A Comprehensive Approach to Sample Preparation for Electron Microscopy and the Assessment of Mitochondrial Morphology in Tissue and Cultured Cells
A Comprehensive Approach to Sample Preparation for Electron Microscopy and the Assessment of Mitochondrial Morphology in Tissue and Cultured Cells
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DOI：
10.1002/adbi.202200202
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发表时间：
2023-05-04
期刊：
影响因子：
3.7
通讯作者：
Abel, E. Dale
中科院分区：
文献类型：
--
作者：
Hinton Jr, Antentor;Katti, Prasanna;Abel, E. Dale
线粒体对细胞的代谢需求和增加的损伤作出反应，部分是通过动态结构变化，包括分裂（碎片化）、融合（不同线粒体的合并）、自噬降解（线粒体自噬）和与内质网（ER）的生物相互作用。线粒体结构和功能关系的高分辨率研究需要快速保存标本，以减少线粒体结构的定量评估加上技术文物。一个实用的方法，用于评估线粒体的精细结构，使用二维和三维高分辨率电子显微镜，和一个系统的方法来测量线粒体的架构，包括体积，长度，超支，嵴形态，和数量和程度的相互作用与ER描述。这些方法用于评估具有高能量需求的细胞和组织中的线粒体结构，包括骨骼肌细胞、小鼠脑组织和果蝇肌肉。在缺失线粒体动力学相关基因的细胞和组织中验证了评估的准确性。
Mitochondria respond to metabolic demands of the cell and to incremental damage, in part, through dynamic structural changes that include fission (fragmentation), fusion (merging of distinct mitochondria), autophagic degradation (mitophagy), and biogenic interactions with the endoplasmic reticulum (ER). High resolution study of mitochondrial structural and functional relationships requires rapid preservation of specimens to reduce technical artifacts coupled with quantitative assessment of mitochondrial architecture. A practical approach for assessing mitochondrial fine structure using two dimensional and three dimensional high-resolution electron microscopy is presented, and a systematic approach to measure mitochondrial architecture, including volume, length, hyperbranching, cristae morphology, and the number and extent of interaction with the ER is described. These methods are used to assess mitochondrial architecture in cells and tissue with high energy demand, including skeletal muscle cells, mouse brain tissue, and Drosophila muscles. The accuracy of assessment is validated in cells and tissue with deletion of genes involved in mitochondrial dynamics.