Recent Advances in Mitochondrial Proteomics

线粒体是细胞能量代谢、生物合成和细胞死亡的调控中心,其功能异常会导致许多疾病的产生。线粒体蛋白质组学研究为解析线粒体的生理功能、探索线粒体相关疾病的发病机制及促进线粒体为靶向的药物研发提供重要理论依据。本文对线粒体蛋白质组学的研究方法,尤其是近年的技术发展以及线粒体蛋白质组学的性质及其应用进行总结,并对其未来的发展前景和挑战进行展望。Mitochondria play a central role in the regulation of cellular energy metabolism,bio-synthesis and cell death.Dysfunction of mitochondria can lead to many diseases.Mitochondrial proteomics provides important theoretical foundation for a systematic understanding of the biological functions of mitochondria,studying the mechanisms of mitochondria-related diseases,and promoting the research and development of mitochondria-targeting drugs.The methodologies,recent technology development,and characteristics and applications of mitochondrial proteomics were reviewed and the challenges and prospects were also discussed.国家自然科学基金(Nos.91313302;90913015;21225523)资助项目~

High-Throughput Assessment of Mitochondrial Fluorescence Labeling at Single-Particle Level

线粒体是真核细胞能量代谢和信号转导的调控中枢,虽然各种灵敏、特异的线粒体荧光标记技术已经被广泛应用于线粒体研究,但仍缺乏单线粒体水平的荧光染色性能评估方法。基于超高灵敏流式检测技术(High sensitivity flow cytometry,HSFCM)能对单个线粒体进行高灵敏、高通量、多参数定量分析的独特优势,本研究发展了一种单线粒体水平的荧光标记高通量评估方法。将携带靶向线粒体绿色荧光蛋白基因的p Ac GFP1-Mito质粒转染至人宫颈癌He La细胞中,用G418筛选出稳定转染细胞,分别从瞬时转染和稳定转染的细胞中提取线粒体。此外,从未转染质粒的正常He La细胞中提取线粒体,分别进行Mito Tracker Green标记以及SYTO 62线粒体DNA染色,应用实验室自行研制的超高灵敏流式检测装置在单线粒体水平对这4种线粒体标记方法的荧光亮度、标记效率和稳定性进行评估。实验结果表明,稳定转染细胞中单个线粒体的绿色荧光蛋白(GFP)荧光亮度为瞬时转染的17.7倍,比Mito Tracker Green标记的线粒体亮度高约两个数量级,且标记稳定性好。本研究为线粒体标记方法的选择提供了一种先进的分析方法。Mitochondria play a central role in the regulation of energy metabolism and signal transduction in eukaryotic cells. Although many fluorescent labeling strategies have been developed for mitochondrial studies,the methods that enable labeling efficiency assessment at the single-mitochondrion level are still lacking. By employing the unique advantages of high sensitivity flow cytometry( HSFCM) in the sensitive,rapid,and quantitative multiparameter analysis of individual mitochondria,here we examined the performance of several different mitochondrial labeling strategies from the perspectives of brightness,labeling ratio,and stability.Mitochondria isolated from He La cells transfected with p Ac GFP1-Mito plasmid upon transient or stable transfections,and mitochondria directly labeled with Mito Tracker Green or SYTO 62 were analyzed by a laboratory-built three-channel HSFCM. Upon the quantitative measurement of fluorescence brightness,it was found that the fluorescence intensity of green fluorescent protein( GFP) in mitochondria isolated from cells with stable transfection was about 17. 7-fold higher than the transient transfection ones,and was approximately two orders of magnitude brighter than mitochondria labeled with Mito Tracker Green. On the other hand,the fluorescence signal of SYTO 62 labeling decreased upon washing,indicating its rapid dissociation rate. The strong fluorescence intensity and good labeling stability make stable transfection an efficient method to label mitochondria. The experimental results demonstrates that HSFCM provides a powerful analytical tool to assess the performance of mitochondrial fluorescence labeling via high throughput single mitochondria analysis.国家自然科学基金项目(Nos.91313302,90913015,21225523)资助~

Mediating effect of self-control between mindfulness experience and sleep disorder

目的 探讨正念经验与睡眠障碍的作用机制。方法 采用匹兹堡睡眠量表(PSQI)、自我控制问卷(SCS)对860名被试进行线上测量。结果 无正念经验组中睡眠障碍的检出率明显高于有正念经验组。自控力在正念经验与睡眠障碍中起部分中介作用。结论 正念经验与较高睡眠质量相关，正念练习可部分通过提高自控力水平发挥作用。</p