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.

环境压力集中在线粒体上，可以触发或抑制细胞死亡。可兴奋的有丝分裂后细胞（例如心脏中的心肌细胞和大脑中的神经元）响应亚致死性有害应激，参与提供保护免受后续损伤的机制。这些保护机制涉及内源性信号传导的激活，可以对氧化剂和与缺氧/复氧（即心脏病发作或中风期间）相关的其他应激产生显着的抵抗力，从而促进细胞生存能力的增强。然而，上游信号机制仍然是一个活跃争论的领域，而末端执行器仍未得到解决。 我们发现，长时间缺氧后的复氧会降低心肌细胞中线粒体通透性转变（mPTP）的活性氧（ROS-）阈值，并且细胞存活率与去极化线粒体的比例呈显着负相关。我们证明，通过不同上游机制发挥作用的多种心脏/神经保护剂均通过限制 mPTP 诱导来促进细胞存活。我们发现，可以通过两种依赖和独立于调节性线粒体肿胀的一般方式触发保护，这些调节性线粒体肿胀通过抑制末端效应器（渗透性过渡孔复合物）上的 GSK-3b 来收敛，从而防止 mPTP。表现出记忆（即“预处理”）的细胞保护是由触发的线粒体肿胀（由于通过流入和/或保留增强 K+ 积累）引起的，导致底物氧化和 ROS 产生增强，导致 PKC 的氧化还原激活，进而抑制 GSK- 3b（通过 Ser-9 磷酸化）。我们得出的结论是，GSK-3b（特别是其失活）是作用于负责心脏保护的末端效应器（线粒体通透性过渡孔）的多个上游信号的主要必需整合点。当细胞保护信号通路被激活时，我们发现 Bcl-2 家族成员将来自 GSK-3b 的信号传递到孔处或附近的靶标上。因此，通过抑制 GSK-3b（通过 Bcl-2 蛋白传递），这些信号通路的汇聚对末端效应器（通透性转换孔复合物）的影响，以限制 mPTP 诱导，是心肌细胞保护的一般机制。 我们提出，旨在抑制主开关激酶 GSK-3b 的临床治疗策略，以保护通透性转换孔复合物免受 mPTP 诱导，可通过预防患者死亡，有效减少心脏病发作或中风发作期间的梗塞范围。心肌细胞和神经元（分别）。 正在检查与年龄相关的缺血预处理能力丧失背后的信号缺陷，这可能会导致与保持健康衰老相关的可测试的临床疗法。 实验正在进行中，以验证 mPTP 对 ROS 的敏感性可以作为衰老过程中线粒体健康度的生物标志物的假设。肌肉干细胞的线粒体适应性可能在一定程度上是与年龄相关的肌少症发生的基础。 健康且营养调整良好的生活方式应有利于平衡的能量供应和需求，与线粒体裂变融合和线粒体自噬生物发生过程相一致，从而导致线粒体群体适当的细胞器周转和能量氧化还原功能。融合裂变动力学扰动和线粒体自噬缺陷导致线粒体质量受损，从而导致线粒体损伤扩散，如能量氧化还原损伤所反映的那样。在这种情况下，由于裂变过程的失败中断了正常的线粒体自噬，受损的线粒体无法被去除和积累。线粒体质量的下降会导致线粒体网络的总体平均能量氧化还原损伤，如从受代谢疾病或裂变蛋白消融影响的器官中分离出的线粒体群体所示。自噬/线粒体自噬和线粒体功能与我们的主要新假设观点直接相关，即由 ATP 柠檬酸裂合酶控制的柠檬酸/AcCoA 循环调节线粒体、细胞质和核区室中的关键乙酰化/脱乙酰化过程，从而在线粒体、细胞质和核区室中具有潜在的核心和统一作用。控制衰老和长寿。我们的结果表明，在多种动物实验模型中已被证明可以延长寿命的多种干预措施都会减少心肌细胞和神经元的整体乙酰化，并对自噬结果产生积极影响。 在线粒体网络的健康状况影响/控制细胞和有机体的健康跨度，从而影响寿命的假设框架内 (1)，我们正在研究线粒体功能和代谢重塑作为高和低跑步能力中年龄的函数（HCR 和 LCR）大鼠，通过近亲繁殖从原始 NIH 远交群体获得。 HCR 和 LCR 大鼠在寿命方面表现出非常显着的差异，这与跑步能力的先天差异相似。 HCR 大鼠的寿命比 LCR 大鼠长 40%，这一特征与在运动条件下使用脂质而不是葡萄糖作为底物的能力有关。我们假设，寿命较长的 HCR 中心肌细胞的呼吸储备能力将高于 LCR 中的心肌细胞，同时线粒体的周转相对增强，从而决定线粒体健康状况的改善。由于线粒体能量和周转不足而导致的更快衰老将导致线粒体群体逐渐受损。 在不同底物组合（葡萄糖、棕榈酸酯或葡萄糖+棕榈酸酯）采用高通量 Seahorse 技术。在同一样本中，使用激光扫描荧光共聚焦成像和电子显微镜对线粒体周转（自噬/线粒体自噬）进行平行研究。 主要结果表明，HCR 心肌细胞中基于脂质（棕榈酸酯）的线粒体呼吸储备高于 LCR 心肌细胞，但两组中从 5 月龄到 17 月龄均有所下降。然而，出人意料的是，呼吸储备此后增加，HCR 中的呼吸储备总是比 LCR 中的增加以及单独使用棕榈酸酯或与葡萄糖组合时增加。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，呼吸储备仍然低得多，在两组中分析的年龄范围内表现出轻微下降。与我们的假设一致，寿命较长的 HCR 中心肌细胞自噬/线粒体自噬的周转率和稳态水平较高。乙酰辅酶 A 水平始终处于 HCR 较低的、可能允许的浓度范围内 (< 1mM)，这与成像和电子显微镜研究一致。有趣的是，HCR-LCR 大鼠的线粒体能量学显示出与心肌细胞相似的底物消耗模式，通过呼吸测量和分析进一步支持了我们的发现。

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