Dataset Information

Ajmaline blocks INa and IKr without eliciting differences between Brugada syndrome patient and control human pluripotent stem cell-derived cardiac clusters.

ABSTRACT: The class Ia anti-arrhythmic drug ajmaline is used clinically to unmask latent type I ECG in Brugada syndrome (BrS) patients, although its mode of action is poorly characterised. Our aims were to identify ajmaline's mode of action in human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes (CMs), and establish a simple BrS hiPSC platform to test whether differences in ajmaline response could be determined between BrS patients and controls. Control hiPSCs were differentiated into spontaneously contracting cardiac clusters. It was found using multi electrode array (MEA) that ajmaline treatment significantly lengthened cluster activation-recovery interval. Patch clamping of single CMs isolated from clusters revealed that ajmaline can block both INa and IKr. Following generation of hiPSC lines from BrS patients (absent of pathogenic SCN5A sodium channel mutations), analysis of hiPSC-CMs from patients and controls revealed that differentiation and action potential parameters were similar. Comparison of cardiac clusters by MEA showed that ajmaline lengthened activation-recovery interval consistently across all lines. We conclude that ajmaline can block both depolarisation and repolarisation of hiPSC-CMs at the cellular level, but that a more refined integrated tissue model may be necessary to elicit differences in its effect between BrS patients and controls.

SUBMITTER: Miller DC

PROVIDER: S-EPMC5727153 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Ajmaline blocks I<sub>Na</sub> and I<sub>Kr</sub> without eliciting differences between Brugada syndrome patient and control human pluripotent stem cell-derived cardiac clusters.

Miller Duncan C DC Harmer Stephen C SC Poliandri Ariel A Nobles Muriel M Edwards Elizabeth C EC Ware James S JS Sharp Tyson V TV McKay Tristan R TR Dunkel Leo L Lambiase Pier D PD Tinker Andrew A

Stem cell research 20171107

The class Ia anti-arrhythmic drug ajmaline is used clinically to unmask latent type I ECG in Brugada syndrome (BrS) patients, although its mode of action is poorly characterised. Our aims were to identify ajmaline's mode of action in human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes (CMs), and establish a simple BrS hiPSC platform to test whether differences in ajmaline response could be determined between BrS patients and controls. Control hiPSCs were differentiated into sponta ...[more]

PMID: 29172153

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Project description:Background The rate of sudden cardiac death (SCD) in Brugada syndrome (BrS) is ≈1%/y. Noninvasive electrocardiographic imaging is a noninvasive mapping system that has a role in assessing BrS depolarization and repolarization abnormalities. This study aimed to analyze electrocardiographic imaging parameters during ajmaline test (AJT). Methods and Results All consecutive epicardial maps of the right ventricle outflow tract (RVOT-EPI) in BrS with CardioInsight were retrospectively analyzed. (1) RVOT-EPI activation time (RVOT-AT); (2) RVOT-EPI recovery time, and (3) RVOT-EPI activation-recovery interval (RVOT-ARI) were calculated. ∆RVOT-AT, ∆RVOT-EPI recovery time, and ∆RVOT-ARI were defined as the difference in parameters before and after AJT. SCD-BrS patients were defined as individuals presenting a history of aborted SCD. Thirty-nine patients with BrS were retrospectively analyzed and 12 patients (30.8%) were SCD-BrS. After AJT, an increase in both RVOT-AT [105.9 milliseconds versus 65.8 milliseconds, P<0.001] and RVOT-EPI recovery time [403.4 milliseconds versus 365.7 milliseconds, P<0.001] was observed. No changes occurred in RVOT-ARI [297.5 milliseconds versus 299.9 milliseconds, P=0.7]. Before AJT no differences were observed between SCD-BrS and non SCD-BrS in RVOT-AT, RVOT-EPI recovery time, and RVOT-ARI (P=0.9, P=0.91, P=0.86, respectively). Following AJT, SCD-BrS patients showed higher RVOT-AT, higher ∆RVOT-AT, lower RVOT-ARI, and lower ∆RVOT-ARI (P<0.001, P<0.001, P=0.007, P=0.002, respectively). At the univariate logistic regression, predictors of SCD-BrS were the following: RVOT-AT after AJT (specificity: 0.74, sensitivity 1.00, area under the curve 0.92); ∆RVOT-AT (specificity: 0.74, sensitivity 0.92, area under the curve 0.86); RVOT-ARI after AJT (specificity 0.96, sensitivity 0.58, area under the curve 0.79), and ∆RVOT-ARI (specificity 0.85, sensitivity 0.67, area under the curve 0.76). Conclusions Noninvasive electrocardiographic imaging can be useful in evaluating the results of AJT in BrS.

Project description:AimsHuman-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have proven valuable for studies in drug discovery and safety, although limitations regarding their structural and electrophysiological characteristics persist. In this study, we investigated the electrophysiological properties of Pluricyte® CMs, a commercially available hiPSC-CMs line with a ventricular phenotype, and assessed arrhythmia incidence by IKr block at the single-cell and 2D monolayer level.Methods and resultsAction potentials were measured at different pacing frequencies, using dynamic clamp. Through voltage-clamp experiments, we determined the properties of INa, IKr, and ICaL. Intracellular Ca2+ measurements included Ca2+-transients at baseline and during caffeine perfusion. Effects of IKr block were assessed in single hiPSC-CMs and 2D monolayers (multi-electrode arrays). Action-potential duration (APD) and its rate dependence in Pluricyte® CMs were comparable to those reported for native human CMs. INa, IKr, and ICaL revealed amplitudes, kinetics, and voltage dependence of activation/inactivation similar to other hiPSC-CM lines and, to some extent, to native CMs. Near-physiological Ca2+-induced Ca2+ release, response to caffeine and excitation-contraction coupling gain characterized the cellular Ca2+-handling. Dofetilide prolonged the APD and field-potential duration, and induced early afterdepolarizations. Beat-to-beat variability of repolarization duration increased significantly before the first arrhythmic events in single Pluricyte® CMs and 2D monolayers, and predicted pending arrhythmias better than action-potential prolongation.ConclusionTaking their ion-current characteristics and Ca2+ handling into account, Pluricyte® CMs are suitable for in vitro studies on action potentials and field potentials. Beat-to-beat variability of repolarization duration proved useful to evaluate the dynamics of repolarization instability and demonstrated its significance as proarrhythmic marker in hiPSC-CMs during IKr block.

Project description:Host defense peptides (HDPs) are an integral part of the innate immune system with both antimicrobial and immunomodulatory activities. Induction of endogenous HDP synthesis is being actively explored as an antibiotic-alternative approach to disease control and prevention. Butyrate, a short-chain fatty acid, and forskolin, a phytochemical, have been shown separately to induce HDP gene expression in human cells. Here, we investigated the ability of butyrate and forskolin to induce the expressions of chicken HDP genes and the genes involved in barrier function such as mucin 2 and claudin 1 both in vitro and in vivo. We further evaluated their efficacy in protecting chickens from Clostridium perfringens-induced necrotic enteritis. Additionally, we profiled the transcriptome and global phosphorylation of chicken HD11 macrophage cells in response to butyrate and forskolin using RNA sequencing and a kinome peptide array, respectively. Our results showed a strong synergy between butyrate and forskolin in inducing the expressions of several, but not all, HDP genes. Importantly, dietary supplementation of butyrate and a forskolin-containing plant extract resulted in significant alleviation of intestinal lesions and the C. perfringens colonization in a synergistic manner in a chicken model of necrotic enteritis. RNA sequencing revealed a preferential increase in HDP and barrier function genes with no induction of proinflammatory cytokines in response to butyrate and forskolin. The antiinflammatory and barrier protective properties of butyrate and forskolin were further confirmed by the kinome peptide array. Moreover, we demonstrated an involvement of inducible cAMP early repressor (ICER)-mediated negative feedback in HDP induction by butyrate and forskolin. Overall, these results highlight a potential for developing butyrate and forskolin, two natural products, as novel antibiotic alternatives to enhance intestinal health and disease resistance in poultry and other animals.

Dataset Information

Ajmaline blocks INa and IKr without eliciting differences between Brugada syndrome patient and control human pluripotent stem cell-derived cardiac clusters.

Publications

Ajmaline blocks I<sub>Na</sub> and I<sub>Kr</sub> without eliciting differences between Brugada syndrome patient and control human pluripotent stem cell-derived cardiac clusters.

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