title: Effect of the long-acting insulin analogues glargine and degludec on cardiomyocyte cell signalling and function.
abstract: BACKGROUND
  The effects of insulin on cardiomyocytes, such as positive inotropic action and glucose uptake are well described. However, in vitro studies comparing long-acting insulin analogues with regard to cardiomyocyte signalling and function have not been systematically conducted.
  
  
  METHODS
  Insulin receptor (IR) binding was assessed using membrane embedded and solubilised IR preparations. Insulin signalling was analysed in adult rat ventricular myocytes (ARVM) and HL-1 cardiac cells. Inotropic effects were examined in ARVM and the contribution of Akt to this effect was assessed by specific inhibition with triciribine. Furthermore, beating-rate in Cor.4U(®) human cardiomyocytes, glucose uptake in HL-1 cells, and prevention from H2O2 induced caspase 3/7 activation in cardiac cells overexpressing the human insulin receptor (H9c2-E2) were analysed. One-way ANOVA was performed to determine significance between conditions.
  
  
  RESULTS
  Insulin degludec showed significant lower IR affinity in membrane embedded IR preparations. In HL-1 cardiomyocytes, stimulation with insulin degludec resulted in a lower Akt(Ser(473)) and Akt(Thr(308)) phosphorylation compared to insulin, insulin glargine and its active metabolite M1 after 5- and 10-min incubation. After 60-min treatment, phosphorylation of Akt was comparable for all insulin analogues. Stimulation of glucose uptake in HL-1 cells was increased by 40-60 %, with a similar result for all analogues. Incubation of electrically paced ARVM resulted for all insulins in a significantly increased sarcomere shortening, contractility- and relaxation-velocity. This positive inotropic effect of all insulins was Akt dependent. Additionally, in Cor.4U(®) cardiomyocytes a 10-20 % increased beating-rate was detected for all insulins, with slower onset of action in cells treated with insulin degludec. H9c2-E2 cells challenged with H2O2 showed a fivefold increase in caspase 3/7 activation, which could be abrogated by all insulins used.
  
  
  CONCLUSIONS
  In conclusion, we compared for the first time the signalling and functional impact of the long-acting insulin analogues insulin glargine and insulin degludec in cardiomyocyte cell models. We demonstrated similar efficacy under steady-state conditions relative to regular insulin in functional endpoint experiments. However, it remains to be shown how these results translate to the in vivo situation.
authors.affiliation: German Diabetes Center, Paul-Langerhans-Group for Integrative Physiology, Auf'm Hennekamp 65, 40225, Düsseldorf, Germany.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Hartmann
authors.affiliation: Institute for Clinical Biochemistry and Pathobiochemistry, German Diabetes Center, 40225, Düsseldorf, Germany.
authors.forename: Sabrina
authors.initials: S
authors.lastname: Overhagen
authors.affiliation: Institute for Clinical Biochemistry and Pathobiochemistry, German Diabetes Center, 40225, Düsseldorf, Germany.|German Center for Diabetes Research (DZD), Neuherberg, 85764, Munich, Germany.|Department of Endocrinology, Ghent University Hospital, 9000, Ghent, Belgium.
authors.forename: D Margriet
authors.initials: DM
authors.lastname: Ouwens
authors.affiliation: German Diabetes Center, Paul-Langerhans-Group for Integrative Physiology, Auf'm Hennekamp 65, 40225, Düsseldorf, Germany.
authors.forename: Silja
authors.initials: S
authors.lastname: Raschke
authors.affiliation: R&D Diabetes Division, Sanofi-Aventis Deutschland GmbH, 65929, Frankfurt, Germany.
authors.forename: Paulus
authors.initials: P
authors.lastname: Wohlfart
authors.affiliation: R&D Diabetes Division, Sanofi-Aventis Deutschland GmbH, 65929, Frankfurt, Germany.
authors.forename: Norbert
authors.initials: N
authors.lastname: Tennagels
authors.affiliation: German Diabetes Center, Paul-Langerhans-Group for Integrative Physiology, Auf'm Hennekamp 65, 40225, Düsseldorf, Germany.
authors.forename: Nina
authors.initials: N
authors.lastname: Wronkowitz
authors.affiliation: German Diabetes Center, Paul-Langerhans-Group for Integrative Physiology, Auf'm Hennekamp 65, 40225, Düsseldorf, Germany. eckel@uni-duesseldorf.de.|German Center for Diabetes Research (DZD), Neuherberg, 85764, Munich, Germany. eckel@uni-duesseldorf.de.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Eckel
mesh_terms: D000818:Animals; D001786:Blood Glucose / Q000187:drug effects*; D003922:Diabetes Mellitus, Type 1 / Q000378:metabolism; D007003:Hypoglycemia / Q000378:metabolism; D007004:Hypoglycemic Agents / Q000494:pharmacology*; D000069036:Insulin Glargine / Q000494:pharmacology*; D049528:Insulin, Long-Acting / Q000494:pharmacology*; D051379:Mice; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D011972:Receptor, Insulin / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*
chemical_list: D001786:Blood Glucose; D007004:Hypoglycemic Agents; D049528:Insulin, Long-Acting; D000069036:Insulin Glargine; C571886:insulin degludec; D011972:Receptor, Insulin
keywords: Cardiac action; Insulin analogues; Insulin degludec; Insulin glargine