Distinguishing pathogenic mutations from background genetic noise in cardiology: The use of large genome databases for genetic interpretation.

Advances in clinical genetic testing have led to increased insight into the human genome, including how challenging it is to interpret rare genetic variation. In some cases, the ability to detect genetic mutations exceeds the ability to understand their clinical impact, limiting the advantage of these technologies. Obstacles in genomic medicine are many and include: understanding the level of certainty/uncertainty behind pathogenicity determination, the numerous different variant interpretation-guidelines used by clinical laboratories, delivering the certain or uncertain result to the patient, helping patients evaluate medical decisions in light of uncertainty regarding the consequence of the findings. Through publication of large publicly available exome/genome databases, researchers and physicians are now able to highlight dubious variants previously associated with different cardiac traits. Also, continuous efforts through data sharing, international collaborative efforts to develop disease-gene-specific guidelines, and computational analyses using large data, will indubitably assist in better variant interpretation and classification. This article discusses the current, and quickly changing, state of variant interpretation resources within cardiovascular genetic research, e.g., publicly available databases and ways of how cardiovascular genetic counselors and geneticists can aid in improving variant interpretation in cardiology.

Cardiac repolarization and depolarization in people with Type 1 diabetes with normal ejection fraction and without known heart disease: a case-control study.

AIMS: To investigate depolarization and repolarization durations in people with Type 1 diabetes, including the relationship to age. METHODS: 855 persons with Type 1 diabetes without known heart disease were included and matched with 1710 participants from a general population study. Clinical examinations, questionnaires and biochemistry were assessed. A 10-second 12-lead ECG was performed and analysed digitally. RESULTS: QTc was longer in people with Type 1 diabetes compared to controls (414±16 vs. 411±19 ms, P <0.001), and particularly so in young people with Type 1 diabetes. The fully adjusted increase was 13.8 ms (95% confidence interval (CI): 8.6-19.0 ms, P <0.001) at age 20 years and 3.4 ms (CI: 1.5-5.3 ms, P<0.001) at age 40 years. The rate-corrected QRSc was increased in people with Type 1 diabetes (97±11 vs. 95±11 ms, P <0.001) and was age-independent (P =0.5). JTc was increased in the young people with Type 1 diabetes (10.7 ms (CI: 5.4-16.0 ms, P <0.001) at age 20 years), but not in older people with Type 1 diabetes (interaction age-diabetes, P <0.01). CONCLUSIONS: For people with Type 1 diabetes, cardiac depolarization is increased at all ages, whereas repolarization is increased only relatively in young people with Type 1 diabetes. Hence, young people with Type 1 diabetes may be more prone to ventricular arrhythmias. The findings contribute to the understanding of sudden cardiac death in young people with Type 1 diabetes.

Hidradenitis suppurativa and electrocardiographic changes: a cross-sectional population study.

BACKGROUND: Hidradenitis suppurativa (HS) is a chronic, inflammatory skin disease, recently associated with metabolic syndrome, subclinical atherosclerosis and increased risk of cardiovascular disease. OBJECTIVES: To investigate the hitherto unknown electrocardiographic (ECG) changes associated with HS, which have recently been associated with significant cardiovascular burden. METHODS: Data were obtained from the cross-sectional Danish General Population Study (GESUS). HS diagnosis was based on a validated self-reported questionnaire; 404 individuals met the HS diagnosis criteria and 19 001 controls without HS were identified. Severity of HS was staged according to a modified Hurley score. The ECG parameters of heart rate (HR), PR interval, QRS duration, JTc interval and QTc interval were obtained from 12-lead resting ECGs. We investigated the difference in means by unpaired t-test or anova. RESULTS: HR was significantly higher [mean difference 2·3 beats per min (bpm), 95% confidence interval (CI) 1·2-3·4; P < 0·01] when adjusting for age and sex, but when adjusting for multivariates, there was no significant difference (0·3 bpm, 95% CI -0·7 to 1·4; P = 0·52). Severe HS was significantly associated with increased HR across all models (2·9 bpm, 95% CI 0·7-5·1; P = 0·01). Mean QRS duration was significantly shorter in the group with mild HS but not in the groups with moderate and severe HS. CONCLUSIONS: Mean resting HR in patients with severe HS was significantly higher compared with controls. Given that resting HR is associated with increased risk of all-cause and cardiovascular mortality, and that patients with HS have increased risk of cardiovascular events, this finding is potentially important, easily testable and intervenable.

Effects of trimethoprim-sulfadiazine and detomidine on the function of equine Kv 11.1 channels in a two-electrode voltage-clamp (TEVC) oocyte model.

The long QT syndrome (LQTS) is a channelopathy that can lead to severe arrhythmia and sudden cardiac death. Pharmacologically induced LQTS is caused by interaction between drugs and potassium channels, especially the Kv 11.1 channel. Due to such interactions, numerous drugs have been withdrawn from the market or are administered with precautions in human medicine. However, some compounds, such as trimethoprim-sulfonamide combinations are still widely used in veterinarian medicine. Therefore, we investigate the effect of trimethoprim-sulfadiazine (TMS), trimethoprim, sulfadiazine, and detomidine on equine-specific Kv 11.1 channels. Kv 11.1 channels cloned from equine hearts were heterologously expressed in Xenopus laevis oocytes, and whole cell currents were measured by two-electrode voltage-clamp before and after drug application. TMS blocked equine Kv 11.1 current with an IC50 of 3.74 mm (95% CI: 2.95-4.73 mm) and affected the kinetics of activation and inactivation. Similar was found for trimethoprim but not for sulfadiazine, suggesting the effect is due to trimethoprim. Detomidine did not affect equine Kv 11.1 current. Thus, equine Kv 11.1 channels are also susceptible to pharmacological block, indicating that some drugs may have the potential to affect repolarization in horse. However, in vivo studies are needed to assess the potential risk of these drugs to induce equine LQTS.

Integration of 60,000 exomes and ACMG guidelines question the role of Catecholaminergic Polymorphic Ventricular Tachycardia-associated variants.

Catecholaminergic Polymorphic Ventricular Tachycardia (CPVT) is a highly lethal cardiac arrhythmia disease occurring during exercise or psychological stress. CPVT has an estimated prevalence of 1:10,000 and has mainly been associated with variants in calcium-regulating genes. Identification of potential false-positive pathogenic variants was conducted by searching the Exome Aggregation Consortium (ExAC) database (n = 60,706) for variants reported to be associated with CPVT. The pathogenicity of the interrogated variants was assessed using guidelines from the American College of Medical Genetics and Genomics (ACMG) and in silico prediction tools. Of 246 variants 38 (15%) variants previously associated with CPVT were identified in the ExAC database. We predicted the CPVT prevalence to be 1:132. The ACMG standards classified 29% of ExAC variants as pathogenic or likely pathogenic. The in silico predictions showed a reduced probability of disease-causing effect for the variants identified in the exome database (p < 0.001). We have observed a large overrepresentation of previously CPVT-associated variants in a large exome database. Based on the frequency of CPVT in the general population, it is less likely that the previously proposed variants are associated with a highly penetrant monogenic form of the disease.

T-wave morphology analysis of competitive athletes.

BACKGROUND: T-wave morphology has been shown to be more sensitive than QT and QTc interval to describe repolarization abnormalities. The electrocardiogram (ECG) performed in athletes may manifest abnormalities, including repolarization alterations. The aim of this study was to investigate the characteristics of T-wave morphology features in athletes. METHODS: Eighty male elite athletes, consisting of 40 Tour de France cyclists (age 27±5years), 40 soccer players (age 26±6years) and 40 healthy men (age 27±5years) were included. RESULTS: Sinus bradycardia, left ventricular (LV) hypertrophy, incomplete right bundle branch block and early repolarization were documented in 25 %, 20%, 13% and 14% of athletes, respectively. ECG criteria for LV hypertrophy in 12-lead ECG were more common in cyclists (35%) than in soccer players (5%), P<0.0001. Cyclists and soccer players had significantly longer RR interval, and repolarization features than the control group. CONCLUSIONS: T-wave morphology of athletes is different from non-athletes, depending of the sport. Decreased potassium current in cardiomyocytes associated with LVH may contribute to these changes.

The effect of sertindole on QTD and TPTE.

OBJECTIVE: Recent research suggests that other surrogate markers than QTc, including QTc dispersion and Tpeak-Tend, may better correlate with cardiac arrhythmia risk. While sertindole significantly prolongs the QTc interval, the effects on other markers of arrhythmia risk, such as QTc dispersion and Tpeak-Tend are unknown. METHOD: Digital 12-lead ECG was recorded at baseline and at steady-state in 37 patients switched to sertindole. ECG was analysed for Fridericia-corrected QT duration (QTcF), QT dispersion and Tpeak-Tend. RESULTS: From a baseline QTcF of 407 +/- 22 ms, mean QTcF prolongation during sertindole treatment was 20 +/- 23 ms, P < 0.01. No effect on QTc dispersion was found (-1 +/- 11 ms; P = 0.41). No increased duration of the Tpeak-Tend interval from baseline was found (+7 +/- 21 ms; P = 0.05). CONCLUSION: These findings might be related to the absence of confirmed Torsade de Pointes (TdP) cases related to sertindole exposure, despite sertindole's QTc prolonging effects.

Classification of the long-QT syndrome based on discriminant analysis of T-wave morphology.

The long QT syndrome (LQTS) is a genetic disorder, typically characterized by a prolonged QT interval in the ECG due to abnormal cardiac repolarization. LQTS may lead to syncopal episodes and sudden cardiac death. Various parameters based on T-wave morphology, as well as the QT interval itself have been shown to be useful discriminators, but no single ECG parameter has been sufficient to solve the diagnostic problem. In this study we present a method for discrimination among persons with a normal genotype and those with mutations in the KCNQ1 (KvLQT1 or LQT1) and KCNH2 (HERG or LQT2) genes on the basis of parameters describing T-wave morphology in terms of duration, asymmetry, flatness and amplitude. Discriminant analyses based on 4 or 5 parameters both resulted in perfect discrimination in a learning set of 36 subjects. In both cases cross-validation of the resulting classifiers showed no misclassifications either.

Left Ventricular Function After Prolonged Exercise in Equine Endurance Athletes.

BACKGROUND: Prolonged exercise in human athletes is associated with transient impairment of left ventricular (LV) function, known as cardiac fatigue. Cardiac effects of prolonged exercise in horses remain unknown. OBJECTIVES: To investigate the effects of prolonged exercise on LV systolic and diastolic function in horses. ANIMALS: Twenty-six horses competing in 120-160 km endurance rides. METHODS: Cross-sectional field study. Echocardiography was performed before and after rides, and the following morning, and included two-dimensional echocardiography, anatomical M-mode, pulsed-wave tissue Doppler imaging, and two-dimensional speckle tracking. Correlation between echocardiographic variables and cardiac troponin I was evaluated. RESULTS: Early diastolic myocardial velocities decreased significantly in longitudinal (baseline: -17.4 ± 2.4cm/s; end of ride: -15.8 ± 3.2cm/s (P = .013); morning after: -15.4 ± 3.0cm/s (P = .0033)) and radial directions (-32.8 ± 3.4cm/s; -28.1 ± 5.8cm/s (P < .001); -26.4 ± 5.5cm/s (P < .001)). Early diastolic strain rates decreased significantly in longitudinal (1.58 ± 0.27s(-1) ; 1.45 ± 0.26s(-1) (P = .036); 1.41 ± 0.25s(-1) (P = .013)) and circumferential directions (2.43 ± 0.29s(-1) ; 1.96 ± 0.46s(-1) (P < .001); 2.11 ± 0.32s(-1) (P < .001)). Systolic variables showed ambiguous results. No correlations with serum cardiac troponin I concentrations were evident. CONCLUSIONS AND CLINICAL IMPORTANCE: Prolonged exercise in horses is associated with impaired LV diastolic function. Reduced ventricular filling persisted for 7-21 hours despite normalization of biochemical indicators of hydration status, indicating that the observed changes were not entirely related to altered preload conditions. The clinical relevance of cardiac fatigue in horses remains uncertain.

Differences in the electrocardiographic QT interval of various breeds of athletic horses during rest and exercise.

OBJECTIVES: Quantitative measurements of cardiac repolarization, defined as the electrocardiographic QT interval, have important diagnostic implications in humans, as irregularities can trigger potentially fatal ventricular tachyarrhythmia. In both humans and horses, cardiac repolarization is influenced to some extent by heart rate, age, body weight (BW), sex, autonomic tone, and environment. In horses, there is substantial inter-breed variation in size and training, and the aims of this study were therefore to determine the best model describing the QT to RR relationship in breeds of various athletic horses and to test for differences in the QT interval. ANIMALS: Ten Icelandic horses, 10 Arabian horses, 10 Thoroughbreds, 10 Standardbreds, six Coldblood trotters, 10 Warmbloods (dressage) and 10 Warmbloods (show jumping). All horses were geldings. METHODS: QT intervals were measured from resting to peak exercise level and plotted against RR intervals. Data points were fitted with relevant regression models, and the effect of breed, BW, and estimated exercise intensity was examined. RESULTS: For all breeds in this study, the QT interval was best described as a function of RR by the piecewise linear regression model. The breed of horse had a significant effect on the model. There was no systematic effect of BW or estimated exercise intensity, but a high inter-horse variability was observed. CONCLUSIONS: The equine QT interval should preferably be corrected for heart rate according to breed. In addition, the results indicate that equine studies of the QT interval must be designed to eliminate the influence of a large inter-horse variation.

Short- and long-term variations in non-linear dynamics of heart rate variability.

OBJECTIVES: The purpose of the study was to investigate the short- and long-term variations in the non-linear dynamics of heart rate variability, and to determine the relationships between conventional time and frequency domain methods and the newer non-linear methods of characterizing heart rate variability. METHODS: Twelve healthy subjects were investigated by 3-h ambulatory ECG recordings repeated on 3 separate days. Correlation dimension, non-linear predictability, mean heart rate, and heart rate variability in the time and frequency domains were measured and compared with the results from corresponding surrogate time series. RESULTS: A small significant amount of non-linear dynamics exists in heart rate variability. Correlation dimensions and non-linear predictability are relatively specific parameters for each individual examined. The correlation dimension is inversely correlated to the heart rate and describes mainly linear correlations. Non-linear predictability is correlated with heart rate variability measured as the standard deviation of the R-R intervals and the respiratory activity expressed as power of the high-frequency band. The dynamics of heart rate variability changes suddenly even during resting, supine conditions. The abrupt changes are highly reproducible within the individual subjects. CONCLUSIONS: The study confirms that the correlation dimension of the R-R intervals is mostly due to linear correlations in the R-R intervals. A small but significant part is due to non-linear correlations between the R-R intervals. The different measures of heart rate variability (correlation dimension, average prediction error, and the standard deviation of the R-R intervals) characterize different properties of the signal, and are therefore not redundant measures. Heart rate variability cannot be described as a single chaotic system. Instead heart rate variability consists of intertwined periods with different non-linear dynamics. It is hypothesized that the heart rate is governed by a system with multiple "strange" attractors.

Recessive Romano-Ward syndrome associated with compound heterozygosity for two mutations in the KVLQT1 gene.

We describe a Swedish family with the proband and his brother suffering from severe Romano-Ward syndome (RWS) associated with compound heterozygosity for two mutations in the KVLQT1 (also known as KCNQ1 and KCNA9) gene (R518X and A525T). The mutations were found to segregate as heterozygotes in the maternal and the paternal lineage, respectively. None of the heterozygotes exhibited clinical long QT syndrome (LQTS). No hearing defects were found in the proband. The data strongly indicates that the compound heterozygosity for R518X and A525T is the cause of an autosomal recessive form of RWS in this family. Our findings support the implication of a higher frequency of gene carriers than previously expected. We suggest that relatives of 'sporadic RWS' patients should be considered potential carriers, at risk of dying suddenly from drug-induced LQTS.

Exaggerated natriuresis and lithium clearance in spontaneously hypertensive rats.

Since hypertension is associated with changes in the handling of various cations (including sodium and lithium) across the cell membrane, the present study investigated the validity of the lithium clearance method in hypertension by comparing two measures of proximal reabsorption. Thus, fractional lithium excretion and transit time (TT)-occlusion time (OT; e-TT/T) were determined successively in the same spontaneously hypertensive rat (SHR, Okamoto strain). The rats were examined both before and after an acute saline load. The results show that the lithium clearance method can be used for the determination of proximal reabsorption in SHR. Utilizing the lithium clearance method, the changes in renal sodium handling underlying the exaggerated natriuresis were investigated in unanaesthetized catheterized rats. It was found that the exaggerated natriuresis was associated with an increased output from the proximal tubule, whereas no difference in distal sodium handling could be detected between SHR and normotensive Wistar-Kyoto rats (WKY).

A biphasic response of urinary prostaglandin E2 excretion to water deprivation in conscious diabetes insipidus Brattleboro rats.

UNLABELLED: The effects of water deprivation on the urinary excretion rate of prostaglandin E2 (PGE2) were examined in conscious Brattleboro rats. In order to study the time course of the changes in the PGE2 excretory rate, urine was collected in 6 periods, CONTROL: 0-1 hour (h.). 1: 3-4.5 h., 8-10 h., III: 12-15 h., IV: 24-28 h. and V: 32-36 h. after removal of water and food. It was found that the PGE2 excretion rate changed in a biphasic pattern. During the first 2 experimental periods it increased. Thereafter it decreased towards the control value. There was an increase in PGE2 excretion with urinary flows down to 3 microliter/(min*100 g b. wt). At further reductions in urinary flow rate, PG excretion decreased towards basal levels.

A single strand conformation polymorphism/heteroduplex (SSCP/HD) method for detection of mutations in 15 exons of the KVLQT1 gene, associated with long QT syndrome.

Congenital long QT syndrome (LQTS) is characterised by prolongation of the QT interval on ECG and cardiac arrhythmias, syncopes and sudden death. A rapid and reliable genetic diagnosis of the disease may be of great importance for diagnosis and treatment of LQTS. Mutations in the KVLQT1 gene, encoding a potassium-channel subunit of importance for the depolarisation of cardiac myocytes, is believed to be associated with 50% of all LQTS cases. Our data confirms that KvLQT1 isoform 1 is encoded by 16 exons, and not 15, as reported previously. We have used genomic DNA sequences to design intronic PCR primers for amplification of 15 exons of KVLQT1 and optimised a non-radioactive single stranded conformation polymorphism/heteroduplex (SSCP/HD) method for detection of mutations in KVLQT1. The sensitivity of the method was 100% when it was tested on 15 in vitro constructed mutants. By multiplexing the PCR amplification of KVLQT1, it is possible to cover all 15 exons in four PCR reactions.

Detection of "noisy" chaos in a time series.

Time series from biological system often displays fluctuations in the measured variables. Much effort has been directed at determining whether this variability reflects deterministic chaos, or whether it is merely "noise". The output from most biological systems is probably the result of both the internal dynamics of the systems, and the input to the system from the surroundings. This implies that the system should be viewed as a mixed system with both stochastic and deterministic components. We present a method that appears to be useful in deciding whether determinism is present in a time series, and if this determinism has chaotic attributes. The method relies on fitting a nonlinear autoregressive model to the time series followed by an estimation of the characteristic exponents of the model over the observed probability distribution of states for the system. The method is tested by computer simulations, and applied to heart rate variability data.

Detection of chaotic determinism in time series from randomly forced maps.

Time series from biological system often display fluctuations in the measured variables. Much effort has been directed at determining whether this variability reflects deterministic chaos, or whether it is merely "noise". Despite this effort, it has been difficult to establish the presence of chaos in time series from biological sytems. The output from a biological system is probably the result of both its internal dynamics, and the input to the system from the surroundings. This implies that the system should be viewed as a mixed system with both stochastic and deterministic components. We present a method that appears to be useful in deciding whether determinism is present in a time series, and if this determinism has chaotic attributes, i.e., a positive characteristic exponent that leads to sensitivity to initial conditions. The method relies on fitting a nonlinear autoregressive model to the time series followed by an estimation of the characteristic exponents of the model over the observed probability distribution of states for the system. The method is tested by computer simulations, and applied to heart rate variability data.

Approximate entropy and point correlation dimension of heart rate variability in healthy subjects.

The contribution of nonlinear dynamics to heart rate variability in healthy humans was examined using surrogate data analysis. Several measures of heart rate variability were used and compared. Heart rates were recorded for three hours and original data sets of 8192 R-R intervals created. For each original data set (n = 34), three surrogate data sets were made by shuffling the order of the R-R intervals while retaining their linear correlations. The difference in heart rate variability between the original and surrogate data sets reflects the amount of nonlinear structure in the original data set. Heart rate variability was analyzed by two different nonlinear methods, point correlation dimension and approximate entropy. Nonlinearity, though under 10 percent, could be detected with both types of heart rate variability measures. More importantly, not only were the correlations between these measures and the standard deviation of the R-R intervals weak, the correlation among the nonlinear measures themselves was also weak (generally less than 0.6). This suggests that in addition to standard linear measures of heart rate variability, the use of multiple nonlinear measures of heart rate variability might be useful in monitoring heart rate dynamics.

[Molecular genetics of the long QT syndrome. Genes causing syncope and sudden death]. / Molekylärgenetik vid långt QT-syndrom. Gener som orsakar svimning och plötslig död.

Molecular genetic studies in congenital long QT syndrome have characterized genes and mechanisms of arrhythmias. At least six genes encoding cardiac potassium and sodium ionic channels have been described with several mutations in each gene. The altered function produces abnormal cardiac repolarization seen on ECG as prolongation of the QT-interval and T-wave abnormalities. This may increase the propensity for ventricular arrhythmias such as Torsade de Pointe, the cause of unexpected syncope and sudden death in young patients. Clinical manifestations vary depending on the genotype present. Gene-specific therapies have recently been tried. Initial therapy of choice for symptomatic and also asymptomatic children is administration of beta-blockers.