Screening for ryanodine receptor type 2 mutations in families with effort-induced polymorphic ventricular arrhythmias and sudden death: early diagnosis of asymptomatic carriers

OBJECTIVES: We sought to establish the role of genetic screening for ryanodine receptor type 2 (RyR2) gene mutations in families with effort-induced polymorphic ventricular arrhythmia (PVA), syncope and juvenile sudden death. BACKGROUND: The RyR2 mutations have been associated with PVA, syncope and sudden death in response to physical or emotional stress. METHODS: We studied 81 subjects (39 males and 42 females; mean age 31 +/- 20 years) belonging to eight families with pathogenic RyR2 mutations. All subjects underwent screening for RyR2 mutations, electrocardiography (ECG), 24-h Holter monitoring, signal-averaged electrocardiography (SAECG), two-dimensional echocardiography and exercise stress testing. Electrophysiologic (EP) study was performed in nine patients. RESULTS: Six different RyR2 mutations were found in eight families. Forty-three family members carried the gene mutation. Of these, 28 (65%) showed effort-induced arrhythmic symptoms or signs and one died suddenly during follow-up. Family history revealed 19 juvenile cases of sudden death during effort or emotion. In two families sharing the same mutation, no subject presented with PVA during the stress test; thus, sudden death and syncope were the only clinical manifestations. The 12-lead ECG was normal in all but two subjects, whereas five patients showed positive late potentials on the SAECG. In 17 (39.5%) of 43 subjects, the two-dimensional echocardiogram revealed localized kinetic abnormalities and mild structural alterations of the right ventricle. The EP study was not able to induce PVA. CONCLUSIONS: The absence of symptoms and PVA on the stress test in more than one-third of carriers of RyR2 mutations, as well as the lack of PVA inducibility by the EP study, underlies the importance of genetic screening for the early diagnosis of asymptomatic carriers and prevention of sudden death.     

Composite polymorphisms in the ryanodine receptor 2 gene associated with arrhythmogenic right ventricular cardiomyopathy

OBJECTIVE: Mutations in the cardiac ryanodine receptor (RYR2) gene have been reported to cause arrhythmogenic right ventricular cardiomyopathy (ARVC). The molecular mechanisms by which genetic modifications lead to ARVC are still not well understood. METHODS: ARVC patients were screened for mutations in the RYR2 gene by denaturing HPLC and DNA sequencing. Single channel measurements were carried out with RyR2 channels purified from explanted hearts of ARVC patients. RESULTS: None of the published RYR2 mutations were found in our ARVC-cohort. However, we identified two single nucleotide polymorphisms (SNPs) in exon 37 of the human RYR2 gene which lead to the amino acid exchanges G1885E and G1886S, respectively. Both SNPs together were found exclusively in 3 out of 85 ARVC patients in a composite heterozygous fashion (genotype T4). This genotype was associated with ARVC (p<0.05) but not with dilated cardiomyopathy (DCM, 79 patients) or none-failing controls (463 blood donors). However, either one of the two SNPs were identified in further 7 ARVC patients, in 11 DCM patients, and in 64 blood donors. The SNP leading to G1886S may create a protein kinase C phosphorylation site in the human RyR2. Single channel recordings at pCa4.3 revealed four conductance states for the RyR2 of genotype T4 and a single open state for the wild type RyR2. At pCa7.7, the lowest subconductance state of the RyR2 channel of genotype T4 persisted with a greatly enhanced open probability indicating a leaky channel. CONCLUSION: The RyR2 channel leak under diastolic conditions could cause SR-Ca2+ depletion, concomitantly arrhythmogenesis and heart failure in a subgroup of ARVC patients of genotype T4. A change in the RyR2 subunit composition due to the combined expression of both SNPs alters the behaviour of the tetrameric channel complex.     

Mutations in the cardiac ryanodine receptor gene (hRyR2) underlie catecholaminergic polymorphic ventricular tachycardia

BACKGROUND: Catecholaminergic polymorphic ventricular tachycardia is a genetic arrhythmogenic disorder characterized by stress-induced, bidirectional ventricular tachycardia that may degenerate into cardiac arrest and cause sudden death. The electrocardiographic pattern of this ventricular tachycardia closely resembles the arrhythmias associated with calcium overload and the delayed afterdepolarizations observed during digitalis toxicity. We speculated that a genetically determined abnormality of intracellular calcium handling might be the substrate of the disease; therefore, we considered the human cardiac ryanodine receptor gene (hRyR2) a likely candidate for this genetically transmitted arrhythmic disorder. METHODS AND RESULTS: Twelve patients presenting with typical catecholaminergic polymorphic ventricular tachycardia in the absence of structural heart abnormalities were identified. DNA was extracted from peripheral blood lymphocytes, and single-strand conformation polymorphism analysis was performed on polymerase chain reaction-amplified exons of the hRyR2 gene. Four single nucleotide substitutions leading to missense mutations were identified in 4 probands affected by the disease. Genetic analysis of the asymptomatic parents revealed that 3 probands carried de novo mutations. In 1 case, the identical twin of the proband died suddenly after having suffered syncopal episodes. The fourth mutation was identified in the proband, in 4 clinically affected family members, and in none of 3 nonaffected family members in a kindred with 2 sudden deaths that occurred at 16 and 14 years, respectively, in the sisters of the proband. CONCLUSIONS: We demonstrated that, in agreement with our hypothesis, hRyR2 is a gene responsible for catecholaminergic polymorphic ventricular tachycardia.     

慢性心力衰竭2型ryanodine受体过度磷酸化机制

慢性心力衰竭（心衰）是严重危害人类健康的重大疾患,约20％的患者在初次诊断后1年内死亡,约80％的患者在8年内死亡。因此,深入研究心衰的病理生理和分子机制,发现更为有效的治疗靶点仍具有十分重要的理论和现实意义。兴奋收缩耦联机制是心脏泵功能的生理基础,研究心衰的分子基础自然会联系到该机制。     

Calcium channel antagonism reduces exercise-induced ventricular arrhythmias in catecholaminergic polymorphic ventricular tachycardia patients with RyR2 mutations

Calcium channel antagonism in RyR2 defects. INTRODUCTION: Recently, gain-of-function mutations of cardiac ryanodine receptor RyR2 gene have been identified as a cause of familial or catecholaminergic polymorphic ventricular tachycardia. We examined the influence of the calcium channel blockers, verapamil and magnesium, on exercise-induced ventricular arrhythmias in patients with RyR2 mutations. METHODS AND RESULTS: Six molecularly defined catecholaminergic polymorphic ventricular tachycardia patients, all carrying a RyR2 mutation and on beta-adrenergic blocker therapy, underwent exercise stress test four times: at baseline, after verapamil and magnesium sulphate infusions, and finally, without interventions. The number of isolated and successive premature ventricular complexes during exercise ranged from 40 to 374 beats (mean 165 beats) at baseline, and was reduced during verapamil by 76+/-17% (P<0.05). Premature ventricular complexes appeared later and at higher heart rate during verapamil than at baseline (119+/-21 vs. 127+/-27 min-1, P<0.05). Magnesium did not inhibit the arrhythmias. Results in the fourth exercise stress test without interventions were similar to those in the first baseline study. CONCLUSIONS: This study provides the first in vivo demonstration that a calcium channel antagonist, verapamil, can suppress premature ventricular complexes and nonsustained ventricular salvoes in catecholaminergic polymorphic ventricular tachycardia caused by RyR2 mutations. Modifying the abnormal calcium handling by calcium antagonists might have therapeutic value.     

Functional heterogeneity of ryanodine receptor mutations associated with sudden cardiac death

OBJECTIVES: Point mutations in the cardiac ryanodine receptor (RyR2) mediate abnormal intracellular Ca(2+) release and are associated with stress-induced ventricular tachycardia (VT), leading to sudden cardiac death (SCD). Although the precise molecular basis of RyR2 dysfunction in SCD remains controversial, there is consensus that the mutations characterised to date all exhibit gain-of-function Ca(2+) release properties following cell stimulation. We investigated the functional impact of a distinct set of SCD-linked RyR2 mutations (L(433)P, N(2386)I, R(176)Q/T(2504)M) on intracellular Ca(2+) handling. METHODS: We expressed full-length recombinant human wild-type (WT) and SCD-linked RyR2 mutations in human embryonic kidney (HEK) cells, and profiled the spatial and amplitude characteristics of caffeine-evoked Ca(2+) release through homo-tetrameric channels in living cells using rapid confocal laser scanning microscopy. RESULTS: Analysis of the precise mode of Ca(2+) release in HEK cells expressing RyR2 mutants demonstrated profound differences when compared with WT channels. The SCD-linked RyR2 mutations characterised in this study exhibited heterogeneous Ca(2+) release profiles, including the novel observation that one of the mutants, (L(433)P), exhibited a marked reduction in sensitivity to channel activation. However, all SCD-linked RyR2 mutations characterised in this study resulted in an increased duration of elevated cytoplasmic Ca(2+) levels following channel activation. CONCLUSIONS: Our live cell-based data demonstrates functional heterogeneity of Ca(2+) release through SCD-linked RyR2 mutants, suggesting that the mechanistic basis of RyR2 dysfunction in SCD may be more complex than previously anticipated. These findings may have profound consequences for the therapeutic modulation of RyR2 in stress-induced VT and SCD.     

Elevated pain threshold in patients with effort-induced angina pectoris and asymptomatic myocardial ischemia during exercise test

The purpose of the present study was to analyze the prevalence of asymptomatic (silent) myocardial ischemia during exercise testing among patients with effort-induced angina pectoris, and further, to compare the pain threshold of patients with symptomatic and asymptomatic myocardial ischemia. A group of 26 patients comprised the study. In half of the patients myocardial ischemia during the exercise testing was silent and in one half it was symptomatic. Asymptomatic myocardial ischemia was defined as an asymptomatic ST-segment depression greater than or equal to 0.1 mV, lasting longer than 60 s during an exercise test. In patients with asymptomatic ischemia the pain thresholds both on toe and finger were significantly higher than in patients with symptomatic ischemia: mean values were 10.1 versus 4.9 mA on the toes, p less than 0.025, and 8.4 versus 2.5 mA on the fingers, p less than 0.01. We conclude that asymptomatic myocardial ischemia during exercise test is seen often in patients with angina pectoris and that this may be due to an increased pain threshold.     

Stabilization of cardiac ryanodine receptor prevents intracellular calcium leak and arrhythmias

Catecholaminergic polymorphic ventricular tachycardia is a form of exercise-induced sudden cardiac death that has been linked to mutations in the cardiac Ca2+ release channel/ryanodine receptor (RyR2) located on the sarcoplasmic reticulum (SR). We have shown that catecholaminergic polymorphic ventricular tachycardia-linked RyR2 mutations significantly decrease the binding affinity for calstabin-2 (FKBP12.6), a subunit that stabilizes the closed state of the channel. We have proposed that RyR2-mediated diastolic SR Ca2+ leak triggers ventricular tachycardia (VT) and sudden cardiac death. In calstabin-2-deficient mice, we have now documented diastolic SR Ca2+ leak, monophasic action potential alternans, and bidirectional VT. Calstabin-deficient cardiomyocytes exhibited SR Ca2+ leak-induced aberrant transient inward currents in diastole consistent with delayed after-depolarizations. The 1,4-benzothiazepine JTV519, which increases the binding affinity of calstabin-2 for RyR2, inhibited the diastolic SR Ca2+ leak, monophasic action potential alternans and triggered arrhythmias. Our data suggest that calstabin-2 deficiency is as a critical mediator of triggers that initiate cardiac arrhythmias.     

室性心律失常的基因治疗进展

近年来,随着以基因治疗为代表的生物治疗在单基因疾病领域取得成功,针对难治性的遗传性和获得性室性心律失常,基因治疗可能成为一种具有广阔应用前景的治疗选择.本文就该领域的研究进展进行概述.     

Unusual clinical presentation in a family with catecholaminergic polymorphic ventricular tachycardia due to a G14876A ryanodine receptor gene mutation

A family was identified, of whom which 11 members were carriers of the G14876A ryanodine 2 receptor mutation. All but 1 were symptomatic at the time of the study. Exercise testing showed bidirectional or polymorphic arrhythmias in 4 patients, whereas in 5 patients, it showed monomorphic or rare minor polymorphic ventricular arrhythmias. Two young patients died suddenly at rest while asleep. This study demonstrates that arrhythmias occurring during exercise stress testing in patients affected by catecholaminergic polymorphic ventricular tachycardia (CPVT) could be minor even in very symptomatic patients. The diagnosis of CPVT must be considered in these patients with a familial history of typical CPVT.     

Reflections on the diseases linked to mutations of the androgen receptor

This review summarizes the most recent information on two pathologies linked to mutations of the androgen receptor, namely, the complete androgen insensitivity syndrome (CAIS) and the spinal and bulbar muscular atrophy (SBMA or Kennedy's disease). Data on the clinical manifestations of the two diseases are presented, together with the most relevant findings on their physiopathology and genetics.