| Background: Treatment and preconditioning with inhibitor of Na~+/H~+ exchanger isoform-1 (NHE1) are effective methods against ischemia/reperfusion injury in mature hearts, which is unclear in immature hearts. Comparing with ischemia preconditioning (IPC), this study was undertaken to illustrate the preconditioning effects of HOE642 (cariporide), an investigational, potent, selective NHE1 inhibitor, in immature hearts of rabbits. Moreover, we investigated the myocardial protective effects of HOE642 administered at different phases of ischemia/reperfusion: pre-ischemia(p), during-ischemia(s) and at reperfusion(r) in isolated immature hearts of rabbits.Methods: After peritoneal Pentobarbital sodium anesthesia, 40 immature hearts isolated from New Zealand white rabbits (2-3 weeks old) were rappidly put on the Langendorff apparatus. Perfused with Krebs-Henseleit (KH) solution for 15min, isolated perfused heart models were established. They were randomly divided into 3 groups: the control group (IR group, n=8),IPC group (n=8), and HOE642 groups (n=24). According to the phase of HOE642 administered, HOE642 groups were randomly divided into 3 groups: HOEp group, HOEs group and HOEr group. In the IR group, hearts were subjected to 60min of normothermic global ischemia plus 60min of reperfusion (I/R) after another 15min of KH solution perfusion; In the IPC group, hearts were preconditioned by 5min of ischemia followed by lOmin of reperfusion before I/R; In the HOEp group, hearts were preconditioned by HOE642 (5Mmol/L) for 15min before I/R; In HOEs group, HOE642 (10m mol/L) was added to St.Thomas cardioplegia; In HOEr group, hearts were perfused with HOE642(5Mmol/L) during the first 15min of reperfusion. The recovery rates of heart functions(LVDP, +dp/dtmax, -dp/dtmax) and coronary artery flow (CAF) in 5min, 15min, 30min, 60min of reperfusion were monitored. At 60min of reperfusion, CK, CK-MB, LDH leakege were measured. After reperfusion, malondialdehyde (MDA), adenosine triphosphate (ATP), myocardial water content (WC) and myocardial intracellular calcium content (Ca) were measured. Myocardial and endothelial structure were observed under transmission electron microscope. Results: 1 Heart function: (1) Recovery rates of LVDP: IPC group and all HOE642 groups had higher recovery rates of LVDP than IR group at all points of reperfusion (PO.05). While, HOEr group had better recovery rates of LVDP compared to IPC at every point of reperfusion (P<0.05). Moreover, at 5min of reperfusion, recovery rates of LVDP in HOEr group is markedly lower than HOEp and HOEs group (P<0.05), though there are no significant differences at 15, 30,60min of reperfusion in each HOE642 group (P>0.05). (2) Recovery rates of +dp/dtmax: IPC group and all HOE642 groups hadhigher recovery rates of +dp/dtmax than IR group at all points of reperfusion (PO.05), except that of 5min of reperfusion in HOEr group. Recovery rates of+dp/dtmax at 5, 15min are not significant different in HOEp group and IPC group (P>0.05), but they are higher in HOEp group than IPC group at 30, 60min of reperfusion (P<0.05). Recovery rates of +dp/dtmax at 30, 60min are not markedly different in each HOE642 group (P>0.05), but they are lower in HOEr group than HOEp and HOEs group at 5,15min of reperfusion (P<0.05).(3) Recovery rates of -dp/dtmax: Recovery rates of -dp/dtmax in all groups had the same changing trend with that of+dp/dtmax, except that HOEr group had lower recovery rates of +dp/dtmax at 15min of reperfusion than HOEp and HOEs group (PO.05).2 Heart rebeating rates: Though ventricular fibrillation occured in two hearts of IR group and one heart of IPC group, there were no significant differences of heart rebeating rates between each group (P>0.05).3 Recovery rates of CAF: HOE642 groups had higher recovery rates of CAF at all points of reperfusion than control (p<0.05), while IPC only incrased it at 5, 15, 30min of reperfusion(p<0.05). There were no significant differences between IPC and HOEp group at 5, 15, 30min of reperfusion (P>0.05), but at 60min, recovery rate of CAF was higher in HOEp group (p<0.05). And recovery rate of CAF in HOEr group was lower at 5min, and was higher at 60min than that in HOEp and HOEs group(P<0.05), with no difference at 15,30min(P>0.05).4 Leakage of myocardial enzyme: Leakage of CK, CK-MB, LDH were significantly lower in HOE642 groups than the control's (p<0.05), while IPConly decreased LDH leakage (PO.05). CK, CK-MB levels were significantly lower in HOEr group than the IPC's (PO.05). There was no significant difference in each HOE642 group (p>0.05).5 Water content (WC): IPC and HOE642 treatment reduced WC compared to control (P<0.05). WC in HOEp group was much lower than IPC (P<0.05). There was no significant difference in each HOE642 group (p>0.05).6 ATP: IPC and HOE642 treatment increased ATP concent compared to control (P<0.05). ATP in HOEp group was much higher than that of IPC (PO.05). Moreover, ATP concent was higher HOEp group than the other two HOE642 groups (pO.05), with no significant difference between HOEs and HOEr (pX).O5).7 MDA: IPC and HOE642 treatment reduced MDA compared to control (PO.05), with no significant difference between each HOE642 group (p>0.05), and also no difference between HOEp group and IPC group (p>0.05).8 Intracellar calcium concent (Ca): IPC and HOE642 treatment reduced Ca compared to control (PO.05). Ca was signicantly lower in HOEp group than IPC's (PO.05), with no significant difference between each HOE642 group (pX).O5).9 Myocardial structure: Under transmission electron microscope, we observed that IPC and HOE642 treatment shew great protection to the structure of mitochondrion, sarcoplasmic reticulum, nucleoli; relieved the edema of myocyte and endothelium; increased glycogen concent compared to control. Moreover, HOEp provides a stronger protection to myocyte and endothelium injury than IPC. And HOEp and HOEs provides stronger protection tomyocyte and endothelium injury than HOEr group.Conclusion: IPC and HOE642 treatment can effectively improve the contractility and compliance of immature myocardium, relieve the edema and injury of myocytes and its organelles, ameliorate the coronary circulation, increase the ATP reserve, reduce the leakage of myocardial enzyme and the MDA concent, decrease the calcium concent of myocardium, exert great protection against ischemia/reperfusion injury in immature myocardium. Moreover, this study demonstrates that HOEp provide a stronger protection than IPC. There are similar protective effects of HOE642 administered at different phases. However, HOE642 administered before reperfusion provides a stronger protection than it does during reperfusion. The most possible mechanism of this protective effection is reducing the intraceller calcium overload. |
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