified NR2F2 mutation and 280 unrelated, sex‑ and ethnicity‑matched healthy individuals as controls were additionally genotyped for NR2F2. The functional effect of the mutation was characterized using a dual‑luciferase reporter assay system. As a result, a novel heterozygous NR2F2 mutation, NM_021005.3: c.288C>A; p.(Cys96*), was identified in a family with BAV, which was transmitted in an autosomal dominant mode with complete penetrance. The nonsense mutation was absent from the 560 control chromosomes. Functional analysis identified that the mutant NR2F2 protein had no transcriptional activity. Furthermore, the mutation disrupted the synergistic transcriptional activation between NR2F2 and transcription factor GATA‑4, another transcription factor that is associated with BAV. These findings suggested NR2F2 as a novel susceptibility gene of human BAV, which reveals a novel molecular pathogenesis underpinning BAV.

Atrial fibrillation (AF) represents the most frequent form of sustained cardiac rhythm disturbance, affecting approximately 1% of the general population worldwide, and confers a substantially enhanced risk of cerebral stroke, heart failure, and death. Increasing epidemiological studies have clearly demonstrated a strong genetic basis for AF, and variants in a wide range of genes, including those coding for ion channels, gap junction channels, cardiac structural proteins and transcription factors, have been identified to underlie AF. Nevertheless, the genetic pathogenesis of AF is complex and still far from completely understood. Here, whole-exome sequencing and bioinformatics analyses of a three-generation family with AF were performed, and after filtering variants by multiple metrics, we identified a heterozygous variant in the ISL1 gene (encoding a transcription factor critical for embryonic cardiogenesis and postnatal cardiac remodeling), NM_002202.2: c.481G > T; p.(Glu161*), which was validated by Sanger sequencing and segregated with autosome-dominant AF in the family with complete penetrance. The nonsense variant was absent from 284 unrelated healthy individuals used as controls. Functional assays with a dual-luciferase reporter assay system revealed that the truncating ISL1 protein lost transcriptional activation on the verified target genes MEF2C and NKX2-5. Additionally, the variant nullified the synergistic transactivation between ISL1 and TBX5 as well as GATA4, two other transcription factors that have been implicated in AF. The findings suggest ISL1 as a novel gene contributing to AF, which adds new insight to the genetic mechanisms underpinning AF, implying potential implications for genetic testing and risk stratification of the AF family members.

Objectives: This study will evaluate the results of parameters measured during a treadmill exercise test to predict marathon performances. Methods: We studied 171 Chinese recreational athletes who participated in marathons or half-marathons (42.2[Formula: see text]k or 21.1[Formula: see text]k, respectively) between October 2016 and December 2017. The participants completed a survey that included questions about demographics and training, and they underwent a treadmill exercise test according to the Bruce protocol. The number of years in training, mean weekly hours of training, mean weekly training volume, and performance time in subsequent marathon events were recorded and analyzed in this study. Results: The total exercise times achieved on the treadmill test were significantly longer for men compared to women ([Formula: see text]). The performance times in the half-marathons were significantly shorter for men compared to women ([Formula: see text]). Training volume was the only independent predictor of total exercise time on the treadmill and performance time in marathons and half-marathons (all [Formula: see text]). The value of the total exercise time on the treadmill to predict performance times in half-marathons ([Formula: see text]) was superior to full marathons ([Formula: see text]) and significantly better in study subjects aged 30–39 years ([Formula: see text], [Formula: see text]) and 40–49 years ([Formula: see text], [Formula: see text]) compared to study subjects aged 20–29 years and 50–59 years. The percentage of decrease in the maximal heart rate (MHR) at the end of one minute of recovery time was negatively correlated with performance times in marathons. Conclusions: The total exercise time achieved during an exhaustive treadmill exercise test and percentage of decrease in the MHR at the end of one minute of recovery time are accessible parameters that can help athletes manage their expectations and adjust their training plans. A large study that includes additional countries is needed to confirm the value of treadmill exercise test results for predicting marathon performance.