In this study, we treated PC12 cells with 0-20 μM amyloid-β peptide （25-35） for 24 hours to induce cytotoxicity, and found that 5-20 μM amyloid-β peptide （25-35） decreased PC12 cell viability, but adenosine tr...In this study, we treated PC12 cells with 0-20 μM amyloid-β peptide （25-35） for 24 hours to induce cytotoxicity, and found that 5-20 μM amyloid-β peptide （25-35） decreased PC12 cell viability, but adenosine triphosphate-sensitive potassium channel activator diazoxide suppressed the decrease in PC12 cell viability induced by amyloid-β peptide （25-35）. Diazoxide protected PC12 cells against amyloid-β peptide （25-35）-induced increases in mitochondrial membrane potential and intracellular reactive oxygen species levels. These protective effects were reversed by the selective mitochondrial adenosine triphosphate-sensitive potassium channel blocker 5-hydroxydecanoate. An inducible nitric oxide synthase inhibitor, Nw-nitro-L-arginine, also protected PC12 cells from amyloid-β peptide （25-35）-induced increases in both mitochondrial membrane potential and intracellular reactive oxygen species levels. However, the H202-degrading enzyme catalase could not reverse the amyloid-β peptide （25-35）-induced increase in intracellular reactive oxygen species. A 24-hour exposure to amyloid-13 peptide （25-35） did not result in apoptosis or necrosis, suggesting that the increases in both mitochondrial membrane potential and reactive oxygen species levels preceded cell death. The data suggest that amyloid-β peptide （25-35） cytotoxicity is associated with adenosine triphosphate-sensitive potassium channels and nitric oxide. Regulation of adenosine triphosphate-sensitive potassium channels suppresses PC12 cell cytotoxicity induced by amyloid-β peptide （25-35）.展开更多
Background Many studies have indicated that hyperpolarizing cardioplegia is responsible for myocardial preservation and researchers have suggested that the adenosine triphosphate-sensitive potassium channels （KATe） ...Background Many studies have indicated that hyperpolarizing cardioplegia is responsible for myocardial preservation and researchers have suggested that the adenosine triphosphate-sensitive potassium channels （KATe） were the end effectors of cardio-protection. But whether mitochondrial KATe plays an important role in hyperpolarizing cardioplegia is not apparent. The present study investigated the effect of hyperpolarizing cardioplegia containing pinacidil （a nonselective KATe opener） on ischemia/repeffusion injury in rat hearts, especially the role of mitochondrial KATe in pinacidil hyperpolarizing cardioplegia. Methods Sprague-Dawley rat hearts were Langendorff-perfused for 20 minutes with Krebs-Henseleit buffer at 37℃ before equilibration. Cardiac arrest was then induced in different treatments： there was no arrest and ischemia in the normal group, the control group were arrested by clamping the aorta, depolarizing caidioplegia （St. Thomas solution containing 16 mmol/L KCI） and hyperpolarizing cardioplegia groups used St. Thomas solution containing 0.05 mmol/L pinacidil and 5 mmol/L KCI to induce cardiac arrest in group hyperkalemic and group pinacidil, in group hyperkalemic ＋ 5-hydroxydecanote （5HD） and Pinacidil ＋ 5HD, 5HD （0.1 retool/L） was added to the above two solutions to block mitochondria KATe channels. Global ischemia was then administrated for 40 minutes at 37℃, followed by 30 minutes of reperfusion. At the end of equilibration and reperfusion, hemodynamics, ultrastructure, and mitochondrial function were measured. Results In the control group, ischemia/reperfusion decreased the left ventricular developed pressure, heart rate, coronary flow, mitochondrial membrane potential, impaired mitochondrial respiratory function, increased reactive oxygen species and left ventricular end diastolic pressure. Damage to myocardial ultrastructure was also evident. Both depolarized arrest and especially hyperpolarized cardioplegia significantly reduced these lesions. 5HD partially blocked the beneficial effects of pinacidil cardioplegia but showing no effects on hyperkalemic arrest. Conclusions Pinacidil cardioplegia provides better cardioprotection with preservation of hemodynamics, ultrastructure, and mitochondrial function than traditional cardioplegia. The mitochondria KATe channels may play an important role in the protection mechanism.展开更多
Backgroud Recent studies in adult hearts have indicated that K ATP channels in the inner mitochondrial membrance are responsible for the protection. And we investigated whether opening of mitochondrial K ATP ...Backgroud Recent studies in adult hearts have indicated that K ATP channels in the inner mitochondrial membrance are responsible for the protection. And we investigated whether opening of mitochondrial K ATP channels (mK ATP ) could provide myocardial protection for immature rabbits and determined its role in cardioprotection Methods Thirty-four 3-4-week-old rabbits, weighing 300-350 g, were divided randomly into five groups: Group Ⅰ (control group, n=8); Group Ⅱ [diazoxide preconditioning group; n=8; the hearts were pretreated with 100 μmol/L diazoxide for 5 minutes followed by 10-minute wash out with Krebs-Henseleit buffer (KHB)]; Group Ⅲ ; Group Ⅲ [diazoxide+5-hydroxydeconate (5-HD) preconditioning group; n=5; the hearts were pretreated with 100 μmol/L diazoxide and 100 μmol/L 5-HD); Group Ⅳ (diazoxide+cardioplegia group; n=8; cardioplegia containing 100 μmol/L diazoxide perfused the hearts for 5 minutes before ischemia); Group Ⅴ (diazoxide+5-HD+cardioplegia group; n=5; the cardioplegia contained 100 μmol/L diazoxide and 100 μmol/L 5-HD) All hearts were excised and connected to langendrff perfusion system and passively perfused with KHB at 38℃ under a pressure of 70 cmH 2O After reperfusion, the recovery rate of left ventricular diastolic pressure (LVDP), ±dp/dt max , coronary flow (CF), the creatinine kinase (CK), lactate dehydrogenase (LDH), aspartate aminotransferase (AST) in coronary sinus venous effluent and the tissue ATP were measured Mitochondria were evaluated semiquantitatively by morphology Results After ischemia and reperfusion (I/R), the two groups that were treated by diazoxide only (Groups Ⅱ and Ⅳ) had a significant improvement in LVDP, ±dp/dt max , and CF recovery AST, LDH, and CK were decreased, and the levels of tissue ATP in the two groups were higher Mitochondria was protected better in Group Ⅳ than in other groups Conclusions Activating mK ATP channels before and during ischemia can similarly protect immature rabbit hearts, and the mechanism is related to the direct protective effect on mitochondria Opening of mK ATP channel during ischemia provides a better protection for mitochondria than it does before ischemia展开更多
Reperfusion therapy must be applied as soon as possible to attenuate the ischemic insult of acute myocardial infarction(AMI).However reperfusion is responsible for additional myocardial damage,which likely involves op...Reperfusion therapy must be applied as soon as possible to attenuate the ischemic insult of acute myocardial infarction(AMI).However reperfusion is responsible for additional myocardial damage,which likely involves opening of the mitochondrial permeability transition pore(mPTP).In reperfusion injury,mitochondrial damage is a determining factor in causing loss of cardiomyocyte function and viability.Major mechanisms of mitochondrial dysfunction include the long lasting opening of mPTPs and the oxidative stress resulting from formation of reactive oxygen species(ROS).Several signaling cardioprotective pathways are activated by stimuli such as preconditioning and postconditioning,obtained with brief intermittent ischemia or with pharmacological agents.These pathways converge on a common target,the mitochondria,to preserve their function after ischemia/reperfusion.The present review discusses the role of mitochondria in cardioprotection,especially the involvement of adenosine triphosphate-dependent potassium channels,ROS signaling,and the mPTP.Ischemic postconditioning has emerged as a new way to target the mitochondria,and to drastically reduce lethal reperfusion injury.Several clinical studies using ischemic postconditioning during angioplasty now support its protective effects,and an interesting alternative is pharmacological postconditioning.In fact ischemic postconditioning and the mPTP desensitizer,cyclosporine A,have been shown to induce comparable protection in AMI patients.展开更多
Background Hypoxic pulmonary hypertension (HPH) is initiated by inhibition of O 2 sensitive, voltage gated (Kv) channels in pulmonary arterial smooth muscle cells (PASMCs) The mechanism of hypoxic pulmonary hyp...Background Hypoxic pulmonary hypertension (HPH) is initiated by inhibition of O 2 sensitive, voltage gated (Kv) channels in pulmonary arterial smooth muscle cells (PASMCs) The mechanism of hypoxic pulmonary hypertension has not yet been fully elucidated The mitochondrial ATP sensitive K + channel (MitoK ATP ) is extremely sensitive to hypoxia, and is a decisive factor in the control of mitochondrial membrane potential (ΔΨ m) This study investigated the changes of cell membrane potential and Kv channel in cultured human pulmonary artery smooth muscle cell (hPASMC) exposed to 24 hour hypoxia, and explored the role of MitoK ATP and ΔΨ m in this condition Methods Fresh human lung tissues were obtained from the patients undergoing a chest operation hPASMCs were isolated, cultured, and divided into 6 groups: ① control group, cultured under normoxia; ② diazoxide group, cultured in normoxia with diazoxide, an opener of MitoK ATP ; ③ 5 HD group, cultured in normoxia with sodium 5 hydroxydecanoate (5 HD), an antagonist of MitoK ATP ; ④ 24 hour hypoxia group; ⑤ 24 hour hypoxia + diazoxide group; and ⑥ 24 hour hypoxia + 5HD group Whole cell patch clamp technique was used to trace the cell membrane K + currents The expressions of cell membrane Kv1 5 mRNA and protein were determined by RT PCR and Western blot technique, respectively The relative changes in mitochondrial potential were tested with rhodamine fluorescence (R 123) technique Results After exposure to diazoxide for 24 hours, the intensity of R 123 fluorescence in normoxic hPASMCs was significantly increased compared with control group ( P <0 05), but there were no significant changes in these tests after the hPASMCs had been exposed to 5 HD for 24 hours Twenty four hour hypoxia or 24 hour hypoxia + diazoxide could markedly increase the intensity of R 123 fluorescence in hPASMC and the changes were more significant in 24 hour hypoxia +diazoxide group than in 24 hour hypoxia group ( P <0 05) although 5 HD could partly weaken the effect of 24 hour hypoxia on the intensity of R 123 fluorescence After exposure to diazoxide for 24 hours, the cell membrane K + currents and the expression of cell membrane Kv1 5 mRNA and protein in normoxic hPASMCs were significantly decreased compared with control group ( P <0 05), but there were no significant changes in these tests after the hPASMCs had been exposed to 5 HD for 24 hours Also, 24 hour hypoxia or 24 hour hypoxia + diazoxide decreased the cell membrane K + currents and the expression of Kv1 5 mRNA and protein ( P <0 05) but the changes were more significant in 24 hour hypoxia + diazoxide group than in 24 hour hypoxia group ( P <0 05) Again, 5 HD could partly weaken the inhibitory effect of 24 hour hypoxia on the cell membrane K + currents and the expression of Kv1 5 mRNA or protein ( P <0 05) Conclusions The opening of MitoK ATP followed by a depolarization of ΔΨ m in hypoxia might contribute to the alterations in the expression of cell membrane Kv1 5 mRNA and protein leading to change in the cell membrane potential of hypoxic hPASMCs This might be a mechanism of the development of hypoxic pulmonary hypertension展开更多
基金supported by the Project Sponsored by Yantai Science and Technology Bureau,China,No.2010232
文摘In this study, we treated PC12 cells with 0-20 μM amyloid-β peptide （25-35） for 24 hours to induce cytotoxicity, and found that 5-20 μM amyloid-β peptide （25-35） decreased PC12 cell viability, but adenosine triphosphate-sensitive potassium channel activator diazoxide suppressed the decrease in PC12 cell viability induced by amyloid-β peptide （25-35）. Diazoxide protected PC12 cells against amyloid-β peptide （25-35）-induced increases in mitochondrial membrane potential and intracellular reactive oxygen species levels. These protective effects were reversed by the selective mitochondrial adenosine triphosphate-sensitive potassium channel blocker 5-hydroxydecanoate. An inducible nitric oxide synthase inhibitor, Nw-nitro-L-arginine, also protected PC12 cells from amyloid-β peptide （25-35）-induced increases in both mitochondrial membrane potential and intracellular reactive oxygen species levels. However, the H202-degrading enzyme catalase could not reverse the amyloid-β peptide （25-35）-induced increase in intracellular reactive oxygen species. A 24-hour exposure to amyloid-13 peptide （25-35） did not result in apoptosis or necrosis, suggesting that the increases in both mitochondrial membrane potential and reactive oxygen species levels preceded cell death. The data suggest that amyloid-β peptide （25-35） cytotoxicity is associated with adenosine triphosphate-sensitive potassium channels and nitric oxide. Regulation of adenosine triphosphate-sensitive potassium channels suppresses PC12 cell cytotoxicity induced by amyloid-β peptide （25-35）.
基金This project was supported by a grant from the National Natural Science Foundation of China （No. 30460132）.
文摘Background Many studies have indicated that hyperpolarizing cardioplegia is responsible for myocardial preservation and researchers have suggested that the adenosine triphosphate-sensitive potassium channels （KATe） were the end effectors of cardio-protection. But whether mitochondrial KATe plays an important role in hyperpolarizing cardioplegia is not apparent. The present study investigated the effect of hyperpolarizing cardioplegia containing pinacidil （a nonselective KATe opener） on ischemia/repeffusion injury in rat hearts, especially the role of mitochondrial KATe in pinacidil hyperpolarizing cardioplegia. Methods Sprague-Dawley rat hearts were Langendorff-perfused for 20 minutes with Krebs-Henseleit buffer at 37℃ before equilibration. Cardiac arrest was then induced in different treatments： there was no arrest and ischemia in the normal group, the control group were arrested by clamping the aorta, depolarizing caidioplegia （St. Thomas solution containing 16 mmol/L KCI） and hyperpolarizing cardioplegia groups used St. Thomas solution containing 0.05 mmol/L pinacidil and 5 mmol/L KCI to induce cardiac arrest in group hyperkalemic and group pinacidil, in group hyperkalemic ＋ 5-hydroxydecanote （5HD） and Pinacidil ＋ 5HD, 5HD （0.1 retool/L） was added to the above two solutions to block mitochondria KATe channels. Global ischemia was then administrated for 40 minutes at 37℃, followed by 30 minutes of reperfusion. At the end of equilibration and reperfusion, hemodynamics, ultrastructure, and mitochondrial function were measured. Results In the control group, ischemia/reperfusion decreased the left ventricular developed pressure, heart rate, coronary flow, mitochondrial membrane potential, impaired mitochondrial respiratory function, increased reactive oxygen species and left ventricular end diastolic pressure. Damage to myocardial ultrastructure was also evident. Both depolarized arrest and especially hyperpolarized cardioplegia significantly reduced these lesions. 5HD partially blocked the beneficial effects of pinacidil cardioplegia but showing no effects on hyperkalemic arrest. Conclusions Pinacidil cardioplegia provides better cardioprotection with preservation of hemodynamics, ultrastructure, and mitochondrial function than traditional cardioplegia. The mitochondria KATe channels may play an important role in the protection mechanism.
文摘Backgroud Recent studies in adult hearts have indicated that K ATP channels in the inner mitochondrial membrance are responsible for the protection. And we investigated whether opening of mitochondrial K ATP channels (mK ATP ) could provide myocardial protection for immature rabbits and determined its role in cardioprotection Methods Thirty-four 3-4-week-old rabbits, weighing 300-350 g, were divided randomly into five groups: Group Ⅰ (control group, n=8); Group Ⅱ [diazoxide preconditioning group; n=8; the hearts were pretreated with 100 μmol/L diazoxide for 5 minutes followed by 10-minute wash out with Krebs-Henseleit buffer (KHB)]; Group Ⅲ ; Group Ⅲ [diazoxide+5-hydroxydeconate (5-HD) preconditioning group; n=5; the hearts were pretreated with 100 μmol/L diazoxide and 100 μmol/L 5-HD); Group Ⅳ (diazoxide+cardioplegia group; n=8; cardioplegia containing 100 μmol/L diazoxide perfused the hearts for 5 minutes before ischemia); Group Ⅴ (diazoxide+5-HD+cardioplegia group; n=5; the cardioplegia contained 100 μmol/L diazoxide and 100 μmol/L 5-HD) All hearts were excised and connected to langendrff perfusion system and passively perfused with KHB at 38℃ under a pressure of 70 cmH 2O After reperfusion, the recovery rate of left ventricular diastolic pressure (LVDP), ±dp/dt max , coronary flow (CF), the creatinine kinase (CK), lactate dehydrogenase (LDH), aspartate aminotransferase (AST) in coronary sinus venous effluent and the tissue ATP were measured Mitochondria were evaluated semiquantitatively by morphology Results After ischemia and reperfusion (I/R), the two groups that were treated by diazoxide only (Groups Ⅱ and Ⅳ) had a significant improvement in LVDP, ±dp/dt max , and CF recovery AST, LDH, and CK were decreased, and the levels of tissue ATP in the two groups were higher Mitochondria was protected better in Group Ⅳ than in other groups Conclusions Activating mK ATP channels before and during ischemia can similarly protect immature rabbit hearts, and the mechanism is related to the direct protective effect on mitochondria Opening of mK ATP channel during ischemia provides a better protection for mitochondria than it does before ischemia
基金Supported by National Institutes of Cardiovascular ResearchRegione Piemonte,PRIN,ex-60% and Compagnia di San Paolo,Italy
文摘Reperfusion therapy must be applied as soon as possible to attenuate the ischemic insult of acute myocardial infarction(AMI).However reperfusion is responsible for additional myocardial damage,which likely involves opening of the mitochondrial permeability transition pore(mPTP).In reperfusion injury,mitochondrial damage is a determining factor in causing loss of cardiomyocyte function and viability.Major mechanisms of mitochondrial dysfunction include the long lasting opening of mPTPs and the oxidative stress resulting from formation of reactive oxygen species(ROS).Several signaling cardioprotective pathways are activated by stimuli such as preconditioning and postconditioning,obtained with brief intermittent ischemia or with pharmacological agents.These pathways converge on a common target,the mitochondria,to preserve their function after ischemia/reperfusion.The present review discusses the role of mitochondria in cardioprotection,especially the involvement of adenosine triphosphate-dependent potassium channels,ROS signaling,and the mPTP.Ischemic postconditioning has emerged as a new way to target the mitochondria,and to drastically reduce lethal reperfusion injury.Several clinical studies using ischemic postconditioning during angioplasty now support its protective effects,and an interesting alternative is pharmacological postconditioning.In fact ischemic postconditioning and the mPTP desensitizer,cyclosporine A,have been shown to induce comparable protection in AMI patients.
基金Thestudywassupportedbya grantfromtheNationalNaturalScienceFoundationofChina (No 3 0 3 70 62 3 )
文摘Background Hypoxic pulmonary hypertension (HPH) is initiated by inhibition of O 2 sensitive, voltage gated (Kv) channels in pulmonary arterial smooth muscle cells (PASMCs) The mechanism of hypoxic pulmonary hypertension has not yet been fully elucidated The mitochondrial ATP sensitive K + channel (MitoK ATP ) is extremely sensitive to hypoxia, and is a decisive factor in the control of mitochondrial membrane potential (ΔΨ m) This study investigated the changes of cell membrane potential and Kv channel in cultured human pulmonary artery smooth muscle cell (hPASMC) exposed to 24 hour hypoxia, and explored the role of MitoK ATP and ΔΨ m in this condition Methods Fresh human lung tissues were obtained from the patients undergoing a chest operation hPASMCs were isolated, cultured, and divided into 6 groups: ① control group, cultured under normoxia; ② diazoxide group, cultured in normoxia with diazoxide, an opener of MitoK ATP ; ③ 5 HD group, cultured in normoxia with sodium 5 hydroxydecanoate (5 HD), an antagonist of MitoK ATP ; ④ 24 hour hypoxia group; ⑤ 24 hour hypoxia + diazoxide group; and ⑥ 24 hour hypoxia + 5HD group Whole cell patch clamp technique was used to trace the cell membrane K + currents The expressions of cell membrane Kv1 5 mRNA and protein were determined by RT PCR and Western blot technique, respectively The relative changes in mitochondrial potential were tested with rhodamine fluorescence (R 123) technique Results After exposure to diazoxide for 24 hours, the intensity of R 123 fluorescence in normoxic hPASMCs was significantly increased compared with control group ( P <0 05), but there were no significant changes in these tests after the hPASMCs had been exposed to 5 HD for 24 hours Twenty four hour hypoxia or 24 hour hypoxia + diazoxide could markedly increase the intensity of R 123 fluorescence in hPASMC and the changes were more significant in 24 hour hypoxia +diazoxide group than in 24 hour hypoxia group ( P <0 05) although 5 HD could partly weaken the effect of 24 hour hypoxia on the intensity of R 123 fluorescence After exposure to diazoxide for 24 hours, the cell membrane K + currents and the expression of cell membrane Kv1 5 mRNA and protein in normoxic hPASMCs were significantly decreased compared with control group ( P <0 05), but there were no significant changes in these tests after the hPASMCs had been exposed to 5 HD for 24 hours Also, 24 hour hypoxia or 24 hour hypoxia + diazoxide decreased the cell membrane K + currents and the expression of Kv1 5 mRNA and protein ( P <0 05) but the changes were more significant in 24 hour hypoxia + diazoxide group than in 24 hour hypoxia group ( P <0 05) Again, 5 HD could partly weaken the inhibitory effect of 24 hour hypoxia on the cell membrane K + currents and the expression of Kv1 5 mRNA or protein ( P <0 05) Conclusions The opening of MitoK ATP followed by a depolarization of ΔΨ m in hypoxia might contribute to the alterations in the expression of cell membrane Kv1 5 mRNA and protein leading to change in the cell membrane potential of hypoxic hPASMCs This might be a mechanism of the development of hypoxic pulmonary hypertension
基金This work was supported by the Natural Science Foundation of Jiangsu Province(No.BK2002138).
文摘利用离体海马脑片缺氧无糖(oxygen-glucose deprivation,OGD)损伤模型,探讨七氟醚预处理对神经细胞的保护作用及陔作用与线粒体内膜ATP敏感钾通道(mitochondrial ATP-sensitive potassium channels,mitoK_(ATP)channels)的关系,随机将脑片用2%、4%、6%七氟醚,以及6%七氟醚复合mitoK_(ATP)通道阻滞剂5-羟基奎酸盐(5-hydroxydecanoic acid,5-HD)预处理30min,观察OGD损伤14min复氧1h期间顺向群峰电位(orthodromic population spike,OPS)的变化,并应用透射电镜观察细胞超微结构的改变。结果表明,与单纯OGD组相比,七氟醚预处理可使海马脑片OPS消失时间明显延长(P＜0．01),使OPS明显恢复,其中4%、6%七氟醚组的恢复率均为71．4%(P＜0．05 vs OGD),相应恢复程度为(61．0±42．3)%和(78．7±21．1)%(P＜0．01),而且6%七氟醚的保护作用可被5-HD取消。OGD组的海马CA1区锥体细胞明显水肿,核膜皱缩、破裂,染色质聚集,线粒体肿胀畸形,嵴断裂或消失,而4%和6%七氟醚组仅见海马CA1区锥体细胞轻度水肿,核膜皱缩不明显,染色质均匀,线粒体轻度肿胀。结果提示,七氟醚预处理对大鼠海马脑片OGD损伤有一定的保护作用,且七氟醚对神经细胞的保护作用与激活mitoK_(ATP)通道有关。
