Jianding Cheng

α1-Syntrophin Mutations Identified in Sudden Infant Death Syndrome Cause an Increase in Late Cardiac Sodium Current

Background—Sudden infant death syndrome (SIDS) is a leading cause of death during the first 6 months after birth. About 5% to 10% of SIDS may stem from cardiac channelopathies such as long-QT syndrome. We recently implicated mutations in &agr;1-syntrophin (SNTA1) as a novel cause of long-QT syndrome, whereby mutant SNTA1 released inhibition of associated neuronal nitric oxide synthase by the plasma membrane Ca-ATPase PMCA4b, causing increased peak and late sodium current (INa) via S-nitrosylation of the cardiac sodium channel. This study determined the prevalence and functional properties of SIDS-associated SNTA1 mutations. Methods and Results—Using polymerase chain reaction, denaturing high-performance liquid chromatography, and DNA sequencing of SNTA1s open reading frame, 6 rare (absent in 800 reference alleles) missense mutations (G54R, P56S, T262P, S287R, T372M, and G460S) were identified in 8 (≈3%) of 292 SIDS cases. These mutations were engineered using polymerase chain reaction–based overlap extension and were coexpressed heterologously with SCN5A, neuronal nitric oxide synthase, and PMCA4b in HEK293 cells. INa was recorded using the whole-cell method. A significant 1.4- to 1.5-fold increase in peak INa and 2.3- to 2.7-fold increase in late INa compared with controls was evident for S287R-, T372M-, and G460S-SNTA1 and was reversed by a neuronal nitric oxide synthase inhibitor. These 3 mutations also caused a significant depolarizing shift in channel inactivation, thereby increasing the overlap of the activation and inactivation curves to increase window current. Conclusions—Abnormal biophysical phenotypes implicate mutations in SNTA1 as a novel pathogenic mechanism for the subset of channelopathic SIDS. Functional studies are essential to distinguish pathogenic perturbations in channel interacting proteins such as &agr;1-syntrophin from similarly rare but innocuous ones.

Alpha1-syntrophin mutations identified in sudden infant death syndrome cause an increase in late cardiac sodium current.

Background—Sudden infant death syndrome (SIDS) is a leading cause of death during the first 6 months after birth. About 5% to 10% of SIDS may stem from cardiac channelopathies such as long-QT syndrome. We recently implicated mutations in &agr;1-syntrophin (SNTA1) as a novel cause of long-QT syndrome, whereby mutant SNTA1 released inhibition of associated neuronal nitric oxide synthase by the plasma membrane Ca-ATPase PMCA4b, causing increased peak and late sodium current (INa) via S-nitrosylation of the cardiac sodium channel. This study determined the prevalence and functional properties of SIDS-associated SNTA1 mutations. Methods and Results—Using polymerase chain reaction, denaturing high-performance liquid chromatography, and DNA sequencing of SNTA1s open reading frame, 6 rare (absent in 800 reference alleles) missense mutations (G54R, P56S, T262P, S287R, T372M, and G460S) were identified in 8 (≈3%) of 292 SIDS cases. These mutations were engineered using polymerase chain reaction–based overlap extension and were coexpressed heterologously with SCN5A, neuronal nitric oxide synthase, and PMCA4b in HEK293 cells. INa was recorded using the whole-cell method. A significant 1.4- to 1.5-fold increase in peak INa and 2.3- to 2.7-fold increase in late INa compared with controls was evident for S287R-, T372M-, and G460S-SNTA1 and was reversed by a neuronal nitric oxide synthase inhibitor. These 3 mutations also caused a significant depolarizing shift in channel inactivation, thereby increasing the overlap of the activation and inactivation curves to increase window current. Conclusions—Abnormal biophysical phenotypes implicate mutations in SNTA1 as a novel pathogenic mechanism for the subset of channelopathic SIDS. Functional studies are essential to distinguish pathogenic perturbations in channel interacting proteins such as &agr;1-syntrophin from similarly rare but innocuous ones.

SCN5A Rare Variants in Familial Dilated Cardiomyopathy Decrease Peak Sodium Current Depending on the Common Polymorphism H558R and Common Splice Variant Q1077del

Obtaining functional data with newly identified rare variants increases certainty that the variant identified is relevant for dilated cardiomyopathy (DCM) causation. Two novel SCN5A rare variants, R222Q and I1835T, segregated with DCM in two families with affected individuals homozygous or heterozygous for the common SCN5A polymorphism H558R. cDNAs with each rare variant were constructed in the common Q1077del or Q1077 splice variant backgrounds with and without the H558R polymorphism and expressed in HEK293 cells. Sodium current (INa) was studied for each using whole‐cell voltage clamp. In the Q1077del background INa densities of R222Q and I1835T were not different from wild type, but the combined variants of R222Q/H558R, I1835T/H558R caused approximately 35% and approximately 30% reduction, respectively, and each showed slower recovery from inactivation. In the Q1077del background R222Q and R222Q/H558R also exhibited a significant negative shift in both activation and inactivation while I1835T/H558R showed a significant negative shift in inactivation that tended to decrease window current. In contrast, expression in the Q1077 background showed no changes in peak INa densities, decay, or recovery from inactivation for R222Q/H558R and I1835T/H558R. We conclude that the biophysical findings, dependent upon common SCN5A variants, provide further evidence that these novel SCN5A rare variants are relevant for DCM. Clin Trans Sci 2010; Volume 3: 287–294

Is sudden unexplained nocturnal death syndrome in Southern China a cardiac sodium channel dysfunction disorder

Sudden unexplained nocturnal death syndrome (SUNDS) remains an enigma to both forensic pathologists and physicians. Previous epidemiological, clinical, and pilot genetic studies have implicated that SUNDS is most likely a disease allelic to Brugada syndrome (BrS). We have performed postmortem genetic testing to address the spectrum and role of genetic abnormalities in the SCN5A-encoded cardiac sodium channel and its several associated proteins in SUNDS victims from Southern China. Genomic DNA extracted from the blood samples of 123 medico-legal autopsy-negative SUNDS cases and 104 sex-, age- and ethnic-matched controls from Southern China underwent comprehensive amino acid coding region mutational analysis for the BrS associated genes SCN5A, SCN1B, SCN2B, SCN3B, SCN4B, MOG1, and GPD1-L using PCR and direct sequencing. We identified a total of 7 unique (4 novel) putative pathogenic mutations (all in SCN5A; V95I, R121Q [2 cases], R367H, R513H, D870H, V1764D, and S1937F) in 8/123 (6.5%) SUNDS cases. Three SCN5A mutations (V95I, R121Q, and R367H) have been previously implicated in BrS. An additional 8 cases hosted rare variants of uncertain clinical significance (SCN5A: V1098L, V1202M, R1512W; SCN1B: V138I [3 cases], T189M [2 cases]; SCN3B: A195T). There were no non-synonymous mutations found in SCN2B, SCN4B, MOG1, or GPD1-L. This first comprehensive genotyping for SCN5A and related genes in the Chinese Han population with SUNDS discovered 13 mutations, 4 of them novel, in 16 cases, which suggests cardiac sodium channel dysfunction might account for the pathogenesis of 7-13% of SUNDS in Southern China.

Sudden unexplained nocturnal death syndrome in Southern China: an epidemiological survey and SCN5A gene screening.

Abstract Based on autopsy data collected in Southern China from 2001–2006, 975 cases of sudden unexplained nocturnal death syndrome (SUNDS) were surveyed. Genetic screening of SCN5A gene encoding the voltage dependent cardiac sodium channel was performed in 74 SUNDS cases. The annual occurrence rate of SUNDS in the area was estimated to be about 1 per 100,000 people. About 80.6% of deaths occurred between the ages of 21 to 40 years and the case number peaked at age 30 years. In 75.4% of cases with witnesses, victims died asleep between 11 PM and 4 AM and they showed predominantly abrupt respiratory distress shortly preceding death. The monthly distribution of emergency fever cases in the area during the same period was positively correlated to that of SUNDS cases (rs = 0.611, P = 0.035). Four polymorphisms in SCN5A were identified in both SUNDS and control groups. Compared with controls, the allele frequency of C5457 and C3666 + 69 were significant higher in SUNDS (P < 0.005) while the genotypes of both 5457CC (P = 0.012, OR = 2.0, 95% CI = 1.3–3.2) and 3666+69CC (P = 0.004, OR = 2.1, 95% CI = 1.3–3.3) in SUNDS cases were significantly higher. This is the first report of an epidemiological survey and SCN5A gene screening in SUNDS in the Han population of China. The genotypes of 5457CC and 3666+69CC in SCN5A gene may be Chinese SUNDS susceptible polymorphisms.

Identification of rare variants of DSP gene in sudden unexplained nocturnal death syndrome in the southern Chinese Han population.

Sudden unexplained nocturnal death syndrome (SUNDS) is a perplexing disorder to both forensic pathologists and clinic physicians. Desmoplakin (DSP) gene was the first desmosomal gene linked to arrhythmogenic right ventricular cardiomyopathy (ARVC) which was associated with sudden death. To identify the genetic variants of the DSP gene in SUNDS in the southern Chinese Han population, we genetically screened the DSP gene in 40 sporadic SUNDS victims, 16 Brugada syndrome (BrS) patients, and 2 early repolarization syndrome (ERS) patients using next generation sequencing (NSG) and direct Sanger sequencing. A total of 10 genetic variants of the DSP gene were detected in 11 cases, comprised of two novel missense mutations (p.I125F and p.D521A) and eight previously reported rare variants. Of eight reported variants, two were previously considered pathogenic (p.Q90R and p.R2639Q), three were predicted in silico to be pathogenic (p.R315C, p.E1357D and p.D2579H), and the rest three were predicted to be benign (p.N1234S, p.R1308Q, and p.T2267S). This is the first report of DSP genetic screening in Chinese SUNDS and Brugada syndrome. Our results imply that DSP mutations contribute to the genetic cause of some SUNDS victims and maybe a new susceptible gene for Brugada syndrome.

Postmortem molecular analysis of KCNQ1, KCNH2, KCNE1 and KCNE2 genes in sudden unexplained nocturnal death syndrome in the Chinese Han population

The etiology of sudden unexplained nocturnal death syndrome (SUNDS) remains unclear. Previous studies have implicated that SUNDS is probably allelic to cardiac sodium channel diseases such as Brugada syndrome. The variation in cardiac potassium channels is the main genetic cause of inherited long QT syndrome (LQTS), which may manifest as syncope and sudden cardiac death without structural disease. We hypothesized that cardiac potassium channel disease may be responsible for certain Chinese SUNDS cases. Genotyping of 4 main LQTS-susceptibility genes (KCNQ1, KCNH2, KCNE1, and KCNE2) was performed here for the first time in SUNDS victims from the Chinese Han population to address the pathogenic cause of some SUNDS using polymerase chain reaction and direct DNA sequencing. 120 sporadic SUNDS cases were enrolled. Genomic DNA was extracted from blood samples. A total of 2 novel non-synonymous mutations and 3 previously reported arrhythmia susceptibility polymorphisms were identified in KCNQ1, KCNH2, KCNE1, and KCNE2. We concluded that the variants in KCNQ1, KCNH2, KCNE1 and KCNE2 genes may be correlated with the occurrence of part of SUNDS cases in southern China.

Postmortem genetic screening of SNPs in RyR2 gene in sudden unexplained nocturnal death syndrome in the southern Chinese Han population

To investigate the genetic variants of the RyR2 gene in sudden unexplained nocturnal death syndrome (SUNDS) in the southern Chinese Han population, we genetically screened 29 of the 105 coding exons of the RyR2 gene associated with catecholaminergic polymorphic ventricular tachycardia (CPVT) and arrhythmogenic right ventricular cardiomyopathy (ARVC) in sporadic SUNDS victims using polymerase chain reaction (PCR) and direct sequencing methods. Genomic DNA was extracted from blood samples of 127 SUNDS cases and 165 healthy unrelated controls. None of the published or novel RyR2 missense mutations were found in 127 SUNDS cases. A total of sixteen genetic variants of the RyR2 gene were identified, comprised of: one novel synonymous coding mutation (c.13710C>A), one novel synonymous rare polymorphism (c.14871C>T), and fourteen previously reported polymorphisms. The genotype and allele frequency of previously reported missense polymorphism c.5656G>A (G1886S) was of no statistical difference between SUNDS cases and controls (x(2)=0.390, P>0.05; x(2)=0.271, P>0.05). This is the first report of genetic phenotype of RyR2 gene of SUNDS in the southern Chinese Han population. Previously reported plausible pathogenic missense polymorphism G1886S may not be an independent predisposition factor of SUNDS in the southern Chinese Han population. The association of genetic variants of the RyR2 gene with SUNDS needs further elucidation.

A case-control study of sudden unexplained nocturnal death syndrome in the southern Chinese Han population.

AbstractTo study the epidemiological characteristics of sudden unexplained nocturnal death syndrome (SUNDS) in the southern Chinese Han population during 2007 to 2013, we gathered 879 SUNDS victims from Dongguan City and in the Longgang District in Shenzhen City as the case group then selected 879 all-cause death cases, adopting a 1:1 pair method, as the control group I and collected 8142 all-cause death cases from the Bao’an District in Shenzhen City as the control group II, simultaneously. Case information collected was statistically analyzed. The annual incidence of SUNDS is 1.02 and 2.23 per 100,000 person-years for Dongguan City and in the Longgang District, respectively. The number of male and female victims is drastically different, with a ratio of 13.92:1, whereas the incidence between the 2 sexes is significantly different (&khgr;2 = 78.734, P < 0.01), with an odds ratio value of 11.32 (95% confidence interval, 5.75–22.28). The age of death of SUNDS cases ranges from 17 to 55 years with a median age of 35 years; furthermore, the difference of distribution of age of death between the SUNDS victims and the all-cause death population is significant (&khgr;2 = 767.12, P < 0.001). The birthplace of SUNDS victims is distributed throughout 27 provinces of China, but the difference between the SUNDS victims and the all-cause death population is not significant (&khgr;2 = 27.273, P > 0.05). The monthly incidence of SUNDS is relatively higher from March to June, whereas the difference of monthly distribution between SUNDS victims and all-cause death population is significant (&khgr;2 = 9.869, P < 0.05), with an odds ratio value of 1.42 (95% confidence interval, 1.14–1.76). Although the majority of SUNDS occurred during midnight sleep, they were mostly discovered from 7 to 9 am once the inmates or spouses woke in the morning. A total of 97.74% of the SUNDS victims were blue-collar factory workers with a high-intensity labor and poor education background. This investigation confirmed the stability of epidemiological characteristics of SUNDS in South China and implicated that risk factors of this fatal disease still exist. The efficient strategy of early identification such as molecular diagnosis for SUNDS is extremely urgently required.

Molecular Autopsy of Desmosomal Protein Plakophilin‐2 in Sudden Unexplained Nocturnal Death Syndrome

Plakophilin‐2 (PKP2) variants could produce a phenotype of Brugada syndrome (BrS), which seems to be most likely the same allelic disorder as some sudden unexplained nocturnal death syndrome (SUNDS). All coding regions of PKP2 gene in 119 SUNDS victims were genetically screened using PCR and direct Sanger sequencing methods. Three novel mutations (p.Ala159Thr, p.Val200Val, and p.Gly265Glu), one novel rare polymorphism (p.Thr723Thr), and eight reported polymorphisms were identified. A compound mutation (p.Ala159Thr and p.Gly265Glu) and a rare polymorphism (p.Thr723Thr) were found in one SUNDS case with absence of the cardiomyopathic features. The detected compound mutation identified in this first investigation of PKP2 genetic phenotype in SUNDS is regarded as the plausible genetic cause of this SUNDS case. The rare incidence of PKP2 mutation in SUNDS (1%) supports the previous viewpoint that SUNDS is most likely an allelic disorder as BrS.