Methylglyoxal accumulation contributes to accelerated brain aging in spontaneously hypertensive rats.

While it is well-acknowledged that neurovascular dysfunction in hypertension is tightly associated with accelerated brain aging, we contend that the deleterious effects of hypertension may extend beyond affecting only the arteries. Methylglyoxal (MG) derived from glycolysis, is involved in the accumulation of advanced glycated end products (AGEs), which are the hallmarks of neurodegenerative disorders. Therefore, the present study aims to firstly investigate the role of MG metabolism in the hypertension-accelerated brain aging process. The results of our study indicate that the levels of MG increase with age in both the plasma and hippocampus of SHRs at 12, 16, and 30 weeks old. AGE methylglyoxal-hydro imidazoline-1 (MG-H1) is primarily localized in astrocytes, while its presence was not observed in neurons and microglia within the hypertensive hippocampus. Our observations also suggest that angiotensin II (Ang II) enhances glucose uptake and glycolysis while reducing the expression of Glo1 in cultured astrocytes. N-acetylcysteine (NAC) was found to counteract the increase in escape latency and inhibit the activation of the AGEs-RAGE axis in 30-week-old SHRs. NAC decreased Iba-1 immunofluorescence intensity, inhibited the levels of pro-inflammatory markers, and enhanced the abundance of anti-inflammatory markers in the hippocampus of SHRs. Moreover, NAC reduced the immunofluorescence signal of 4HNE and increased the content of GSH and SOD in SHRs. Finally, NAC was observed to inhibit apoptosis in the hippocampus of SHRs. Collectively, we firstly showed the enhanced accumulation of MG in the hypertensive brain, whereas the clearance of MG by NAC treatment mitigated the aging process and attenuated AGEs generation, neuroinflammation, and oxidative damage.

A novel KLF13 mutation underlying congenital patent ductus arteriosus and ventricular septal defect, as well as bicuspid aortic valve.

Recently, mutations in the Kruppel-like factor 13 (KLF13) gene encoding a Kruppel-like transcription factor have been reported to cause congenital heart disease (CHD). However, due to pronounced genetic heterogeneity, the mutational spectrum of KLF13 in other cohorts of cases suffering from distinct types of CHD remain to be ascertained. In the present investigation, by Sanger sequencing of KLF13 in 316 unrelated cases affected by different forms of CHD, a new mutation in heterozygous status, NM_015995.3: c.430G>T; p.(Glu144*), was detected in an index patient affected with patent ductus arteriosus (PDA) and ventricular septal defect (VSD), as well as bicuspid aortic valve (BAV), with a mutation frequency of ~0.32%. Genetic investigation of the available family members of the proband demonstrated that the truncating mutation co-segregated with CHD. The nonsense mutation was not observed in 400 unrelated volunteers without CHD who were enrolled as control subjects. Quantitative biological measurements with dual luciferase reporters revealed that Glu144*-mutant KLF13 did not transactivate the downstream genes vascular endothelial growth factor A and natriuretic peptide A. In addition, the mutation abrogated the synergistic transcriptional activation between KLF13 and T-box transcription factor 5, a well-established CHD-causing gene. In conclusion, the present study indicates that genetically defective KLF13 contributes to familial PDA and VSD, as well as BAV, which expands the phenotypic spectrum linked to KLF13, and reveals a novel molecular pathogenesis of the disease, providing a new molecular target for the early prophylaxis and individualized treatment of CHD.

SOX17 Loss-of-Function Mutation Underlying Familial Pulmonary Arterial Hypertension.

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 ß-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.

Detection and functional characterization of a novel MEF2A variation responsible for familial dilated cardiomyopathy.

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.

A novel TBX5 mutation predisposes to familial cardiac septal defects and atrial fibrillation as well as bicuspid aortic valve.

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.

A New TBX5 Loss-of-Function Mutation Contributes to Congenital Heart Defect and Atrioventricular Block.

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.

Identification and functional characterization of KLF5 as a novel disease gene responsible for familial dilated cardiomyopathy.

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.

A New ISL1 Loss-of-Function Mutation Predisposes to Congenital Double Outlet Right Ventricle.

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.225C>G; 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.

TBX1 loss-of-function mutation contributes to congenital conotruncal defects.

Conotruncal defects (CTDs) account for ~30% of all types of congenital heart disease and contribute to increased morbidity and mortality rates. Increasing evidence suggests that genetic risk factors are involved in the pathogenesis of CTDs. Mutations in a number of genes, including the TBX1 gene that codes for a T-box transcription factor essential for normal cardiovascular development, may contribute to the development of CTD. CTDs are genetically heterogeneous and the genetic defects responsible for CTDs in the majority of patients remain unknown. The present study sequenced the coding regions and splicing junction boundaries of TBX1 in 136 patients with CTDs and 300 matched healthy individuals. The disease-causing potential of the identified TBX1 sequence variation was evaluated using MutationTaster, PolyPhen-2, SIFT and PROVEN software. The functional characteristics of the mutant TBX1 gene were defined using a dual-luciferase reporter assay system. A novel heterozygous TBX1 mutation, p.S233Y, was identified in a patient with transposition of the great arteries (TGA) and a ventricular septal defect. This mutation was absent in the 300 controls and altered the amino acid produced, serine, which is evolutionarily conserved across several species, and was predicted to be pathogenic in silico. Luciferase assays conducted in COS-7 cells demonstrated that the newly identified TBX1 mutation was associated with significantly diminished transcriptional activation of the ANF promoter compared with the wild-type TBX1. To the best of our knowledge, the present study is the first to associate a TBX1 loss-of-function mutation with enhanced susceptibility to TGA, which adds significant insight to the molecular mechanism of TGA.

miR-218 Involvement in Cardiomyocyte Hypertrophy Is Likely through Targeting REST.

MicroRNAs (miRNAs) have been identified as key players in cardiomyocyte hypertrophy, which is associated with significant risks of heart failure. However, many microRNAs are still not recognized for their functions in pathophysiological processes. In this study, we evaluated effects of miR-218 in cardiomyocyte hypertrophy using both in vitro and in vivo models. We found that miR-218 was evidently downregulated in a transverse aortic constriction (TAC) mouse model. Overexpression of miR-218 is sufficient to reduce hypertrophy, whereas the suppression of miR-218 aggravates hypertrophy in primary cardiomyocytes induced by isoprenaline (ISO). In addition, we identified RE1-silencing transcription factor (REST) as a novel target of miR-218; it negatively regulated the expression of REST in hypertrophic cardiomyocytes and the TAC model. These results showed that miR-218 plays a crucial role in cardiomyocyte hypertrophy, likely via targeting REST, suggesting a potential candidate target for interfering hypertrophy.

A novel HAND2 loss-of-function mutation responsible for tetralogy of Fallot.

Congenital heart disease (CHD), the most common type of developmental abnormality, is associated with substantial morbidity and mortality in humans worldwide. The basic helix-loop-helix transcription factor, heart and neural crest derivatives expressed 2 (HAND2), has been demonstrated to be crucial for normal cardiovascular development in animal models. However, whether a genetically defective HAND2 contributes to congenital heart disease (CHD) in humans remains to be explored. In this study, the entire coding region and splicing boundaries of the HAND2 gene were sequenced in a cohort of 145 unrelated patients with CHD. A total of 200 unrelated, ethnically-matched healthy individuals used as controls were also genotyped for HAND2. The functional effect of the mutant HAND2 was characterized in contrast to its wild-type counterpart by using a dual-luciferase reporter assay system. As a result, a novel heterozygous HAND2 mutation, p.L47P, was identified in a patient with tetralogy of Fallot (TOF). The misense mutation, which altered the amino acid conserved evolutionarily among species, was absent in 400 control chromosomes. Functional analyses unveiled that the mutant HAND2 had a significantly decreased transcriptional activity. Furthermore, the mutation markedly reduced the synergistic activation between HAND2 and GATA4 or NKX2.5, other two cardiac key transcription factors involved in the pathogenesis of CHD. To the best of our knowledge, this study is the first to report the association of a HAND2 loss-of-function mutation with an increased vulnerability to TOF in humans, which provides novel insight into the molecular mechanism underpinning CHD, suggesting potential implications for the genetic counseling of families with CHD.

TBX20 loss-of-function mutation associated with familial dilated cardiomyopathy.

BACKGROUND: Dilated cardiomyopathy (DCM) is a major cause of congestive heart failure, sudden cardiac death and cardiac transplantation. Aggregating evidence highlights the genetic origin of DCM. However, DCM is a genetically heterogeneous disorder, and the genetic components underlying DCM in most cases remain unknown. METHODS: The coding regions and splicing junction sites of the TBX20 gene were sequenced in 120 unrelated patients with idiopathic DCM. The available close relatives of the index patient carrying an identified mutation and 300 unrelated ethnically matched healthy individuals used as controls were genotyped for TBX20. The functional characteristics of the mutant TBX20 were assayed in contrast to its wild-type counterpart by using a dual-luciferase reporter assay system. RESULTS: A novel heterozygous TBX20 mutation, p.F256I, was identified in a family with DCM transmitted in an autosomal dominant fashion, which co-segregated with DCM in the family with complete penetrance. The missense mutation was absent in 600 control chromosomes and the altered amino acid was completely conserved evolutionarily among various species. Functional assays revealed that the mutant TBX20 had significantly diminished transcriptional activity. Furthermore, the mutation markedly reduced the synergistic activation of TBX20 with NKX2-5 or GATA4. CONCLUSIONS: This study links TBX20 loss-of-function mutation to idiopathic DCM in humans for the first time, providing novel insight into the molecular mechanism underpinning DCM.

Effects of antiarrhythmic peptide 10 on acute ventricular arrhythmia.

OBJECTIVE: To observe the effects antiarrhythmic peptide 10 (AAP10) aon acute ventricular arrhythmia and the phosphorylation state of ischemic myocardium connexin. METHODS: Acute total ischemia and partial ischemia models were established by ceasing perfusion and ligating the left anterior descending coronary artery in SD rats. The effects of AAP10 (1 mg/L) on the incidence rate of ischemia-induced ventricular arrhythmia were observed. The ischemic myocardium was sampled to detect total-Cx43 and NP-Cx43 by immunofluorescent staining and western blotting. the total-Cx43 expression was detected through image analysis system by semi-quantitative analysis. RESULTS: AAP10 could significantly decrease the incidence of ischemia-induced ventricular tachycardia and ventricular fibrillation. During ischemic stage, total ischemia (TI) and AAP10 total ischemia (ATI) groups were compared with partial ischemia (PI) and AAP10 partial ischemia (API) groups. The rates of incidence for arrhythmia in the ATI and API groups (10% and 0%) were lower than those in the TI and PI groups (60% and 45%). The difference between the two groups was statistically significant (P=0.019, P=0.020). The semi-quantitative analysis results of the ischemic myocardium showed that the total-Cx43 protein expression distribution areas for TI, ATI, PI and API groups were significantly decreased compared with the control group. On the other hand, the NP-Cx43 distribution areas of TI, ATI, PI and API groups were significantly increased compared with the control group (P>0.05). AAP10 could increase the total-Cx43 expression in the ischemic area and decrease the NP-Cx43 expression. Western blot results were consistent with the results of immunofluorescence staining. CONCLUSIONS: AAP10 can significantly decrease the rate of incidence of acute ischemia-induced ventricular tachycardia and ventricular fibrillation. Acute ischemic ventricular arrhythmias may have a relationship with the decreased phosphorylation of Cx43 induced by ischemia. AAP10 may stimulate the phosphorylation of Cx43 by increasing the total-Cx43 expression and decreasing the NP-Cx43 expression in the ischemic area, so as to decrease ventricular arrhythmia.

PITX2 Loss-of-Function Mutation Contributes to Congenital Endocardial Cushion Defect and Axenfeld-Rieger Syndrome.

Congenital heart disease (CHD), the most common type of birth defect, is still the leading non-infectious cause of infant morbidity and mortality in humans. Aggregating evidence demonstrates that genetic defects are involved in the pathogenesis of CHD. However, CHD is genetically heterogeneous and the genetic components underpinning CHD in an overwhelming majority of patients remain unclear. In the present study, the coding exons and flanking introns of the PITX2 gene, which encodes a paired-like homeodomain transcription factor 2essential for cardiovascular morphogenesis as well as maxillary facial development, was sequenced in 196 unrelated patients with CHD and subsequently in the mutation carrier's family members available. As a result, a novel heterozygous PITX2 mutation, p.Q102X for PITX2a, or p.Q148X for PITX2b, or p.Q155X for PITX2c, was identified in a family with endocardial cushion defect (ECD) and Axenfeld-Rieger syndrome (ARS). Genetic analysis of the pedigree showed that the nonsense mutation co-segregated with ECD and ARS transmitted in an autosomal dominant pattern with complete penetrance. The mutation was absent in 800 control chromosomes from an ethnically matched population. Functional analysis by using a dual-luciferase reporter assay system revealed that the mutant PITX2 had no transcriptional activity and that the mutation eliminated synergistic transcriptional activation between PITX2 and NKX2.5, another transcription factor pivotal for cardiogenesis. To our knowledge, this is the first report on the association of PITX2 loss-of-function mutation with increased susceptibility to ECD and ARS. The findings provide novel insight into the molecular mechanisms underpinning ECD and ARS, suggesting the potential implications for the antenatal prophylaxis and personalized treatment of CHD and ARS.

TBX5 loss-of-function mutation contributes to familial dilated cardiomyopathy.

The cardiac T-box transcription factor TBX5 is crucial for proper cardiovascular development, and mutations in TBX5 have been associated with various congenital heart diseases and arrhythmias in humans. However, whether mutated TBX5 contributes to dilated cardiomyopathy (DCM) remains unclear. In this study, the coding exons and flanking introns of the TBX5 gene were sequenced in 190 unrelated patients with idiopathic DCM. The available family members of the index patient carrying an identified mutation and 200 unrelated ethnically matched healthy individuals used as controls were genotyped for TBX5. The functional characteristics of the mutant TBX5 were explored in contrast to its wild-type counterpart by using a dual-luciferase reporter assay system. As a result, a novel heterozygous TBX5 mutation, p.S154A, was identified in a family with DCM inherited in an autosomal dominant pattern, which co-segregated with DCM in the family with complete penetrance. The missense mutation was absent in 400 control chromosomes and the altered amino acid was completely conserved evolutionarily across various species. Functional assays revealed that the mutant TBX5 had significantly decreased transcriptional activity. Furthermore, the mutation markedly diminished the synergistic activation of TBX5 with NKX2-5 or GATA4, other two transcription factors causatively linked to DCM. This study firstly associates TBX5 loss-of-function mutation with enhanced susceptibility to DCM, providing novel insight into the molecular mechanisms of DCM, and suggesting the potential implications in the development of new treatment strategies for this common form of myocardial disorder.

TBX20 loss-of-function mutation contributes to double outlet right ventricle.

Congenital heart disease (CHD), the most prevalent birth defect in humans worldwide, is still a leading non­infectious cause of infant morbidity and mortality. Increasing evidence demonstrates that genetic risk factors play a key role in the pathogenesis of CHD, and more than 50 genes have been linked to various types of CHD. Nevertheless, CHD is a heterogeneous disorder and the genetic components underpinning CHD in an overwhelming majority of cases remain unknown. In the present study, the entire coding exons and flanking introns of the TBX20 gene, which codes for a T-box transcription factor essential for the proper development of the heart, were sequenced in a cohort of 146 unrelated patients with CHD. The available relatives of the index patient harboring an identified mutation and 200 unrelated ethnically matched healthy individuals used as the controls were also genotyped for TBX20. The functional characteristics of the TBX20 mutation were assayed by using a dual-luciferase reporter assay system. As a result, a novel heterozygous TBX20 mutation, p.R143W, was identified in an index patient with double outlet right ventricle (DORV). Genetic analyses of the pedigree of the proband revealed that in the family, the mutation co-segregated with DORV transmitted in an autosomal dominant pattern with complete penetrance. The missense mutation was absent in 400 control chromosomes and the altered amino acid was completely conserved evolutionarily across species. Functional analysis revealed that mutant TBX20 had a significantly diminished transcriptional activity compared with its wild-type counterpart. To the best of our knowledge, this study is the first to report the association of TBX20 loss-of-function mutation with increased susceptibility to DORV in humans, which provides novel insight into the molecular mechanisms responsible for CHD, suggesting potential implications for the antenatal prophylaxis of CHD.

GATA5 loss-of-function mutation in familial dilated cardiomyopathy.

Dilated cardiomyopathy (DCM), the most common form of primary myocardial disease, is an important cause of sudden cardiac death and heart failure and is the leading indication for heart transplantation in children and adults worldwide. Recent studies have revealed a strong genetic basis for idiopathic DCM, with many distinct genes causally implicated. Nevertheless, DCM is a genetically heterogeneous disorder and the genetic determinants underlying DCM in a substantial proportion of patients remain unclear. In this study, the whole coding exons and flanking introns of the GATA binding protein 5 (GATA5) gene, which codes for a zinc-finger transcription factor essential for cardiovascular development and structural remodeling, were sequenced in 130 unrelated patients with idiopathic DCM. The available relatives of the index patient carrying an identified mutation and 200 unrelated ethnically matched healthy individuals used as the controls were genotyped for GATA5. The functional characteristics of the mutant GATA5 were analyzed in contrast to its wild-type counterpart by using a dual-luciferase reporter assay system. As a result, a novel heterozygous GATA5 mutation, p.G240D, was identified in a family with DCM inherited in an autosomal dominant pattern, which co-segregated with DCM in the family with complete penetrance. The missense mutation was absent in 400 reference chromosomes and the altered amino acid was completely conserved evolutionarily across species. Functional analyses revealed that the GATA5 mutant was associated with significantly diminished transcriptional activity. This study firstly links GATA5 mutation to DCM, which provides novel insight into the molecular mechanisms of DCM, suggesting a potential molecular target for the prenatal prophylaxis and allele-specific treatment of DCM.

A novel NKX2.5 loss-of-function mutation associated with congenital bicuspid aortic valve.

Bicuspid aortic valve (BAV) is the most common form of congenital cardiovascular defect in humans and is associated with substantial morbidity and mortality. Emerging evidence demonstrates that genetic risk factors play an important role in the pathogenesis of BAV. However, BAV is a genetically heterogenous disorder, and the genetic defects underpinning BAV in most patients remain to be identified. In the present study, the coding exons and flanking introns of the NKX2.5 gene, which encodes a homeodomain-containing transcription factor essential for the normal development of the aortic valve, were sequenced in 142 unrelated patients with BAV. The available relatives of the mutation carrier and 200 unrelated healthy subjects used as controls were also genotyped for NKX2.5. The functional characteristics of the mutation were delineated by using a dual-luciferase reporter assay system. As a result, a novel heterozygous NKX2.5 mutation, p.K192X, was identified in a family with BAV transmitted in an autosomal dominant pattern. The nonsense mutation was absent in 400 control chromosomes. Functional analyses revealed that the mutant NKX2.5 had no transcriptional activity compared with its wild-type counterpart. Furthermore, the mutation abolished the synergistic transcriptional activation between NKX2.5 and GATA5, another transcription factor crucial for the aortic valvular morphogenesis. In conclusion, this study is the first to link an NKX2.5 loss-of-function mutation to enhanced susceptibility to human BAV, providing novel insight into the molecular mechanism of BAV and suggesting potential implications for genetic counseling and clinical care of families presenting with BAV.