| Background For present, coronary artery heart disease(CAD) especial acute myocardial infarction(AMI) has become main cause of mortality in the world. For patients with AMI, it is important to reopen the obstructed coronary arteries timely and effectively to save ischemic myocardium and to reduce the infarcted myocardial size. Thrombolytic therapy and percutaneous coronary intervention(PCI) are two common useful methods. However, reperfusion itself can cause myocardial cell damage, which is called ischemia reperfusion injury. Unfortunately, this injury is still unclear and lack of effective clinical therapeutic methods.Adrenergic nerve system is divided into sympathetic nervous system and parasympathetic nervous system. Under normal circumstances, these two systems coordinate with each other. When coronary artery is occluded, the body is in a state of stress and circulating catecholamine hormones including norepinephrine and epinephrine are increased by exciting sympathetic nervous system. The catecholamine are deleterious to heart by promoting cell necrosis and apoptosis. And this toxicity will be enhanced in the state of myocardial reperfusion. Beta receptor blockers can effectively interdict the activation of sympathetic nervous system, resist the myocardial toxicity of catecholamine and improve the ventricular remodeling. The numbers of beta receptors on myocardium increase after ischemia/reperfusion processing especially beta receptor one. By treatment of bisoprolol, a highly selective beta blocker, it can be found that myocardial protective adipocyte factor, adiponectin, is increased in rats’serum and myocardium. Thus, it may suggest that bisoprolol may have myocardial protective.Phosphatidylinositol-3 kinase/serine-threonine kinase/glycogen synthase kinase 3b(PI3K/AKT/GS3K-b) is an important signaling pathway which plays a key role in the apoptosis, survival and proliferation. Activated PI3K can activate Akt, which will further activate GS3K-b, endothelial nitric oxide synthase and heat shock protein to generate myocardial protective effect. Drug preconditioning treatment can activate cardiac intracellular PI3K/AKT/GS3K-b pathway, which may play an important role in myocardial protection. Objective To investigate the protective effect of bisoprolol, a highly selective beta blocker, preconditioning on myocardial ischemia reperfusion injury in rats by the medium of PI3K/AKT/GS3K-b.Methods1.Experiment one:Thirty rats were randomly divided into 5 groups(n=6):sham group(S), ischemia/reperfusion group(I/R group), little dose bisoprolol group(LBiso), middle dose bisoprolol group(MBiso) and high dose bisoprolol group(HBiso). Evans blue and TTC staining were used to evaluate the extent of myocardial infarction which could suggest the protective effect of bisoprolol on myocardium. Optimal dose of bisoprolol was selected based on extent of myocardial infarction. The numbers of incidence, cardioversion and death caused by ischemia-reperfusion associated malignant arrhythmia were compared in each group.2.Experiment two:Sixty rats were randomly divided into 4 groups (n=15):sham group(S), ischemia reperfusion group(I/R), bisoprolol group(Biso) and bisoprolol+LY294002 group(Biso+LY). LY294002 is an specific inhibitor to PI3K. Enzyme-linked immuno sorbent assay method was used to measure the levels of serum cardiac damage biomarkers(cTnI and CK-MB). Evans blue and TTC staining were used to evaluate the extent of myocardial infarction. Western blot was used to evaluate the levels of protein kinase B(total-AKT), phosphorylation of AKT(p-AKT), glycogen synthase protein kinase-3b(total-GS3K-b) and phosphorylation GS3K-b(p-GS3K-b) in myocardial tissue. TdT-mediated dUTP nick end labeling was used to determinate the rates of apoptosis. These methods were used to certify the role of PI3K/AKT/GS3K-b pathway in the protective effect of bisoprolol. The numbers of incidence, cardioversion and death caused by ischemia-reperfusion associated malignant arrhythmia were compared in each groups in experience two and should be also compared in total S group, total I/R group and total Biso group(Biso group, LBiso group, MBiso group and HBiso group).3.The actual finishing experiment of rat numbers in each groups should be counted and analyzed to ensure no significant difference under each groups.Results1.Nighty rats were enrolled in our study(30 in experiment one and 60 in experiment two). The number of death rat were nine. The actual finishing experiment of rat numbers in S group, I/R group, LBiso group, MBiso group and HBiso group in experiment one were 6,4,5,6, and 6, respectively. The actual finishing experiment of rat numbers in S group, I/R group, Biso group, and Biso+LY group in experiment two were 15,11,15, and 13, respectively. No significant difference could be found in each groups (P>0.05)2.Experiment one:Least area of infarction was shown in the MBiso group (IA/AAR=34.6±6.9%) compared with other dose bisoprolol groups(LBiso group: IA/AAR=40.9±6.6% and HBiso group:IA/AAR=45.4±5.0%)(p<0.05). Heart rates were significantly decreased by using bisoprolol (LBiso:386.5±47.4bpm, MBiso: 364.2±72.4bpm and HBiso 349.6 ±66.4bpm) (p<0.05), while no significant difference were found in different dose of bisoprolol groups(p>0.05). In the aspects of MA, unless rats in LBiso group showed similar results in incidence (times),cardioversion(times) and death(number) with I/R group(2,1,1,respectively vs. 3,1,2,respectively), rats in MBiso group(0,0, 0respectively) and HBiso group (0,0, O,respectively) showed better results than I/R group.3.Experiment two:In the areas of infarction(Biso group:IA/AAR=39.6±5.9%, Biso+Ly group:IA/AAR=46.3±7.4% and I/R group IA/AAR=51.1±5.1%), rates of myocardial apoptosis(Biso group:Al=17.7±3.2%, Biso+LY group:A1=24.3±2.7% and I/R group:Al=28.9+2.1%) and levels of cardiac injury biomarkers(Biso group: cTnI=235.8±41.3pg/ml and CK-MB=18.3±3.7ng/ml, Biso+LY group:cTnI=303.1 ±44.2pg/ml and CK-MB=21.2±7.3ng/ml and I/R group:cTnI=431.6±53.7pg/ml and CK-MB=25.9±7.3ng/ml), Biso group and Biso+LY group showed better results than I/R group(p<0.05). In Biso group the activities of AKT and GS3K-b were higher than other groups(Biso group:p-AKT/AKT=0.85±0.11, p-GSK3-b/GSK3-b=0.91± 0.21 and I/R group:p-AKT/AKT=0.55±0.07, p-GSK3-b/GSK3-b=0.57+0.07) (p<0.05). The PI3K inhibitor LY294002 could significantly decrease the activity of AKT and GS3K-b (Biso+LY group:p-AKT/AKT=0.27±0.08, p-GSK3-b/GSK3-b =0.31±0.05) (p<0.05). In the aspects of MA, rats in Biso group showed better results in incidence (times) and death(number) with I/R group(3,0, respectively vs.8,4, respectively). The electrical protection of bisoprolol could partly be weaken by the treatment of LY294002.ConclusionLHigh dose(100mg/kg/day), middle dose(10mg/kg/day) and little dose(lmg/kg/day) of bisoprolol have protective effects on myocardium in SD rats, meanwhile middle dose showed optimal curative effects(p<0.05).2.Middle dose of bisoprolol treatment can alleviate the cardiac injury biomarkers (cTn1 and CK-MB) and decrease the rate of myocardial apoptosis(p<0.05).3.Bisoprolol preconditioning may have myocardial electrical protection by reduction of incidence, increase of cardioversion and decrease of death cause by ischemia-reperfusion associated malignant arrhythmia in rats.4.The enhancement in the activity of PI3K/AKT/GS3K-b pathway may partly participate in the myocardial protective effects of bisoprolol. |
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