title: JTV519 (K201) reduces sarcoplasmic reticulum Ca²⁺ leak and improves diastolic function in vitro in murine and human non-failing myocardium.
abstract: OBJECTIVE
  Ca²⁺ leak from the sarcoplasmic reticulum (SR) via ryanodine receptors (RyR2s) contributes to cardiomyocyte dysfunction. RyR2 Ca²⁺ leak has been related to RyR2 phosphorylation. In these conditions, JTV519 (K201), a 1,4-benzothiazepine derivative and multi-channel blocker, stabilizes RyR2s and decrease SR Ca²⁺ leak. We investigated whether JTV519 stabilizes RyR2s without increasing RyR2 phosphorylation in mice and in non-failing human myocardium and explored underlying mechanisms.
  
  
  METHODS
  SR Ca²⁺ leak was induced by ouabain in murine cardiomyocytes. [Ca²⁺]-transients, SR Ca²⁺ load and RyR2-mediated Ca²⁺ leak (sparks/waves) were quantified, with or without JTV519 (1 µmol·L⁻¹). Contribution of Ca²⁺ -/calmodulin-dependent kinase II (CaMKII) was assessed by KN-93 and Western blot (RyR2-Ser(2814) phosphorylation). Effects of JTV519 on contractile force were investigated in non-failing human ventricular trabeculae.
  
  
  RESULTS
  Ouabain increased systolic and diastolic cytosolic [Ca²⁺](i) , SR [Ca²⁺], and SR Ca²⁺ leak (Ca²⁺ spark (SparkF) and Ca²⁺ wave frequency), independently of CaMKII and RyR-Ser(2814) phosphorylation. JTV519 decreased SparkF but also SR Ca²⁺ load. At matched SR [Ca²⁺], Ca²⁺ leak was significantly reduced by JTV519, but it had no effect on fractional Ca²⁺ release or Ca²⁺ wave propagation velocity. In human muscle, JTV519 was negatively inotropic at baseline but significantly enhanced ouabain-induced force and reduced its deleterious effects on diastolic function.
  
  
  CONCLUSIONS
  JTV519 was effective in reducing SR Ca²⁺ leak by specifically regulating RyR2 opening at diastolic [Ca²⁺](i) in the absence of increased RyR2 phosphorylation at Ser(2814) , extending the potential use of JTV519 to conditions of acute cellular Ca²⁺ overload.
authors.affiliation: Division of Cardiology, Medical University of Graz, Austria.
authors.forename: M
authors.initials: M
authors.lastname: Sacherer
authors.forename: S
authors.initials: S
authors.lastname: Sedej
authors.forename: P
authors.initials: P
authors.lastname: Wakuła
authors.forename: M
authors.initials: M
authors.lastname: Wallner
authors.forename: M A
authors.initials: MA
authors.lastname: Vos
authors.forename: J
authors.initials: J
authors.lastname: Kockskämper
authors.forename: P
authors.initials: P
authors.lastname: Stiegler
authors.forename: M
authors.initials: M
authors.lastname: Sereinigg
authors.forename: D
authors.initials: D
authors.lastname: von Lewinski
authors.forename: G
authors.initials: G
authors.lastname: Antoons
authors.forename: B M
authors.initials: BM
authors.lastname: Pieske
authors.forename: F R
authors.initials: FR
authors.lastname: Heinzel
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D003971:Diastole; D006352:Heart Ventricles / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D051379:Mice; D009206:Myocardium / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism; D010042:Ouabain / Q000494:pharmacology; D010766:Phosphorylation; D019837:Ryanodine Receptor Calcium Release Channel / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D013599:Systole; D013841:Thiazepines / Q000494:pharmacology*
chemical_list: D019837:Ryanodine Receptor Calcium Release Channel; D013841:Thiazepines; C109183:K201 compound; D010042:Ouabain; D002118:Calcium