Effect of the long-acting insulin analogues glargine and degludec on cardiomyocyte cell signalling and function

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.