Effects And Mechanisms Of Aminoguanidine On Ischemia/Reperfusion Injury Myocardium In Rats

Myocardial ischemia/reperfusion injury is a commonly occurred perioperative cardiac problem. Several pathogenic mechanisms have been proposed to explain the myocardial damage that occurs during myocardial ischemia/reperfusion injury. In recent years, Oxidative stress has been shown to play a pivotal role in the pathogenesis of ischemia/reperfusion injury. It has been demonstarted that scavenging the free radical could protect the myocardium against ischemia reperfusion injury, ameliorate myocardial function. Apoptosis (programmed cell death) is the cellular basis in oxidative stress induced by ischemia/reperfusion injury. Mitochondria as oxidative stress sensor and death signal integrator play a key role in initiating and mediating apoptosis of cardiomyocytes suffered from ischemia/reperfusion damage.As an important chemical mediator and cellular signal molecular, nitric oxide has been attracting the attention of every researcher in medical domain. Many studies have examined the role of nitric oxide in physiological and pathophysiological process. Recently, it has been demonstarted that ischemia/ reperfusion leads to the activation of nitric oxide participate in the regulation of hemodynamic and microcirculatory changes in organic ischemia/reperfusion. Aminoguanidine, a selective inhibitor of iNOS, can significantly inhibit the production of NO. Unfortunately, it is controversial whether NO is protective or deleterious against myocardial ischemia/reperfusion injury so far. Additionally, there is a continuing debate as to the effects of inhibitors of NOS on myocardial ischemia/reperfusion injury.In the present study, we investigated the effects of aminoguanidine on ischemia/reperfusion injury myocardium and explored the possible mechanisms.PartⅠEffects of Aminoguanidine on the free radical in Ischemia/Reperfusion Injury Myocardium in RatsObjective: To investigate effects of aminoguanidine on ischemia/reperfusion (I/R) injury myocardium in rats and clarify the possible mechanism from oxidative stress pathway.Methods: Rat myocardial I/R injury was induced by occluding the left main coronary artery for 30 min and reperfusing for 2h. Aminoguanidine (100mg/kg) was administration by intraperitoneal injection at 10min before ischemia, Sham myocardial ischemia rats (sham-operated rats) were used as controls. The cardiac function indexes such as the left ventricular developed pressure (LVDP), the left ventricular end diastolic pressure (LVDEP) and±dp/dtmax were recorded at the end of reperfusion. The lactate dehydrogenase (LDH), creatine kinase (CK), inducible nitric oxide synthase (iNOS) activities and the content of nitric oxide (NO) in serum were respectively measured. The activity of superoxide dismutase (SOD) and the content of malondialdehyde (MDA) in the cardiomyocytes were respectively measured. The pathological changes of myocardium mitochondria were observed by electronmicroscope.Results:1. Compared with those of sham, the LVDP,±dp/dtmax were significantly decreased in I/R group at the end of reperfusion (P<0.05, P<0.01), the LVDEP was markedly increased (P<0.05), and the activities of LDH and CK were markedly enhanced (P<0.01). Compared with that of I/R group, the LVDP and±dp/dtmax were significantly increased, the LVDEP and activities of LDH, CK were markedly decreased in AG group (P<0.05).2. Compared with those of sham, the activity of SOD was markedly decreased and the activity of iNOS was increased in I/R group (P<0.01), and the contents of MDA, NO were increased in I/R group (P<0.01). Compared with those of I/R group, the activity of SOD was significantly enhanced and the activity of iNOS and the contents of MDA and NO were markedly decreased in AG group (P<0.05, P<0.01).3. The pathological changes of cardiomyocytes induced by I/R injury, such as mitochondrial membrane swelling, mitochondrial matrix swelling, cristae chaos, disruption, dissolution or disappearance; myofiber swelling, myofibrillar lysis, disorganized; glycogen granule reduction, were significantly alleviated by AG.Conclusion:It could be concluded that aminoguanidine has a beneficial myocardial protection against the ischemia/reperfusion injury in rats. It is considered that the protection is related to scavenging the free radical, decreasing the content of NO, diminishing oxidative stress, protecting cardiomyocytes from peroxynitrite injury.PartⅡEffects of Aminoguanidine on the function of mitochondria and apoptosis in Ischemia/ Reperfusion Injury Myocardium in RatsObjective: To investigate effects of aminoguanidine on ischemia/reperfusion (I/R) injury myocardium in rats and clarify the possible molecular mechanisms from the the function of mitochondria and apoptosis.Methods: Rat myocardium I/R injury was induced by occluding the left main coronary artery for 30 min and reperfusing for 2h. Aminoguanidine (100mg/kg) was administration by intraperitoneal injection at 10min before ischemia, Sham myocardial ischemia rats (sham-operated rats) were used as controls. The cardiac function indexes such as the left ventricular developed pressure (LVDP), the left ventricular end diastolic pressure (LVDEP) and±dp/dtmax were recorded at the end of reperfusion. The mitochondria were prepared by differential centrifugation. The activities of SOD, ATPase and the content of malondialdehyde (MDA) in myocardial mitochondria were respectively measured. The swelling and activity of mitochondria were respectively determined. The apoptotic rate of cardiomyocytes was evaluated by Flow Cytometry. The positive expressions of Bcl-2, Bax in cardiomy- ocytes were respectively detected by immunohistochemistry.Results:1. Compared with those of sham, the LVDP,±dp/dtmax were significantly decreased in I/R group at the end of reperfusion (P<0.01); the LVDEP was markedly increased (P<0.05). Compared with those of I/R group, the LVDP and±dp/dtmax were significantly increased, the LVDEP was markedly decreased in AG group (P<0.05, P<0.01).2. In the I/R group, the activities of SOD, ATPase were decreased and the content of MDA was increased in myocardial mitochondria (P<0.01, P<0.05); the swelling of mitochondria was markedly increased and the activity of mitochondria was decreased (P<0.01). Compared with those of I/R group, the activities of SOD, ATPase and myocardium mitochondria were significantly increased, the mitochondria swelling and MDA content were markedly decreased in AG group (P<0.05,P<0.01).3. The apoptotic rate of cardiomyocytes in I/R group (15.38±0.48) was significantly higher than that of sham group (5.18±0.33)(P<0.01). The apoptotic rate of cardiomyocytes in AG group (13.55±0.93) was markedly lower than that of I/R group (P<0.01). The positive expression of Bcl-2 and Bax protein were markedly increased in I/R group, respectively (P<0.01), Compared with those of I/R group, the expression of Bax was significantly decreased and the expression of Bcl-2 was markedly increased in AG group (P<0.01).Conclusion:It could be concluded that aminoguanidine has a beneficial myocardial protection against the ischemia/reperfusion injury in rats. It is considered that the protection is related to diminishing oxidative stress, ameliorating the function of myocardium mitochondria, upregulating the expression of anti-apoptotic protein Bcl-2, downregulating the expression of pro-apoptotic protein Bax.