Linford Nancy J, Bilgir Ceyda, Ro Jennifer, Pletcher Scott D
Department of Molecular and Integrative Physiology, University of Michigan, Michigan, USA.
J Vis Exp. 2013 Jan 7(71):50068. doi: 10.3791/50068.
Aging is a phenomenon that results in steady physiological deterioration in nearly all organisms in which it has been examined, leading to reduced physical performance and increased risk of disease. Individual aging is manifest at the population level as an increase in age-dependent mortality, which is often measured in the laboratory by observing lifespan in large cohorts of age-matched individuals. Experiments that seek to quantify the extent to which genetic or environmental manipulations impact lifespan in simple model organisms have been remarkably successful for understanding the aspects of aging that are conserved across taxa and for inspiring new strategies for extending lifespan and preventing age-associated disease in mammals. The vinegar fly, Drosophila melanogaster, is an attractive model organism for studying the mechanisms of aging due to its relatively short lifespan, convenient husbandry, and facile genetics. However, demographic measures of aging, including age-specific survival and mortality, are extraordinarily susceptible to even minor variations in experimental design and environment, and the maintenance of strict laboratory practices for the duration of aging experiments is required. These considerations, together with the need to practice careful control of genetic background, are essential for generating robust measurements. Indeed, there are many notable controversies surrounding inference from longevity experiments in yeast, worms, flies and mice that have been traced to environmental or genetic artifacts(1-4). In this protocol, we describe a set of procedures that have been optimized over many years of measuring longevity in Drosophila using laboratory vials. We also describe the use of the dLife software, which was developed by our laboratory and is available for download (http://sitemaker.umich.edu/pletcherlab/software). dLife accelerates throughput and promotes good practices by incorporating optimal experimental design, simplifying fly handling and data collection, and standardizing data analysis. We will also discuss the many potential pitfalls in the design, collection, and interpretation of lifespan data, and we provide steps to avoid these dangers.
衰老现象几乎在所有已被研究的生物体中都会导致生理机能持续衰退，进而致使身体机能下降，患病风险增加。个体衰老在种群层面表现为年龄依赖性死亡率上升，通常在实验室中通过观察大量年龄匹配个体的寿命来衡量这一指标。旨在量化基因或环境操纵对简单模式生物寿命影响程度的实验，在理解不同分类群中保守的衰老方面以及启发延长哺乳动物寿命和预防与年龄相关疾病的新策略方面取得了显著成功。黑腹果蝇作为一种模式生物，因其寿命相对较短、饲养方便且遗传学操作简便，成为研究衰老机制的理想选择。然而，包括特定年龄生存率和死亡率在内的衰老人口统计学指标极易受到实验设计和环境中哪怕微小变化的影响，因此在衰老实验期间需要严格遵循实验室规范。这些因素，再加上对基因背景进行严格控制的需求，对于获得可靠的测量结果至关重要。事实上，围绕酵母、蠕虫、果蝇和小鼠寿命实验的推断存在许多显著争议，这些争议可追溯到环境或基因假象（1 - 4）。在本方案中，我们描述了一套经过多年使用实验室小瓶测量果蝇寿命优化的程序。我们还介绍了dLife软件的使用，该软件由我们实验室开发，可从（http://sitemaker.umich.edu/pletcherlab/software）下载。dLife通过纳入最佳实验设计、简化果蝇处理和数据收集以及标准化数据分析，提高了通量并促进了良好操作规范。我们还将讨论寿命数据设计、收集和解释中的许多潜在陷阱，并提供避免这些风险的步骤。
