2+ ihandling proteins changes identified, an enhanced I NaL seems to be a major contributor to the electrophysiological and Ca

Cardiomyocytes derived from human embryonic stem cells constitute a promising cell source for the regeneration of damaged hearts. The assessment of their in vitro functional properties is mandatory to envisage appropriate cardiac cell-based therapies. In this study, we characterized human embryonic stem cell-derived cardiomyocytes over a 3-month period, using patch-clamp or intracellular recordings to assess their functional maturation and reverse transcriptase-polymerase chain reaction to evaluate the expression of ion channel-encoding subunits. I to1 and I K1 , the transient outward and inward rectifier potassium currents, were present in cardiomyocytes only, whereas the rapid delayed rectifier potassium current (I Kr ), pacemaker current (I f ), and L-type calcium current (I Ca,L ) could be recorded both in undifferentiated human embryonic stem cells and in cardiomyocytes. Most of the currents underwent developmental maturation in cardiomyocytes, as assessed by modifications in current density (I to1 , I K1 , and I Ca,L ) and properties (I f ). Ion-channel mRNAs were always present when the current was recorded. Intracellular recordings in spontaneously beating clusters of cardiomyocytes revealed changes in action potential parameters and in response to pharmacological tools according to time of differentiation. In summary, human embryonic stem cell-derived cardiomyocytes mature over time during in vitro differentiation, approaching an adult phenotype.

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I(f) is present in human LVMs. Its electrophysiological characteristics resemble those previously described in hypertrophied rat LVMs and suggest that I(f) could be an arrhythmogenic mechanism in patients with severe heart failure.

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3-Iodothyronamine T1AM is a novel endogenous thyroid hormone derivative that activates the G protein-coupled receptor known as trace anime-associated receptor 1 (TAAR1). In the isolated working rat heart and in rat cardiomyocytes, T1AM produced a reversible, dose-dependent negative inotropic effect (e.g., 27+/-5, 51+/-3, and 65+/-2% decrease in cardiac output at 19, 25, and 38 microM concentration, respectively). An independent negative chronotropic effect was also observed. The hemodynamic effects of T1AM were remarkably increased in the presence of the tyrosine kinase inhibitor genistein, whereas they were attenuated in the presence of the tyrosine phosphatase inhibitor vanadate. No effect was produced by inhibitors of protein kinase A, protein kinase C, calcium-calmodulin kinase II, phosphatidylinositol-3-kinase, or MAP kinases. Tissue cAMP levels were unchanged. In rat ventricular tissue, Western blot experiments with antiphosphotyrosine antibodies showed reduced phosphorylation of microsomal and cytosolic proteins after perfusion with synthetic T1AM; reverse transcriptase-polymerase chain reaction experiments revealed the presence of transcripts for at least 5 TAAR subtypes; specific and saturable binding of [125I]T1AM was observed, with a dissociation constant in the low micromolar range (5 microM); and endogenous T1AM was detectable by tandem mass spectrometry. In conclusion, our findings provide evidence for the existence of a novel aminergic system modulating cardiac function.

IntroductionHypertrophic cardiomyopathy (HCM) is a cause of sudden arrhythmic death, but the understanding of its pro-arrhythmic mechanisms and an effective pharmacological treatment are lacking. HCM electrophysiological remodelling includes both increased inward and reduced outward currents, but their role in promoting repolarisation abnormalities remains unknown. The goal of this study is to identify key ionic mechanisms driving repolarisation abnormalities in human HCM, and to evaluate anti-arrhythmic effects of single and multichannel inward current blocks.MethodsExperimental ionic current, action potential (AP) and Ca2 +-transient (CaT) recordings were used to construct populations of human non-diseased and HCM AP models (n = 9118), accounting for inter-subject variability. Simulations were conducted for several degrees of selective and combined inward current block.ResultsSimulated HCM cardiomyocytes exhibited prolonged AP and CaT, diastolic Ca2 + overload and decreased CaT amplitude, in agreement with experiments. Repolarisation abnormalities in HCM models were consistently driven by L-type Ca2 + current (ICaL) re-activation, and ICaL block was the most effective intervention to normalise repolarisation and diastolic Ca2 +, but compromised CaT amplitude. Late Na+ current (INaL) block partially abolished repolarisation abnormalities, with small impact on CaT. Na+/Ca2 + exchanger (INCX) block effectively restored repolarisation and CaT amplitude, but increased Ca2 + overload. Multichannel block increased efficacy in normalising repolarisation, AP biomarkers and CaT amplitude compared to selective block.ConclusionsExperimentally-calibrated populations of human AP models identify ICaL re-activation as the key mechanism for repolarisation abnormalities in HCM, and combined INCX, INaL and ICaL block as effective anti-arrhythmic therapies also able to partially reverse the HCM electrophysiological phenotype.