Although aberrant reactivation of embryonic gene programs is intricately linked to pathological heart disease, the transcription factors driving these gene programs remain ill-defined. Here we report that increased calcineurin/Nfat signalling and decreased miR-25 expression integrate to re-express the basic helix-loop-helix (bHLH) transcription factor dHAND (also known as Hand2) in the diseased human and mouse myocardium. In line, mutant mice overexpressing Hand2 in otherwise healthy heart muscle cells developed a phenotype of pathological hypertrophy. Conversely, conditional gene-targeted Hand2 mice demonstrated a marked resistance to pressure-overload-induced hypertrophy, fibrosis, ventricular dysfunction and induction of a fetal gene program. Furthermore, in vivo inhibition of miR-25 by a specific antagomir evoked spontaneous cardiac dysfunction and sensitized the murine myocardium to heart failure in a Hand2-dependent manner. Our results reveal that signalling cascades integrate with microRNAs to induce the expression of the bHLH transcription factor Hand2 in the postnatal mammalian myocardium with impact on embryonic gene programs in heart failure.

Background: Lipid-soluble thiamine precursors have a much higher bioavailability than genuine thiamine and therefore are more suitable for therapeutic purposes. Benfotiamine (S-benzoylthiamine Omonophosphate), an amphiphilic S-acyl thiamine derivative, prevents the progression of diabetic complications, probably by increasing tissue levels of thiamine diphosphate and so enhancing transketolase activity. As the brain is particularly sensitive to thiamine deficiency, we wanted to test whether intracellular thiamine and thiamine phosphate levels are increased in the brain after oral benfotiamine administration.

Rationale:The mutation A341V in the S6 transmembrane segment of KCNQ1, the ␣-subunit of the slowly activating delayed-rectifier K ؉ (I Ks ) channel, predisposes to a severe long-QT1 syndrome with sympathetictriggered ventricular tachyarrhythmias and sudden cardiac death.Objective: Several genetic risk modifiers have been identified in A341V patients, but the molecular mechanisms underlying the pronounced repolarization phenotype, particularly during ␤-adrenergic receptor stimulation, remain unclear. We aimed to elucidate these mechanisms and provide new insights into control of cAMPdependent modulation of I Ks . Methods and Results:We characterized the effects of A341V on the I Ks macromolecular channel complex in transfected Chinese hamster ovary cells and found a dominant-negative suppression of cAMP-dependent Yotiao-mediated I Ks upregulation on top of a dominant-negative reduction in basal current. Phosphomimetic substitution of the N-terminal position S27 with aspartic acid rescued this loss of upregulation. Western blot analysis showed reduced phosphorylation of KCNQ1 at S27, even for heterozygous A341V, suggesting that phosphorylation defects in some (mutant) KCNQ1 subunits can completely suppress I Ks upregulation. Functional analyses of heterozygous KCNQ1 WT:G589D and heterozygous KCNQ1 WT:S27A, a phosphorylation-inert substitution, also showed such suppression. Immunoprecipitation of Yotiao with KCNQ1-A341V (in the presence of KCNE1) was not different from wild-type. Key Words: ion channels Ⅲ long-QT syndrome Ⅲ potassium Ⅲ torsade de pointes T he slowly activating delayed-rectifier K ϩ current (I Ks ) contributes importantly to cardiac repolarization. In large mammals, including humans, it has a small amplitude under basal isolated-myocyte conditions but forms a sizable repolarization reserve that is recruited when the action potential duration (APD) prolongs and during ␤-adrenergic receptor (␤AR) stimulation. 1,2 I Ks is carried by a macromolecular channel complex consisting of a homotetramer of pore-forming ␣-subunits encoded by KCNQ1 (Kv7.1), KCNE1 ␤-subunits, 3,4 and the regulatory A-kinase anchoring protein Yotiao, which binds to the KCNQ1 C-terminus. 5 There are also multiple interactions with other proteins. During ␤AR stimulation, when cAMP levels rise, phosphorylation of KCNQ1 at N-terminal position S27 is controlled by protein kinase A (PKA) and protein phosphatase 1 that are localized to the complex by Yotiao, thereby providing local control of I Ks enhancement. 5 Anchored PKA also phosphorylates Yotiao itself, thereby further enhancing I Ks . 6 An intact C-terminus of KCNE1 is critical for the PKA-dependent upregulation of I Ks . 7 Congenital defects (long-QT syndrome types 1 and 5; LQT1, LQT5; Jervell and Lange-Nielsen syndrome), pharmacological inhibition, 8 and acquired loss of I Ks , 9 can all result in QT-interval prolongation and enhanced susceptibility to ventricular tachyarrhythmias, notably torsade de pointes. These arrhythmias occur predominantly during conditions of exerc...

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Aims Mutation type, location, dominant-negative IKs reduction, and possibly loss of cyclic adenosine monophosphate (cAMP)-dependent IKs stimulation via protein kinase A (PKA) influence the clinical severity of long QT syndrome type 1 (LQT1). Given the malignancy of KCNQ1-p.A341V, we assessed whether mutations neighbouring p.A341V in the S6 channel segment could also increase arrhythmic risk. Methods and results Clinical and genetic data were obtained from 1316 LQT1 patients [450 families, 166 unique KCNQ1 mutations, including 277 p.A341V-positive subjects, 139 patients with p.A341-neighbouring mutations (91 missense, 48 non-missense), and 900 other LQT1 subjects]. A first cardiac event represented the primary endpoint. S6 segment missense variant characteristics, particularly cAMP stimulation responses, were analysed by cellular electrophysiology. p.A341-neighbouring mutation carriers had a QTc shorter than p.A341V carriers (477 ± 33 vs. 490 ± 44 ms) but longer than the remaining LQT1 patient population (467 ± 41 ms) (P < 0.05 for both). Similarly, the frequency of symptomatic subjects in the p.A341-neighbouring subgroup was intermediate between the other two groups (43% vs. 73% vs. 20%; P < 0.001). These differences in clinical severity can be explained, for p.A341V vs. p.A341-neighbouring mutations, by the p.A341V-specific impairment of IKs regulation. The differences between the p.A341-neighbouring subgroup and the rest of LQT1 mutations may be explained by the functional importance of the S6 segment for channel activation. Conclusion KCNQ1 S6 segment mutations surrounding p.A341 increase arrhythmic risk. p.A341V-specific loss of PKA-dependent IKs enhancement correlates with its phenotypic severity. Cellular studies providing further insights into IKs-channel regulation and knowledge of structure-function relationships could improve risk stratification. These findings impact on clinical management.