Figure 4.

GSK3β inhibition at reoxygenation reverses the activation of IP₃R induced by HR in adult cardiomyocytes. (a) Cell viability measured with propidium iodide revealed that SB21 clearly protected adult cardiomyocytes against reoxygenation injury (n=4 distinct experiments per group); mean±S.E.M. of cell death is shown as percentage. *P<0.05. (b) GSK3β activity quantified by western immunoblotting of phospho-GSK3β (Tyr₂₁₆, active form) under hypoxic conditions. Results were normalized with total GSK3β (n=4–5 distinct experiments per group). *P<0.05. (c) Immunoblotting of relative expression of phosphor-Ser₁₇₅₆ IP₃R normalized with total IP₃R during HR injury. Results expressed as fold versus basal are means±S.E.M. *P<0.05 (n=4 distinct experiments). (d) In situ protein interactions using PLA in adult cardiomyocytes revealed that SB21 (6 μM) significantly reduced the interaction of GSK3β with IP₃R induced by HR. Results are expressed as means±S.E.M. of 8–10 distinct experiments (each experiments represent the average of 4–8 adult cardiomyocytes). *P<0.05