Teoh Jean-Sébastien, Soh Ming S, Byrne Joseph J, Neumann Brent
Neuroscience Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne VIC 3800, Australia.
Neuroscience Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne VIC 3800, Australia.;
J Vis Exp. 2019 Jul 5(149). doi: 10.3791/59978.
Defining the cellular mechanisms underlying disease is essential for the development of novel therapeutics. A strategy frequently used to unravel these mechanisms is to introduce mutations in candidate genes and qualitatively describe changes in the morphology of tissues and cellular organelles. However, qualitative descriptions may not capture subtle phenotypic differences, might misrepresent phenotypic variations across individuals in a population, and are frequently assessed subjectively. Here, quantitative approaches are described to study the morphology of tissues and organelles in the nematode Caenorhabditis elegans using laser scanning confocal microscopy combined with commercially available bio-image processing software. A quantitative analysis of phenotypes affecting synapse integrity (size and integrated fluorescence levels), muscle development (muscle cell size and myosin filament length), and mitochondrial morphology (circularity and size) was performed to understand the effects of genetic mutations on these cellular structures. These quantitative approaches are not limited to the applications described here, as they could readily be used to quantitatively assess the morphology of other tissues and organelles in the nematode, as well as in other model organisms.
确定疾病背后的细胞机制对于新型疗法的开发至关重要。一种常用于揭示这些机制的策略是在候选基因中引入突变，并定性描述组织和细胞器形态的变化。然而，定性描述可能无法捕捉到细微的表型差异，可能会误代表型在群体中个体间的变化，并且经常是主观评估的。在此，描述了定量方法，以使用激光扫描共聚焦显微镜结合市售的生物图像处理软件来研究线虫秀丽隐杆线虫组织和细胞器的形态。对影响突触完整性（大小和积分荧光水平）、肌肉发育（肌肉细胞大小和肌球蛋白丝长度）和线粒体形态（圆形度和大小）的表型进行了定量分析，以了解基因突变对这些细胞结构的影响。这些定量方法并不局限于此处所述的应用，因为它们可以很容易地用于定量评估线虫以及其他模式生物中其他组织和细胞器的形态。
