AIM: To investigate the effect of diazoxide (D) postconditioning on Cardiac function and mito-chondrial cardiolipin in isolated rat heart and to explore the protective effect of ATP sensitive potassium channel on diazo-xide postconditioning myocardium.METHODS: The myocardial ischemia/reperfusion injury model in isolated rat hearts was established by Langendorff apparatus.The isolated rat hearts were randomized into 4 groups ( n=8): control group ( control) , myocardial ischemia/reperfusion injury group ( I/R) , diazoxide postconditioning group ( I/R+D) , 5-hydroxy decanoic acid (5-HD) plus diazoxide postconditioning group (I/R+5-HD+D).The hearts in each group were started with 20 min perfusion for equilibration.The hearts in control group perfused for 70 min;The hearts in I/R group was global ischemia for 40 min after ischemia reperfusion at 4℃ST.Thomas cardioplegia, then reperfusion for 30 min;The hearts in I/R+D group were treated with diazoxide (50μmol/L) in K-H perfusion for 5 min after global ischemia for 40 min, then reperfusion for 25 min;The hearts in I/R+5-HD+D group were treated with 5-HD (100μmol/L) in K-H perfusion for 5 min before diazoxide postconditioning, then reperfusion for 20 min.The heart rate, coronary outflow volume, heart func-tion, myocardial enzymes and myocardial mitochondrial cardiolipin at the end of perfusion in each group were determined. RESULTS:Compared with control group and I/R+D group, the heart rate, the concentration of heart phospholipid and the coronary outflow volume were reduced, the heart function was significantly impaired the contents of myocardial enzymes were increased in I/R group.However, no significant difference between I/R group and I/R+5-HD+D group was ob-served.CONCLUSION:The diazoxide postconditioning protects the myocardium by increasing mitochondrial cardiolipin content, reducing the release of myocardial enzymes, improving heart function and reducing myocardial reperfusion injury. The myocardial protective effect of diazoxide is completely blocked by 5-hydroxy decanoic acid.

Objective To evaluate the myocardial protective effect of hyperpolarized heart arrest at different temperature during cardiopulmonary bypass CPB.Methods Eighteen dogs were randomly allocated to be infused with St.Thomas solution containing 50?mol/L pinacidil and 5mmol/L K + at 37℃ (group A) or 4℃ (group B), or the standard St.Thomas solution containing K + 16mmol/L at 4℃ (group C) respectively, through aortic root after aortic clamping . The global surgical ischemia lasted 60min with the declamping of 20min .The activities of serum myocardial enzymes, and levels of lipid peroxide and adenine nucleotide of myocardium were measured.Results All paramaters showed to occur serious ischemic and reperfusion damages during the whole procedures in group C, while there were the mild damages in two hyperpolarized groups, especially in group B.Conclusions Myocardial protective effect of hyperpolarized arrest induced with pinacidil, an ATP sensitive potassium channel opener, is superior to that of traditional hyperkalemic cardioplegia , which is much more prominent in hypothermic state.

Objective To investigate the effects of diazoxide postconditioning on myocardial ischemiareperfusion (I/R) injury in isolated rat hearts. Methods Male SD rats weighing 250-300 g were anesthetized with intraperitoneal pentobarbital 40 mg/kg. Their hearts were excised and perfused in a Langendorff apparatus with K-H solution saturated with 95% O2-5% CO2 at 37 ℃. Sixty-four isolated rat hearts were randomized into 4 groups after 20 min of equilibration (n = 16 each): group control (group C), group I/R, group diazoxide postconditioning (group D) and group mito-KATP channel blocker 5-hydroxydecanoate (5-HD) + diazoxide postconditioning (group 5-HD + D). Group C received continuous perfusion for another 70 min. In group I/R, D and 5-HD + D, the hearts underwent 40 min of iscbemia followed by 30 min of reperfusion. 4 ℃ ST. Thomas cardioplegic solution was administered prior to ischemia in group I/R. Group D received 5 min of perfusion with K-H solution containing 50μ mol/L diazoxide at 5 min of reperfusion. Group 5-HD + D received 5 min of perfusion with K-H solution containing 50 μmol/L 5-HD before reperfusion with diazoxide. Eight hearts were taken at the end of equilibration and reperfusion and the indexes of cardiac function were recorded. Then the mitochondria were extracted for determination of mitochondrial membrane potential (MMP), reactive oxygen species (ROS) release, and respiratory function indexes. Results Compared with group C, MMP was significantly decreased, ROS release was significantly increased, mitochondrial respiratory function and cardiac function declined at the end of reperfusion in the other three groups ( P ＜ 0.05 or 0.01 ). MMP was significantly increased, ROS release was significantly decreased, mitochondrial respiratory function and cardiac function were improved in group D compared with group I/R and 5-HD + D.Conclusion Diazoxide postconditioning can reduce myocardial I/R injury in rats via the opening of mito-KATPchannels and improving the mitochondrial function.

Objective To investigate the effects of pyrrolidine dithiocarbamate (PDTC) on plasma concentrations of proinflammatory cytokines and myocardial energy metabolism induced by cardiopulmonary bypass (CPB) .Methods Twelve healthy mongrel dogs of both sexes weighing 13.5-17.5 kg were randomly divided into 2 groups (n = 6 in each group) : PDTC group and control group. The animals were anesthetized with intraperitoneal 2.5% pentobarbital sodium 25 mg?kg-1, intubated and mechanically ventilated. In PDTC group PDTC 30 mg?kg-1 was given i.v. after tracheal intubation while in control group normal saline was given instead of PDTC. Aorta was clamped for 60 min and then undamped for 60 min reperfusion during CPB. Blood samples were taken before (baseline), 30 and 60 min after aortic clamping and 30 and 60 min after aortic unclamping for determination of plasma concentrations of TNF-? and IL-1?. Myocardial tissue was obtained before and 60 min after aortic clamping and 60 min after aortic unclamping for determination of myocardial content of adenine nucleotide (ATP, ADP, AMP, TAN, EC) and expression of ICAM-1 protein.Results Plasma TNF-? concentration was increased after aortic cross-clamping as compared to the baseline value before clamping in both groups but the TNF-? concentration was significantly lower in PDTC group than in control group (P

AIM: To study the protective effect of hyperpolarized cardioplegic arrest on reperfused rat heart performance and to investigate the role of mitochondrial ATP-sensitive K+ channels(mitoKATP) opening in the protection of hyperpolarized cardioplegia against ischemia/reperfusion damage.METHODS: Forty Sprague-Dawley rats were randomized into five groups(n=8 in each group): control group(Con);depolarized arrest group(D);hyperpolarized arrest group(H);depolarized cardioplegia with 5-hydroxydecanoate(5-HD) group(5HD+D);hyperpolarized cardioplegia with 5-HD group(5HD+H).The rat hearts were quickly removed to Langendorff apparatus.The heart perfusion was performed for 20 min with 37 ℃ Krebs-Henseleit buffer balanced with gas mixture(O2∶CO2=95%∶5%) at 5.8 kPa perfusion pressure,then cardial arrest was induced by different cardioplegic solution.Hearts were subjected to ischemia at 37 ℃ for 40 min followed by 30 min reperfusion.(1) The hemodynamics was detected at recovery after 30 min reperfusion.(2) Before ischemia and at the end-reperfusion,tissue was harvested for mitochondrial isolation and ultrastructure was observed by transmission electron microscopy(TEM).(3) Production of reactive oxygen species(ROS) was also determined at different time points.RESULTS:(1) Compared with end-equilibration,30 min reperfusion caused significant differences in left ventricular developed pressure(LADP),left ventricular end-diastolic pressure(LVEDP),double product(DP),heart rate (HR),coronary flow(CF)(P

Objective To evaluate the effects of ischemic postconditioning on mitochondrial cardiolipin synthesis during myocardial ischemia-reperfusion (Ⅰ/R) in rats in vitro.Methods Healthy male Sprague-Dawley rats,aged 16-20 weeks,weighing 250-350 g,were used in the study.The animals were anesthetized with intraperitoneal pentobarbital sodium 40 mg/kg and received intraperitoneal heparin 250 U/kg.Their hearts were excised and retrogradely perfused in a Langendorff apparatus.Sixty-four isolated rat hearts were randomly divided into 4 groups (n =16 each) using a random number table:control group (group C),Ⅰ/R group,ischemic postconditioning group (group IPO) and 5-hydroxydecanoate (5-HD) plus ischemic postconditioning group (group 5-HD + IPO).After 20 min of equilibration,the hearts were continuously perfused with K-H solution for 70 min in group C.In Ⅰ/R group,after 20 min of equilibration,the hearts were continuously perfused with 4 ℃ ST.Thomas cardioplegic solution 10 ml/kg,and exposed to 40 min of ischemia followed by reperfusion with oxygenated K-H solution at 37 ℃ for 30 min.In group IPO,after 20 min of equilibration,the hearts were subjected to 6 cycles of 10 s reperfusion followed by 10 s ischemia starting from 40 min of ischemia,and then were reperfused with oxygenated K-H solution at 37 ℃ for 28 min.In group 5-HD + IPO,after 20 min of equilibration,the hearts were perfused with K-H solution containing 100 μmol/L 5-HD (mitochondrial ATP-sensitive potassium channel blocker) for 5 min starting from 40 min of ischemia,and then the other procedures were similar to those previously described in group IPO.At 20 min of equilibration (T1) and 30 min of reperfusion (T2),HR,left ventricular developed pressure (LVDP),left ventricular end-diastolic pressure (LVEDP) and coronary flow (CF) were recorded.The coronary effluent 2 ml was collected for detection of lactic dehydrogenase (LDH) and creatine kinase (CK) activities.The mitochondria were extracted for determination of cardiolipin content.Results HR,LVDP,and CF were significantly lower,LVEDP was higher,and the LDH and CK activities in coronary effluent were higher at T2 than at T1 in the four groups.Compared with group C,HR,LVDP and CF were significantly decreased,LVEDP was increased,and the LDH and CK activities in coronary effluent were increased at T2 in the other three groups.Compared with Ⅰ/R group,HR,LVDP and CF were significantly increased,LVEDP was decreased,and the LDH and CK activities in coronary effluent were decreased at T2 in IPO group.Compared with IPO group,HR,LVDP and CF were significantly decreased,LVEDP was increased,and the LDH and CK activities in coronary effluent were increased at T2 in 5-HD+IPO group.Conclusion The mechanism by which ischemic postconditioning reduces myocardial Ⅰ/R injury is related to opening of mitochondrial ATP sensitive potassium channels and increasing mitochondrial cardiolipin synthesis in rats.

Aim To discuss whether specific mitochon-drial ATP-sensitive potassium channel opener diazoxide ( DZ ) postconditioning activates RISK signaling path-way to protect isolated rat hearts against ischemica reperfusion injury ( IRI ) . Methods Langendorff de-vice was used to establish rat in vitro model of myocar-dial ischemia reperfusion. SD rats were randomly di-vided into normal group ( NOR ) , control group ( CON ) , diazoxide after treatment group ( DZ ) , and LY294002 antagonistic nitrogen Triazine group ( DZ +LY) , with 8 cases in each. The following was com-pared:①whether heart function of each group changed at the end of equilibration and reperfusion; ② at the end of myocardial perfusion and separation, protein was extracted, and protein kinase B ( PKB / Akt ) , P70S6 kinase (P70S6K), endothelial nitric oxide syn-thase ( eNOS) phosphorylation level of expression were analysed by Western blot. Results ① Indicators of changes in heart function: for DZ group at the end of reperfusion , HR , CF , LVDP , LVEDP , +d p/d tmax and -dp/dtmax were significantly better than those in CON group and DZ + LY group ( P <0.01 ) , but worse than those in NOR group ( P <0.01 ); there was no statistical difference in cardioac function at the end of equilibration. ② For DZ group at the end of reperfu-sion Akt, P70S6K, eNOS phosphorylation level of ex-pression were significantly higher than those in NOR group, CON group, and DZ + LY group (P<0.01). There was no difference in expression level of ERK1/2 phosphorylation ( P >0.05 ) . Conclusion Diazoxide postconditioning through the activation of RISK signa-ling pathway can protect isolated rat hearts against is-chemia reperfusion injury.

ObjectiveTo investigate the effect ofpinacidil hyperpolarizaed arrest on p38 mitogen-activited protein kinase (p38MAPK) during myocardial ischemia-reperfusion (I/R) in isolated rat hearts.MethodsFortyeight male SD rats weighting 250-300 g were randomly divided into 6 groups( n =8 each): natural arrest group (group A) ; St.Thomas group (group B) ; pinacidil hyperpolarization arrest group (group C) ;5-hydroxydecanoate (5-HD) group (group D);HMR-1098 group(group E) and 5-HD + HMR-1098 group(group F).Langendorff reperfusion model was established and K-H solution was retrogradely perfused for 15 min.In group A the hearts were arrested naturally afar perfusion was stopped; in group B St.Thomas solution was perfused; in group C pinacidil hyperpolarization solution was perfused; in the other three groups,K-H solution was perfused to isolated rat hearts for 10 min followed by 5 min 5-HD (group D) or HMR-1098(group E) or 5-HD and HMR-1098(group F) perfusion,then hyperpolarization arrest solution was given in each group.Each hearts suffered 60 min ischemia after arrest followed by 30 min K-H solution reperfusion.Coronary flow(CF),HR,left ventricular developed pressure( LVDP),left ventricular systolic pressure(LVSP) and the maximum rate of pressure rise (dp/dtmax) were measured at the end of 15 min K-H solution perfusion and at 20 min of reperfusion.Myocardial phosphatic and nonphosphatic p38MAPK expression was determined by Western blot at 30 min of reperfusion.ResultsCompared with group C,CF,HR,LVDP,LVSP and dp/dtmax were significantly decreased at 20 min of reperfusion and phosphatic p38MAPK expression was down-regulated,non-phosphatic p38MAPK expression was up-regulated at 30 min of reperfusion in groups A,B,D,E and F (P ＜ 0.05).Compared with group E,CF,HR,LVDP,LVSP and dp/ dtmax were significantly decreased at 20 min of reperfusion and phosphatic p38MAPK expression was down-regulated,non-phosphatic p38MAPK expression was up-regulated at 30 min of reperfusion in groups D and F ( P ＜0.05).ConclusionHyperpolarized arrest induced by pinacidil can improve cardiac function following myocardial I/R injury by up-regulating phosphatic p38MAPK expression,down-regulating non-phosphatic p38 MAPK expression and mitochondrial potassium channel is more important than membranous one during the regulation of phosphatic p38MAPK expression.

Objective To evaluate the role of nuclear factor-E2 related factor 2 (Nrf2)-antioxidant response element (ARE) pathway in cardio-protection by ischemic or pinacidil postconditioning ( IP,PP) against ischemia-reperfusion (I/R) injury in isolated rat hearts.Methods Fifty-six male SD rats of both sexes weighing 200-250 g were anesthetized with intraperitoneal amobarbital sodium.The isolated rat hearts were perfused in a Langendorff apparatus with Krebs-Hensleit buffer (K-H).Fifty-six isolated rat hearts with I/R injury were randomly divided into 7 groups ( n =8 each):normal control group (group C) ; group I/R; group IP and group PP1-4 postconditioning with 4 different concentrations of pinacidil.After 20 min of equilibration,the perfusion was suspended for 40 min (global ischemia) followed by 60 min of reperfusion in group I/R.In group IP after 40 min of global ischemia,the isolated hearts underwent 6 cycles of 10 s reperfusion and 10 s ischemia followed by 58 min of reperfusion.In group PP1-4 at the end of 40 min of global ischemia,the isolated hearts were perfused with K-H containing pinacidil 5,10,30 and 50μmol/L for 5 min respectively followed by 55 min reperfusion with regular K-H.Left ventricular developed pressure (LVDP) and LVEDP were measured immediately before global ischemia and at the end of 60 min reperfusion.Myocardial specimens were obtained at the end of reperfusion for detection of Nrf2,quinopeoxidoreductase (NQO1),HO-1 and SOD1 mRNA (by RT-PCR) and protein (by Western blot) expression.Results I/R significantly up-regulated Nrf2,NQO1,HO-1 and SODI mRNA and protein expression,decreased LVDP and increased LVEDP in group I/R as compared with group C.IP and 30,50 μmol/L pinacidil postconditioning further significantly increased Nrf2,NQO1,HO-1 and SOD1 mRNA and protein expression and IP,5,10,30,50 μmol/L pinacidil postconditioning significantly increased LVDP and decreased LVEDP as compared with group I/R.Conclusion Ischemic or pinacidil postconditioning can attenuate I/R injury by activating Nrf2-ARE pathway in isolated rat hearts.

Objective To investigate the effect of hydrogen sulfide postconditioning on the systolic function of left ventricle during myocardial ischemia-reperfusion (IR) in rats. Methods Part Ⅰ Adult male SD rats weighing 200-250 g were anesthetized with pentobarbital 40 mg/kg and heparin 250 U/kg. Their hearts were excised and perfused in a Langendorff apparatus with K-H solution saturated with 95% O2-5% CO2 at 37 ℃. Forty isolated rat hearts were randomly divided into 5 groups ( n = 8 each) after 20 min of equilibration: control group (group C); IR group; sodium hydrosulfide 1,10, 100 μmol/L postconditioning group (group SP1, SP10, SP100 ).In group Cthe hearts were perfused continuously for another 100 min. In group IR, the hearts were reperfused for 60 min after 40 min ischemia induced by 10 ml/kg ST. Thomas solution. In group SP1 , SP10 and SP100 the hearts were perfused with K-H solution containing sodium hydrosulfide 1, 10, 100 μmol/L for 2 min before reperfusion.LVDP and ± dp/dtmax were recorded at the end of equilibration and reperfusion. Part Ⅱ Cardiomyocytes were isolated from the male SD rats (weighing 200-250 g) and then cultured in CO2 incubator for 4 h. Sixty-four dishes of cultured myocytes were randomly divided into 4 groups( n = 16 each): control group (group C), hypoxia/reoxygenation group (group HR), hydrogen sulfide postconditioning group (group SP) and hypoxia postconditioning group (group HP). Group C were cultured continuously for 2 h. Group HR, SP and HP were exposed to 1 h hypoxia (95%N2-5%CO2 ) followed by 1 h reoxygenation. In group SP 10 μmol/L sodium hydrosulfide was added and the myocytes were then incubated for 2 min before reoxygenation. In group HP the cultured myocytes were expased to 3 min reoxygenation followed by 3 min hypoxia for 3 times before the 1 h reoxygenation. Mitochondrial membrane potential and F-actin expression were determined. Results Part Ⅰ Compared with group C, LVDP and ± dp/dtmax were significantly decreased at the end of reperfusion in group IR (P ＜ 0.05), while no significant difference was found in group SP1 , SP10 and SP100(P ＞0.05). Compared with group IR, LVDP and ± dp/dtmax were significantly increased in group SP ( P ＜ 0.05). There was no significant difference in LVDP and ± dp/dtmax among group SP1, SP10 and SP100(P ＞0.05). Part H Compared with group C, the mitochondrial membrane potential was significantly decreased in group HR and HP, and the expression of F-actin was significantly up-regulated in group HR, SP and HP ( P ＜ 0.05). Compared with group HR, the mitochondrial membrane potential was significantly increased and the expression of F-actin up-regulated in group SP and HP ( P ＜ 0.05 ). There were no significant difference in the mitochondrial membrane potential and expression of F-actin between group SP and HP ( P ＞0.05).Conclusion Hydrogen sulfide postconditioning can improve left ventricular systolic function during IR in rats by stabilizing mitochondrial membrane potential and promoting aggregation of F-actin.