| Objective: The role of simvastatin in attenuating myocardial no-reflow (NR) is lack of enough evidence, and the potential mechanisms are need to be explored. Therefore, experimental models of myocardial NR after ischemia-reperfusion were made by left coronary artery ligation. This study sought to assess the effects of short-term use of high-dose simvastatin on myocardial NR and heart function in 28 days later. We also observed the changes of the activity of endothelial nitric oxide synthases (eNOS), inducible nitric oxide synthases (iNOS), nuclear factor (NF)-KB and myeloperoxidase (MPO), and the content of nitric oxide (NO) and malonaldehyde (MDA). Hence, the main objective of this study is to assess the effect of simvastatin on myocardial NR and explore the possible potential mechanisms. Finally, we sought to suggest the possible target for improving NR in clinical condition.Methods: 69 male wistar rats were randomized into Sham group (n=18), Control group (n=25), and Sim group (n=26). The rats in Sim group were pretreated with simvastatin 20mg/kg/day for 3 days, and the myocardial NR model was induced with 90 minutes occlusion of the left coronary artery followed by 120 minutes reperfusion in Control and Sim group, the rats in Sham group accepted thoracotomy without coronary artery ligation. After surgery, a part of the rats were assigned to acute experiment to assess the role of simvastatin in myocardial NR and the potential mechanisms, and the other part were assigned to chronic experiment to observe the heart function and left ventricular remodeling in 28 days later. The indexes in acute experiment include:①Area parameters: area at risk/area of left ventricular (RA/LVA) stands for the extent of ischemia, area of NR/area at risk (NA/RA) stands for the extent of NR and area of myocardial infarcton/area at risk (MIA/RA) stands for the extent of myocardial infarction.②Myocardium homogenate was made to determine the activity of eNOS, iNOS and MPO, and the content of NO and MDA.③Immunohistochemistry of myocardium was done to determine the activity of NF-kB p65 in cadiocytes and arteriole. The indexes in chronic experiment include:①Indexes of haemodynamics: heart rate (HR), systolic blood pressure (SBP), diastolic blood pressure (DBP), left ventricular systolic pressure (LVSP), left ventricular end-diastolic pressure (LVEDP), LV+dp/dt and LV-dp/dt.②Picrosirius red stain was done to determine left ventricular wall thickness in myocardial infarction area (MILVWT), interventricular septum thickness (IVST), collagen volume fraction (CVF) and cross sectional area (CSA).Results:1. The results of acute experiment(1) The extent of ischemia, NR and myocardial infarction between Control group and Sim group: There was no significant difference between the two groups about the extent of ischemia (44.22±7.97% vs 39.98±3.75%, P=NS); the extent of NR in Sim group was significantly smaller than Control group (34.10±7.05% vs 52.09±6.89%, P＜0.01), and the extent of myocardial infarction in Sim group was smaller than Control group (78.80±7.60 % vs 90.13±5.72%, P＜0.05).(2) The activity oftNOS, eNOS and iNOS and the content of NO in myocardium: There was no significant difference among the three groups about tNOS; the actiyity of iNOS and the content of NO in Control and Sim group were higher than Sham group, the activity of eNOS in Control group was significantly lower than Sham group, while there was no significantly difference between Sim group and Sham group about the activity of eNOS; the activity of iNOS in Sim group was significantly lower than Control group (5.02±1.64U/rag vs 9.19±2.89U/rag, P＜0.01), the activity of eNOS in Sim group was significantly higher than Control group (7.08±1.74 U/mg vs 3.72±0.98 U/mg, P＜0.07), while the content of NO in Sire group was lower than Control group (586.21±126.97nmol/g vs 744.49±137.53 nmol/g, P＜0.05).(3) The activity of MPO and the content of MDA in myocardium: The activity of MPO and the content of MDA in Control and Sim group were higher than Sham group; the activity of MPO and the content of MDA in Sire group were lower than Control group (257.72±93.43 U/g vs 384.10±40.68 U/g, 72.10±18.56nmol/mg vs111.84±38.58 nmol/mg, respectively P＜0.05).(4) The results of NF-kB p65 immunohistochemistry test: There were no significant differences among the three groups about the positive index of NF-kB p65 in cadiocytes of interventricular septum; the positive index of NF-kB p65 in cadiocytes and arteriole at left ventricular wall round the area of myocardial infarction in Control and Sire group were significantly higher than Sham group; while the positive index of NF-kB p65 in cadiocytes and arteriole at left ventricular wall round the area of myocardial infarction in Sire group was lower than Control group (21.59±10.51% vs 34.32±9.55%, 27.27±13.19% vs 44.91±15.06%, respectively P＜0.05).2. The results of chronic experiment(1) Hemodynamic measurements in 28 days later: There was no significant differences among the three groups about HR; SBP,DBP,LVSP,LV+dp/dt and LV-dp/dt in Control and Sim group were lower than Sham group, while LVEDP in Control and Sim group was higher than Sham group; LV+dp/dt and LV-dp/dt in Sim group were higher than Control group (7871.13±851.53 mmHg/s vs 6555.34±859.63 mmHg/s, 6352.87±743.19 mmHg/s vs 5253.87±638.21 mmHg/s, respectively P＜0.05), while LVEDP in Sim group was lower than Control group (7.06±2.48 mmHg vs 9.65±2.17 mmHg, P＜0.05), and there were no significant differences between Sim and Control group about SBP, DBP and LVSR(2) The results of picrosirius red stain: There were no significant differences among the three groups about IVST and CVF in non-myocardial infarction area; MILVWT in Control and Sim group was significantly smaller than Sham group, CVF in myocardial infarction area and CSA in Control and Sim group were larger than Sham group; MILVWT in Sim group was larger than Control group (1.79±0.32mm vs 1.36±0.28mm, P＜0.05), while MICVF and CSA in Sim group were significantly smaller than Control group (61.11±5.74% vs 80.64±9.07%, 796.48±102.70μm~2 vs 1056.734±130.54μm~2, respectively P＜0.01).Conclusion:①Areas of impaired perfusion within the previously ischemic myocardium were seen and myocardial NR was occurred after 90 minutes occlusion of the left coronary artery followed by 120 minutes reperfusion.②Simvastatin could improve myocardium perfusion after ischemia-reperfusion, and attenuate NR.③Simvastatin might attenuate myocardial NR through improving endothelium dysfunction, inhibiting inflammation response, inhibiting neutrophil activation and reducing the production of Reactive oxygen species.④Simvastatin could improve heart function and left ventricular remodeling after ischemia-reperfusion in 28 days later, that maybe the result of improving myocardium per fusion and attenuating NR. |
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