Figure 4. Nobiletin is cardioprotective via PER2 and reverses midazolam induced ROS production.

Mice were treated i.p. with vehicle (solutol in 0.9% NaCl [ratio 1:100]), nobiletin in solutol:0.9%NaCl (ratio 1:100; 1mg/kg), midazolam (200mg/kg) or midazolam + nobiletin 2 hours prior to myocardial ischemia. Myocardial ischemia consisted of 60 min of ischemia followed by 120 minutes of reperfusion. For ROS (H₂O₂) measurements reperfusion consisted of 15 minutes after 60 min of ischemia. Infarct sizes were measured by double staining with Evan’s blue and triphenyl-tetrazolium chloride. Infarct sizes are expressed as the percent of the area at risk (AAR) that underwent infarction. Serum troponin I concentrations were measured by enzyme-linked immunosorbent assay (ELISA). (A) ROS production determined by H₂O₂ measurement in the area at risk of wildtype mice. (B) Infarct sizes as the percent of AAR and serum troponin I levels in wildtype mice treated with vehicle or nobiletin (1mg/kg) 2 hours prior to myocardial ischemia. (C) Infarct sizes as the percent of AAR and serum troponin I levels in Per2^−/− mice treated with vehicle or nobiletin (1mg/kg) 2 hours prior to myocardial ischemia. (D) Representative infarct staining from wildtype and Per2^−/− mice; (n=7–8; mean±SD; p<0.05; NOB=nobiletin).