Impact of Administration Time and Kv7 Subchannels on the Cardioprotective Efficacy of Kv7 Channel Inhibition.

PURPOSE: The mechanism of cardioprotection by Kv7.1-5 (KCNQ1-5) channels inhibition by XE991 is unclear. We examined the impact of administration time on the cardioprotective efficacy of XE991, the involvement of key pro-survival kinases, and the importance of the Kv7 subchannels. METHODS: Isolated perfused rat hearts were divided into five groups: 1) vehicle, 2) pre-, 3) post- or 4) pre- and post-ischemic administration of XE991 or 5) chromanol 293B (Kv7.1 inhibitor) followed by infarct size quantification. HL-1 cells undergoing simulated ischemia/reperfusion were exposed to either a) vehicle, b) pre-, c) per-, d) post-ischemic administration of XE991 or pre-, per- and post-ischemic administration of e) XE991, f) Chromanol 293B or g) HMR1556 (Kv7.1 inhibitor). HL-1 cell injury was evaluated by propidium iodide/Hoechst staining. Pro-survival kinase activation of Akt, Erk and STAT3 in XE991-mediated HL-1 cell protection was evaluated using phosphokinase inhibitors. Kv7 subtype expression was examined by RT-PCR and qPCR. RESULTS: XE991, but not Chromanol 293B, reduced infarct size and improved hemodynamic recovery in all isolated heart groups. XE991 protected HL-1 cells when administered during simulated ischemia. Minor activation of the survival kinases was observed in cells exposed to XE991 but pharmacological inhibition of kinase activation did not reduce XE991-mediated protection. Kv7 subchannels 1-5 were all present in rat hearts but predominately Kv7.1 and Kv7.4 were present in HL-1 cells and selective Kv7.1 did not reduce ischemia/reperfusion injury. CONCLUSION: The cardioprotective efficacy of XE991 seems to depend on its presence during ischemia and early reperfusion and do not rely on RISK (p-Akt and p-Erk) and SAFE (p-STAT3) pathway activation. The protective effect of XE991 seems mainly mediated through the Kv7.4 subchannel.

Untargeted metabolomics reveals a mild impact of remote ischemic conditioning on the plasma metabolome and α-hydroxybutyrate as a possible cardioprotective factor and biomarker of tissue ischemia.

INTRODUCTION: Remote ischemic conditioning (RIC) is a maneuver by which short non-lethal ischemic events are applied on distant organs or limbs to reduce ischemia and reperfusion injuries caused by e.g. myocardial infarct. Although intensively investigated, the specific mechanism of this protective phenomenon remains incompletely understood and in particular, knowledge on the role of small metabolites is scarce. OBJECTIVES: In this study, we aimed to study perturbations in the plasma metabolome following RIC and gain insight into metabolic changes by the intervention as well as to identify potential novel cardio-protective metabolites. METHODS: Blood plasma samples from ten healthy males were collected prior to and after RIC and tested for bioactivity in a HL-1 based cellular model of ischemia-reperfusion damage. Following this, the plasma was analyzed using untargeted LC-qTOF-MS and regulated metabolites were identified using univariate and multivariate statistical analysis. Results were finally verified in a second plasma study from the same group of volunteers and by testing a metabolite ester in the HL-1 cell model. RESULTS: The analysis revealed a moderate impact on the plasma metabolome following RIC. One metabolite, α-hydroxybutyrate (AHB) however, stood out as highly significantly upregulated after RIC. AHB might be a novel and more sensitive plasma-biomarker of transient tissue ischemia than lactate. Importantly, it was also found that a cell permeable AHB precursor protects cardiomyocytes from ischemia-reperfusion damage. CONCLUSION: Untargeted metabolomics analysis of plasma following RIC has led to insight into metabolism during RIC and revealed a possible novel metabolite of relevance to ischemic-reperfusion damage.