Effects of heptanol preconditioning on structure,function and Cx43 content of mitochondria in rabbit model of myocardial ischemia/reperfusion injury

AIM: To investigate the effects of heptanol preconditioning on the changes of structure, function and connexin 43 (Cx43) content in mitochondria in a rabbit model of myocardial isehemia-reperfusion (IR) injury. METHODS: In anesthetized open-chest rabbits, the left anterior descending artery (LAD) was occluded for 30 min and reperfused for 4 h. Sixty-four rabbits were randomly divided into 4 groups (n=16 in each group): sham operation group (sham group), ischemia-reperfusion group (IR group), ischemic preconditioning group (IP group) and heptanol preconditioning group (HT group). All rabbits in the 4 groups were killed 4 h after reperfusion. Myocardial infarct size was determined at the end of the experiment. Mitochondria was isolated by centrifugations. The ultrastructural changes of the mitochondria were observed under electronic microscope. The mitochondrial membrane potential, Ca2+ concentration, MDA content and SOD activity of myocardial mitochondria were also examined. The content of mitochondria Cx43 was detected by Western blotting. RESULTS: Compared to IR group, the myocardial infarct size was significantly reduced in IP (18.97%±2.80%) and HT (19.97%±3.80%) groups, the damage of mitoehondrial ultrastructure was milder (P<0.05), mitochondrial membrane potential was significantly higher and Ca2+ concentration was much lower (P<0.01) in IP group and HT group. No significant difference of MDA content and SOD activity in myocardial mitochondria between IR group and HT group was found. However, MDA content were much lower and SOD activity was significantly higher in IP group as compared to IR group (P<0.01). Compared to sham group, the mitochondria Cx43 expression was distinctly decreased compared to IR group (P<0.05) and no significant difference was found between IP group and HT group (P>0.05). CONCLUSION: Heptanol preconditioning protects myocardium from ischemia-reperfusion injury. The mechanism may be related to increasing in mitochondrial membrane potential, alleviating Ca2+ overload in myocardial mitochondria and attenuating the decrease in mitochondria Cx43 expression induced by isehemia-reperfusion.     

Role of apelin in rat myocardial ischemic preconditioning and ischemia/ reperfusion injury

AIM: To study the alteration and role of apelin in myocardial ischemic preconditioning and ischemia/reperfusion injury of rats.METHODS: Forty-five SD rats were randomly divided into three groups: ischemia/reperfusion group (IR),ischemia pre-conditioning group (IP) and sham operation group.ECG was continuously used to evaluate the score of arrhythmias.The protein levels of apelin-36 in myocardium and plasma were detected by radioimmunoassay.The expression of apelin was observed by immunohistochemistry.RESULTS: (1) The scores of arrhythmias in IP group (2.1±0.5) was only 58.3% of IR group (3.6±0.8) ( P<0.01).(2) The apelin-36 protein level of plasma and myocardium in IR group were respectively lower by 36.1% and 45.6% than those in SH group (P<0.01),and those in IP group were lower by 23.8% and 24.7% than those in SH group (P<0.01),but higher than those in IR group (18.9% and 38.5%,respectively,P<0.05).(3) The staining absorbance of apelin in IR,SH and IP group was (7.87±2.41),(22.53±2.54) and (14.23±2.15),respectively.There were significant differences between IR and SH (P<0.01) and between IP group and SH group (P<0.05).(4) The scores of arrhythmias in IP and IR were negatively correlated with the protein level of apelin-36 in myocardium (r= 0.847,P <0.01).CONCLUSION: The down-regulation of apelin-36 in the plasma and myocardium of rats indicates that apelin has an important role in myocardial ischemic preconditioning and ischemia/reperfusion injury.     

Effects of ischemic preconditioning on oxidative stress and mitochondrial function in young and old rat myocardium with ischemia/reperfusion

AIM: To study the protective effect of ischemia preconditioning (IPC) on ischemia/reperfusion (IR)-damaged myocardium in young and old rats. METHODS: Male Wistar rats aged at 3 months (young) and 20 months (old) were used to establish myocardial IPC model and IR model with the method of Langendorff heart perfusion. The rats were divided into young ischemia/reperfusion (YIR) group, young ischemic preconditioning (YPC) group, old ischemia/reperfusion (OIR) group and old ischemic preconditioning (OPC) group. Transmission electron microscopy was used to observe the ultrastructural changes of myocardial tissue and myocardial mitochondria. The myocardial infarction area was determined by TTC staining. The lactate dehydrogenase (LDH) content in coronary effluent fluid and the levels of superoxide dismutase (SOD) and malondialdehyde (MDA) in myocardial tissues were detected by the method of colorimetry. The levels of nitrated and carbonylated proteins in myocardial tissue were measured by ELISA. The myocardial cell apoptosis was analyzed by TUNEL assay. The mitochondrial respiratory function and mitochondrial permeability transition pore opening induced by calcium load were evaluated by oxygen electrode method. RESULTS: Compared with YIR group, the myocardial infarction area in YPC group was obviously smaller, SOD activity in myocardial tissues increased, LDH activity in coronary effluent fluid and the content of MDA decreased, and the levels of nitrated and carbonylated proteins in the cardiac tissues reduced. In YPC group, the mitochondrial membrane structure appeared intact, cristae of the mitochondria showed close arrangement, and the matrix was compressed under the electron microscope. Myocardial mitochondrial respiratory control rate, state Ⅲ oxygen consumption and the P/O ratio in YIR group all significantly increased, proton leak decreased, mitochondrial swelling induced by calcium distinctly reduced, and myocardial apoptosis rate declined. No significant difference of the above indexes between OIR group and OPC group was observed. Compared with YPC group, myocardial ultrastructural damage increased clearly, cardiac oxidative stress increased, mitochondrial respiratory function declined, and cell apoptosis and necrosis increased in OPC group. CONCLUSION: Ischemic preconditioning has protective effect against myocardial IR injury in young rat hearts, while old rat hearts were less sensitive to ischemic preconditioning, leading to bluntness of cardioprotection with IPC in aging hearts. This may be related to mitochondrial injury and severe cellular apoptosis caused by increase of cardiac oxidative stress levels in the aging ischemic preconditioning heart.     

Role of nitric oxide in ischemia/reperfusion injury and ischemic preconditioning

AIM: To clarify the role of nitric oxide(NO) in ischemic preconditioning(IP) and its effects on apoptosis. METHODS: Seventy-two male Wistar rats were divided into the following six groups:ischemia/reperfusion (IR) group,IP group,IR+L-arg group,IP+L-arg group,IR+L-NAME group and IP+L-NAME group,The following changes were measured:cardiac hemodynamic parameters,infarct size,PMNs counting myocardial MPO activity and TUNEL staining.RESULTS: ①L-arg significantly attenuated ischemia/reperfusion-induced heart injury,reduced PMNs infiltration and cardiomyocyte apoptosis.②L-NAME also significantly reduced infarct size,PMNs infiltration and cardiomyocyte apoptosis compared with IR group,however,L-NAME aggravated ischemia/reperfusions-induced cardiac functional injury.③L-arg or L-NAME did not significantly alter the protective effect of ischemic preconditioning. CONCLUSION: Increased production of endogenous NO before prolonged ischemic period can protect hearts and inhibit apoptosis.L-NAME can inhibit iNOS activity and ONOO^- production in reperfusion period to protect heart.     

Effects of non-wounded ischemic preconditioning on ischemia-reperfusion injury in isolated rat hearts

AIM:To observe the protective effect of non-wounded ischemic preconditioning on ischemic/reperfusion injury in isolated rat hearts. METHODS: 25 male SD rats, weighting (250±30) g, were randomly divided into three groups: control group (C,n=8), anoxia/reoxygenation group (A,n=8) and non-wounded legs ischemic preconditioning group (N-WIP,n=9).Hearts were isolated from rats and perfused on a Langendorff apparatus with a normal Krebs-Henseleit buffer (saturation 95% O₂+5% CO₂) at a constant pressure (8.33 kPa) and temperature (37 ℃) in C group; Following 15 min equilibration, hearts were subjected to 15 min of global ischemia and 15 min reperfusion (37℃) in A group; Rats were subjected to non-wounded leg repeated-brief ischemic preconditioning, and then treated in procedure similar to A group in N-WIP group.The activities of superoxide dismutase (SOD) and Ca²⁺-Mg²⁺-ATPase, malondialdehyde (MDA) content of efflux from coronary vessel and myocardium, myocardium monophasic action potential and contractile force were measured before ischemia, 15 minutes after ischemia and 5, 15 minutes after reperfusion. RESULTS:Compared with A group, non-wounded legs ischemic preconditioning reduced the incidence of reperfusion arrhythmias (P＜0.05), decreased the content of MDA of myocardium (P＜0.01), enhanced the activities of SOD (P＜0.01) and stabilized myocardial membranous potential,the activity of Ca²⁺-Mg²⁺-ATPase and contractile function. CONCLUSION:These results indicate that non-wounded leg ischemic preconditioning has a protective effect on ischemia-reperfusion injury in isolated rat hearts. The mechanism may be related to the strength of antioxidation, the stability of Ca²⁺-Mg²⁺-ATPase activity and membranous structure in myocardium.     

Effects of norepinephrine preconditioning and ischemic preconditioning on apoptosis and Bcl-2, Bax expression in rat myocardial cells during myocardial ischemic reperfusion

AIM: To study the effects of norepinephrine preconditioning(NE-P) and ischemic preconditioning (IP)on apoptosis and Bcl-2, Bax expression in rat myocardial cells in myocardial ischemic reperfusion (I/R). METHODS: The model of rat ischemic-reperfusion was used to conduct NE-preconditioning. Apoptotic myocytes were detected with TUNEL. Bcl-2, Bax expression were detected with immunohistochemistry. RESULTS: The rate of apoptosis cells in I/R group was higher, the rate of apoptosis cells in NE-P group and IP was lower significantly than that in I/R group(P＜0.01). The expression of Bcl-2 in I/R group was lower, but the expression of Bax was higher, the expression of Bcl-2 in NE-P group was higher significantly than that in I/R group(P＜0.01), the expression of Bax in NE-P group was lower than that in I/R group(P＜0.01). There was no significantly difference between NE-P and IP group in the above parameters (P＞0.05). CONCLUSION: NE-P reduced myocyte apoptosis by I/R in rats; The expression of Bcl-2 ,Bax genes played an important role in myocardial apoptosis.     

Effect of Fas/FasL on late reperfusion of acute myocardial infarction and the potential oxidative stress mechanism in canine

AIM: To discuss the effect of Fas/FasL on the late reperfusion of acute myocardial infarction (AMI) and the potential oxidative stress mechanism. METHODS:Eighteen anesthetized dogs were randomly divided into three groups: late reperfusion group (n=6): ligated the coronary for 6 h, followed by reperfusion for 6 h; permanent ischemia group (n=6): after pericardium were opened for 6 h, ligated the coronary for 6 h, and did not reperfuse; control group (n=6): did not ligate the coronary but operation last for 12 h. Infarction brim myocardial Fas/FasL was detected by immunohistochemistry. Apoptosis index (AI) was detected by TUNEL. SOD and GR activity and MDA content were detected by colorimetry. RESULTS:The expression of Fas/FasL and apoptosis index were significantly higher in permanent ischemia group and late reperfusion group than those in control group (P<0.05, P<0.01), and the difference between them was also significant (P<0.05). SOD and GR activities were lower in permanent ischemia group and late reperfusion group than those in control group (P<0.05, P<0.01). The MDA contents in permanent ischemia group and late reperfusion group were higher than that in control group (P<0.05). CONCLUSION:The late reperfusion of AMI promotes the expression of Fas/FasL and myocardial apoptosis, and it may be due to oxidative stress mechanism.     

Dexamethasone pretreatment attenuates reperfusion arrhythmia in rats

AIM: To investigate the effect of dexamethasone (DEX) preconditioning on reperfusion arrhythmia. METHODS: 46 Sprague-Dawley rats were divided randomly into DEX and control (CON) group, the rats were pretreated with DEX or sodium chloride before their hearts were separated for Langendorff perfusion and for ischemia/reperfusion. The reperfusion arrhythmias were observed dynamically after 60 min reperfusion following 30 min ischemia. The expression of HSP72 in myocardium was examined by Western blotting and immunohistochemistry at reperfusion 60 min. The levels of malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px) and the activities of Na+-K+-ATP ase, Ca2+-Mg2+-ATPase on myocardial plasma membrane were detected. RESULTS: Compared with control group, the accumulated points and persistence time of ventricular arrhythmia were reduced significantly in DEX group (P<0.05), the expression of HSP72 was significant upregulated (P<0.05), the level of MDA was reduced significantly, the activities of SOD, CAT, GSH-Px and Na+-K+-ATPase were significantly higher (P<0.05). CONCLUSION: Dexamethasone pretreatment markedly reduces the reperfusion ventricular arrhythmias in rats, which may be attributed to upregulation of HSP72, SOD, CAT, GSH-Px , Na+-K+-ATPase and inhibition of lipid peroxidation.     

Ischemic postconditioning reduces myocardial damage in rats suffering from limb ischemia-reperfusion

AIM: To investigate the protective effect of limb ischemic postconditioning on the myocardial damage in the rats suffering from limb ischemia-reperfusion (LIR). METHODS: Wistar rats were randomly divided into control group (C group), ischemia-reperfusion group (IR group) and ischemic post-conditioning group (IR+IPostC group). For conducting ischemic postconditioning, the rats in IR+IPostC group underwent 5 min of ischemia and 5 min of reperfusion on their hind limbs repeatedly after 4 h of ischemia, and then, 4 h of reperfusion was applied. The activity of superoxide dismutase (SOD), xanthine oxidase (XOD) and myeloperoxidase (MPO) was measured. The levels of malonaldehyde (MDA) in plasma and myocardial tissues, the levels of creatine kinase (CK), creatine kinase MB (CK-MB), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), α-hydroxybutyrate dehydrogenase (α-HBDH) and myocardial troponin I (cTnI) were also detected. The changes of ultrastructure in the myocardium were observed under electron microscope. RESULTS: Compared with C group,the levels of CK-MB, AST, LDH,α-HBDH and cTnI were all increased in IR and IR+IPostC groups. The levels of MDA and XOD also increased (P<0.05), but the activity of SOD decreased (P<0.05). However, compared with group IR, the levels of CK-MB, AST, LDH, α-HBDH and cTnI decreased (P<0.05) in IR+IPostC group.The levels of MDA and XOD also decreased (P<0.05), but the activity of SOD increased (P<0.05). Under electron microscope, the cardiac myofibrils arranged neatly, light and dark bands were clear, the mitochondrial cristae arranged closely and neatly, and the mitochondrial matrix densification was observed in C group. However, the cardiac fiber arrangement was disordered or disappeared, stromal edema was obvious, most or all mitochondrial cristae and membrane became fusion or disappeared, mitochondrial vacuolization and decrease in glycogen were obvious in IR group. In IR+IPostC group, the pathological changes mentioned above were attenuated somewhat than those in IR group. CONCLUSION: Ischemic postconditioning protects rat myocardium under limb ischemia-reperfusion.     

Effects of glucagon-like peptide-1 on myocardial ischemia-reperfusion/hypoxia-reoxygenation injury in rats

AIM: To investigate the effects of glucagon-like peptide-1 (GLP-1) on myocardial ischemia-reperfusion (IR)/hypoxia-reoxygenation (HR) injury in rats. METHODS: Sprague-Dawley rats were randomly divided into 5 groups: sham group, IR group and IR+GLP-1 (0.03 nmol/L, 0.16 nmol/L and 0.30 nmol/L) groups. IR group and IR+GLP-1 group were subject to 30 min of ischemia and 3 h of reperfusion. The myocardial infarct size, the ultrastructural changes of the myocardial tissues, the apoptosis of the cardiomyocytes, the activity of superoxide dismutase (SOD) and the concentration of malondialdehyde (MDA) were detected. Primarily cultured cardiomyocytes were divided into 5 groups at random: control group, HR group and HR+GLP-1 (1 μmol/L, 5 μmol/L and 10 μmol/L) groups. The morphology and apoptosis of the cardiomyocytes were observed. The levels of lactate dehydrogenase (LDH),MDA,SOD,reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) in different groups were detected. RESULTS: Compared with IR group, the myocardial infarct size and cardiomyocyte apoptosis were remarkably reduced, mitochondrial ultrastructures were improved, the activity of SOD was increased and the concentration of MDA was decreased in IR+GLP-1 (0.03 nmol/L, 0.16 nmol/L and 0.30 nmol/L) groups. Compared with HR group, GLP-1 (1 μmol/L, 5 μmol/L and 10 μmol/L) preconditioning significantly decreased the myocardial injury, increased SOD activity, decreased MDA concentration and ROS production, and heightened MMP in a dose-dependent manner. CONCLUSION: GLP-1 protects cardiomyocytes from IR/HR injury, which may be partially due to the effects of anti-oxidative mechanism and the function of mitochondrial protection.     

Panax quinquefolium saponins protect rat myocardium from ischemia/reperfusion injury through attenuating excessive endoplasmic reticulum stress

AIM: To investigate whether excessive endoplasmic reticulum stress (ERS) is involved in the protective mechanism of Panax quinquefolium saponins (PQS) against ischemia/reperfusion (I/R) injury in rat myocardium. METHODS: The model of myocardial I/R injury in vivo was made by ligating the left anterior descending artery for 45 min followed by 24 h of reperfusion in SD rats. The hemodynamics and serum content of cardiac troponin T (cTnT) were measured. The myocardial infarct size was measured by Evans blue and 2, 3, 5-triphenyltetrazolium chloride (TTC) staining. Cardiomyocyte apoptosis was detected using in situ TDT-mediated dUTP nick end labeling (TUNEL). The protein levels of glucose-regulated protein 78 (GRP78), calreticulin (CRT), C/EBP homologous protein (CHOP), caspase-12, apoptosis-associated proteins Bax and Bcl-2 were determined by Western blotting. RESULTS: Compared with I/R group, the mean arterial pressure in PQR+IR group was decreased by 32.0%, and left ventricular±dp/dtmax was increased by 64.0% and 35.0%, respectively.The serum content of cTnT was decreased by 53.3%, the percentage of area of necrosis (AN)/area at risk (AAR) was reduced by 65.5% and the apoptosis rate was decreased by 54.9%.The myocardial pathological changes were improved. Bcl-2 protein expression was increased by 110.0% and that of Bax was decreased by 47.8%. CRT protein expression was decreased by 43.4 %, CHOP protein expression and the protein level of cleaved caspase-12 were decreased by 38.6% and 23.7% in PQS+I/R group. CONCLUSION: PQS alleviates I/R injury in myocardium by inhibition of excessive ERS.     

Significance and changes of serum interleukin-8 and 10 in human myocardium ischemic preconditioning

AIM: To explore the mechanism of myocardium protection after ischemia/reperfusion (I/R) injury by preconditioning with ischemia in human. METHODS: Thirty-six patients underwent valve replacement were divided into ischemic preconditioning group (IP group, 20 cases) and non-ischemic preconditioning group (control group, 16 cases) according to whether they were given single cycle reperfusion before cardioplegia or not. Serum levels of interleukin-8 and 10 were measured with ELISA. Expressions of myocardial Bcl-2 and caspase-3 were analyzed. RESULTS: The inflammatory factors IL-8 and IL-10 increased to the highest level in serum at 6 h after declamping and recovered to normal level on 5 d after declamping. On 6 h, 1 d and 2 d after declamping, serum level of IL-8 was significantly lower in IP group than that in control group (P<0.05), but serum level of IL-10 was higher in IP group (P<0.05). Expression of myocardial Bcl-2 and caspase-3 increased in both groups after reperfusion, and Bcl-2 was lower in the control group than that in IP group while the level of caspase-3 was higher (P<0.05). Expression of myocardial Bcl-2 had positive correlation with IL-10 and negative correlation with IL-8. CONCLUSION: Ischemic preconditioning has the effect of protection of human myocardial cells after ischemia/reperfusion injury through decreasing systemic inflammatory response following ischemia reperfusion injury.     

Effect of pretreatment with 11,12-EET on myocardial ischemia/reperfusion injury in rats

AIM: To examine the effect of pretreatment with low-concentration of 11, 12-epoxyeicosatrienoic acid(EET) on myocardial ischemia/reperfusion injury in rats. METHODS: After tracheotomy, myocardial ischemia/reperfusion was produced by occlusion and release of the left anterior descending artery(LAD) of the rats. Ischemic preconditioning(IP) was made by two times of ischemia(5 min)/reperfusion(5 min). The experiment was conducted in three groups: control,IP and pretreatment with 11,12-EET(6.24×10^-8 mol/L), and each group was subdivided into two subgroups:A,the rats were subjected to ischemia(10 min)/reperfusion(10 min) and arrhythmias during the whole periods were monitored; The rats in B were subjected to ischemia(60 min)/reperfusion(30 min) and arrhythmias, cardiac funtion and myocardial infarction size were documented. RESULTS: Both IP and pretreatment with 11,12-EET could protect the heart against arrhythmias, cardiac disfunction and myocardial infarction. CONCLUSION: Pretreatment with 11,12-EET had protective effect on myocardium in case of ischemia/reperfusion, which was similar to ischemic preconditioning.     

Protective effect of delayed ischemic preconditioning on renal ischemia/reperfusion injury via induction of hypoxia inducible factor

AIM: To investigate the effects of delayed ischemic preconditioning on renal ischemia-reperfusion injury in mice and to study the role of hypoxia inducible factor 1α(HIF-1α). METHODS: Male C57/BL6N mice were randomly divided into 3 groups: sham operation group(sham), ischemia/reperfusion group(IR) and ischemic preconditioning group(IPC). Thirty-minute ischemia was induced by clamping renal bilateral pedicles followed by reperfusion in IR group. Fifteen-minute pre-ischemia was performed 4 days before IR in IPC group. Serum creatinine(Scr), blood urea nitrogen(BUN), kidney morphology and apoptosis were observed at different time points following reperfusion. The expression of HIF-1α in the renal tissues was evaluated by the methods of immunohistochemistry and Western blotting analysis. The mRNA expression of vascular endothelial growth factor(VEGF) and glucose transporter-1(Glut-1) was detected by real-time quantitative RT-PCR.RESULTS: Compared with IR group at 24 h following reperfusion, acute tubulointerstitial injury was significantly relieved in IPC group. The levels of Scr and BUN, and apoptosis of tubular epithelial cells were also decreased in IPC group. Nuclear expression of HIF-1α was higher in IPC group than that in IR group. The mRNA expression of VEGF and Glut-1, the target genes of HIF-1, was also increased significantly in IPC group. CONCLUSION: Delayed ischemic preconditioning attenuates both morphologic and functional injuries induced by renal ischemia/reperfusion. This protective effect may be related to the increased expression of hypoxia inducible factor.     

Effects of urocortin-I preconditioning on myocardial mitochondrial respiratory function and enzyme activity in rats

ATM: To investigate the influence of urocortin-I (Ucn I) preconditioning on the myocardial mitochondrial respiratory function and enzyme activity in the rats with ischemia reperfusion, and to observe the changes of ATP content in the myocardial cells. METHODS: (1) The healthy male Sprague-Dawley rats were randomly divided into 4 groups:normal group (Nor group), ischemia reperfusion group (IR group), Ucn I preconditioning group (Ucn I group), 5-hydroxy acid (5-HD)+Ucn I group. Langendorff perfusion was used to establish the in vitro model of cardiac ischemia reperfusion. At the end of the balance (T₁), before ischemia (T₂) and at the end of the reperfusion (T₃) respectively, the myocardial mitochondria was extracted, the mitochondrial respiratory function and respiratory enzyme activity in each group were determined. (2) The method of MPA isolated heart perfusion was used to isolate myocardial cells of the adult rats. After cultured for 24 h, myocardial cells were divided into 4 groups:Nor group, hypoxia/reoxygenation group (I/R group), Ucn I group, 5-HD+Ucn I group. Hypoxia/reoxygenation model of myocardial cells was established. At the end of reoxygenation, the changes of myocardial ATP content were measured by high performance liquid chromatography.RESULTS: (1) Compared with T₁, T₂ time points, the respiratory function (state 3 respiratory rate, respiratory control rate) and NADH oxidase, succinate oxidase and cytochrome C oxidase activities at T₃ time point were significantly decreased (P<0.05) in all groups except Nor group. At T₃ time point, the myocardial mitochondrial respiratory function and respiratory enzyme activity in Ucn I group were superior to 5-HD+Ucn I group and IR group (P<0.05), but was inferior to Nor group (P<0.05). At T₃ time point, the respiratory function of myocardial mitochondria and respiratory enzyme activities (NADH oxidase, succinate oxidase) in 5-HD+Ucn I group were better than those in IR group (P<0.05), but no statistical difference of the cytochrome C oxidase activity between the 2 groups was observed. The respiratory function and 3 kinds of respiratory enzyme activities at T₁, T₂ time points had no statistical change. (2) At the end of the reoxygenation, the myocardial ATP content in Nor group was higher than that in other groups (P<0.01). The myocardial ATP contents in I/R group and 5-HD+Ucn I group were lower than that in Ucn I group (P<0.05). In additon, 5-HD+Ucn I group was higher ATP content compared with I/R group (P<0.05). CONCLUSION: Ucn I preconditioning attenuates the ischemia/reperfusion induced damages of myocardial mitochondrial respiratory function and respiratory enzyme activity, thus ensuring the myocardial ATP contents under the condition of hypoxia/reoxygenation.     

Role of heme oxygenase-1 in the ischemic preconditioning of isolated rat heart

AIM: To investigate the influence of ischemic preconditioning on heart function, the activities of lactate dehydrogenase (LDH), malondialdehyde (MDA), and heme oxygenase-1 (HO-1) after ischemia/reperfusion in isolated rat heart. METHODS: The model of Langendorff was used in isolated rat heart perfusion. Ischemic preconditioning protocol: stopping perfusion for 5 minutes and reperfusion for 5 minutes, repeating three times. Ischemia protocol: stopping perfusion for 40 minutes and reperfusion for 20 minutes. Indexes of heart function were recorded in control M8, ischemia and reperfusion group (IR), and ischemic preconditioning group (IPC). The content of LDH of coronary effluent was measured. Moreover, the content of MDA and activity of HO-1 in myocardium were also measured. RESULTS: The recovery percentage of heart function in IPC group was significantly higher than that in IR group (P<0.01) and the activity of heme oxygenase-1 also increased significantly (P<0.05). CONCLUSION: The contents of LDH and MDA significantly decreased in IPC group compared with IR group. The increase in heme oxygenase-1 activity might be involved in the protective effect of ischemic preconditioning on ischemic/reperfused rat heart.     

Effect of ischemic preconditioning on myocardial Bcl-2 expression and mitochondrial structure during heart valve replacement surgery

AIM: To investigate the effect of ischemic preconditioning (IP) on myocardial Bcl-2 expression and mitochondrial structure during heart valve replacement surgery under cardiopulmonary bypass. METHODS: Fifty-four patients were prospectively randomized to receive or not ischemic preconditioning (IP) before cold cardioplegic arrest. Ischemic preconditioning in the IP patients (n=22) was induced by a single 2-min ischemia followed by 3-min reperfusion just before aortic clamping and cold crystalloid cardioplegia for myocardial protection. The control group (n=32) received no ischemic preconditioning before cold cardioplegic arrest. The levels of ejection fraction (EF), fractional shortening(FS) and stroke volume (SV) in both groups were measured and compared. troponin T (c-TnT) level, Bcl-2 protein expression and microscopic changes of myocardial mitochondrial structure were recorded for each group before and after surgery. RESULTS: The level of EF, FS and SV in IP group was higher than those in control group (P<0.05). No significant difference in preoperative c-TnT levels between two groups was observed. The level of c-TnT in IP group was lower than that in control group and with a declining trend over time of 6 h, 24 h, 48 h, 72 h and 5 d after surgery, respectively. The preoperative positive unit of Bcl-2 expression between two groups showed no statistical difference (P> 0.05). Postoperatively, the positive unit of Bcl-2 expression in IP group was 19.85±5.88, significantly increased as compared to the preoperative value (P<0.05). In control group, the positive unit of Bcl-2 expression was 14.17±3.39, showed no statistically significant difference to the preoperative value (P>0.05). Postoperative Bcl-2 expression between two groups showed a significant difference (P<0.05). In the control group, microscopic observation revealed swollen mitochondrion, with a hardly visible or disrupted membrane for some mitochondrion; mitochondrial crista were obviously dissolved and loose with a large number of vacuoles formation. However in IP group, myocardial mitochondrion appeared with intact membrane, concentrated mitochondrial cristae with high electron density and no vacuoles formation was observed. CONCLUSION: IP may up-regulate the expression of myocardial anti-apoptotic protein Bcl-2 to protect the mitochondrion, thus protecting cardiocytes and cardiac functions.     

Effect of ischemic preconditioning on myoelectricity and the structure of myocardial gap junction during heart valve replacement surgery

AIM: To investigate the cardio-protective mechanism of ischemic preconditioning (IP) during heart valve replacement of the perspective of architectural changes of myocardial gap junction. METHODS: Fifty-four patients were prospectively randomized to receive or not ischemic preconditioning before cold cardioplegic arrest. The IP protocol in IP patients (n=22) consisted of a single 2-minute ischemia followed by 3-minute reperfusion just before aortic clamping and cold crystalloid cardioplegia for myocardial protection. The control group (n=32) received no ischemic preconditioning prior to cold cardioplegic arrest. The parameters including arrhythmias occurrence, Cx43 expression (immunohistochemistry SABC method) and myocardial structure and intercalated discs observed under electronic microscope were recorded before and after surgery in each group. RESULTS: In IP group, one case (4.55%) of ventricular arrhythmia (sporadic ventricular premature beat), 11 cases (50.00%) of supraventricular arrhythmia (atrial fibrillation, atrial flutter, supraventricular tachycardia, atrioventricular block) and 10 cases (45.50%) of ischemic ST-T changes were observed. In control group, there were 14 cases (43.75%) of ventricular arrhythmia (ventricular premature beat, tachycardia), 18 cases (56.25%) of supraventricular tachycardia and 28 cases (87.50%) of ischemic ST-T changes. No statistical difference in preoperative positive unit of Cx43 expression between the two groups was found (P>0.05). Postoperatively, the positive unit of Cx43 expression in IP group was 16.15±4.40, but the difference was not significant compared to the preoperative value (P>0.05). In control group, Cx43 expression was 11.92±1.26, significantly lower than that of the preoperative value (P<0.05). Cx43 expression between the two postoperative groups showed a significant difference (P<0.05). In control group, electronic microscopic observation revealed disrupted intercalated discs, with some partially or even totally ruptured and disintegrated. Enormous necrotic structural changes of myocardial fibers were also observed, including swelling, dissolution and disorganization of myofilaments and fibers, widening of the Z striae and disorganization. However in IP group, the intercalated discs appeared intact, continuous with normal myocardial structure. CONCLUSION: IP maintains normal expression of the myocardial junctional gap protein Cx43, which preserves a seamless intercellular gap junction and a normal myocardial electric conduction activity.     

Expression of calcium sensing receptor during ischemia/reperfusion myocardial damage and relationship between CaSR and injury of myocardium

AIM: To investigate the expression of calcium sensing receptor(CaSR) during myocardial injury induced by ischemia/reperfusion and disclose the relationship between CaSR and myocardial ischemia/reperfusion. METHODS: The experimental model was established by the 30 min ligating and 1 h, 2 h, 4 h, 6 h reperfusing the left descending coronary artery (LAD) in rats. Wistar rats were randomly divided into 5 groups: sham group, ischemia/reperfusion 1 h, 2 h, 4 h, 6 h groups (I/R 1 h, 2 h, 4 h, 6 h group). CaSR mRNA expression was detected by RT-PCR. Left ventricular function was recorded. The levels of plasma lactate dehydrogenase (LDH), alondialdehyde (MDA) and superoxide dismutase (SOD) were measured. The change of ultrastructure in the ischemia/reperfusion myocardium of rats was observed by electron microscopy. RESULTS: LVSP,±dp/dtmax and SOD activity decreased gradually with the reperfusion time prolonged. LDH and MDA peaked at 2 h. The ultramicro-structural injury at the 1 h and 2 h was more serious than that at 4 h and 6 h. The expression of CaSR increased significantly after reperfusion of 1 h and 2 h, and decreased after 4 h and 6 h. CONCLUSION: The increased expression of CaSR mRNA and serious injure of myocardium were observed. CaSR may be associated with the myocardial ischemia/reperfusion injury.     

Effects of spermine on myocardial ischemia/ reperfusion injury in rats

AIM: To observe the effect of exogenous spermine (low concentration) on myocardial ischemia/reperfusion injury in rats.METHODS: 40 Wistar rats were randomly divided into 4 groups: sham- operation group (Sham), ischemic reperfusion group (I/R), spermine group (Sp) and natural saline group (NS). The model of ischemic/reperfusion injury was established by ligating rat coronary artery. In Sp group, spermine (0.5 mmol/L, 2 mL/kg) was injected slowly into rat vein. During the process, we recorded the electrocardiogram and the LV functional parameters, assayed the levels of SOD, LDH, NO and MDA in serum, and examined the ultrastructure of the myocardium. RESULTS: In I/R group, the incidence of arrhythmia was 90%, myocardial ultrastructure was injured seriously, values of LVSP and ±dp/dtmax decreased, levels of LDH, NO and MDA increased while SOD activity decreased (P<0.05 or P<0.01, compared with Sham group). Compared with I/R and NS group, all those indexes in Sp group changed significantly (P<0.05 or P<0.01). CONCLUSION: Exogenous spermine alleviates myocardial ischemia/ reperfusion injury in rats. The mechanism may be related to its antioxidant effect and relieving the injury caused by oxygen free radical.