NO信号通路在慢性缺氧心肌线粒体生物合成中的调控机制研究
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摘要
背景与目的
心肌慢性缺氧是临床常见的病理生理过程,可导致心肌产生一系列代谢适应性改变,与线粒体的数量和功能密切相关的线粒体生物合成过程的适应性改变可能是其中重要的环节,但慢性缺氧对心肌线粒体生物合成影响的机制目前尚不完全清楚。一氧化氮(NO)可能参与了多种细胞线粒体生物合成的调控,其在慢性缺氧心肌线粒体生物合成中的作用尚不明了。深入研究NO与慢性缺氧心肌线粒体的适应性改变和调控之间的关系,将有望阐明缺氧状态下心肌的病理生理改变机制,为进一步增强缺氧状态下的心肌保护,提高临床治疗效果提供新的思路。本课题分别在先天性心脏病心肌组织标本和体外培养慢性缺氧心肌细胞模型上,对心肌细胞的线粒体生物合成和NO信号通路变化进行形态学和分子生物学研究,探讨NO在慢性缺氧状态下对心肌线粒体生物合成的调控作用机制。研究方法
本研究共分为三部分:
第一部分:选取紫绀型和非紫绀型先天性心脏病患者各10例,留取手术中切除的肥厚右室流出道组织,通过电镜观察分析标本心肌细胞超微结构,运用实时荧光PCR、RT-PCR及Western blot等方法比较心肌组织mtDNA拷贝数,细胞色素c氧化酶I (COX I)、过氧化物酶体增生物激活受体γ辅激活因子-1α(PGC-1α)、核呼吸因子1(NRF1)、线粒体转录因子A(Tfam)和三型NOS的mRNA或(和)蛋白水平,检测NOS酶活性,并对PGC-1α、eNOS蛋白水平与患者末梢血氧饱和度SaO2进行相关性分析。
第二部分:体外培养乳鼠心肌细胞,根据不同干预因素将心肌细胞分为对照组,NO供体SNAP组,SNAP+NO清除剂氧合血红蛋白(OxyHb)组及SNAP+可溶性鸟苷酸环化酶(sGC)抑制剂(ODQ)组。应用线粒体荧光探针在激光共聚焦显微镜下观察各组细胞线粒体量的变化,应用实时荧光PCR、RT- PCR及Western blot等方法分别比较各组细胞mtDNA拷贝数,COX I、PGC-1α、NRF1和Tfam的mRNA水平以及COXI蛋白水平等。
第三部分:通过1%O2浓度培养48小时建立体外培养乳鼠心肌细胞慢性缺氧模型,根据不同干预方式将心肌细胞分为对照组,慢性缺氧组,慢性缺氧+NOS抑制剂L-NAME组,慢性缺氧+OxyHb组以及慢性缺氧+ODQ组。检测细胞培养液上清NO浓度和心肌细胞NOS蛋白水平,应用线粒体荧光探针观察各组细胞线粒体量的变化,应用实时荧光PCR、RT- PCR及Western blot等方法分别比较各组细胞mtDNA拷贝数,COX I、PGC-1α、NRF1和Tfam的mRNA水平以及COX I蛋白水平等。
研究结果
本研究主要结果如下:
1.紫绀型先天性心脏病右室流出道心肌组织中线粒体体密度Vv、数密度Nv明显高于非紫绀组(p<0.05, p<0.01),mtDNA拷贝数显著高于非紫绀组(p<0.01),COX I、PGC-1α、NRF1和Tfam的mRNA水平均显著高于非紫绀组(p<0.01),COX I、eNOS蛋白表达显著高于非紫绀组(p<0.05, p<0.05),心肌组织PGC-1αmRNA水平与SaO2呈显著的负相关(r=-0.75, p<0.01),eNOS蛋白水平与SaO2呈显著的负相关(r=-0.64, p<0.01),PGC-1αmRNA水平与eNOS蛋白水平呈显著的正相关(r=0.58, p<0.01);
2. (1) NO供体SNAP干预组心肌细胞线粒体荧光强度、mtDNA拷贝数、COXI mRNA和蛋白水平均明显高于对照组(p<0.01, p<0.01, p<0.01, p<0.01),与线粒体生物合成相关的信号分子PGC-1α、NRF1和Tfam mRNA水平也高于对照组(p<0.01, p<0.01, p<0.05);
(2)与SNAP组比较,SNAP+OxyHb组线粒体荧光强度、mtDNA拷贝数、COXI mRNA均降低(p<0.01, p<0.01, p<0.05),PGC-1α和NRF1 mRNA水平也降低(p<0.01, p<0.05);
(3)与SNAP组比较,SNAP+ODQ组线粒体荧光强度、mtDNA拷贝数和COXI mRNA水平均降低(p<0.05, p<0.01, p<0.05),PGC-1αmRNA水平也降低(p<0.05);
3. (1)慢性缺氧心肌细胞培养液上清NO浓度明显高于对照组(p<0.01),iNOS和eNOS蛋白水平明显高于对照组(p<0.05, p<0.05);慢性缺氧心肌细胞线粒体荧光强度、mtDNA拷贝数、COXI mRNA水平均明显高于对照组(p<0.01, p<0.01, p<0.01),与线粒体生物合成相关的信号分子PGC-1α、NRF1和Tfam mRNA水平也高于对照组(p<0.01, p<0.05, p<0.01);
(2)与慢性缺氧组比较,慢性缺氧+ L-NAME组线粒体荧光强度、COXI mRNA和蛋白水平均降低(p<0.01, p<0.01, p<0.05),PGC-1α和Tfam mRNA水平也降低(p<0.05,p<0.01);
(3)与慢性缺氧组比较,慢性缺氧+ OxyHb组PGC-1αmRNA水平降低(p<0.05),其它指标无明显变化;
(4)与慢性缺氧组比较,慢性缺氧+ODQ组线粒体荧光强度、mtDNA拷贝数、COXI mRNA和蛋白水平均降低(p<0.01, p<0.05, p<0.01, p<0.05),PGC-1α和Tfam mRNA水平也降低(p<0.01, p<0.05)。
结论
本课题分别在先天性心脏病心肌组织标本和体外培养慢性缺氧心肌细胞模型上,对心肌细胞的线粒体生物合成和NO信号通路变化进行形态学和分子生物学研究,探讨NO在慢性缺氧状态下对心肌线粒体生物合成的调控作用机制。结果发现:
1.紫绀型先天性心脏病右室流出道心肌组织中线粒体数量增多, mtDNA拷贝数增多,线粒体编码的COXI蛋白mRNA和蛋白水平升高。说明紫绀型先天性心脏病心肌线粒体生物合成明显增多;同时,紫绀型先天性心脏病右室流出道心肌组织中eNOS蛋白水平升高,并且与线粒体生物合成呈显著的正相关,提示其参与了线粒体生物合成的调节;
2. NO供体在体外可以促进培养心肌细胞的线粒体生物合成,该过程可能通过sGC-cGMP途径介导;
3.慢性缺氧的培养心肌细胞中iNOS和eNOS蛋白表达升高,NO合成增多,同时线粒体的生物合成上调;NO可能通过NOS-NO-sGC-cGMP途径参与慢性缺氧心肌细胞线粒体生物合成的调节。
Background
Myocardial chronic hypoxia is a common clinical pathophysiological process and leads to a series of myocardial metabolic adaptive changes. Mitochondrial biogenesis which controls the quantity and quality of mitochondria may plays an important role in myocardial metabolic adaptation. However, the effect of chronic hypoxia in myocardial mitochondrial biogenesis has not been fully understood. Nitric oxide (NO) may be involved in the regulation of mitochondria biogenesis in a variety of mammalian cell lines, but its role in cardiac mitochondrial biogenesis during chronic hypoxia remains unknown. Research on the relationship between NO and myocardial mitochondrial biogenetic adaptation during chronic hypoxia may help to clarify the mechanism of chronic hypoxic adaptation and provide some new ideas to enhance myocardial protection in health and patients. To better clarify the regulatory mechanisms of NO pathway in cardiac mitochondrial biogenesis during chronic hypoxia, we performed a detailed morphologic and molecular analysis on heart tissue samples from patients with congenital heart disease and cultured cardiomyocytes exposed to chronic hypoxia.
Methods
In the first part of this study, samples from the right ventricular outflow tract myocardium were taken from patients with cyanotic(n=10) and acyanotic(n=10) congenital heart diseases. The ultrastructure of samples was analyzed by transmission electron microscope. MtDNA copy number, cytochrome c oxidase I (COX I), peroxisome proliferators activated receptorγcoactivator-1α(PGC-1α), nuclear respiratory factor 1 (NRF1), mitochondrial transcription factor A (Tfam) and three nitric oxide synthase(NOS) isoforms transcript and(or) protein levels were detected by real-time PCR , RT-PCR and Western blot respectively. The activity levels of NOS were also analyzed.
In the second part, cultured neonatal rat cardiomyocytes were treated with SNAP (NO donor), oxygenated hemoglobin (OxyHb, NO scavenger) or ODQ (soluble guanylate cyclase inhibitor). Mitochondrial specific fluorescence probe was used for detecting the mitochondrial mass, real-time PCR, RT-PCR and Western blot were used for detecting the expression of related gene and protein.
In the last part, neonatal rat cardiomyocytes were cultured in 1%O2 for 48h and treated with L-NAME(NOS inhibitor),OxyHb or ODQ. The NO production in cardiomyocytes, mitochondrial mass, related gene and protein expression were also detected.
Results
1. Mitochondrial volume density (Vv) and numerical density (Nv) were significantly elevated in patients with cyanotic compared to acyanotic congenital heart disease (p<0.05, p<0.01). Elevated mtDNA and up-regulated COXI, PGC-1α, NRF1 and Tfam mRNA levels were observed in cyanotic patients (p<0.01). Levels of COXI and eNOS protein were significantly higher in the myocardium of cyanotic patients than those of acyanotic (p<0.05, p<0.05). Levels of PGC-1αtranscript and eNOS protein inversely correlated with SaO2(r=-0.75, p<0.01 and r=-0.64, p<0.01, respectively). Levels of PGC-1αtranscript correlated with the levels of eNOS(r=0.58, p<0.01).
2. (1) In SNAP-treated cardiomyocytes, the mitochondria fluorescence intensity, copy number of mtDNA, levels of COXI mRNA and protein were significantly higher than those of control group (p <0.01, p <0.01, p <0.01, p <0.01). Mitochondrial biogenesis related signaling molecules PGC-1α, and NRF1 Tfam mRNA levels were also elevated (p <0.01, p <0.01, p <0.05).
(2) Comparing with the SNAP group, the mitochondria fluorescence intensity, copy number of mtDNA and COXI mRNA decreased in SNAP + OxyHb group (p <0.01, p <0.01, p <0.05), and levels of PGC-1αand NRF1 mRNA had also decreased (p <0.01, p <0.05).
(3) Comparing with the SNAP group, the mitochondria fluorescence intensity, mtDNA copy number and COXI mRNA levels decreased in the SNAP + ODQ group (p <0.05, p <0.01, p <0.05), and PGC-1αmRNA levels have also decreased (p < 0.05).
3. (1) Comparing with the control group, NO production, eNOS and iNOS protein levels were significantly higher in cardiomyocytes exposed to chronic hypoxia (p <0.01, p <0.05, p <0.05). The mitochondria fluorescence intensity, mtDNA copy number and COXI mRNA levels were significantly higher (p <0.01, p <0.01, p <0.01). Mitochondrial biogenesis related signaling molecules PGC-1α, and NRF1 Tfam mRNA levels were also elevated in cardiomyocytes exposed to chronic hypoxia (p <0.01, p <0.05, p <0.01).
(2) Comparing with the chronic hypoxia group, the mitochondria fluorescence intensity, COXI mRNA and protein levels decreased (p <0.01, p <0.01, p <0.05), and PGC-1αand Tfam mRNA levels have also decreased (p <0.05, p <0.01) in chronic hypoxia + L-NAME group.
(3) Comparing with the chronic hypoxia group, levels of PGC-1αmRNA decreased in chronic hypoxia + OxyHb group (p <0.05). No significant changes were observed in other parameters.
(4) Comparing with the chronic hypoxia group, the mitochondria fluorescence intensity, mtDNA copy number, COXI mRNA and protein levels decreased (p <0.01, p <0.05, p <0.01, p <0.05), levels of PGC-1αand Tfam mRNA were also decreased (p <0.01, p <0.05) in chronic hypoxia + ODQ group.
Conclusions
This study based on a detailed morphologic and molecular analysis on heart tissue samples from congenital heart disease patients and cultured cardiomyocytes exposed to chronic hypoxia. The results indicated that:
1. The mitochondrial mass, mtDNA copy number, mitochondrial encoded COXI mRNA and protein levels were increased in the right ventricular outflow tract myocardium of congenital heart disease with cyanosis, suggesting that mitochondrial biogenesis is activated in myocardium in congenital heart disease with cyanosis. Meanwhile, levels of eNOS protein were also elevated in cyanotic group, and were significantly positive correlated with mitochondrial biogenesis, which implied that eNOS may be involved in the regulation of mitochondrial biogenesis.
2. NO donor can promote the mitochondrial biogenesis in cultured cardiomyocytes, possibly mediated by sGC-cGMP pathway.
3. NO production, eNOS and iNOS protein expression as well as mitochondrial biogenesis were up-regulated in cardiomyocytes exposed to chronic hypoxia. NO may regulate mitochondrial biogenesis through NOS-NO-sGC-cGMP pathway in cardiomyocytes exposed to chronic hypoxia.
心肌慢性缺氧是临床常见的病理生理过程,可导致心肌产生一系列代谢适应性改变,与线粒体的数量和功能密切相关的线粒体生物合成过程的适应性改变可能是其中重要的环节,但慢性缺氧对心肌线粒体生物合成影响的机制目前尚不完全清楚。一氧化氮(NO)可能参与了多种细胞线粒体生物合成的调控,其在慢性缺氧心肌线粒体生物合成中的作用尚不明了。深入研究NO与慢性缺氧心肌线粒体的适应性改变和调控之间的关系,将有望阐明缺氧状态下心肌的病理生理改变机制,为进一步增强缺氧状态下的心肌保护,提高临床治疗效果提供新的思路。本课题分别在先天性心脏病心肌组织标本和体外培养慢性缺氧心肌细胞模型上,对心肌细胞的线粒体生物合成和NO信号通路变化进行形态学和分子生物学研究,探讨NO在慢性缺氧状态下对心肌线粒体生物合成的调控作用机制。研究方法
本研究共分为三部分:
第一部分:选取紫绀型和非紫绀型先天性心脏病患者各10例,留取手术中切除的肥厚右室流出道组织,通过电镜观察分析标本心肌细胞超微结构,运用实时荧光PCR、RT-PCR及Western blot等方法比较心肌组织mtDNA拷贝数,细胞色素c氧化酶I (COX I)、过氧化物酶体增生物激活受体γ辅激活因子-1α(PGC-1α)、核呼吸因子1(NRF1)、线粒体转录因子A(Tfam)和三型NOS的mRNA或(和)蛋白水平,检测NOS酶活性,并对PGC-1α、eNOS蛋白水平与患者末梢血氧饱和度SaO2进行相关性分析。
第二部分:体外培养乳鼠心肌细胞,根据不同干预因素将心肌细胞分为对照组,NO供体SNAP组,SNAP+NO清除剂氧合血红蛋白(OxyHb)组及SNAP+可溶性鸟苷酸环化酶(sGC)抑制剂(ODQ)组。应用线粒体荧光探针在激光共聚焦显微镜下观察各组细胞线粒体量的变化,应用实时荧光PCR、RT- PCR及Western blot等方法分别比较各组细胞mtDNA拷贝数,COX I、PGC-1α、NRF1和Tfam的mRNA水平以及COXI蛋白水平等。
第三部分:通过1%O2浓度培养48小时建立体外培养乳鼠心肌细胞慢性缺氧模型,根据不同干预方式将心肌细胞分为对照组,慢性缺氧组,慢性缺氧+NOS抑制剂L-NAME组,慢性缺氧+OxyHb组以及慢性缺氧+ODQ组。检测细胞培养液上清NO浓度和心肌细胞NOS蛋白水平,应用线粒体荧光探针观察各组细胞线粒体量的变化,应用实时荧光PCR、RT- PCR及Western blot等方法分别比较各组细胞mtDNA拷贝数,COX I、PGC-1α、NRF1和Tfam的mRNA水平以及COX I蛋白水平等。
研究结果
本研究主要结果如下:
1.紫绀型先天性心脏病右室流出道心肌组织中线粒体体密度Vv、数密度Nv明显高于非紫绀组(p<0.05, p<0.01),mtDNA拷贝数显著高于非紫绀组(p<0.01),COX I、PGC-1α、NRF1和Tfam的mRNA水平均显著高于非紫绀组(p<0.01),COX I、eNOS蛋白表达显著高于非紫绀组(p<0.05, p<0.05),心肌组织PGC-1αmRNA水平与SaO2呈显著的负相关(r=-0.75, p<0.01),eNOS蛋白水平与SaO2呈显著的负相关(r=-0.64, p<0.01),PGC-1αmRNA水平与eNOS蛋白水平呈显著的正相关(r=0.58, p<0.01);
2. (1) NO供体SNAP干预组心肌细胞线粒体荧光强度、mtDNA拷贝数、COXI mRNA和蛋白水平均明显高于对照组(p<0.01, p<0.01, p<0.01, p<0.01),与线粒体生物合成相关的信号分子PGC-1α、NRF1和Tfam mRNA水平也高于对照组(p<0.01, p<0.01, p<0.05);
(2)与SNAP组比较,SNAP+OxyHb组线粒体荧光强度、mtDNA拷贝数、COXI mRNA均降低(p<0.01, p<0.01, p<0.05),PGC-1α和NRF1 mRNA水平也降低(p<0.01, p<0.05);
(3)与SNAP组比较,SNAP+ODQ组线粒体荧光强度、mtDNA拷贝数和COXI mRNA水平均降低(p<0.05, p<0.01, p<0.05),PGC-1αmRNA水平也降低(p<0.05);
3. (1)慢性缺氧心肌细胞培养液上清NO浓度明显高于对照组(p<0.01),iNOS和eNOS蛋白水平明显高于对照组(p<0.05, p<0.05);慢性缺氧心肌细胞线粒体荧光强度、mtDNA拷贝数、COXI mRNA水平均明显高于对照组(p<0.01, p<0.01, p<0.01),与线粒体生物合成相关的信号分子PGC-1α、NRF1和Tfam mRNA水平也高于对照组(p<0.01, p<0.05, p<0.01);
(2)与慢性缺氧组比较,慢性缺氧+ L-NAME组线粒体荧光强度、COXI mRNA和蛋白水平均降低(p<0.01, p<0.01, p<0.05),PGC-1α和Tfam mRNA水平也降低(p<0.05,p<0.01);
(3)与慢性缺氧组比较,慢性缺氧+ OxyHb组PGC-1αmRNA水平降低(p<0.05),其它指标无明显变化;
(4)与慢性缺氧组比较,慢性缺氧+ODQ组线粒体荧光强度、mtDNA拷贝数、COXI mRNA和蛋白水平均降低(p<0.01, p<0.05, p<0.01, p<0.05),PGC-1α和Tfam mRNA水平也降低(p<0.01, p<0.05)。
结论
本课题分别在先天性心脏病心肌组织标本和体外培养慢性缺氧心肌细胞模型上,对心肌细胞的线粒体生物合成和NO信号通路变化进行形态学和分子生物学研究,探讨NO在慢性缺氧状态下对心肌线粒体生物合成的调控作用机制。结果发现:
1.紫绀型先天性心脏病右室流出道心肌组织中线粒体数量增多, mtDNA拷贝数增多,线粒体编码的COXI蛋白mRNA和蛋白水平升高。说明紫绀型先天性心脏病心肌线粒体生物合成明显增多;同时,紫绀型先天性心脏病右室流出道心肌组织中eNOS蛋白水平升高,并且与线粒体生物合成呈显著的正相关,提示其参与了线粒体生物合成的调节;
2. NO供体在体外可以促进培养心肌细胞的线粒体生物合成,该过程可能通过sGC-cGMP途径介导;
3.慢性缺氧的培养心肌细胞中iNOS和eNOS蛋白表达升高,NO合成增多,同时线粒体的生物合成上调;NO可能通过NOS-NO-sGC-cGMP途径参与慢性缺氧心肌细胞线粒体生物合成的调节。
Background
Myocardial chronic hypoxia is a common clinical pathophysiological process and leads to a series of myocardial metabolic adaptive changes. Mitochondrial biogenesis which controls the quantity and quality of mitochondria may plays an important role in myocardial metabolic adaptation. However, the effect of chronic hypoxia in myocardial mitochondrial biogenesis has not been fully understood. Nitric oxide (NO) may be involved in the regulation of mitochondria biogenesis in a variety of mammalian cell lines, but its role in cardiac mitochondrial biogenesis during chronic hypoxia remains unknown. Research on the relationship between NO and myocardial mitochondrial biogenetic adaptation during chronic hypoxia may help to clarify the mechanism of chronic hypoxic adaptation and provide some new ideas to enhance myocardial protection in health and patients. To better clarify the regulatory mechanisms of NO pathway in cardiac mitochondrial biogenesis during chronic hypoxia, we performed a detailed morphologic and molecular analysis on heart tissue samples from patients with congenital heart disease and cultured cardiomyocytes exposed to chronic hypoxia.
Methods
In the first part of this study, samples from the right ventricular outflow tract myocardium were taken from patients with cyanotic(n=10) and acyanotic(n=10) congenital heart diseases. The ultrastructure of samples was analyzed by transmission electron microscope. MtDNA copy number, cytochrome c oxidase I (COX I), peroxisome proliferators activated receptorγcoactivator-1α(PGC-1α), nuclear respiratory factor 1 (NRF1), mitochondrial transcription factor A (Tfam) and three nitric oxide synthase(NOS) isoforms transcript and(or) protein levels were detected by real-time PCR , RT-PCR and Western blot respectively. The activity levels of NOS were also analyzed.
In the second part, cultured neonatal rat cardiomyocytes were treated with SNAP (NO donor), oxygenated hemoglobin (OxyHb, NO scavenger) or ODQ (soluble guanylate cyclase inhibitor). Mitochondrial specific fluorescence probe was used for detecting the mitochondrial mass, real-time PCR, RT-PCR and Western blot were used for detecting the expression of related gene and protein.
In the last part, neonatal rat cardiomyocytes were cultured in 1%O2 for 48h and treated with L-NAME(NOS inhibitor),OxyHb or ODQ. The NO production in cardiomyocytes, mitochondrial mass, related gene and protein expression were also detected.
Results
1. Mitochondrial volume density (Vv) and numerical density (Nv) were significantly elevated in patients with cyanotic compared to acyanotic congenital heart disease (p<0.05, p<0.01). Elevated mtDNA and up-regulated COXI, PGC-1α, NRF1 and Tfam mRNA levels were observed in cyanotic patients (p<0.01). Levels of COXI and eNOS protein were significantly higher in the myocardium of cyanotic patients than those of acyanotic (p<0.05, p<0.05). Levels of PGC-1αtranscript and eNOS protein inversely correlated with SaO2(r=-0.75, p<0.01 and r=-0.64, p<0.01, respectively). Levels of PGC-1αtranscript correlated with the levels of eNOS(r=0.58, p<0.01).
2. (1) In SNAP-treated cardiomyocytes, the mitochondria fluorescence intensity, copy number of mtDNA, levels of COXI mRNA and protein were significantly higher than those of control group (p <0.01, p <0.01, p <0.01, p <0.01). Mitochondrial biogenesis related signaling molecules PGC-1α, and NRF1 Tfam mRNA levels were also elevated (p <0.01, p <0.01, p <0.05).
(2) Comparing with the SNAP group, the mitochondria fluorescence intensity, copy number of mtDNA and COXI mRNA decreased in SNAP + OxyHb group (p <0.01, p <0.01, p <0.05), and levels of PGC-1αand NRF1 mRNA had also decreased (p <0.01, p <0.05).
(3) Comparing with the SNAP group, the mitochondria fluorescence intensity, mtDNA copy number and COXI mRNA levels decreased in the SNAP + ODQ group (p <0.05, p <0.01, p <0.05), and PGC-1αmRNA levels have also decreased (p < 0.05).
3. (1) Comparing with the control group, NO production, eNOS and iNOS protein levels were significantly higher in cardiomyocytes exposed to chronic hypoxia (p <0.01, p <0.05, p <0.05). The mitochondria fluorescence intensity, mtDNA copy number and COXI mRNA levels were significantly higher (p <0.01, p <0.01, p <0.01). Mitochondrial biogenesis related signaling molecules PGC-1α, and NRF1 Tfam mRNA levels were also elevated in cardiomyocytes exposed to chronic hypoxia (p <0.01, p <0.05, p <0.01).
(2) Comparing with the chronic hypoxia group, the mitochondria fluorescence intensity, COXI mRNA and protein levels decreased (p <0.01, p <0.01, p <0.05), and PGC-1αand Tfam mRNA levels have also decreased (p <0.05, p <0.01) in chronic hypoxia + L-NAME group.
(3) Comparing with the chronic hypoxia group, levels of PGC-1αmRNA decreased in chronic hypoxia + OxyHb group (p <0.05). No significant changes were observed in other parameters.
(4) Comparing with the chronic hypoxia group, the mitochondria fluorescence intensity, mtDNA copy number, COXI mRNA and protein levels decreased (p <0.01, p <0.05, p <0.01, p <0.05), levels of PGC-1αand Tfam mRNA were also decreased (p <0.01, p <0.05) in chronic hypoxia + ODQ group.
Conclusions
This study based on a detailed morphologic and molecular analysis on heart tissue samples from congenital heart disease patients and cultured cardiomyocytes exposed to chronic hypoxia. The results indicated that:
1. The mitochondrial mass, mtDNA copy number, mitochondrial encoded COXI mRNA and protein levels were increased in the right ventricular outflow tract myocardium of congenital heart disease with cyanosis, suggesting that mitochondrial biogenesis is activated in myocardium in congenital heart disease with cyanosis. Meanwhile, levels of eNOS protein were also elevated in cyanotic group, and were significantly positive correlated with mitochondrial biogenesis, which implied that eNOS may be involved in the regulation of mitochondrial biogenesis.
2. NO donor can promote the mitochondrial biogenesis in cultured cardiomyocytes, possibly mediated by sGC-cGMP pathway.
3. NO production, eNOS and iNOS protein expression as well as mitochondrial biogenesis were up-regulated in cardiomyocytes exposed to chronic hypoxia. NO may regulate mitochondrial biogenesis through NOS-NO-sGC-cGMP pathway in cardiomyocytes exposed to chronic hypoxia.
引文
1. Kolar F., Ostadal B. Molecular mechanisms of cardiac protection by adaptation to chronic hypoxia [J]. Physiol Res, 2004, 53 Suppl 1: S3-13.
2. Fitzpatrick C. M., Shi Y., Hutchins W. C., et al. Cardioprotection in chronically hypoxic rabbits persists on exposure to normoxia: role of NOS and KATP channels [J]. Am J Physiol Heart Circ Physiol, 2005, 288(1): H62-68.
3. Lynn E. G., Lu Z., Minerbi D., et al. The regulation, control, and consequences of mitochondrial oxygen utilization and disposition in the heart and skeletal muscle during hypoxia [J]. Antioxid Redox Signal, 2007, 9(9): 1353-1361.
4. McLeod C. J., Pagel I., Sack M. N. The mitochondrial biogenesis regulatory program in cardiac adaptation to ischemia--a putative target for therapeutic intervention [J]. Trends Cardiovasc Med, 2005, 15(3): 118-123.
5. Massion P. B., Pelat M., Belge C., et al. Regulation of the mammalian heart function by nitric oxide [J]. Comp Biochem Physiol A Mol Integr Physiol, 2005, 142(2): 144-150.
6. Lacza Z., Pankotai E., Csordas A., et al. Mitochondrial NO and reactive nitrogen species production: does mtNOS exist? [J]. Nitric Oxide, 2006, 14(2): 162-168.
7. Clementi E., Nisoli E. Nitric oxide and mitochondrial biogenesis: a key to long-term regulation of cellular metabolism [J]. Comp Biochem Physiol A Mol Integr Physiol, 2005, 142(2): 102-110.
8. Burley D. S., Ferdinandy P., Baxter G. F. Cyclic GMP and protein kinase-G in myocardial ischaemia-reperfusion: opportunities and obstacles for survival signaling [J]. Br J Pharmacol, 2007, 152(6): 855-869.
9. Saraiva R. M., Hare J. M. Nitric oxide signaling in the cardiovascular system: implications for heart failure [J]. Curr Opin Cardiol, 2006, 21(3): 221-228.
10. Wang S., Yan L., Wesley R. A., et al. Nitric oxide increases tumor necrosis factor production in differentiated U937 cells by decreasing cyclic AMP [J]. J Biol Chem, 1997, 272(9): 5959-5965.
11. Ma P., Cui X., Wang S., et al. Nitric oxide post-transcriptionally up-regulates LPS-induced IL-8 expression through p38 MAPK activation [J]. J Leukoc Biol, 2004, 76(1): 278-287.
12. Zhang J., Wang S., Wesley R. A., et al. Adjacent sequence controls the response polarity of nitric oxide-sensitive Sp factor binding sites [J]. J Biol Chem, 2003, 278(31): 29192-29200.
13. Johnson D. T., Harris R. A., French S., et al. Tissue heterogeneity of the mammalian mitochondrial proteome [J]. Am J Physiol Cell Physiol, 2007, 292(2): C689-697.
14. Weibel E. R., Staubli W., Gnagi H. R., et al. Correlated morphometric and biochemical studies on the liver cell. I. Morphometric model, stereologic methods, and normal morphometric data for rat liver [J]. J Cell Biol, 1969, 42(1): 68-91.
15. Nouette-Gaulain K., Malgat M., Rocher C., et al. Time course of differential mitochondrial energy metabolism adaptation to chronic hypoxia in right and left ventricles [J]. Cardiovasc Res, 2005, 66(1): 132-140.
16. Cervos Navarro J., Kunas R. C., Sampaolo S., et al. Heart mitochondria in rats submitted to chronic hypoxia [J]. Histol Histopathol, 1999, 14(4): 1045-1052.
17. Trinei M., Berniakovich I., Pelicci P. G., et al. Mitochondrial DNA copy number is regulated by cellular proliferation: a role for Ras and p66(Shc) [J]. Biochim Biophys Acta, 2006, 1757(5-6): 624-630.
18. Cai J. H., Deng S., Kumpf S. W., et al. Validation of rat reference genes for improved quantitative gene expression analysis using low density arrays [J]. Biotechniques, 2007, 42(4): 503-512.
19. Okamoto K., Shaw J. M. Mitochondrial morphology and dynamics in yeast and multicellular eukaryotes [J]. Annu Rev Genet, 2005, 39: 503-536.
20. Meeusen S., McCaffery J. M., Nunnari J. Mitochondrial fusion intermediates revealed in vitro [J]. Science, 2004, 305(5691): 1747-1752.
21. Collins T. J., Berridge M. J., Lipp P., et al. Mitochondria are morphologically and functionally heterogeneous within cells [J]. Embo J, 2002, 21(7): 1616-1627.
22. Rube D. A., van der Bliek A. M. Mitochondrial morphology is dynamic and varied [J]. Mol Cell Biochem, 2004, 256-257(1-2): 331-339.
23. Chowdhury U. K., Sathia S., Ray R., et al. Histopathology of the right ventricular outflow tract and its relationship to clinical outcomes and arrhythmias in patients with tetralogy of Fallot [J]. J Thorac Cardiovasc Surg, 2006, 132(2): 270-277.
24.孙桂媛石玉秀,祝淑文等.法乐四联症心肌纤维扫描电镜的观察[J].中华胸心外科杂志, 1998, 14(3): 139-141.
25.李刚,王湘,季军等.法乐四联症右室流出道心肌超微结构特点及其临床意义[J].中华实用外科杂志, 2006, 23(9): 1036-1037.
26.李晓峰李仲智,郎志奇等.法洛四联症右室流出道心肌超微结构特点[J].实用儿科临床杂志, 2004, 19(7): 554-557.
27. Anderson S., Bankier A. T., Barrell B. G., et al. Sequence and organization of the human mitochondrial genome [J]. Nature, 1981, 290(5806): 457-465.
28. Garesse R., Vallejo C. G. Animal mitochondrial biogenesis and function: a regulatory cross-talk between two genomes [J]. Gene, 2001, 263(1-2): 1-16.
29. Stiburek L., Hansikova H., Tesarova M., et al. Biogenesis of eukaryotic cytochrome c oxidase [J]. Physiol Res, 2006, 55 Suppl 2: S27-41.
30. Fontanesi F., Soto I. C., Horn D., et al. Assembly of mitochondrial cytochrome c-oxidase, a complicated and highly regulated cellular process [J]. Am J Physiol Cell Physiol, 2006, 291(6): C1129-1147.
31. Puigserver P., Wu Z., Park C. W., et al. A cold-inducible coactivator of nuclear receptors linked to adaptive thermogenesis [J]. Cell, 1998, 92(6): 829-839.
32. Kelly D. P., Scarpulla R. C. Transcriptional regulatory circuits controlling mitochondrial biogenesis and function [J]. Genes Dev, 2004, 18(4): 357-368.
33. Lehman J. J., Barger P. M., Kovacs A., et al. Peroxisome proliferator-activated receptor gamma coactivator-1 promotes cardiac mitochondrial biogenesis [J]. J Clin Invest, 2000, 106(7): 847-856.
34. Wu H., Kanatous S. B., Thurmond F. A., et al. Regulation of mitochondrial biogenesis in skeletal muscle by CaMK [J]. Science, 2002, 296(5566): 349-352.
35. Mootha V. K., Lindgren C. M., Eriksson K. F., et al. PGC-1alpha-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes [J]. Nat Genet, 2003, 34(3): 267-273.
36. Scarpulla R. C. Nuclear control of respiratory gene expression in mammalian cells [J]. J Cell Biochem, 2006, 97(4): 673-683.
37. Irrcher I., Adhihetty P. J., Joseph A. M., et al. Regulation of mitochondrial biogenesis in muscle by endurance exercise [J]. Sports Med, 2003, 33(11): 783-793.
38. Irrcher I., Adhihetty P. J., Sheehan T., et al. PPARgamma coactivator-1alpha expressionduring thyroid hormone- and contractile activity-induced mitochondrial adaptations [J]. Am J Physiol Cell Physiol, 2003, 284(6): C1669-1677.
39. Suliman H. B., Carraway M. S., Welty-Wolf K. E., et al. Lipopolysaccharide stimulates mitochondrial biogenesis via activation of nuclear respiratory factor-1 [J]. J Biol Chem, 2003, 278(42): 41510-41518.
40. Weitzel J. M., Iwen K. A., Seitz H. J. Regulation of mitochondrial biogenesis by thyroid hormone [J]. Exp Physiol, 2003, 88(1): 121-128.
41. Kang D., Hamasaki N. Mitochondrial transcription factor A in the maintenance of mitochondrial DNA: overview of its multiple roles [J]. Ann N Y Acad Sci, 2005, 1042: 101-108.
42. Kang D., Kim S. H., Hamasaki N. Mitochondrial transcription factor A (TFAM): roles in maintenance of mtDNA and cellular functions [J]. Mitochondrion, 2007, 7(1-2): 39-44.
43. May-Panloup P., Vignon X., Chretien M. F., et al. Increase of mitochondrial DNA content and transcripts in early bovine embryogenesis associated with upregulation of mtTFA and NRF1 transcription factors [J]. Reprod Biol Endocrinol, 2005, 3: 65.
44. Corno A. F., Milano G., Samaja M., et al. Chronic hypoxia: a model for cyanotic congenital heart defects [J]. J Thorac Cardiovasc Surg, 2002, 124(1): 105-112.
45. Lahiri S., Roy A., Baby S. M., et al. Oxygen sensing in the body [J]. Prog Biophys Mol Biol, 2006, 91(3): 249-286.
46.任晓莉李瑞兰.动态监测血乳酸对先天性心脏病心内直视手术患儿的临床意义[J].心肺血管病杂志, 2007, 26(3): 174-175.
47. Marin-Garcia J., Ananthakrishnan R., Agrawal N., et al. Mitochondrial gene expression during bovine cardiac growth and development [J]. J Mol Cell Cardiol, 1994, 26(8): 1029-1036.
48. Costa L. E., Boveris A., Koch O. R., et al. Liver and heart mitochondria in rats submitted to chronic hypobaric hypoxia [J]. Am J Physiol, 1988, 255(1 Pt 1): C123-129.
49. Piel D. A., Khan A. R., Waibel R., et al. Chronic hypoxemia increases myocardial cytochrome oxidase [J]. J Thorac Cardiovasc Surg, 2005, 130(4): 1101-1106.
50. Rasbach K. A., Schnellmann R. G. PGC-1alpha over-expression promotes recoveryfrom mitochondrial dysfunction and cell injury [J]. Biochem Biophys Res Commun, 2007, 355(3): 734-739.
51. Shinde S. B., Save V. C., Patil N. D., et al. Impairment of mitochondrial respiratory chain enzyme activities in tetralogy of Fallot [J]. Clin Chim Acta, 2007, 377(1-2): 138-143.
52. MacMicking J., Xie Q. W., Nathan C. Nitric oxide and macrophage function [J]. Annu Rev Immunol, 1997, 15: 323-350.
53. Massion P. B., Feron O., Dessy C., et al. Nitric oxide and cardiac function: ten years after, and continuing [J]. Circ Res, 2003, 93(5): 388-398.
54. Alderton W. K., Cooper C. E., Knowles R. G. Nitric oxide synthases: structure, function and inhibition [J]. Biochem J, 2001, 357(Pt 3): 593-615.
55. Elfering S. L., Sarkela T. M., Giulivi C. Biochemistry of mitochondrial nitric-oxide synthase [J]. J Biol Chem, 2002, 277(41): 38079-38086.
56. Massion P. B., Dessy C., Desjardins F., et al. Cardiomyocyte-restricted overexpression of endothelial nitric oxide synthase (NOS3) attenuates beta-adrenergic stimulation and reinforces vagal inhibition of cardiac contraction [J]. Circulation, 2004, 110(17): 2666-2672.
57. Valdez L. B., Zaobornyj T., Alvarez S., et al. Heart mitochondrial nitric oxide synthase. Effects of hypoxia and aging [J]. Mol Aspects Med, 2004, 25(1-2): 49-59.
58. Seddon M., Shah A. M., Casadei B. Cardiomyocytes as effectors of nitric oxide signalling [J]. Cardiovasc Res, 2007, 75(2): 315-326.
59. Davidson S. M., Duchen M. R. Effects of NO on mitochondrial function in cardiomyocytes: Pathophysiological relevance [J]. Cardiovasc Res, 2006, 71(1): 10-21.
60. Jung F., Palmer L. A., Zhou N., et al. Hypoxic regulation of inducible nitric oxide synthase via hypoxia inducible factor-1 in cardiac myocytes [J]. Circ Res, 2000, 86(3): 319-325.
61. Coulet F., Nadaud S., Agrapart M., et al. Identification of hypoxia-response element in the human endothelial nitric-oxide synthase gene promoter [J]. J Biol Chem, 2003, 278(47): 46230-46240.
62. Forkel J., Chen X., Wandinger S., et al. Responses of chronically hypoxic rat hearts to ischemia: KATP channel blockade does not abolish increased RV tolerance to ischemia[J]. Am J Physiol Heart Circ Physiol, 2004, 286(2): H545-551.
63. Grilli A., De Lutiis M. A., Patruno A., et al. Effect of chronic hypoxia on inducible nitric oxide synthase expression in rat myocardial tissue [J]. Exp Biol Med (Maywood), 2003, 228(8): 935-942.
64. Thompson L. P., Dong Y. Chronic hypoxia decreases endothelial nitric oxide synthase protein expression in fetal guinea pig hearts [J]. J Soc Gynecol Investig, 2005, 12(6): 388-395.
65. Zaobornyj T., Gonzales G. F., Valdez L. B. Mitochondrial contribution to the molecular mechanism of heart acclimatization to chronic hypoxia: role of nitric oxide [J]. Front Biosci, 2007, 12: 1247-1259.
66. Ghafourifar P., Sen C. K. Mitochondrial nitric oxide synthase [J]. Front Biosci, 2007, 12: 1072-1078.
67. Poot M., Zhang Y. Z., Kramer J. A., et al. Analysis of mitochondrial morphology and function with novel fixable fluorescent stains [J]. J Histochem Cytochem, 1996, 44(12): 1363-1372.
68. Livak K. J., Schmittgen T. D. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method [J]. Methods, 2001, 25(4): 402-408.
69. Frazier A. E., Kiu C., Stojanovski D., et al. Mitochondrial morphology and distribution in mammalian cells [J]. Biol Chem, 2006, 387(12): 1551-1558.
70. Zong H., Ren J. M., Young L. H., et al. AMP kinase is required for mitochondrial biogenesis in skeletal muscle in response to chronic energy deprivation [J]. Proc Natl Acad Sci U S A, 2002, 99(25): 15983-15987.
71. Leary S. C., Shoubridge E. A. Mitochondrial biogenesis: which part of "NO" do we understand? [J]. Bioessays, 2003, 25(6): 538-541.
72. Cerra M. C., Pellegrino D. Cardiovascular cGMP-generating systems in physiological and pathological conditions [J]. Curr Med Chem, 2007, 14(5): 585-599.
73. Bellamy T. C., Wood J., Goodwin D. A., et al. Rapid desensitization of the nitric oxide receptor, soluble guanylyl cyclase, underlies diversity of cellular cGMP responses [J]. Proc Natl Acad Sci U S A, 2000, 97(6): 2928-2933.
74. Navarro A., Torrejon R., Bandez M. J., et al. Mitochondrial function andmitochondria-induced apoptosis in an overstimulated rat ovarian cycle [J]. Am J Physiol Endocrinol Metab, 2005, 289(6): E1101-1109.
75. Nagy G., Barcza M., Gonchoroff N., et al. Nitric oxide-dependent mitochondrial biogenesis generates Ca2+ signaling profile of lupus T cells [J]. J Immunol, 2004, 173(6): 3676-3683.
76. Baris O., Savagner F., Nasser V., et al. Transcriptional profiling reveals coordinated up-regulation of oxidative metabolism genes in thyroid oncocytic tumors [J]. J Clin Endocrinol Metab, 2004, 89(2): 994-1005.
77. Carew J. S., Nawrocki S. T., Xu R. H., et al. Increased mitochondrial biogenesis in primary leukemia cells: the role of endogenous nitric oxide and impact on sensitivity to fludarabine [J]. Leukemia, 2004, 18(12): 1934-1940.
78.艾珣,顾玉东.培养人脐静脉内皮细胞慢性缺血缺氧模型[J].中华手外科杂志, 2001, 17(4): 233-236.
79.孙胜,高文祥,廖卫公.神经细胞化学缺氧预处理模型的建立[J].第三军医大学学报, 2007, 29(21): 2100-2101.
80. Roy S., Khanna S., Bickerstaff A. A., et al. Oxygen sensing by primary cardiac fibroblasts: a key role of p21(Waf1/Cip1/Sdi1) [J]. Circ Res, 2003, 92(3): 264-271.
81. Li H. T., Long C. S., Rokosh D. G., et al. Chronic hypoxia differentially regulates alpha 1-adrenergic receptor subtype mRNAs and inhibits alpha 1-adrenergic receptor-stimulated cardiac hypertrophy and signaling [J]. Circulation, 1995, 92(4): 918-925.
82. Kacimi R., Long C. S., Karliner J. S. Chronic hypoxia modulates the interleukin-1beta-stimulated inducible nitric oxide synthase pathway in cardiac myocytes [J]. Circulation, 1997, 96(6): 1937-1943.
83. Newby D., Marks L., Lyall F. Dissolved oxygen concentration in culture medium: assumptions and pitfalls [J]. Placenta, 2005, 26(4): 353-357.
84. Qing M., Gorlach A., Schumacher K., et al. The hypoxia-inducible factor HIF-1 promotes intramyocardial expression of VEGF in infants with congenital cardiac defects [J]. Basic Res Cardiol, 2007, 102(3): 224-232.
1. Lee S, Kim S, Sun X, et al. Cell cycle-dependent mitochondrial biogenesis and dynamics in mammalian cells. Biochem Biophys Res Commun. 2007,25; 357(1):111-117.
2. WeberK, Bruck P, Mikes Z, et al. Glucocorticoid hormone stimulates mitochondrial biogenesis specifically in skeletal muscle. Endocrinology . 2002, 143(1): 177-84.
3. Nisoli E, Carruba MO. Nitric oxide and mitochondrial biogenesis. 2006. J Cell Sci. 119:2855-62.
4. Yaffe, M P.. The cutting edge of mitochondrial fusion. Nat. Cell Biol. 2003,5:497-499.
5. Meeusen S, McCaffery J M , Nunnari J.. Mitochondrial fusion intermediates revealed in vitro. Science, 2004, 305:1747-1752.
6. Okamoto K, Shaw J M. Mitochondrial morphology and dynamics in yeast and multicellular eukaryotes. Annu. Rev. Genet. 2005, 39: 503-536.
7. Rube DA, vander Bliek, AM. Mitochondrial motphology is dynamic and varied. Mol. Cell. Biochem. 2004, 256/257:331-339.
8. Civitarese AE, Carling S, Heilbronn LK,et al.Calorie Restriction Increases Muscle Mitochondrial Biogenesis in Healthy Humans. PLoS Med. 2007, 6;4(3):e76
9. Chabi B, Adhihetty PJ, Ljubucuc V, et al. How is Mitochondrial Biogenesis Affected in Mitochondrial Disease? Med. Sci. Sports Exerc, 2005, 37(12): 2102-2110.
10. Carew JS, Nawrocki ST, Xu RH, et al. Increased mitochondrial biogenesis in primary leukemia cells: the role of endogenous nitric oxide and impact on sensitivity to fludarabine. Leukemia. 2004 , 18(12): 1934-40.
11. Kamal MA, French SW. Drug-induced increased mitochondrial biogenesis in a liver biopsy. xp-Mol-Pathol. 2004 ,77(3): 201-4.
12. Mootha VK, Lindgren CM, Eriksson KF, et al. PGC-1alpha-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat.Genet.2003, 34:267-73
13. Ukropcova B, Sereda O, de Jonge L,et al. Family history of diabetes links impaired substrate switching and reduced mitochondrial content in skeletal muscle. Diabetes. 2007, 56(3):720-7
14. M. Lemoine, M. Kim, V. Ratziu,et al. Liver mitochondrial function and biogenesis in patients with NAFLD: hepatic expression of PGC-1alpha, COX2 and Cox4. Journal of Hepatology, 2007, 46(1):S272-S273.
15. Nisoli E, Clementi E, Carruba MO,et al. Defective mitochondrial biogenesis: a hallmark of the high cardiovascular risk in the metabolic syndrome? Circ Res. 2007 , 100(6):795-806.
16. Libin Cui, Hyunkyung Jeong, Fran Borovecki, et al. Transcriptional Repression of PGC-1αby Mutant Huntingtin Leads to Mitochondrial Dysfunction and Neurodegeneration. Cell, 2006, 127(1):59-69.
17. Gomez C, Bandez MJ, Navarro A. Pesticides and impairment of mitochondrial function in relation with the parkinsonian syndrome. Front Biosci. 2007,12:1079-93.
18. Sardiello M, Tripoli G, Romito A, et al. Energy biogenesis: one key for coordinating two genomes. Trends Genet. 2005,21:12-16.
19. Butow RA, Avadhani NG. Mitochondrial signaling: the retrograde response. Mol. Cell. 2004, 14:1-15.
20. Kelly DP, Scarpulla RC. Transcriptional regulatory circuits controlling mitochondrial biogenesis and function. Genes Dev. 2004, 18:357-68.
21. Garesse R, Vallejo CG. Animal mitochondrial biogenesis and function: a regulatory cross-talk between two genomes. Gene. 2001, 263:1-16.
22. Wickner W, Schekman R. Protein translocation across biological membranes. Science. 2005, 310:1452-1456.
23. Lin J, Handschin C, Spiegelman BM. Metabolic control through the PGC-1 family of transcription coactivators. Cell Metab.2005, 1:361-70
24. Puigserver P, Spiegelman BM. Peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PGC-1 alpha): transcriptional coactivator and metabolic regulator. Endocr Rev. 2003,24:78-90.
25. Puigserver P,Wu Z, Park CW, et al. A cold-inducible coactivator of nuclear receptors linked to adaptive thermogenesis. Cell. 1998,92:829-39.
26. Finck B N, Kelly D P. PGC-1 coactivators: inducible regulators of energy metabolism in health and disease. J of Clin Invest. 2006, 116, 615-622.
27. Puigserver P, Adelmant G,Wu Z, et al. Activation of PPARgamma coactivator-1through transcription factor docking. Science .1999, 286:1368-71.
28. Monsalve M,Wu Z, Adelmant G, et al. Direct coupling of transcription and mRNA processing through the thermogenic coactivator PGC-1. Mol Cell. 2000,6:307-16.
29. Knutti D , Kressler D , Kralli A. Regulation of the transcriptional coactivator PGC21 via MAPK2sensitive interaction with a repressor. Proc Natl Acad Sci USA. 2001 , 98 (17) :9713-9718.
30. Wu Z, Puigserver P, Andersson U, et al. Mechanisms controlling mitochondrial biogenesis and respiration through the thermogenic coactivator PGC-1. Cell .1999, 98:115-24.
31. Mootha VK, Lindgren CM, Eriksson KF, et al. Nat.Genet. 2003, 34:267-73.
32. Vega RB, Huss JM, Kelly DP. The coactivator PGC-1 cooperates with peroxisome proliferator-activated receptor alpha in transcriptional control of nuclear genes encoding mitochondrial fatty acid oxidation enzymes. Mol Cell Biol. 2000,20:1868-76.
33. Cartoni R, Leger B, Hock MB, et al. Mitofusins 1/2 and ERRalpha expression are increased in human skeletal muscle after physical exercise. J Physiol. 2005,567:349-58.
34. Soriano FX, Liesa M, Bach D, et al. Evidence for a Mitochondrial Regulatory Pathway Defined by Peroxisome Proliferator-Activated Receptor-gamma Coactivator-1(alpha), Estrogen-Related Receptor-alpha, and Mitofusin 2. Diabetes. 2006, 55:1783-91.
35. Zong H, Ren JM, Young LH, et al. AMP kinase is required for mitochondrial biogenesis in skeletal muscle in response to chronic energy deprivation. Proc Natl Acad Sci USA. 2002, 99:15983-87.
36. Kukidome D, Nishikawa T, Sonoda K, et al. Activation of AMP-activated protein kinase reduces hyperglycemia-induced mitochondrial reactive oxygen species production and promotes mitochondrial biogenesis in human umbilical vein endothelial cells. Diabetes . 2006,55:120-27.
37. Wu H, Kanatous S B, Thurmond F A, et al. Regulation of mitochondrial biogenesis in skeletal muscle by CaMK. Science. 2002,296: 349-352.
38. Lin J, Wu H, Tarr PT, et al. Transcriptional co-activator PGC-1_ drives the formation of slow-twitch muscle fibres. Nature .2002b.418: 797-801.
39. Lee J, Kim CH, Simon DK, et al. Mitochondrial cyclic AMP response element-binding protein (CREB) mediates mitochondrial gene expression and neuronal survival. J BiolChem. 2005,280:40398-401.
40. Nisoli E, Falcone S, Tonello C. Mitochondrial biogenesis by NO yields functionally active mitochondria in mammals.Proc Natl Acad Sci USA. 2004, 23;101(47):16507-12.
41. Nisoli E, Clementi E, Paolucci C, et al. Mitochondrial biogenesis in mammals: the role of endogenous nitric oxide. Science. 2003,299:896-99.
42. Suliman HB, Carraway MS, Tatro LG, et al. A new activating role for CO in cardiac mitochondrial biogenesis. J Cell Sci. 2007,120(Pt 2):299-308.
43. Puigserver P , Adelmant G, Wu Z, et al. Activation of PPARgamma coactivator21 through transcription factor docking. Science . 1999 , 286(5443) :1368-1371.
44. Puigserver P, Rhee J, Lin J, et al. Cytokine stimulation of energy expenditure through p38 MAP kinase activation of PPARgamma coactivator-1. Mol Cell. 2001, 8:971-82.
45. Fan M, Rhee J , StPierre J , et al. Suppression of mitochondrial respiration through recruitment of p160 myb binding protein to PGC21alpha : Modulation by p38 MAPK. Genes Dev . 2004 , 18 (3):278-289.
1. Salzman AL , Menconi MJ , Unno N , et al. Nitric oxide dilates tight junction and depletes ATP in cultured Caco-2BBe intestinal epthelial monolayers. Am J Physio.1995 ,268 :361-373.
2.杨贵远,赵克森,刘杰.一氧化氮降低失血性休克血管反应性的机制.中国病理生理杂志.2000,16:1163-1166.
3. Hierholzer C , Billiar TR. Molecular mechanisms in t he early phase of hemorrhagic shock. Langenbecks Arch Surg , 2001 ,386 :302-308.
4. Yang J, Wu LJ, Tashiro S, et al .Nitric oxide activated by p38 and NF-kappaB facilitates apoptosis and cell cycle arrest under oxidative stress in evodiamine-treated human melanoma A375-S2 cells. Free Radic Res. 2008 Jan;42[1]:1-11.
5. Aquilano K, Filomeni G, Di Renzo L, et al .Reactive oxygen and nitrogen species are involved in sorbitol-induced apoptosis of human erithroleukaemia cells K562.Free Radic Res. 2007 Apr;41[4]:452-460.
6. Moncada S. Nitric oxide and cell respiration: physiology and pathology.Verh K Acad Geneeskd Belg. 2000;62[3]:171- 181.
7. Mason MG, Nicholls P,Wilson MT, et al. Nitric oxide inhibition of respiration involves both competitive [heme] and noncompetitive [copper] binding to cytochrome c oxidase. Proc Natl Acad Sci USA. 2006;103:708–713.
8. Sarti P, Forte E, Mastronicola D, et al. Nitric oxide and cytochrome oxidase: mechanisms of inhibition and NO degradation. Biochem Biophys Res Commun. 2000;274:183–187.
9. Jang B, Han S. Biochemical properties of cytochrome c nitrated by peroxynitrite. Biochimie. 2006;88:53–58.
10. Batthyany C, Souza JM, Duran R, et al. Time course and site[s] of cytochrome c tyrosine nitration by peroxynitrite. Biochemistry. 2005;44:8038–8046.
11. Pearce LL, Epperly MW, Greenberger JS, et al. Identification of respiratory complexes I and II as mitochondrial sites of damage following exposure to ionizing radiation and nitric oxide. Nitric Oxide. 2001;5:128–136.
12. Sharpe MA, Cooper CE. Interaction of peroxynitrite with mitochondrial cytochrome oxidase: catalytic production of nitric oxide and irreversible inhibition of enzyme activity. J Biol Chem. 1998;273:30961–30972.
13. Zhang J, Jin B, Li L, et al. Nitric oxide-induced persistent inhibition and nitrosylation of active site cysteine residues of mitochondrial cytochrome-c oxidase in lung endothelial cells. Am J Physiol. 2005;288:840–849.
14. Han D, Canali R, Garcia J, et al. Sites and mechanisms of aconitase inactivation by peroxynitrite: modulation by citrate and glutathione. Biochemistry. 2005;44: 11986–11996.
15. Nisoli E , Clementi E , Paolucci C , et al. Mitochondrial biogenesis in mammals : The role of endogenous nit ric oxide. Science. 2003,299:896-899.
16. Van Gurp M,Festjens N,van Loo G, et a1.Mitochondrial intermembrane proteins in cell death.Biochem Biophys Res Commun. 2003,304:487-497.
17. Shi Y. Mechanisms of caspase activation and inhibition during apoptosis. Mol Cell, 2002, 9: 459-470.
18. Balakirev MY, Khramtsov VV, Zimmer G. Modulation of the mitochondrial permeability transition by nitric oxide. Eur J Biochem. 1997;246:710–718.
19. Costa ADT, Garlid KD, West IC, et a1. Protein kinase G transmits the cardioprotective signal from cytosol to mitochondria. Circ Res. 2005;97:329–336.
20. Takuma K, Phuagphong P, Lee E, et a1.Antiapoptotic effect of cGMP in cultured astrocytes: inhibition by cGMPdependent protein kinase of mitochondrial permeable transition pore. J Biol Chem 2001;276:48093–48099.
21. Juhaszova M, Zorov DB, Kim SH, et al. Glycogen synthase kinase-3beta mediates convergence of protection signaling to inhibit the mitochondrial permeability transition pore. J Clin Invest. 2004;113:1535–1549.
22. Palacios-Callender M, Hollis V, Frakich N, et al.Cytochrome c oxidase maintains mitochondrial respiration during partial inhibition by nitric oxide. J Cell Sci. 2007,120[Pt 1]:160-165.
23. Borutaite V, Brown GC. Nitric oxide induces apoptosis via hydrogen peroxide, but necrosis via energy and thiol depletion. Free Radic Biol Med. 2003; 35: 1457–1468.
24. Poderoso JJ, Carreras MC, Schopfer F, et al. The reaction of nitric oxide with ubiquinol:kinetic properties and biological significance. Free Radic Biol Med. 1999;26:925–935.
25. Grivennikova VG, Vinogradov AD.Generation of superoxide by the mitochondrial Complex I.Biochim Biophys Acta. 2006, 1757[5-6]:553-561.
26. Chinopoulos C, Adam-Vizi V. FEBS J. Calcium, mitochondria and oxidative stress in neuronal pathology. Novel aspects of an enduring theme. 2006, 273[3]:433-450.
27. Waring P.Redox active calcium ion channels and cell death. Arch Biochem Biophys. 2005, 434[1]:33-42.
28. Brookes PS, Zhang J, Dai L, et al.Increased sensitivity of mitochondrial respiration to inhibition by nitric oxide in cardiac hypertrophy.J Mol Cell Cardiol. 2001, 33[1]:69-82.
29. Brunori M. Nitric oxide moves myoglobin centre stage. Trends Biochem Sci. 2001, 26:209–210.
30. Flogel U, Merx MW, Godecke A, et a1. Myoglobin: a scavenger of bioactive NO. Proc Natl Acad Sci USA. 2001,98:735–740.
31. Shiva S, Brookes PS, Patel RP , et al. Nitric oxide partitioning into mitochondrial membranes and the control of respiration at cytochrome c oxidase.Proc Natl Acad Sci USA. 2001, 98[13]:7212-7217.
32. Pearce LL, Kanai AJ, Birder LA, et a1.The catabolic fate of nitric oxide: the nitric oxide oxidase and peroxynitrite reductase activities of cytochrome oxidase. J Biol Chem. 2002, 277:13556–13562.
33. Li W, Jue T, Edwards J, et a1.Changes in NO bioavailability regulate cardiac O2 consumption: control by intramitochondrial SOD2 and intracellular myoglobin. Am J Physiol. 2004, 286: H47–54.
34. Shiva S, Huang Z, Grubina R, , et al. Deoxymyoglobin is a nitrite reductase that generates nitric oxide and regulates mitochondrial respiration. Circ Res. 2007, 100:654–661.
35. Crawford JH, Isbell TS, Huang Z, et al. Hypoxia, red blood cells, and nitrite regulate NO-dependent hypoxic vasodilation. Blood. 2006, 107:566–574.
36. Ilangovan G, Osinbowale S, Bratasz A, et al. Heat shock regulates the respiration of cardiac H9c2 cells through upregulation of nitric oxide synthase. Am J Physiol Cell Physiol. 2004, 287: 1472–1481.
37. French S, Giulivi C, Balaban RS. Nitric oxide synthase in porcine heart mitochondria:evidence for low physiological activity. Am J Physiol Heart Circ Physiol. 2001,80:H2863–2867.
38. Boveris A, Valdez LB, Zaobornyj T, et a1.Mitochondrial metabolic states regulate nitric oxide and hydrogen peroxide diffusion to the cytosol. Biochim Biophys Acta. 2006,1757:535–542.
39. Steppan J, Ryoo S, Schuleri KH, et al. Arginase modulates myocardial contractility by a nitric oxide synthase 1-dependent mechanism. Proc Natl Acad Sci USA. 2006,103: 4759–4764.
40. Lim G, Venetucci L, Eisner DA, et al. Does nitric oxide modulate cardiac ryanodine receptor function? Implications for excitation-contraction coupling.Cardiovasc Res. 2008, 77[2]:256-264.
41. Bendall JK, Damy T, Ratajczak P, et al. Role of myocardial neuronal nitric oxide synthase-derived nitric oxide in beta-adrenergic hyporesponsiveness after myocardial infarction-induced heart failure in rat. Circulation. 2004,110[16]:2368-2375.
42. Walsh EK, Huang H, Wang Z, et al.Control of myocardial oxygen consumption in transgenic mice overexpressing vascular eNOS. Am J Physiol Heart Circ Physiol. 2004, 287[5]:H2115-121.
43. Kinugawa S, Wang Z, Kaminski PM, et al. Limited exercise capacity in heterozygous manganese superoxide dismutase gene-knockout mice: roles of superoxide anion and nitric oxide.Circulation. 2005, 111[12]:1480-1486.
44. Khan SA, Skaf MW, Harrison RW, et al. Nitric oxide regulation of myocardial contractility and calcium cycling: independent impact of neuronal and endothelial nitric oxide synthases.Circ Res. 2003, 92[12]:1322-1329.
45. Sanders DB, Larson DF, Hunter K, et al. Comparison of tumor necrosis factor-alpha effect on the expression of iNOS in macrophage and cardiac myocytes.Perfusion. 2001, 16[1]:67-74.
46. Monnet X, Ghaleh B, Lucats L, et al. Phenotypic adaptation of the late preconditioned heart: myocardial oxygen consumption is reduced. Cardiovasc Res. 2006;70:391–398.
47. Champion HC, Skaf MW, Hare JM. Role of nitric oxide in the pathophysiology of heart failure. Heart Fail Rev. 2003, 8[1]:35-46.
2. Fitzpatrick C. M., Shi Y., Hutchins W. C., et al. Cardioprotection in chronically hypoxic rabbits persists on exposure to normoxia: role of NOS and KATP channels [J]. Am J Physiol Heart Circ Physiol, 2005, 288(1): H62-68.
3. Lynn E. G., Lu Z., Minerbi D., et al. The regulation, control, and consequences of mitochondrial oxygen utilization and disposition in the heart and skeletal muscle during hypoxia [J]. Antioxid Redox Signal, 2007, 9(9): 1353-1361.
4. McLeod C. J., Pagel I., Sack M. N. The mitochondrial biogenesis regulatory program in cardiac adaptation to ischemia--a putative target for therapeutic intervention [J]. Trends Cardiovasc Med, 2005, 15(3): 118-123.
5. Massion P. B., Pelat M., Belge C., et al. Regulation of the mammalian heart function by nitric oxide [J]. Comp Biochem Physiol A Mol Integr Physiol, 2005, 142(2): 144-150.
6. Lacza Z., Pankotai E., Csordas A., et al. Mitochondrial NO and reactive nitrogen species production: does mtNOS exist? [J]. Nitric Oxide, 2006, 14(2): 162-168.
7. Clementi E., Nisoli E. Nitric oxide and mitochondrial biogenesis: a key to long-term regulation of cellular metabolism [J]. Comp Biochem Physiol A Mol Integr Physiol, 2005, 142(2): 102-110.
8. Burley D. S., Ferdinandy P., Baxter G. F. Cyclic GMP and protein kinase-G in myocardial ischaemia-reperfusion: opportunities and obstacles for survival signaling [J]. Br J Pharmacol, 2007, 152(6): 855-869.
9. Saraiva R. M., Hare J. M. Nitric oxide signaling in the cardiovascular system: implications for heart failure [J]. Curr Opin Cardiol, 2006, 21(3): 221-228.
10. Wang S., Yan L., Wesley R. A., et al. Nitric oxide increases tumor necrosis factor production in differentiated U937 cells by decreasing cyclic AMP [J]. J Biol Chem, 1997, 272(9): 5959-5965.
11. Ma P., Cui X., Wang S., et al. Nitric oxide post-transcriptionally up-regulates LPS-induced IL-8 expression through p38 MAPK activation [J]. J Leukoc Biol, 2004, 76(1): 278-287.
12. Zhang J., Wang S., Wesley R. A., et al. Adjacent sequence controls the response polarity of nitric oxide-sensitive Sp factor binding sites [J]. J Biol Chem, 2003, 278(31): 29192-29200.
13. Johnson D. T., Harris R. A., French S., et al. Tissue heterogeneity of the mammalian mitochondrial proteome [J]. Am J Physiol Cell Physiol, 2007, 292(2): C689-697.
14. Weibel E. R., Staubli W., Gnagi H. R., et al. Correlated morphometric and biochemical studies on the liver cell. I. Morphometric model, stereologic methods, and normal morphometric data for rat liver [J]. J Cell Biol, 1969, 42(1): 68-91.
15. Nouette-Gaulain K., Malgat M., Rocher C., et al. Time course of differential mitochondrial energy metabolism adaptation to chronic hypoxia in right and left ventricles [J]. Cardiovasc Res, 2005, 66(1): 132-140.
16. Cervos Navarro J., Kunas R. C., Sampaolo S., et al. Heart mitochondria in rats submitted to chronic hypoxia [J]. Histol Histopathol, 1999, 14(4): 1045-1052.
17. Trinei M., Berniakovich I., Pelicci P. G., et al. Mitochondrial DNA copy number is regulated by cellular proliferation: a role for Ras and p66(Shc) [J]. Biochim Biophys Acta, 2006, 1757(5-6): 624-630.
18. Cai J. H., Deng S., Kumpf S. W., et al. Validation of rat reference genes for improved quantitative gene expression analysis using low density arrays [J]. Biotechniques, 2007, 42(4): 503-512.
19. Okamoto K., Shaw J. M. Mitochondrial morphology and dynamics in yeast and multicellular eukaryotes [J]. Annu Rev Genet, 2005, 39: 503-536.
20. Meeusen S., McCaffery J. M., Nunnari J. Mitochondrial fusion intermediates revealed in vitro [J]. Science, 2004, 305(5691): 1747-1752.
21. Collins T. J., Berridge M. J., Lipp P., et al. Mitochondria are morphologically and functionally heterogeneous within cells [J]. Embo J, 2002, 21(7): 1616-1627.
22. Rube D. A., van der Bliek A. M. Mitochondrial morphology is dynamic and varied [J]. Mol Cell Biochem, 2004, 256-257(1-2): 331-339.
23. Chowdhury U. K., Sathia S., Ray R., et al. Histopathology of the right ventricular outflow tract and its relationship to clinical outcomes and arrhythmias in patients with tetralogy of Fallot [J]. J Thorac Cardiovasc Surg, 2006, 132(2): 270-277.
24.孙桂媛石玉秀,祝淑文等.法乐四联症心肌纤维扫描电镜的观察[J].中华胸心外科杂志, 1998, 14(3): 139-141.
25.李刚,王湘,季军等.法乐四联症右室流出道心肌超微结构特点及其临床意义[J].中华实用外科杂志, 2006, 23(9): 1036-1037.
26.李晓峰李仲智,郎志奇等.法洛四联症右室流出道心肌超微结构特点[J].实用儿科临床杂志, 2004, 19(7): 554-557.
27. Anderson S., Bankier A. T., Barrell B. G., et al. Sequence and organization of the human mitochondrial genome [J]. Nature, 1981, 290(5806): 457-465.
28. Garesse R., Vallejo C. G. Animal mitochondrial biogenesis and function: a regulatory cross-talk between two genomes [J]. Gene, 2001, 263(1-2): 1-16.
29. Stiburek L., Hansikova H., Tesarova M., et al. Biogenesis of eukaryotic cytochrome c oxidase [J]. Physiol Res, 2006, 55 Suppl 2: S27-41.
30. Fontanesi F., Soto I. C., Horn D., et al. Assembly of mitochondrial cytochrome c-oxidase, a complicated and highly regulated cellular process [J]. Am J Physiol Cell Physiol, 2006, 291(6): C1129-1147.
31. Puigserver P., Wu Z., Park C. W., et al. A cold-inducible coactivator of nuclear receptors linked to adaptive thermogenesis [J]. Cell, 1998, 92(6): 829-839.
32. Kelly D. P., Scarpulla R. C. Transcriptional regulatory circuits controlling mitochondrial biogenesis and function [J]. Genes Dev, 2004, 18(4): 357-368.
33. Lehman J. J., Barger P. M., Kovacs A., et al. Peroxisome proliferator-activated receptor gamma coactivator-1 promotes cardiac mitochondrial biogenesis [J]. J Clin Invest, 2000, 106(7): 847-856.
34. Wu H., Kanatous S. B., Thurmond F. A., et al. Regulation of mitochondrial biogenesis in skeletal muscle by CaMK [J]. Science, 2002, 296(5566): 349-352.
35. Mootha V. K., Lindgren C. M., Eriksson K. F., et al. PGC-1alpha-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes [J]. Nat Genet, 2003, 34(3): 267-273.
36. Scarpulla R. C. Nuclear control of respiratory gene expression in mammalian cells [J]. J Cell Biochem, 2006, 97(4): 673-683.
37. Irrcher I., Adhihetty P. J., Joseph A. M., et al. Regulation of mitochondrial biogenesis in muscle by endurance exercise [J]. Sports Med, 2003, 33(11): 783-793.
38. Irrcher I., Adhihetty P. J., Sheehan T., et al. PPARgamma coactivator-1alpha expressionduring thyroid hormone- and contractile activity-induced mitochondrial adaptations [J]. Am J Physiol Cell Physiol, 2003, 284(6): C1669-1677.
39. Suliman H. B., Carraway M. S., Welty-Wolf K. E., et al. Lipopolysaccharide stimulates mitochondrial biogenesis via activation of nuclear respiratory factor-1 [J]. J Biol Chem, 2003, 278(42): 41510-41518.
40. Weitzel J. M., Iwen K. A., Seitz H. J. Regulation of mitochondrial biogenesis by thyroid hormone [J]. Exp Physiol, 2003, 88(1): 121-128.
41. Kang D., Hamasaki N. Mitochondrial transcription factor A in the maintenance of mitochondrial DNA: overview of its multiple roles [J]. Ann N Y Acad Sci, 2005, 1042: 101-108.
42. Kang D., Kim S. H., Hamasaki N. Mitochondrial transcription factor A (TFAM): roles in maintenance of mtDNA and cellular functions [J]. Mitochondrion, 2007, 7(1-2): 39-44.
43. May-Panloup P., Vignon X., Chretien M. F., et al. Increase of mitochondrial DNA content and transcripts in early bovine embryogenesis associated with upregulation of mtTFA and NRF1 transcription factors [J]. Reprod Biol Endocrinol, 2005, 3: 65.
44. Corno A. F., Milano G., Samaja M., et al. Chronic hypoxia: a model for cyanotic congenital heart defects [J]. J Thorac Cardiovasc Surg, 2002, 124(1): 105-112.
45. Lahiri S., Roy A., Baby S. M., et al. Oxygen sensing in the body [J]. Prog Biophys Mol Biol, 2006, 91(3): 249-286.
46.任晓莉李瑞兰.动态监测血乳酸对先天性心脏病心内直视手术患儿的临床意义[J].心肺血管病杂志, 2007, 26(3): 174-175.
47. Marin-Garcia J., Ananthakrishnan R., Agrawal N., et al. Mitochondrial gene expression during bovine cardiac growth and development [J]. J Mol Cell Cardiol, 1994, 26(8): 1029-1036.
48. Costa L. E., Boveris A., Koch O. R., et al. Liver and heart mitochondria in rats submitted to chronic hypobaric hypoxia [J]. Am J Physiol, 1988, 255(1 Pt 1): C123-129.
49. Piel D. A., Khan A. R., Waibel R., et al. Chronic hypoxemia increases myocardial cytochrome oxidase [J]. J Thorac Cardiovasc Surg, 2005, 130(4): 1101-1106.
50. Rasbach K. A., Schnellmann R. G. PGC-1alpha over-expression promotes recoveryfrom mitochondrial dysfunction and cell injury [J]. Biochem Biophys Res Commun, 2007, 355(3): 734-739.
51. Shinde S. B., Save V. C., Patil N. D., et al. Impairment of mitochondrial respiratory chain enzyme activities in tetralogy of Fallot [J]. Clin Chim Acta, 2007, 377(1-2): 138-143.
52. MacMicking J., Xie Q. W., Nathan C. Nitric oxide and macrophage function [J]. Annu Rev Immunol, 1997, 15: 323-350.
53. Massion P. B., Feron O., Dessy C., et al. Nitric oxide and cardiac function: ten years after, and continuing [J]. Circ Res, 2003, 93(5): 388-398.
54. Alderton W. K., Cooper C. E., Knowles R. G. Nitric oxide synthases: structure, function and inhibition [J]. Biochem J, 2001, 357(Pt 3): 593-615.
55. Elfering S. L., Sarkela T. M., Giulivi C. Biochemistry of mitochondrial nitric-oxide synthase [J]. J Biol Chem, 2002, 277(41): 38079-38086.
56. Massion P. B., Dessy C., Desjardins F., et al. Cardiomyocyte-restricted overexpression of endothelial nitric oxide synthase (NOS3) attenuates beta-adrenergic stimulation and reinforces vagal inhibition of cardiac contraction [J]. Circulation, 2004, 110(17): 2666-2672.
57. Valdez L. B., Zaobornyj T., Alvarez S., et al. Heart mitochondrial nitric oxide synthase. Effects of hypoxia and aging [J]. Mol Aspects Med, 2004, 25(1-2): 49-59.
58. Seddon M., Shah A. M., Casadei B. Cardiomyocytes as effectors of nitric oxide signalling [J]. Cardiovasc Res, 2007, 75(2): 315-326.
59. Davidson S. M., Duchen M. R. Effects of NO on mitochondrial function in cardiomyocytes: Pathophysiological relevance [J]. Cardiovasc Res, 2006, 71(1): 10-21.
60. Jung F., Palmer L. A., Zhou N., et al. Hypoxic regulation of inducible nitric oxide synthase via hypoxia inducible factor-1 in cardiac myocytes [J]. Circ Res, 2000, 86(3): 319-325.
61. Coulet F., Nadaud S., Agrapart M., et al. Identification of hypoxia-response element in the human endothelial nitric-oxide synthase gene promoter [J]. J Biol Chem, 2003, 278(47): 46230-46240.
62. Forkel J., Chen X., Wandinger S., et al. Responses of chronically hypoxic rat hearts to ischemia: KATP channel blockade does not abolish increased RV tolerance to ischemia[J]. Am J Physiol Heart Circ Physiol, 2004, 286(2): H545-551.
63. Grilli A., De Lutiis M. A., Patruno A., et al. Effect of chronic hypoxia on inducible nitric oxide synthase expression in rat myocardial tissue [J]. Exp Biol Med (Maywood), 2003, 228(8): 935-942.
64. Thompson L. P., Dong Y. Chronic hypoxia decreases endothelial nitric oxide synthase protein expression in fetal guinea pig hearts [J]. J Soc Gynecol Investig, 2005, 12(6): 388-395.
65. Zaobornyj T., Gonzales G. F., Valdez L. B. Mitochondrial contribution to the molecular mechanism of heart acclimatization to chronic hypoxia: role of nitric oxide [J]. Front Biosci, 2007, 12: 1247-1259.
66. Ghafourifar P., Sen C. K. Mitochondrial nitric oxide synthase [J]. Front Biosci, 2007, 12: 1072-1078.
67. Poot M., Zhang Y. Z., Kramer J. A., et al. Analysis of mitochondrial morphology and function with novel fixable fluorescent stains [J]. J Histochem Cytochem, 1996, 44(12): 1363-1372.
68. Livak K. J., Schmittgen T. D. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method [J]. Methods, 2001, 25(4): 402-408.
69. Frazier A. E., Kiu C., Stojanovski D., et al. Mitochondrial morphology and distribution in mammalian cells [J]. Biol Chem, 2006, 387(12): 1551-1558.
70. Zong H., Ren J. M., Young L. H., et al. AMP kinase is required for mitochondrial biogenesis in skeletal muscle in response to chronic energy deprivation [J]. Proc Natl Acad Sci U S A, 2002, 99(25): 15983-15987.
71. Leary S. C., Shoubridge E. A. Mitochondrial biogenesis: which part of "NO" do we understand? [J]. Bioessays, 2003, 25(6): 538-541.
72. Cerra M. C., Pellegrino D. Cardiovascular cGMP-generating systems in physiological and pathological conditions [J]. Curr Med Chem, 2007, 14(5): 585-599.
73. Bellamy T. C., Wood J., Goodwin D. A., et al. Rapid desensitization of the nitric oxide receptor, soluble guanylyl cyclase, underlies diversity of cellular cGMP responses [J]. Proc Natl Acad Sci U S A, 2000, 97(6): 2928-2933.
74. Navarro A., Torrejon R., Bandez M. J., et al. Mitochondrial function andmitochondria-induced apoptosis in an overstimulated rat ovarian cycle [J]. Am J Physiol Endocrinol Metab, 2005, 289(6): E1101-1109.
75. Nagy G., Barcza M., Gonchoroff N., et al. Nitric oxide-dependent mitochondrial biogenesis generates Ca2+ signaling profile of lupus T cells [J]. J Immunol, 2004, 173(6): 3676-3683.
76. Baris O., Savagner F., Nasser V., et al. Transcriptional profiling reveals coordinated up-regulation of oxidative metabolism genes in thyroid oncocytic tumors [J]. J Clin Endocrinol Metab, 2004, 89(2): 994-1005.
77. Carew J. S., Nawrocki S. T., Xu R. H., et al. Increased mitochondrial biogenesis in primary leukemia cells: the role of endogenous nitric oxide and impact on sensitivity to fludarabine [J]. Leukemia, 2004, 18(12): 1934-1940.
78.艾珣,顾玉东.培养人脐静脉内皮细胞慢性缺血缺氧模型[J].中华手外科杂志, 2001, 17(4): 233-236.
79.孙胜,高文祥,廖卫公.神经细胞化学缺氧预处理模型的建立[J].第三军医大学学报, 2007, 29(21): 2100-2101.
80. Roy S., Khanna S., Bickerstaff A. A., et al. Oxygen sensing by primary cardiac fibroblasts: a key role of p21(Waf1/Cip1/Sdi1) [J]. Circ Res, 2003, 92(3): 264-271.
81. Li H. T., Long C. S., Rokosh D. G., et al. Chronic hypoxia differentially regulates alpha 1-adrenergic receptor subtype mRNAs and inhibits alpha 1-adrenergic receptor-stimulated cardiac hypertrophy and signaling [J]. Circulation, 1995, 92(4): 918-925.
82. Kacimi R., Long C. S., Karliner J. S. Chronic hypoxia modulates the interleukin-1beta-stimulated inducible nitric oxide synthase pathway in cardiac myocytes [J]. Circulation, 1997, 96(6): 1937-1943.
83. Newby D., Marks L., Lyall F. Dissolved oxygen concentration in culture medium: assumptions and pitfalls [J]. Placenta, 2005, 26(4): 353-357.
84. Qing M., Gorlach A., Schumacher K., et al. The hypoxia-inducible factor HIF-1 promotes intramyocardial expression of VEGF in infants with congenital cardiac defects [J]. Basic Res Cardiol, 2007, 102(3): 224-232.
1. Lee S, Kim S, Sun X, et al. Cell cycle-dependent mitochondrial biogenesis and dynamics in mammalian cells. Biochem Biophys Res Commun. 2007,25; 357(1):111-117.
2. WeberK, Bruck P, Mikes Z, et al. Glucocorticoid hormone stimulates mitochondrial biogenesis specifically in skeletal muscle. Endocrinology . 2002, 143(1): 177-84.
3. Nisoli E, Carruba MO. Nitric oxide and mitochondrial biogenesis. 2006. J Cell Sci. 119:2855-62.
4. Yaffe, M P.. The cutting edge of mitochondrial fusion. Nat. Cell Biol. 2003,5:497-499.
5. Meeusen S, McCaffery J M , Nunnari J.. Mitochondrial fusion intermediates revealed in vitro. Science, 2004, 305:1747-1752.
6. Okamoto K, Shaw J M. Mitochondrial morphology and dynamics in yeast and multicellular eukaryotes. Annu. Rev. Genet. 2005, 39: 503-536.
7. Rube DA, vander Bliek, AM. Mitochondrial motphology is dynamic and varied. Mol. Cell. Biochem. 2004, 256/257:331-339.
8. Civitarese AE, Carling S, Heilbronn LK,et al.Calorie Restriction Increases Muscle Mitochondrial Biogenesis in Healthy Humans. PLoS Med. 2007, 6;4(3):e76
9. Chabi B, Adhihetty PJ, Ljubucuc V, et al. How is Mitochondrial Biogenesis Affected in Mitochondrial Disease? Med. Sci. Sports Exerc, 2005, 37(12): 2102-2110.
10. Carew JS, Nawrocki ST, Xu RH, et al. Increased mitochondrial biogenesis in primary leukemia cells: the role of endogenous nitric oxide and impact on sensitivity to fludarabine. Leukemia. 2004 , 18(12): 1934-40.
11. Kamal MA, French SW. Drug-induced increased mitochondrial biogenesis in a liver biopsy. xp-Mol-Pathol. 2004 ,77(3): 201-4.
12. Mootha VK, Lindgren CM, Eriksson KF, et al. PGC-1alpha-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat.Genet.2003, 34:267-73
13. Ukropcova B, Sereda O, de Jonge L,et al. Family history of diabetes links impaired substrate switching and reduced mitochondrial content in skeletal muscle. Diabetes. 2007, 56(3):720-7
14. M. Lemoine, M. Kim, V. Ratziu,et al. Liver mitochondrial function and biogenesis in patients with NAFLD: hepatic expression of PGC-1alpha, COX2 and Cox4. Journal of Hepatology, 2007, 46(1):S272-S273.
15. Nisoli E, Clementi E, Carruba MO,et al. Defective mitochondrial biogenesis: a hallmark of the high cardiovascular risk in the metabolic syndrome? Circ Res. 2007 , 100(6):795-806.
16. Libin Cui, Hyunkyung Jeong, Fran Borovecki, et al. Transcriptional Repression of PGC-1αby Mutant Huntingtin Leads to Mitochondrial Dysfunction and Neurodegeneration. Cell, 2006, 127(1):59-69.
17. Gomez C, Bandez MJ, Navarro A. Pesticides and impairment of mitochondrial function in relation with the parkinsonian syndrome. Front Biosci. 2007,12:1079-93.
18. Sardiello M, Tripoli G, Romito A, et al. Energy biogenesis: one key for coordinating two genomes. Trends Genet. 2005,21:12-16.
19. Butow RA, Avadhani NG. Mitochondrial signaling: the retrograde response. Mol. Cell. 2004, 14:1-15.
20. Kelly DP, Scarpulla RC. Transcriptional regulatory circuits controlling mitochondrial biogenesis and function. Genes Dev. 2004, 18:357-68.
21. Garesse R, Vallejo CG. Animal mitochondrial biogenesis and function: a regulatory cross-talk between two genomes. Gene. 2001, 263:1-16.
22. Wickner W, Schekman R. Protein translocation across biological membranes. Science. 2005, 310:1452-1456.
23. Lin J, Handschin C, Spiegelman BM. Metabolic control through the PGC-1 family of transcription coactivators. Cell Metab.2005, 1:361-70
24. Puigserver P, Spiegelman BM. Peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PGC-1 alpha): transcriptional coactivator and metabolic regulator. Endocr Rev. 2003,24:78-90.
25. Puigserver P,Wu Z, Park CW, et al. A cold-inducible coactivator of nuclear receptors linked to adaptive thermogenesis. Cell. 1998,92:829-39.
26. Finck B N, Kelly D P. PGC-1 coactivators: inducible regulators of energy metabolism in health and disease. J of Clin Invest. 2006, 116, 615-622.
27. Puigserver P, Adelmant G,Wu Z, et al. Activation of PPARgamma coactivator-1through transcription factor docking. Science .1999, 286:1368-71.
28. Monsalve M,Wu Z, Adelmant G, et al. Direct coupling of transcription and mRNA processing through the thermogenic coactivator PGC-1. Mol Cell. 2000,6:307-16.
29. Knutti D , Kressler D , Kralli A. Regulation of the transcriptional coactivator PGC21 via MAPK2sensitive interaction with a repressor. Proc Natl Acad Sci USA. 2001 , 98 (17) :9713-9718.
30. Wu Z, Puigserver P, Andersson U, et al. Mechanisms controlling mitochondrial biogenesis and respiration through the thermogenic coactivator PGC-1. Cell .1999, 98:115-24.
31. Mootha VK, Lindgren CM, Eriksson KF, et al. Nat.Genet. 2003, 34:267-73.
32. Vega RB, Huss JM, Kelly DP. The coactivator PGC-1 cooperates with peroxisome proliferator-activated receptor alpha in transcriptional control of nuclear genes encoding mitochondrial fatty acid oxidation enzymes. Mol Cell Biol. 2000,20:1868-76.
33. Cartoni R, Leger B, Hock MB, et al. Mitofusins 1/2 and ERRalpha expression are increased in human skeletal muscle after physical exercise. J Physiol. 2005,567:349-58.
34. Soriano FX, Liesa M, Bach D, et al. Evidence for a Mitochondrial Regulatory Pathway Defined by Peroxisome Proliferator-Activated Receptor-gamma Coactivator-1(alpha), Estrogen-Related Receptor-alpha, and Mitofusin 2. Diabetes. 2006, 55:1783-91.
35. Zong H, Ren JM, Young LH, et al. AMP kinase is required for mitochondrial biogenesis in skeletal muscle in response to chronic energy deprivation. Proc Natl Acad Sci USA. 2002, 99:15983-87.
36. Kukidome D, Nishikawa T, Sonoda K, et al. Activation of AMP-activated protein kinase reduces hyperglycemia-induced mitochondrial reactive oxygen species production and promotes mitochondrial biogenesis in human umbilical vein endothelial cells. Diabetes . 2006,55:120-27.
37. Wu H, Kanatous S B, Thurmond F A, et al. Regulation of mitochondrial biogenesis in skeletal muscle by CaMK. Science. 2002,296: 349-352.
38. Lin J, Wu H, Tarr PT, et al. Transcriptional co-activator PGC-1_ drives the formation of slow-twitch muscle fibres. Nature .2002b.418: 797-801.
39. Lee J, Kim CH, Simon DK, et al. Mitochondrial cyclic AMP response element-binding protein (CREB) mediates mitochondrial gene expression and neuronal survival. J BiolChem. 2005,280:40398-401.
40. Nisoli E, Falcone S, Tonello C. Mitochondrial biogenesis by NO yields functionally active mitochondria in mammals.Proc Natl Acad Sci USA. 2004, 23;101(47):16507-12.
41. Nisoli E, Clementi E, Paolucci C, et al. Mitochondrial biogenesis in mammals: the role of endogenous nitric oxide. Science. 2003,299:896-99.
42. Suliman HB, Carraway MS, Tatro LG, et al. A new activating role for CO in cardiac mitochondrial biogenesis. J Cell Sci. 2007,120(Pt 2):299-308.
43. Puigserver P , Adelmant G, Wu Z, et al. Activation of PPARgamma coactivator21 through transcription factor docking. Science . 1999 , 286(5443) :1368-1371.
44. Puigserver P, Rhee J, Lin J, et al. Cytokine stimulation of energy expenditure through p38 MAP kinase activation of PPARgamma coactivator-1. Mol Cell. 2001, 8:971-82.
45. Fan M, Rhee J , StPierre J , et al. Suppression of mitochondrial respiration through recruitment of p160 myb binding protein to PGC21alpha : Modulation by p38 MAPK. Genes Dev . 2004 , 18 (3):278-289.
1. Salzman AL , Menconi MJ , Unno N , et al. Nitric oxide dilates tight junction and depletes ATP in cultured Caco-2BBe intestinal epthelial monolayers. Am J Physio.1995 ,268 :361-373.
2.杨贵远,赵克森,刘杰.一氧化氮降低失血性休克血管反应性的机制.中国病理生理杂志.2000,16:1163-1166.
3. Hierholzer C , Billiar TR. Molecular mechanisms in t he early phase of hemorrhagic shock. Langenbecks Arch Surg , 2001 ,386 :302-308.
4. Yang J, Wu LJ, Tashiro S, et al .Nitric oxide activated by p38 and NF-kappaB facilitates apoptosis and cell cycle arrest under oxidative stress in evodiamine-treated human melanoma A375-S2 cells. Free Radic Res. 2008 Jan;42[1]:1-11.
5. Aquilano K, Filomeni G, Di Renzo L, et al .Reactive oxygen and nitrogen species are involved in sorbitol-induced apoptosis of human erithroleukaemia cells K562.Free Radic Res. 2007 Apr;41[4]:452-460.
6. Moncada S. Nitric oxide and cell respiration: physiology and pathology.Verh K Acad Geneeskd Belg. 2000;62[3]:171- 181.
7. Mason MG, Nicholls P,Wilson MT, et al. Nitric oxide inhibition of respiration involves both competitive [heme] and noncompetitive [copper] binding to cytochrome c oxidase. Proc Natl Acad Sci USA. 2006;103:708–713.
8. Sarti P, Forte E, Mastronicola D, et al. Nitric oxide and cytochrome oxidase: mechanisms of inhibition and NO degradation. Biochem Biophys Res Commun. 2000;274:183–187.
9. Jang B, Han S. Biochemical properties of cytochrome c nitrated by peroxynitrite. Biochimie. 2006;88:53–58.
10. Batthyany C, Souza JM, Duran R, et al. Time course and site[s] of cytochrome c tyrosine nitration by peroxynitrite. Biochemistry. 2005;44:8038–8046.
11. Pearce LL, Epperly MW, Greenberger JS, et al. Identification of respiratory complexes I and II as mitochondrial sites of damage following exposure to ionizing radiation and nitric oxide. Nitric Oxide. 2001;5:128–136.
12. Sharpe MA, Cooper CE. Interaction of peroxynitrite with mitochondrial cytochrome oxidase: catalytic production of nitric oxide and irreversible inhibition of enzyme activity. J Biol Chem. 1998;273:30961–30972.
13. Zhang J, Jin B, Li L, et al. Nitric oxide-induced persistent inhibition and nitrosylation of active site cysteine residues of mitochondrial cytochrome-c oxidase in lung endothelial cells. Am J Physiol. 2005;288:840–849.
14. Han D, Canali R, Garcia J, et al. Sites and mechanisms of aconitase inactivation by peroxynitrite: modulation by citrate and glutathione. Biochemistry. 2005;44: 11986–11996.
15. Nisoli E , Clementi E , Paolucci C , et al. Mitochondrial biogenesis in mammals : The role of endogenous nit ric oxide. Science. 2003,299:896-899.
16. Van Gurp M,Festjens N,van Loo G, et a1.Mitochondrial intermembrane proteins in cell death.Biochem Biophys Res Commun. 2003,304:487-497.
17. Shi Y. Mechanisms of caspase activation and inhibition during apoptosis. Mol Cell, 2002, 9: 459-470.
18. Balakirev MY, Khramtsov VV, Zimmer G. Modulation of the mitochondrial permeability transition by nitric oxide. Eur J Biochem. 1997;246:710–718.
19. Costa ADT, Garlid KD, West IC, et a1. Protein kinase G transmits the cardioprotective signal from cytosol to mitochondria. Circ Res. 2005;97:329–336.
20. Takuma K, Phuagphong P, Lee E, et a1.Antiapoptotic effect of cGMP in cultured astrocytes: inhibition by cGMPdependent protein kinase of mitochondrial permeable transition pore. J Biol Chem 2001;276:48093–48099.
21. Juhaszova M, Zorov DB, Kim SH, et al. Glycogen synthase kinase-3beta mediates convergence of protection signaling to inhibit the mitochondrial permeability transition pore. J Clin Invest. 2004;113:1535–1549.
22. Palacios-Callender M, Hollis V, Frakich N, et al.Cytochrome c oxidase maintains mitochondrial respiration during partial inhibition by nitric oxide. J Cell Sci. 2007,120[Pt 1]:160-165.
23. Borutaite V, Brown GC. Nitric oxide induces apoptosis via hydrogen peroxide, but necrosis via energy and thiol depletion. Free Radic Biol Med. 2003; 35: 1457–1468.
24. Poderoso JJ, Carreras MC, Schopfer F, et al. The reaction of nitric oxide with ubiquinol:kinetic properties and biological significance. Free Radic Biol Med. 1999;26:925–935.
25. Grivennikova VG, Vinogradov AD.Generation of superoxide by the mitochondrial Complex I.Biochim Biophys Acta. 2006, 1757[5-6]:553-561.
26. Chinopoulos C, Adam-Vizi V. FEBS J. Calcium, mitochondria and oxidative stress in neuronal pathology. Novel aspects of an enduring theme. 2006, 273[3]:433-450.
27. Waring P.Redox active calcium ion channels and cell death. Arch Biochem Biophys. 2005, 434[1]:33-42.
28. Brookes PS, Zhang J, Dai L, et al.Increased sensitivity of mitochondrial respiration to inhibition by nitric oxide in cardiac hypertrophy.J Mol Cell Cardiol. 2001, 33[1]:69-82.
29. Brunori M. Nitric oxide moves myoglobin centre stage. Trends Biochem Sci. 2001, 26:209–210.
30. Flogel U, Merx MW, Godecke A, et a1. Myoglobin: a scavenger of bioactive NO. Proc Natl Acad Sci USA. 2001,98:735–740.
31. Shiva S, Brookes PS, Patel RP , et al. Nitric oxide partitioning into mitochondrial membranes and the control of respiration at cytochrome c oxidase.Proc Natl Acad Sci USA. 2001, 98[13]:7212-7217.
32. Pearce LL, Kanai AJ, Birder LA, et a1.The catabolic fate of nitric oxide: the nitric oxide oxidase and peroxynitrite reductase activities of cytochrome oxidase. J Biol Chem. 2002, 277:13556–13562.
33. Li W, Jue T, Edwards J, et a1.Changes in NO bioavailability regulate cardiac O2 consumption: control by intramitochondrial SOD2 and intracellular myoglobin. Am J Physiol. 2004, 286: H47–54.
34. Shiva S, Huang Z, Grubina R, , et al. Deoxymyoglobin is a nitrite reductase that generates nitric oxide and regulates mitochondrial respiration. Circ Res. 2007, 100:654–661.
35. Crawford JH, Isbell TS, Huang Z, et al. Hypoxia, red blood cells, and nitrite regulate NO-dependent hypoxic vasodilation. Blood. 2006, 107:566–574.
36. Ilangovan G, Osinbowale S, Bratasz A, et al. Heat shock regulates the respiration of cardiac H9c2 cells through upregulation of nitric oxide synthase. Am J Physiol Cell Physiol. 2004, 287: 1472–1481.
37. French S, Giulivi C, Balaban RS. Nitric oxide synthase in porcine heart mitochondria:evidence for low physiological activity. Am J Physiol Heart Circ Physiol. 2001,80:H2863–2867.
38. Boveris A, Valdez LB, Zaobornyj T, et a1.Mitochondrial metabolic states regulate nitric oxide and hydrogen peroxide diffusion to the cytosol. Biochim Biophys Acta. 2006,1757:535–542.
39. Steppan J, Ryoo S, Schuleri KH, et al. Arginase modulates myocardial contractility by a nitric oxide synthase 1-dependent mechanism. Proc Natl Acad Sci USA. 2006,103: 4759–4764.
40. Lim G, Venetucci L, Eisner DA, et al. Does nitric oxide modulate cardiac ryanodine receptor function? Implications for excitation-contraction coupling.Cardiovasc Res. 2008, 77[2]:256-264.
41. Bendall JK, Damy T, Ratajczak P, et al. Role of myocardial neuronal nitric oxide synthase-derived nitric oxide in beta-adrenergic hyporesponsiveness after myocardial infarction-induced heart failure in rat. Circulation. 2004,110[16]:2368-2375.
42. Walsh EK, Huang H, Wang Z, et al.Control of myocardial oxygen consumption in transgenic mice overexpressing vascular eNOS. Am J Physiol Heart Circ Physiol. 2004, 287[5]:H2115-121.
43. Kinugawa S, Wang Z, Kaminski PM, et al. Limited exercise capacity in heterozygous manganese superoxide dismutase gene-knockout mice: roles of superoxide anion and nitric oxide.Circulation. 2005, 111[12]:1480-1486.
44. Khan SA, Skaf MW, Harrison RW, et al. Nitric oxide regulation of myocardial contractility and calcium cycling: independent impact of neuronal and endothelial nitric oxide synthases.Circ Res. 2003, 92[12]:1322-1329.
45. Sanders DB, Larson DF, Hunter K, et al. Comparison of tumor necrosis factor-alpha effect on the expression of iNOS in macrophage and cardiac myocytes.Perfusion. 2001, 16[1]:67-74.
46. Monnet X, Ghaleh B, Lucats L, et al. Phenotypic adaptation of the late preconditioned heart: myocardial oxygen consumption is reduced. Cardiovasc Res. 2006;70:391–398.
47. Champion HC, Skaf MW, Hare JM. Role of nitric oxide in the pathophysiology of heart failure. Heart Fail Rev. 2003, 8[1]:35-46.