Objective Ischemic preconditioning (IPC) is an important endogenous protective mechanism for ischaemia-reperfusion (I/R) injury. Currently, it is believed that opening of the mitochondrial ATP-sensitive potassium channel (mitoKATP channel) plays an important role in IPC. Diazoxide is considered as a mitoKATP agonist that is capable of activating endogenous protective mechanism of the myocardium to antagonise myocardial ischaemic and hypoxic injury. In this study, we applied a rat isolated perfused heart model to observe the expression characteristics of peroxisome proliferator-activated receptor γ coactivator-1 -1α (PGC-1α) and ultrastructural changes of mitochondria after IPC and diazoxide preconditioning (DPC), in order to explore the protective role and mechanism of IPC and DPC during IR injury of the heart.
Methods 30 Wistar rats were used to establish Langendorff isolated perfused heart model. Rats were randomly divided into five groups, six in each group: (1) I/R group: after 30 min of equilibration perfusion, heart was subjected to 30 min of ischaemia and 1 h of reperfusion. (2) IPC group: after 10 min of equilibration perfusion, heart was subjected to two times of 5 min ischaemia and 5 min reperfusion, followed by a 30 min of ischaemia and 1 h of reperfusion. (3) DPC group: after 10 min of equilibration perfusion, heart was given two times of K-H perfusion solution containing diazoxide (100 μmol/l) for 5 min then non-diazoxide K-H perfusion solution for 5 min, followed by 30 min of ischaemia and 1 h of reperfusion. (4) Blank control group: equal amount of saline was used instead of diazoxide. Perfusion procedure was the same as the DPC group. (5) Dimethyl sulfoxide (DMSO) group: DMSO was applied instead of diazoxide, and perfusion procedure was the same as the DPC group. Cardiac apex muscle was cut for frozen section. Immunohistochemistry staining of PGC-1α was performed and average absorbance was calculated. Electron microscope was used for Flameng scoring of the myocardial mitochondria.
Results The average absorbance values of PGC-1α were respectively: I/R group (3.88±1.72), IPC group (10.94±5.23), DPC group (8.40±3.64), blank control group (3.55±1.56) and DMSO group (4.16±0.52). The expression of PGC-1α was significantly increased in IPC and DPC groups and the differences were statistically significant compared to the I/R, blank control and DMSO groups, that is, p<0.01 for IPC group and p<0.05 for DPC group. However, there was no significant difference between the IPC and DPC groups (p>0.05). Flameng score: IPC group (0.44±0.13), DPC group (0.47±0.10), I/R group (1.78±0.14), blank control group (1.70±0.03) and DMSO group (1.68±0.06). Flameng score of the IPC and DPC groups had statistically significant difference as compared to the I/R group, blank control group and DMSO group (p<0.01), but no significant difference was detected between the IPC and DPC groups (p>0.05).
Conclusion IPC and DPC have a protective effect on myocardial mitochondria, and their mechanism in action may be related to activation and over-expression of PGC-1α.