Humans ; Mutation ; NAV1.5 Voltage-Gated Sodium Channel/genetics*

Cardiomyopathy, Dilated ; genetics ; Humans ; NAV1.5 Voltage-Gated Sodium Channel ; Sodium Channels ; genetics

Heart ; Humans ; NAV1.5 Voltage-Gated Sodium Channel/metabolism* ; Protein Processing, Post-Translational

OBJECTIVETo investigate the association of desmoplakin with the distribution and function of Nav1.5 by RNA silencing technology in HL-1 cells.

METHODSHL-1 cells with desmoplakin expression suppression by RNA silencing were examined for desmoplakin and Nav1.5 protein expressions by Western blotting, and the distribution and co-location of desmoplakin and Nav1.5 protein were detected by immunofluorescence staining. Patch-clamp recording was applied to analyze the changes in whole-cell sodium current after desmoplakin silencing.

RESULTSCompared with the untreated group and negative control group, the cells with desmoplakin silencing showed obviously reduced expressions of desmoplakin and Nav1.5 proteins. Co-localization of desmoplakin and Nav1.5 was detected at cell-cell contact in untreated and control conditions, and desmoplakin expression silencing induced a drastic redistribution of Nav1.5 with decreased peak current density (156.3∓6.2 vs 41.8∓3.1, n=6, P<0.05), a shift in voltage dependence of steady-state inactivation (-42 mV vs -61 mV, n=5, P<0.05), and prolonged time of recovery from inactivation.

CONCLUSIONDesmoplakin silencing caused redistribution of Nav1.5 protein and also changes in its electrophysiological properties in HL-1 cells.

Animals ; Cell Line ; Desmoplakins ; genetics ; metabolism ; Gene Silencing ; Mice ; Mutation ; Myocytes, Cardiac ; metabolism ; NAV1.5 Voltage-Gated Sodium Channel ; metabolism

Recent several studies have shown that the genetic variation of SCN5A is related with atrioventricular conduction block (AVB); no study has yet been published in Koreans. Therefore, to determine the AVB-associated genetic variation in Korean patients, we investigated the genetic variation of SCN5A in Korean patients with AVB and compared with normal control subjects. We enrolled 113 patients with AVB and 80 normal controls with no cardiac symptoms. DNA was isolated from the peripheral blood, and all exons (exon 2-exon 28) except the untranslated region and exon-intron boundaries of the SCN5A gene were amplified by multiplex PCR and directly sequenced using an ABI PRISM 3100 Genetic Analyzer. When a variation was discovered in genomic DNA from AVB patients, we confirmed whether the same variation existed in the control genomic DNA. In the present study, a total of 7 genetic variations were detected in 113 AVB patients. Of the 7 variations, 5 (G87A-A29A, intervening sequence 9-3C>A, A1673G-H558R, G3578A-R1193Q, and T5457C-D1819D) have been reported in previous studies, and 2 (C48G-F16L and G3048A-T1016T) were novel variations that have not been reported. The 2 newly discovered variations were not found in the 80 normal controls. In addition, G298S, G514C, P1008S, G1406R, and D1595N, identified in other ethnic populations, were not detected in this study. We found 2 novel genetic variations in the SCN5A gene in Korean patients with AVB. However, further functional study might be needed.
Atrioventricular Block ; DNA ; Exons ; Genetic Variation ; Humans ; Introns ; Multiplex Polymerase Chain Reaction ; NAV1.5 Voltage-Gated Sodium Channel ; Untranslated Regions

Recent several studies have shown that the genetic variation of SCN5A is related with atrioventricular conduction block (AVB); no study has yet been published in Koreans. Therefore, to determine the AVB-associated genetic variation in Korean patients, we investigated the genetic variation of SCN5A in Korean patients with AVB and compared with normal control subjects. We enrolled 113 patients with AVB and 80 normal controls with no cardiac symptoms. DNA was isolated from the peripheral blood, and all exons (exon 2-exon 28) except the untranslated region and exon-intron boundaries of the SCN5A gene were amplified by multiplex PCR and directly sequenced using an ABI PRISM 3100 Genetic Analyzer. When a variation was discovered in genomic DNA from AVB patients, we confirmed whether the same variation existed in the control genomic DNA. In the present study, a total of 7 genetic variations were detected in 113 AVB patients. Of the 7 variations, 5 (G87A-A29A, intervening sequence 9-3C>A, A1673G-H558R, G3578A-R1193Q, and T5457C-D1819D) have been reported in previous studies, and 2 (C48G-F16L and G3048A-T1016T) were novel variations that have not been reported. The 2 newly discovered variations were not found in the 80 normal controls. In addition, G298S, G514C, P1008S, G1406R, and D1595N, identified in other ethnic populations, were not detected in this study. We found 2 novel genetic variations in the SCN5A gene in Korean patients with AVB. However, further functional study might be needed.
Atrioventricular Block ; DNA ; Exons ; Genetic Variation ; Humans ; Introns ; Multiplex Polymerase Chain Reaction ; NAV1.5 Voltage-Gated Sodium Channel ; Untranslated Regions

OBJECTIVEThree long QT syndrome(LQTS) pedigrees were brought together for genetic diagnosis by using short tandem repeat(STR) markers.

METHODSGenomic DNA was extracted from blood samples. STR markers (D7S1824, D7S2439, D7S483, D3S1298, D3S1767, D3S3521) in or spanning the HERG and SCN5A gene were amplified; the haplotype analysis for LQTS was performed.

RESULTSClinical diagnosis showed that 15 are LQTS patients (3 died) and 11 are probable patients. Linkage analysis showed that LQTS patients are linked with the SCN5A gene in family 1, HERG is linked with the disease in family 2 and 3. Fourteen gene carriers were identified, 2 patients and 7 probable patients were excluded.

CONCLUSIONLinkage analysis using STR markers can serve as useful tool for presymptomatic diagnosis.

ERG1 Potassium Channel ; Ether-A-Go-Go Potassium Channels ; Female ; Genetic Linkage ; Haplotypes ; Humans ; Long QT Syndrome ; genetics ; Male ; NAV1.5 Voltage-Gated Sodium Channel ; Pedigree ; Potassium Channels ; genetics ; Potassium Channels, Voltage-Gated ; Sodium Channels ; genetics ; Tandem Repeat Sequences

OBJECTIVETo detect the expression of voltage-gated Na(+) channel (NaCh) isoforms in rat sinoatrial node and explore their functions.

METHODSExpressions of NaCh isoforms Nav1.1, Nav1.2, Nav1.3, Nav1.5, Nav1.6 and Nav1.7 in the rat sinoatrial node were detected by immunohistochemistry. The functional roles of the NaChs were tested by observing the effect of tetrodotoxin, a specific blocker of NaChs, on the intrinsic heart rate of isolated rat working heart.

RESULTSThe tetrodotoxin- sensitive neuronal isoforms Nav1.1, Nav1.6 and Nav1.7 as well as the tetrodotoxin-resistant cardiac isoform Nav1.5 were present in the rat sinoatrial node, and the neuronal isoforms were more abundant than Nav1.5 (P<0.05). The selective blockade of tetrodotoxin-sensitive isoforms (presumably Nav1.1, Nav1.6 and Nav1.7) by 100 nmol/L tetrodotoxin scarcely affected the intrinsic heart rate (0.5-/+2.9%, P>0.05) while blockade of tetrodotoxin-resistant isoform (presumably Nav1.5) by 2 micromol/L tetrodotoxin resulted in an obvious decline in the intrinsic heart rate (22.1-/+2.1%, P<0.001).

CONCLUSIONSNav1.1, Nav1.5, Nav1.6 and Nav1.7 are all present in rat sinoatrial node. Although neuronal isoforms are more abundant, Nav1.5 seems to contribute more to activity of the sinoatrial node.

Animals ; Heart Rate ; drug effects ; physiology ; Immunohistochemistry ; Ion Channel Gating ; drug effects ; physiology ; Male ; NAV1.1 Voltage-Gated Sodium Channel ; NAV1.5 Voltage-Gated Sodium Channel ; NAV1.6 Voltage-Gated Sodium Channel ; Nerve Tissue Proteins ; biosynthesis ; Protein Isoforms ; biosynthesis ; Rats ; Sinoatrial Node ; drug effects ; metabolism ; physiology ; Sodium Channels ; biosynthesis ; Tetrodotoxin ; pharmacology

BACKGROUNDMutations in the cardiac sodium channel gene (SCN5A) may lead to a broad spectrum of familial arrhythmias, including long QT syndrome (LQTS), idiopathic ventricular fibrillation (IVF), and isolated cardiac conduction diseases. Recent studies have shown that polymorphisms in the SCN5A gene also play an important role in the manifestation of disorders involving cardiac excitability. In this study, we investigated the polymorphisms of the SCN5A gene in Han Chinese and its relation to Brugada syndrome (BS).

METHODSGenomic DNA was isolated from 120 unrelated healthy volunteers and 48 unrelated Brugada syndrome patients by means of standard procedures. All exons including the putative splicing sites of the SCN5A gene were amplified by PCR and sequenced directly or after subcloning using an ABI Prism 377 DNA sequencer.

RESULTSA total of 5 single nucleotide polymorphisms (SNPs) were identified in the Han Chinese population, including 3 novel ones: G87A(A29A), 4245 + 82A > G, and G6174A. The allele frequencies of each SNP in the Han Chinese population were as follows: G87A (A29A) 27.5%, A1673G (H558R) 10.4%, 4245 + 82A > G 32.8%, C5457T (D1819D) 41.3%, and G6174A 44.9%. S1102Y and 10 other SNPs identified in other ethnic populations were not detected in this study. There was no significant difference in the allele frequency of A1673G (H558R) between different ethnic populations (all P > 0.5). On the other hand, the allele frequency of C5457T (D1819D) among Han Chinese was similar to its frequency among Japanese (P > 0.5), but higher than that among Americans (P < 0.005). The allele G1673 (R558) was over-represented in BS patients compared to controls (P < 0.005), but there was no significant difference in genotype frequencies at this locus. There were also no differences in either the allele or genotype frequencies of the 4 other identified SNPs when comparing BS patients with healthy controls.

CONCLUSIONSThe distribution of SCN5A SNPs may vary between different ethnicities. The polymorphism of A1673G might be associated with BS and may contribute to a susceptibility to BS in Han Chinese.

Case-Control Studies ; China ; ethnology ; Gene Frequency ; Humans ; NAV1.5 Voltage-Gated Sodium Channel ; Polymorphism, Single Nucleotide ; Sodium Channels ; genetics ; Syndrome ; Ventricular Fibrillation ; genetics

OBJECTIVEBrugada syndrome is an inherited channelopathy that characterized by ST-segment elevation in the right precordial lead (V(1)-V(3)) on the electrocardiogram with or without right bundle branch block and related with high risk of sudden cardiac death and structurally normal hearts. The first and only gene linked to this disease is SCN5A, a gene encodes for alpha subunit of the cardiac sodium channel. The objective of this study is to explore SCN5A gene mutations in Chinese patients with Brugada syndrome.

METHODSFour patients diagnosed as Brugada syndrome and nine patients with suspected Brugada syndrome were chosen for the study. The exons in the functional regions of SCN5A gene were amplified with polymerase chain reaction and the amplified products were sequenced with Sanger method. If a mutation was identified, patient's family members were also screened.

RESULTSTwo heterozygous mutations were found in one family diagnosed as Brugada syndrome. One missense mutation was a G-->A transition in the first nucleotide of codon 95 in SCN5A gene exon 3, which was predicted to result in substitution of Valine with Isoleucine (V95I). The other missense mutation was a C-->T transition in the second nucleotide of codon 1649 in SCN5A gene exon 28, which was predicted to result in substitution of Alanine with Valine (A1649V). A heterozygous mutation was identified in one family suspected to have the disease. The mutation was a three nucleotides (TCT) deletion that caused Phenylalanine deletion in codon 1617 in SCN5A gene exon 28. The three mutations were not detected in 100 control chromosomes.

CONCLUSIONSMutation in SCN5A gene is one of the causes of Brugada syndrome in Chinese. Three novel SCN5A gene mutations were identified in Chinese with Brugada syndrome, which expands the spectrum of SCN5A mutations associated with the disease.

Adolescent ; Adult ; Aged ; Brugada Syndrome ; genetics ; Case-Control Studies ; Exons ; genetics ; Humans ; Male ; Middle Aged ; Muscle Proteins ; genetics ; Mutation ; NAV1.5 Voltage-Gated Sodium Channel ; Sodium Channels ; genetics