Novel and deleterious nucleotide variations in the HAND1 gene probably affect miRNA target sites and protein function in pediatric patients with congenital heart disease.

BACKGROUND: Congenital heart disease (CHD) is the most prevalent developmental defect and principal cause of infant mortality and affects cardiac and large blood vessel structures in approximately 1% of live births worldwide. To date, numerous studies have related critical genetic dysfunctions to the pathogenesis of CHDs. However, the genetic basis underlying CHD remains largely unknown. In the present study, we investigated the association of nucleotide variations in coding and noncoding regions of the HAND1 gene with the risk of CHD. The HAND1 gene, encoding a helix-loop-helix transcription factor, is particularly relevant for mechanisms underlying CHD since it plays a significant role in heart development. METHODS AND RESULTS: The genomic DNA of 150 unrelated pediatric patients with CHD was screened by PCR-SSCP and direct sequencing. Four novel and heterozygous missense mutations were identified in the first exon, with three causing amino acid substitutions (p.Val149Met, p.Tyr142His, and p.Leu146Met). In-silico analysis also indicated their deleterious impact on protein structure and function. In addition, we identified five novel nucleotide variants in the 3'UTR region (c.*461, c.*342, c.*529, c.*448, c.*593), potentially altering the target sites of miRNAs. These changes include the loss of certain target sites and the acquisition of new ones. CONCLUSIONS: These findings confirm the phenotypic association between CHDs and HAND1 mutations and can pave the way for developing new preventive and therapeutic strategies.

Novel nucleotide variations in the thrombomodulin (THBD) gene involved in coagulation pathways can increase the risk of recurrent pregnancy loss (RPL).

Recurrent pregnancy loss (RPL) is a common but complex complication in fertility conditions, affecting about 15-20% of couples. Although several causes have been proposed for RPL, it occurs in about 35-60% of cases without a known explanation. A strong assumption is that genetic factors play a role in the etiology and pathophysiology of PRL. Therefore, several genes are proposed as candidates in the pathogenesis of RPL. The current study aimed to investigate the effects of nucleotide changes in the THBD (thrombomodulin) gene as an RPL-related candidate gene. This gene encodes a cell receptor for thrombin and is involved in reproductive loss in RPL cases. Its involvement in the natural anticoagulant system has been extensively studied. By genetic screening of the entire coding and noncoding regions of the THBD gene, we found twenty-seven heterozygous and homozygous nucleotide changes. Ten of them led to amino acid substitutions, seven variants were identified in the promoter region, and eight of them occurred in 3'UTR. Potentially, the pathogenicity effects of these variations on THBD protein were evaluated by several prediction tools. The numerous genomic variations prompted noticeable modifications of the protein's structural and functional properties. Furthermore, in-silico scores were consistent with deleterious effects for these mutations. The results of this study provide genetic information that will be useful in the future for clinicians, scientists, and students to understand the unknown causes of RPL better. It may also pave the way for developing diagnostic/prognostic approaches to help treat PRL patients.

Mitochondrial mutations in protein coding genes of respiratory chain including complexes IV, V, and mt-tRNA genes are associated risk factors for congenital heart disease.

Most studies aiming at unraveling the molecular events associated with cardiac congenital heart disease (CHD) have focused on the effect of mutations occurring in the nuclear genome. In recent years, a significant role has been attributed to mitochondria for correct heart development and maturation of cardiomyocytes. Moreover, numerous heart defects have been associated with nucleotide variations occurring in the mitochondrial genome, affecting mitochondrial functions and cardiac energy metabolism, including genes encoding for subunits of respiratory chain complexes. Therefore, mutations in the mitochondrial genome may be a major cause of heart disease, including CHD, and their identification and characterization can shed light on pathological mechanisms occurring during heart development. Here, we have analyzed mitochondrial genetic variants in previously reported mutational genome hotspots and the flanking regions of mt-ND1, mt-ND2, mt-COXI, mt-COXII, mt-ATPase8, mt-ATPase6, mt-COXIII, and mt-tRNAs (Ile, Gln, Met, Trp, Ala, Asn, Cys, Tyr, Ser, Asp, and Lys) encoding genes by polymerase chain reaction-single stranded conformation polymorphism (PCR-SSCP) in 200 patients with CHD, undergoing cardiac surgery. A total of 23 mitochondrial variations (5 missense mutations, 8 synonymous variations, and 10 nucleotide changes in tRNA encoding genes) were identified and included 16 novel variants. Additionally, we showed that intracellular ATP was significantly reduced (P=0.002) in CHD patients compared with healthy controls, suggesting that the mutations have an impact on mitochondrial energy production. Functional and structural alterations caused by the mitochondrial nucleotide variations in the gene products were studied in-silico and predicted to convey a predisposing risk factor for CHD. Further studies are necessary to better understand the mechanisms by which the alterations identified in the present study contribute to the development of CHD in patients.

Mutational and bioinformatics analysis of the NKX2.1 gene in a cohort of Iranian pediatric patients with congenital hypothyroidism (CH).

Congenital hypothyroidism (CH) occurs with a relatively alarming prevalence in infants, and if not diagnosed and treated in time, it can have devastating consequences for the development of the nervous system. CH is associated with genetic changes in several genes that encode transcription factors responsible for thyroid development, including mutations in the NK2 homeobox 1 (NKX2.1) gene, which encodes the thyroid transcription factor-1 (TTF-1). Although CH is frequently observed in pediatric populations, there is still a limited understanding of the genetic factors and molecular mechanisms contributing to this disease. The sequence of the NKX2.1 gene was investigated in 75 pediatric patients with CH by polymerase chain reaction (PCR), single-stranded conformation polymorphism (SSCP), and direct DNA sequencing. Four missense heterozygous variations were identified in exon 3 of the NKX2.1 gene, including three novel missense variations, namely c.708A>G, p.Gln202Arg; c.713T>G, p.Tyr204Asp; c.833T>G, p.Tyr244Asp, and a previously reported variant rs781133468 (c.772C>G, p.His223Gln). Importantly, these variations occur in highly conserved residues of the TTF-1 DNA-binding domain and were predicted by bioinformatics analysis to alter the protein structure, with a probable alteration in the protein function. These results indicate that nucleotide changes in the NKX2.1 gene may contribute to CH pathogenesis.

Mutational Analysis of the VPREB1 Gene of Pre-BCR Complex in a Cohort of Sporadic Pediatric Patients With B-Cell Acute Lymphoblastic Leukemia.

During bone marrow B-cell development, the pre-B-cell receptor is formed by the association of the immunoglobulin heavy chain with a surrogate light chain, which is encoded by the VPREB1, and λ5 genes. It is known that pre-BCR signaling signifies a critical checkpoint at the pre-B-cell stage. Thus, failure pre-BCR signaling is proposed as a critical factor for the development of B-cell acute lymphoblastic leukemia (B-ALL). B­ALL is the most common pediatric cancer and is one of the leading causes of death in children. Until now, several molecular analyses were performed for genomic alterations in B-ALL, but for genomic analysis of the VPREB1 gene and its rare variations, limited studies have been conducted. In this study, using polymerase chain reaction and direct sequencing of 88 pediatric patients with B-ALL, we investigated the genomic region of the VPREB1 gene to find sequence variations of this gene. Our study presented ten homozygous and heterozygous point mutations and heterozygous nucleotide deletions, in the VPREB1 gene in 36 boys and 32 girls' patients. Our Bioinformatics assay results presented that these variations may alter the RNA folding, protein structure, and therefore probable effect on the protein function. These results propose that nucleotide changes probably contribute to B-ALL pathogenesis.

New insights into short peptides derived from the collagen NC1 α1, α2, and α3 (IV) domains: An experimental and MD simulations study.

Proteolytically cleavage of the collagen NC1 α1 to α3 (IV) domains leads to antiangiogenic proteins called Arresten, Canstatin, and Tumstatin, respectively. The research identified that the two overlapping peptides derived from Tumstatin are more effective than other fragments and amino acids L78, V82, and D84 are essential for their activity. In the present study, the efficacy of a nine amino acid peptide derived from Tumstatin (Tum), containing amino acids L78, V82, and D84 was compared to the corresponding sequence in Arresten (Ars) and Canstatin (Can) in vitro and in vivo. Moreover, CD spectroscopy, MD, and docking simulations were performed to evaluate the structure and the interaction of peptides to integrin αvß3, respectively. Results demonstrated that peptides inhibit viability, migration, and tube formation in vitro, as well as the growth of tumor in vivo and Canstatin-derived peptide was more potent than others. CD measurement and DSSP calculation revealed that Can had more coil conformation. According to MD simulations, Can had more fluctuation, less intramolecular interactions, and less structural compactness compared to Tum and Ars. It can be assumed that amino acid variations lead to a more flexible and loose structure compared to the other peptides. The Canstatin-derived peptide interacts with the integrin αvß3 extremely close to RGD binding site by the most negative binding energy and more interactions. In conclusion, we for the first time identified an active peptide derived from Canstatin and showed that the sequence affected structure and thereby interaction of peptide to its receptor.

Novel Point Mutations in 3'-Untranslated Region of GATA4 Gene Are Associated with Sporadic Non-syndromic Atrial and Ventricular Septal Defects.

OBJECTIVE: Transcription factor GATA4 has significant roles in embryonic heart development. Mutations of GATA4 appear to be responsible for a wide variety of congenital heart defects (CHD). Despite the high prevalence of GATA4 mutations in CHD phenotypes, extensive studies have not been performed. The 3'-untranslated region (3'-UTR) of the GATA4 gene comprises regulatory motifs and microRNA binding sites that are critical for the appropriate gene expression, nuclear transportation, and regulation of translation, and stability of mRNA. This study aimed to evaluate the association between mutations in the 3'-UTR of the GATA4 gene and CHD risk among Iranian patients. METHODS: We analyzed the coding region of exon 6 and the whole 3'-UTR of GATA4 in DNA isolated from 175 blood samples of CHD patients and 115 unrelated healthy individuals. The functional importance of the observed GATA4 mutations was evaluated using a variety of bioinformatics algorithms for assessment of nonsynonymous mutations and those observed in miRNA binding sites of 3'-UTR. RESULTS: Twenty-one point mutations including one missense mutation (c.511A>G: p.Ser377Gly) in exon 6 and 20 nucleotide variations in 3'-UTR of GATA4 gene were identified in 65 of the 175 CHD patients. In our patients, we identified 12 novel sequence alterations and 8 single nucleotide polymorphisms in the 3'-UTR of GATA4. Most of them had statistically significant differences between CHD patients and controls. CONCLUSION: Our results suggest that 3'-UTR variations of the GATA4 gene probably change microRNA binding sites and present an additional molecular risk factor for the susceptibility of CHD.

Mutational screening through comprehensive bioinformatics analysis to detect novel germline mutations in the APC gene in patients with familial adenomatous polyposis (FAP).

BACKGROUND: Familial adenomatous polyposis (FAP) as a colon cancer predisposition syndrome is an autosomal-dominant inherited condition and is diagnosed by the progress of hundreds or thousands of adenomatous colonic polyps in the colon. This study aims at the nature and effect of Adenomatous Polyposis Coli (APC) gene mutations in FAP tumorigenesis. METHODS: The genetic screening of 59 FAP Iranian patients in 10 families was performed by polymerase chain reactions and the direct sequencing of the entire coding exons of the APC gene. To do linkage haplotype analysis and multiplex PCR-based microsatellite examination, six short tandem repeat loci were selected in this gene. To evaluate and predict the potentially deleterious effects, comprehensive bioinformatics pathogenicity assays were used. RESULTS: A total of 12 germline heterozygous and homozygous nucleotide variations were identified. They included two missense mutations, four nonsense mutations, which would lead to the truncated and nonfunctional protein products, four synonymous or silent variations, and two nucleotide deletions of 1 to 5 bp or frameshift mutations. In addition, three novel heterozygous nonsense mutations were found in exons 10, 14, and 15 of the gene. There was also p.Arg653Met as a novel heterozygote mutation in exon 14 of the gene. CONCLUSIONS: Bioinformatics analysis and three-dimensional structural modeling predicted that these missense and nonsense mutations generally are associated with the deleted or truncated domains of APC and have functional importance and mainly affected the APC protein. These findings may provide evidence for the progress of potential biomarkers and help to understand the role of the APC gene in FAP.

Relationship of hypomethylation CpG islands in interleukin-6 gene promoter with IL-6 mRNA levels in patients with coronary atherosclerosis.

Introduction: Atherosclerosis is the important cause of most cardiovascular diseases, with high prevalence and mortality. Atherosclerosis is not only a lipid metabolism disorder but also recently is defined as a chronic inflammatory disease. Several studies showed that interleukin-6 (IL-6) is involved in the pathogenesis of atherosclerosis. The aim of the present study is the examination of IL6 mRNA Levels and hypomethylation of IL6 promoter in atherosclerosis patients. Methods: In this assay, a total of 35 cases with atherosclerosis and 30 controls were enrolled. RNA and DNA were isolated from the peripheral blood of all samples. Mean IL6 gene expression was determined by RT-PCR and methylation status at six CpG motifs in IL6 promoter was determined using bisulfite genomic sequencing. Results: Real Time-PCR analysis results showed the mean IL6 RNA level in atherosclerosis patients candidate for CABG (coronary artery bypass grafting) was significantly higher than controls (P value = 0.01). Also, the upstream CpG motifs (-1038 to -952) in IL6 promoter were predominantly unmethylated in patients than in the controls (P value = 0.01). Conclusion: These findings suggest that an increase in IL-6 gene expression and its DNA hypomethylation promoter are associated with atherosclerosis patient's candidate for CABG surgery.

Novel Point Mutations in Mitochondrial MT-CO2 Gene May Be Risk Factors for Coronary Artery Disease.

A wide range of genetic and environmental interactions are involved in the development of coronary artery disease (CAD). Considerable evidence suggests that mitochondrial DNA mutations are associated with heart failure. In this work, we examined the possible mutations in hotspot mitochondrial genes and their association with Iranian patients with coronary artery disease. In this case-control study, nucleotide variations were investigated in 109 patients with coronary atherosclerosis and 105 control subjects with no family history of cardiovascular disease. The molecular analysis of related mitochondrial genes was performed by polymerase chain reaction sequencing. Our results showed 25 nucleotide variations (10 missense mutations, 9 synonymous polymorphisms, and 6 variants in tRNA genes) that for the first time were presented in coronary artery disease. Our results suggest that novel heteroplasmic m.8231 C>A mutation is involved in CAD (p = 0.007). These nucleotide variations suggest the role of mitochondrial mutations as a predisposing factor which in combination with environmental risk factors may affect the pathogenesis of coronary atherosclerosis. So, further investigation is needed for a better understanding of the pathogenesis and predisposing effects of these variations on the disease.

Novel Point Mutations of CITED2 Gene Are Associated with Non-familial Congenital Heart Disease (CHD) in Sporadic Pediatric Patients.

CITED2 is a cardiac transcription factor that plays a critical role in cardiac development. Gene mutations in CITED2 lead to a series of cardiac malformations and congenital heart defects (CHD). Congenital heart disease generally refers to defects in the heart's structure or function and often seen in many forms such as ventricular septal defects (VSDs), atrial septal defects (ASDs), and tetralogy of Fallot (TOF). However, the mechanisms involved in these mutations are poorly understood. The aim of the present study was to evaluate the mutations of the CITED2 gene in pediatric patients with congenital heart defects. We studied the potential impact of sequence variations of the CITED2 gene in a cohort of 150 patients with non-familial CHD and 98 control individuals by polymerase chain reaction-single-stranded conformation polymorphism (PCR-SSCP) and subsequently direct sequencing. We identified seven novel CITED2 nucleotide changes. Four of these alterations were found in the coding region (c.716insG, c.389A>G, c.450G>C and c.512-538del27) and were only seen in our patients, and not detected in the control group. These mutations are leading to changes in the amino acid sequence in the position of p.Gly236fs, p.Asn125Ser, p.Gln145His, and p.Ser170-Gly178del, respectively. Other variations are located in the 5'UTR region of the gene (c.-43C>T, c.-64C>T and c.-90A>G). CITED2 gene mutations in control subjects were not observed. Our Bioinformatics assay results showed that these novel mutations alter the RNA folding, protein structure, and, therefore, probable effect on the protein function and may play a significant role in the development of congenital heart diseases.

A novel heteroplasmic mutation in mitochondrial tRNAArg gene associated with non-dystrophic myotonias.

Non-dystrophic myotonias (NDM) are rare diseases caused by defects in skeletal muscle chloride and sodium ion channels. It is well established that high-energy consuming tissues such as muscular and nervous systems are exclusively dependent on the ATP generation by mitochondria. The mitochondrial dysfunction, which is caused by mitochondrial DNA mutations, played an important role in the pathogenesis of non-dystrophic myotonias. The purpose of this study is to identify mitochondrial tRNA mutations in non-dystrophic myotonias patients. In this study, 45 Iranian patients with non-dystrophic myotonia were investigated for intracellular ATP content and the mutation screening in all the mitochondrial tRNA genes by DNA sequencing. Our findings showed that lymphocyte intracellular ATP is significantly decreased in NDM patients compared with control subjects (p = 0.001). We found nine mutations in mitochondrial tRNA genes, including m.4454 T > C (in the TψC loop of tRNAMet), m.5568 A > G (tRNATrp), m.5794 T > C (in the anticodon loop of tRNACys), novel m.10438 A > T, and m.10462 T > C (in anticodon loop and ACC stem of tRNAArg), m.12308 A > G (tRNALeu(CUN)) and m.15907 A > G, m.15924 A > G, and m.15928 G > A (in the anticodon stem of tRNAThr) in 31 NDM patients. These results suggest that novel m.10438 A > T mutation is involved in NDM patients and reinforces the significant association between this mutation in mitochondrial tRNAArg Gene and NDM patients (p = 0.008).

Association of Pathogenic Missense and Nonsense Mutations in Mitochondrial COII Gene with Familial Adenomatous Polyposis (FAP).

Nuclear genetic mutations have been extensively investigated in solid tumors. However, the role of the mitochondrial genome remains uncertain. Since the metabolism of solid tumors is associated with aerobic glycolysis and high lactate production, tumors may have mitochondrial dysfunctions. Familial adenomatous polyposis (FAP) is a rare form| of colorectal cancer and an autosomal dominant inherited condition that is characterized by the progress of numerous adenomatous polyps in the rectum and colon. The present study aimed at understanding the nature and effect of mitochondrial cytochrome c oxidase subunit 2 (COII) gene mutations in FAP tumorigenesis. Fifty-six (26 familial and 30 sporadic) FAP patients and 60 normal controls were enrolled in this study. COII point mutations were evaluated by PCR and direct sequencing methods, and a total of 7 mtDNA mutations were detected (3 missense, 1 nonsense, and 3 synonymous variations). Novel non-synonymous COII gene mutations were mostly in heteroplasmic state. These mutations change amino acid residues in the N-terminal and C-terminal regions of COXII. Bioinformatics analysis and three-dimensional structural modeling predicted that these missense and nonsense mutations have functional importance, and mainly affected on cytochrome c oxidase (complex IV). Also, FAP patients carried a meaningfully higher prevalence of mutations in the COII gene in comparison with healthy controls (P <0.001). Analysis of cancer-associated mtDNA mutation could be an invaluable tool for molecular assessment of FAP so that these findings can be helpful for the development of potential new biomarkers in the diagnosis of cancer for future clinical assessments.

The association between TNP2 gene polymorphisms and Iranian infertile men with varicocele: A case-control study.

BACKGROUND: Numerous researches have provided great evidence that revealed the relationship between varicocele and sperm DNA damage. OBJECTIVE: Because of the crucial role of nuclear transition proteins (TPs) in sperm DNA condensation and integrity, this case-control study was designed to study TNP2 gene nucleotide variations in Iranian patients with varicocele. MATERIALS AND METHODS: PCR-SSCP and DNA sequencing were used to search for mutations in exons 1 & 2 of the TNP2 gene in 156 infertile patients with varicocele and 150 fertile men. RESULTS: The results of sequencing showed three variants at positions c.301C░>░T (p.R101C), c.391C░>░T (p.R131░W), and g.IVS1-26G░>░C (rs8043625) of TNP2 gene. It was found that varicocele risk in men who have the CC genotype of g.IVS1-26G░>░C SNP is higher than those who don't have these genotypes (according to Co-dominant model, Dominant model, Recessive model, and Over-dominant model). The haplotype-based analysis showed that (C/C/T) and (C/T/T) haplotypes were a risk factor of in patients with varicocele compared to controls (OR░=░3.278, p░=░0.000 and OR░=░9.304, p░=░0.038, respectively). CONCLUSION: Because of the significant difference in the genotype and allele frequencies of g.IVS1-26G░>░C SNP in the intronic region of TNP2 in patients with varicocele compared with controls and also because of the high conservation of this SNP position during evolution, this SNP may be involved in some important processes associated with the expression of this gene like mRNA splicing, but the exact mechanism is not clear.

Protective effect of Berberis vulgaris on Fenton reaction-induced DNA cleavage.

OBJECTIVE: Berberis vulgaris contains antioxidants that can inhibit DNA cleavage. The purpose of this study was to evaluate the antioxidant and protective activity of B. vulgaris on DNA cleavage. MATERIALS AND METHODS: In this study, the antioxidant capacity of B. vulgaris was investigated using DPPH and its protective effect was evaluated on pBR322 plasmid and lymphocyte genomic DNA cleavage induced by Fenton reaction, by DNA electrophoresis. RESULTS: Aqueous extract of B. vulgaris presented dual behavior with a potent antioxidant activity at 0.25and 0.75mg/ml for pBR322 plasmid and lymphocyte genomic DNA, respectively, but a pro-oxidant activity was observed at higher concentrations. CONCLUSION: Our results indicated that B. vulgaris extract an inhibit Fenton reaction-induced DNA cleavage and oxidative cleavage of double-stranded DNA assay is a powerful technique that can be used to determine the antioxidant and pro-oxidant properties of a compound on cellular components such as DNA.

The association of renin-angiotensinogen system genes polymorphisms and idiopathic recurrent pregnancy loss.

The most common complication of pregnancy is idiopathic recurrent pregnancy loss (RPL). To identify the contribution of gene polymorphisms to this condition, we evaluated the association between RPL and the angiotensinogen (AGT), angiotensin receptor 1 (AGTR1) and Angiotensinogen converting enzyme (ACE). In this case-control study, the frequency of AGT (rs4762 and rs699), AGTR1 (rs5186) and ACE insertion/deletion (rs4340) polymorphisms in 202 idiopathic RPL women was compared with 210 women with no history of abortion, using tetra-primer ARMS-PCR. Polymorphisms were analysed by logistic regression analysis according to inheritance models. The CT genotype of AGT rs4762, the CC genotype of AGT rs699 and the AC genotype of AGTR1 rs5186 in a co-dominant inheritance model were associated with idiopathic RPL (OR = 1.63, 95% CI = 1.07-2.49 of CT versus CC; OR = 5.97, 95% CI = 1.28-27.82 of CC versus TT; and OR = 1.99, 95% CI = 1.22-3.07 of AC versus AA). The allele frequency of AGT rs699 and AGTR1 rs5186 polymorphisms, but not AGT rs4762 and ACE rs4340 polymorphisms were significantly different between women with RPL patients and controls (p = 0.020, p = 0.003, p = 0.105 and p = 0.065, respectively). These results show that there is a significant relationship between AGT (rs699) and AGTR1 (rs5186) polymorphisms and idiopathic RPL in the Iranian population.

Novel Point Mutations in the NKX2.5 Gene in Pediatric Patients with Non-Familial Congenital Heart Disease.

Background and objective: Congenital heart disease (CHD) is the most common birth abnormality in the structure or function of the heart that affects approximately 1% of all newborns. Despite its prevalence and clinical importance, the etiology of CHD remains mainly unknown. Somatic and germline mutations in cardiac specific transcription factor genes have been identified as the factors responsible for various forms of CHD, particularly ventricular septal defects (VSDs), tetralogy of Fallot (TOF), and atrial septal defects (ASDs). p. NKX2.5 is a homeodomain protein that controls many of the physiological processes in cardiac development including specification and proliferation of cardiac precursors. The aim of our study was to evaluate the NKX2.5 gene mutations in sporadic pediatric patients with clinical diagnosis of congenital heart malformations. Materials and methods: In this study, we investigated mutations of the NKX2.5 gene's coding region in 105 Iranian pediatric patients with non-familial CHD by polymerase chain reaction-single stranded conformation polymorphism (PCR-SSCP) and direct sequencing. Results: We observed a total of four mutations, of which, two were novel DNA sequence variants in the coding region of exon 1 (c. 95 A > T and c. 93 A > T) and two others were previously reported as single-nucleotide polymorphisms (SNPs), namely rs72554028 (c. 2357 G > A) and rs3729753 (c. 606 G > C) in exon 2. Further, observed mutations are completely absent in normal healthy individuals (n = 92). Conclusion: These results suggest that NKX2.5 mutations are highly rare in CHD patients. However, in silico analysis proves that c.95 A > T missense mutation in NKX2.5 gene is probably pathogenic and may be contributing to the risk of sporadic CHD in the Iranian population.

Identification of a novel non-sense mutation in TBX5 gene in pediatric patients with congenital heart defects.

Introduction: Congenital heart diseases (CHDs) are structural cardiovascular malformations that arise from abnormal development of the heart during the prenatal life. Mutations in the TBX5 gene, encoding T-box transcription factor, are a major cause of CHD. To evaluate the TBX5 mutations in hotspot exons in sporadic pediatric patients with CHD phenotypes, analytical case/control study performed in an Iranian cohort of unrelated patients with clinical diagnosis of congenital heart malformations. Methods: We investigated TBX5 coding exons 4, 5, 6 and 7 in 95 sporadic patients with CHD phenotypes and compared to 82 healthy controls using PCR-SSCP and DNA sequencing approaches. Results: We report here on a novel and heterozygote Non-sense mutation in exon 5 of TBX5, E128X (G14742T), in two Iranian children. This mutation locates inside the T-box and both of pediatric patients carrying this novel mutation suffer from severe heart malformations. The G14742T mutation leads to the substitution of glutamic acid (E) by stop codon (X) at residue 128, an evolutionarily conserved position in T-box as well as in other species. The non-sense mutation of E128X was predicted to be pathogenic by Mutation Taster and Polyphen software programs. Conclusion: TBX5 E128X mutation results in a translational premature stop. This type of mutation results in a shortened protein that may function improperly and which cannot bind to other transcription factors; therefore, haploinsufficiency of TBX5 protein is presumably causing the severe cardiac malformations in these patients.

Novel and heteroplasmic mutations in mitochondrial tRNA genes in Brugada syndrome.

BACKGROUND: Brugada syndrome (BrS) is a rare cardiac arrhythmia characterized by sudden death associated with electrocardiogram patterns characterized by incomplete right bundle-branch block and ST-segment elevations in the anterior precordial leads. This syndrome predominantly is seen in younger males with structurally normal hearts. Mitochondrial variants particularly mt-tRNA mutations, are hot spots that lead to cardiological disorders. Previous studies have shown that mutations in mitochondrial tRNA genes play an important causal or modifying role in BrS. The present study aims to evaluate the involvement of mitochondrial tRNA genes in arrhythmogenic BrS. METHODS: In this study, 40 Iranian patients were investigated for the presence of the mutations in 6 mitochondrial tRNA genes (tRNA Ile, Met, Gln, Asn, Ala and Trp) by PCR-SSCP analysis. RESULTS: There were 4 mutations in tRNA genes, that for first time, were found in BrS patients and these mutations were not in controls. Three of them were heteroplasmic and located in tRNAGln (T4377A) and tRNAMet (G4407A and C4456T) which were assessed as pathogenic mutations. A homo-plasmic variant (5580T > C) in tRNATrp gene was located within the junction region between tRNATrp and tRNAAla genes. This mutation may disturb the processing of mt-tRNATrp. CONCLUSIONS: The results of this study suggest that mutations in mitochondrial tRNA genes might lead to deficiencies in translational process of critical proteins of the respiratory chain and potentially lead to BrS in Iranian subjects. (Cardiol J 2018; 25, 1: 113-119).

Molecular Analysis of rs2070744 and rs1799983 Polymorphisms of NOS3 Gene in Iranian Patients With Multiple Sclerosis.

INTRODUCTION: Multiple Sclerosis (MS) is a disease of central nervous system that mainly causes lesions or plaques in the spinal cord and brain. The purpose of this study was to analyze the relation between c.-813C>T (rs2070744) and c.894G>T (rs1799983) polymorphisms of NOS3 gene and MS in Iranian patients. METHODS: A total of 78 patients with MS and 80 healthy controls were screened for NOS3 (rs2070744 and rs1799983) Single Nucleotide Polymorphisms (SNPs) by tetra-primer multiplex ARMS-PCR and PCR-RFLP. RESULTS: Genotype frequencies of the c.-813C>T polymorphism in patients compared to controls were as follows: 53.8% to 80.0% for TT genotype, 41.0% to 18.8% for TC genotype, and 5.1% versus 1.2% for CC genotype (P=0.001). The frequencies of GG genotype was 57.7% and 78.8% and for GT genotype of c.894G>T polymorphism in patients compared to control subjects was 42.3% and 21.2%, respectively (P=0.004). CONCLUSION: Our results indicate that the studied NOS3 polymorphisms may be associated with MS in Iranian patients.