Control of mitochondrial fitness and damage

控制线粒体健康和损伤

• 批准号: 9348196
• 负责人: Steven Sollott
• 金额: $ 80.59万
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9348196
• 关键词: ATP Citrate (pro-S)-Lyase; Ablation; Acetyl Coenzyme A; Acetylation; Affect; Age; Age-Months; Aging; Agreement; Area; Autophagocytosis; BCL-2 Protein; BCL2 gene; Biogenesis; Biological Markers; Biological Preservation; Brain; CCRL2 gene; Cardiac; Cardiac Myocytes; Cell Death; Cell Survival; Cells; Cessation of life; Citrates; Clinical; Clinical Treatment; Complex; Consumption; Cytoprotection; Deacetylation; Defect; Development; Electron Microscopy; Exercise; Exhibits; Experimental Animal Model; Failure; Family member; Fluorescence; Genus Hippocampus; Glucose; Goals; Health; Health Status; Heart; Hypoxia; Image; Impairment; Inbreeding; Infarction; Injury; Intervention; Ischemic Preconditioning; Lasers; Lead; Life; Life Style; Lipids; Longevity; Measurement; Memory; Metabolic; Metabolic Diseases; Mitochondria; Mitochondrial Swelling; Mitotic; Muscle satellite cell; Myocardial Infarction; Nature; Neurons; Neuroprotective Agents; Nuclear; Organ; Organelles; Organism; Outcome; Oxidants; Oxidation-Reduction; Palmitates; Pattern; Permeability; Phosphorylation; Phosphotransferases; Population; Process; Production; Proteins; Protocols documentation; Rattus; Reactive Oxygen Species; Resistance; Respiration; Role; Running; Sampling; Scanning; Signal Pathway; Signal Transduction; Stress; Stroke; Stroke prevention; Technology; United States National Institutes of Health; age related; base; burden of illness; design; energy balance; healthy aging; improved; insight; mitochondrial fitness; mitochondrial permeability transition pore; novel therapeutics; oxidant stress; oxidation; preconditioning; prevent; research study; respiratory; response; sarcopenia; trait; treatment strategy

Environmental stresses converge on the mitochondria that can trigger or inhibit cell death. Excitable, post-mitotic cells (such as cardiac myocytes in heart, and neurons in brain), in response to sub-lethal noxious stress engage mechanisms affording protection from subsequent insults. These protection mechanisms involve activation of endogenous signaling which can confer significant resistance to oxidant and other stresses associated with hypoxia/reoxygenation (i.e., during a heart attack or stroke), which promotes the enhanced capacity for cell survival. However, the upstream signaling mechanisms have remained an area of active debate, and the end effector(s) have remained unsolved. We show that reoxygenation after prolonged hypoxia reduces the reactive oxygen species- (ROS-) threshold for the mitochondrial permeability transition (mPTP) in cardiac myocytes, and that cell survival is steeply negatively correlated with the fraction of depolarized mitochondria. We demonstrate that a wide variety of cardio/neuroprotective agents acting via distinct upstream mechanisms all promote cell survival by limiting mPTP induction. We found that protection can be triggered in 2 general ways dependent and independent of regulatory mitochondrial swelling which converge via inhibition of GSK-3b on the end effector, the permeability transition pore complex, preventing the mPTP. Cell protection exhibiting a memory (i.e., "preconditioning") results from triggered mitochondrial swelling (due to enhanced K+ accumulation via influx and/or retention) causing enhanced substrate oxidation and ROS production, leading to redox activation of PKC which in turn inhibits GSK-3b (via phosphorylation of ser-9). We concluded that GSK-3b (and specifically its inactivation) is a major, required integration point for a multitude of upstream signals acting on an end-effector responsible for cardioprotection (the mitochondrial permeability transition pore). When cell protection signaling pathways are activated, we found that the Bcl-2 family members relay the signal from GSK-3b onto a target at or in close proximity to the pore. Thus, the effect of the convergence of these signaling pathways via inhibition of GSK-3b, relayed through Bcl-2 proteins, on the end effector, the permeability transition pore complex, to limit mPTP induction, is the general mechanism of cardiomyocyte protection. We propose that clinical treatment strategies designed to inhibit the master switch kinase, GSK-3b, to protect the permeability transition pore complex from mPTP induction, would be effective to reduce the size of infarction during episodes of heart attack or stroke by preventing the death of cardiac myocytes and neurons (respectively). Signaling defects underlying the age-assocciated loss of the capacity for ischemic preconditioning are being examined which could lead to testable clinical therapies relevant to the preservation of healthy aging. Experiments are ongoing examining the hypothesis that the sensitivity of mPTP to ROS can serve as a biomarker of mitochondrial fitness during aging. The mitochondrial fitness of muscle stem cells may in part underlie the development of age-related sarcopenia. A healthy and nutritionally well-adjusted lifestyle should favor balanced energy supply and demand in concert with mitochondrial fission-fusion and mitophagy-biogenesis processes leading to proper organelle turnover and energy-redox function of the mitochondrial population. Perturbed fusion-fission dynamics and defective mitophagy result in impaired mitochondrial quality and consequently in spread mitochondrial damage as reflected by energetic-redox impairment. In this scenario damaged mitochondria cannot be removed and accumulate due to failure of the fission process interrupting normal mitophagy. Degradation of mitochondrial quality results in the overall average energetic-redox impairment of the mitochondrial network, as shown by mitochondrial populations isolated from organs affected by metabolic disease or by ablation of fission proteins. Autophagy/mitophagy, and mitochondrial function relate directly to our main new hypothetical idea that citrate/AcCoA cycling controlled by ATP citrate lyase regulates key acetylation/deacetylation processes in the mitochondrial, cytoplasmic and nuclear compartments and thereby has a potentially central and unifying role in the control of aging and longevity. Our results suggest that diverse interventions that have been proven to prolong lifespan in multiple animal experimental models all decrease global acetylation in cardiomyocytes and neurons and positively affect the autophagy outcome. Within the hypothetical framework that the health status of the mitochondrial network affects/governs the health span of cells and the organism, thus influencing longevity (1), we are studying mitochondrial function and metabolic remodeling as a function of age in high and low running capacity (HCR and LCR) rats, obtained by inbreeding from an original NIH outbred colony. HCR and LCR rats exhibit a very significant difference in lifespan that parallels the innate differences in running capacity. The HCR exhibit a 40% longer lifespan than LCR rats, a trait associated with a higher capacity to use lipids than glucose as substrate, under exercising conditions. We hypothesized that the respiratory reserve capacity of the cardiac cell in the longer-living HCR will be higher than in LCR, along with relatively enhanced turnover of mitochondria thus determining improved mitochondrial health. Faster aging due to deficit in mitochondrial energetics and turnover will result in a progressively damaged mitochondrial population. The mitochondrial respiratory reserve (quantified as the difference in respiratory flux under maximal uncoupling minus baseline conditions) of isolated cardiac myocytes from HCR-LCR rats at 5, 17 and 24 months of age, was analyzed under different substrate combinations (glucose, palmitate, or glucose+palmitate) with high throughput Seahorse technology. In the same samples, parallel studies of mitochondrial turnover (autophagy/mitophagy) using laser-scanning fluorescence confocal imaging and electron microscopy were performed. Main results show that lipid (palmitate)-based mitochondrial respiratory reserve is higher in HCR than LCR cardiomyocytes but decreases from 5 to 17 months of age in both groups. However, and unexpectedly, respiratory reserve increased thereafter, always more in HCR than LCR and with palmitate alone or in combination with glucose. Our experimental protocol enabled us to dissect the effect of glucose on respiratory reserve which remained much lower exhibiting a slight decrease over the age range analyzed in both groups. In agreement with our hypothesis, the turnover as well as steady state level of autophagy/mitophagy in cardiomyocytes was higher in the longer-living HCRs. The acetyl-CoA levels were always in the lower, likely permissive, concentration range (< 1mM) for HCR, in agreement with the imaging and electron microscopy studies. Interestingly, mitochondrial energetics in HCR-LCR rats showed a similar pattern of substrate consumption as in cardiomyocytes, further supporting our findings with respiration measurements and analyses.

环境应力会在线粒体上汇聚，该线粒体会触发或抑制细胞死亡。可激发的，可激发的有丝分裂后细胞（例如心脏中的心肌细胞和大脑中的神经元），以响应亚洲性有害压力的响应，从而使机制具有保护，从而可以保护免受随后的侮辱。这些保护机制涉及激活内源性信号传导，该信号传导可以赋予对氧化剂的显着抗性和与缺氧/重氧相关的其他应力（即心脏病发作或中风期间），从而促进了细胞存活的增强能力。但是，上游信号传导机制仍然是主动争论的领域，末端效应子仍未解决。 我们表明，长时间缺氧后的重氧降低了心肌细胞中线粒体通透性过渡（MPTP）的活性氧（ROS-）阈值，并且该细胞的存活与地下细胞体的脱极层的分数陡峭地呈负相关。我们证明，通过不同上游机制作用的各种心脏/神经保护剂都通过限制MPTP诱导来促进细胞存活。我们发现，可以以两种一般方式触发保护，并独立于调节线粒体肿胀，这些线粒体肿胀通过抑制GSK-3B对末端效应器的抑制（渗透性过渡孔复合物）融合，从而防止MPTP。细胞保护表现出记忆（即“预处理”）是由触发的线粒体肿胀引起的（由于涌入和/或保留的K+积累增强而导致底物氧化和ROS产生增强，从而导致PKC的氧化还原激活，从而导致PKC的氧化还原激活，从而通过Ser-3b抑制了GSK-3B（通过Ser-9的磷酸化）。我们得出的结论是，GSK-3B（尤其是其失活的）是主要的，所需的集成点，用于作用于负责心脏保护的最终效应器（线粒体渗透性过渡孔）的众多上游信号。当激活细胞保护信号通路时，我们发现Bcl-2家族成员将信号从GSK-3B传递到孔或近距离接近孔的目标。因此，通过抑制GSK-3B的抑制，通过Bcl-2蛋白继电器对末端效应子（渗透性过渡孔复合物）限制MPTP诱导的融合的影响，是心肌细胞保护的一般机制。 我们建议旨在抑制主开关激酶GSK-3B的临床治疗策略，以保护渗透性过渡孔复合物免受MPTP诱导的影响，将有效地减少心脏病发作或中风期间梗塞的大小，从而通过防止心肌细胞和神经元的死亡（分别可以防止心脏肌细胞和神经元）（分别防止）。 正在检查与年龄相关的缺血性预处理损失背后的信号缺陷，这可能会导致与保存健康衰老有关的可测试临床疗法。 实验正在进行研究以下假设：MPTP对ROS的敏感性可以作为衰老过程中线粒体适应性的生物标志物。肌肉干细胞的线粒体适应性可能部分是与年龄有关的肌肉减少症的发展。 健康且营养良好的生活方式应与线粒体裂变融合和线粒体生物发生过程一致均衡能量供求，从而导致线粒体种群的适当细胞器更换和能量 - 复杂功能。扰动的融合膜动力学和有缺陷的线粒体会导致线粒体质量受损，从而导致线粒体损伤所反映出的散布线粒体损伤。在这种情况下，由于裂变过程中断正常线粒体的失败，无法去除线粒体并积累。线粒体质量的降解会导致线粒体网络的总体平均能量 - 雷克斯损伤，如从由代谢性疾病影响的器官或裂变蛋白消融的器官中分离出的线粒体种群所示。自噬/线粒体和线粒体功能直接与我们的主要新假设观念有关，即由ATP柠檬酸盐裂解酶控制的柠檬酸/ACCOA循环可以调节线粒体细胞质质量和细胞囊腔和核隔室的关键乙酰化/脱乙酰化过程，并具有在中心和统一的控制中。我们的结果表明，已证明在多种动物实验模型中延长寿命的各种干预措施都降低了心肌细胞和神经元中的全球乙酰化，并积极影响自噬结果。 在假设的框架内，线粒体网络的健康状况会影响/控制细胞和有机体的健康跨度，从而影响寿命（1），我们正在研究线粒体功能和代谢重塑作为年龄在高和低跑步能力（HCR和LCR）大鼠中的年龄的函数，并通过Inbrede colony获得了近来的Nih。 HCR和LCR大鼠在寿命上表现出非常显着的差异，与跑步能力的先天差异相同。在运动条件下，HCR比LCR大鼠的寿命比LCR大鼠长40％，该特征与脂质的使用能力高于葡萄糖作为底物。我们假设，较长生命的HCR中心脏细胞的呼吸储备能力将比LCR高，并且线粒体的周转相对增强，从而确定了改善的线粒体健康。由于线粒体能量学和周转率的不足导致的更快的老化将导致线粒体人群逐渐受损。 在5、17和24个月大的HCR-LCR大鼠中分离的心肌细胞的线粒体呼吸疗法（在最大解开量不合偶联下的呼吸通量差异）下，分析了不同的底物组合（葡萄糖，棕榈，棕榈酸盐，棕榈酸盐，或棕榈酸盐+棕榈酸盐），并使用高纤维蛋白透过覆盖型技术分析。在相同的样品中，进行了线粒体周转（自噬/线粒体）的平行研究，使用激光扫描荧光共聚焦成像和电子显微镜进行了平行研究。 主要结果表明，HCR中基于脂质的线粒体呼吸储备高于LCR心肌细胞，但两组均从5至17个月降低到17个月。然而，出乎意料的是，此后呼吸储备增加，在HCR中总是比单独使用棕榈酸盐或与葡萄糖结合使用。我们的实验方案使我们能够剖析葡萄糖对呼吸储备的影响，葡萄糖在两组分析的年龄范围内表现出略有下降。与我们的假设一致，在较长的HCR中，心肌细胞中自噬/线皮的稳态水平较高。与成像和电子显微镜研究一致的乙酰基-COA水平总是在较低的，可能允许的浓度范围（<1mm）中。有趣的是，HCR-LCR大鼠中的线粒体能量表现出与心肌细胞相似的底物消耗模式，进一步通过呼吸测量和分析来支持我们的发现。

项目成果

Blueberry-enriched diet protects rat heart from ischemic damage.

• DOI: 10.1371/journal.pone.0005954
• 发表时间: 2009-06-18
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Ahmet I;Spangler E;Shukitt-Hale B;Juhaszova M;Sollott SJ;Joseph JA;Ingram DK;Talan M
• 通讯作者: Talan M

Paracrine effects of hypoxic fibroblast-derived factors on the MPT-ROS threshold and viability of adult rat cardiac myocytes.

缺氧成纤维细胞衍生因子对成年大鼠心肌细胞 MPT-ROS 阈值和活力的旁分泌作用。

• DOI: 10.1152/ajpheart.91443.2007
• 发表时间: 2008
• 期刊: American journal of physiology. Heart and circulatory physiology
• 作者: Shivakumar,K;Sollott,SJ;Sangeetha,M;Sapna,S;Ziman,B;Wang,S;Lakatta,EG
• 通讯作者: Lakatta,EG

Role of glycogen synthase kinase-3beta in cardioprotection.

• DOI: 10.1161/circresaha.109.197996
• 发表时间: 2009-06-05
• 期刊: Circulation research
• 影响因子: 20.1
• 作者: Juhaszova M;Zorov DB;Yaniv Y;Nuss HB;Wang S;Sollott SJ
• 通讯作者: Sollott SJ

Mitochondrial health, the epigenome and healthspan.

• DOI: 10.1042/cs20160002
• 发表时间: 2016-08-01
• 期刊: Clinical science (London, England : 1979)
• 作者: Aon MA;Cortassa S;Juhaszova M;Sollott SJ
• 通讯作者: Sollott SJ

Analysis of mitochondrial 3D-deformation in cardiomyocytes during active contraction reveals passive structural anisotropy of orthogonal short axes.

对主动收缩期间心肌细胞线粒体 3D 变形的分析揭示了正交短轴的被动结构各向异性。

• DOI: 10.1371/journal.pone.0021985
• 发表时间: 2011
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Yaniv,Yael;Juhaszova,Magdalena;Wang,Su;Fishbein,KennethW;Zorov,DmitryB;Sollott,StevenJ
• 通讯作者: Sollott,StevenJ