Abstract
Epidemiological and preclinical studies have pointed out a correlation between hyperglycemia and increasing risk of heart failure and cardiac death. In cardiomyocytes, hyperglycemia has been shown to alter Ca2 + signalling via CaMKII. Yet, the underlying mechanisms are still unclear. To determine the molecular basis of high glucose-mediated Ca2 + mishandling. Ventricular cardiomyocytes were isolated from control and Epac2-KO mice. Parallel experiments were repeated in human cardiomyocytes differentiated from induced pluripotent stem cells (h-iPSC-CM). Ca2 + signaling was studied in cells loaded with a fluorescent Ca2 + dye and treated with high glucose (HG) ± ESI-05 (Epac2 inhibitor). We assessed ryanodine receptor (RyR2) phosphorylation state by western blot and measured single RyR2 channel activity incorporated into bilayers. HG-mediated SR Ca2 + leak and pro-arrhythmic events were prevented with Epac2 blocker (ESI-05) in normal mice cells and with Epac2 deletion (Epac2-KO) (Ca2 + sparks frequency N /100 μm/s: 0.23 ± 0.15 for HG + ESI-05, 0.09 ± 0.04 for Epac2-KO + HG vs. 0.16 ± 0.06 for normal glucose, P = N.S.). HG treatment increased RyR2 phosphorylation at the CaMKII site (S2814, ∼ 29% increase, P < 0.05) leading to a dramatic increase in RyR2 open probability (P0 = 0.76 ± 0.07 in HG vs. 0.22 ± 0.07 in NG, P < 0.01) fully prevented with ESI-05 (P0 = 0.15 ± 0.03, P = N.S.). Similarly, in h-iPSC-CM, chronic HG (1 week) increased the percentage of cells presenting pro-arrhythmic events (85% of cells). Electrically evoked [Ca2 + ]i transients were decreased in h-IPSC-CM (F/F0 = 2.10 ± 0.07 vs. 2.94 ± 0.17, P < 0.01) and associated to higher RyR-S2814 phosphorylation. Those effects were blunted by ESI-05. Our data suggests that HG alters Ca2 + signalling via Epac2–mediated pro-arrhythmic events due a CaMKII-dependent increase of RyR2 activity. Overtime, this newly described mechanism lowers systolic Ca2 + release as seen in diabetic-associated cardiomyopathy.
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