Department of Physics, ‡Department of Chemistry, and §Grossman Institute of Neuroscience, Quantitative Biology and Human Behavior, University of Chicago , Chicago, Illinois 60637, United States.
Nano Lett. 2018 Feb 14;18(2):1351-1359. doi: 10.1021/acs.nanolett.7b05040. Epub 2018 Jan 17.
We describe a new method to measure viscosity within subcellular organelles of a living cell using nanorheology. We demonstrate proof of concept by measuring viscosity in lysosomes in multiple cell types and disease models. The lysosome is an organelle responsible for the breakdown of complex biomolecules. When different lysosomal proteins are defective, they are unable to break down specific biological substrates, which get stored within the lysosome, causing about 70 fatal diseases called lysosomal storage disorders (LSDs). Although the buildup of storage material is critical to the pathology of these diseases, methods to monitor cargo accumulation in the lysosome are lacking for most LSDs. Using passive particle tracking nanorheology and fluorescence recovery after photobleaching, we report that viscosity in the lysosome increases significantly during cargo accumulation in several LSD models. In a mammalian cell culture model of Niemann Pick C, lysosomal viscosity directly correlates with the levels of accumulated cholesterol. We also observed increased viscosity in diverse LSD models in Caenorhabditis elegans, revealing that lysosomal viscosity is a powerful reporter with which to monitor substrate accumulation in LSDs for new diagnostics or to assay therapeutic efficacy.
我们描述了一种使用纳米流变学测量活细胞亚细胞器内粘度的新方法。我们通过在多种细胞类型和疾病模型中测量溶酶体的粘度来证明其概念验证。溶酶体是负责分解复杂生物分子的细胞器。当不同的溶酶体蛋白出现缺陷时，它们就无法分解特定的生物底物，这些底物在溶酶体中积累，导致大约 70 种致命疾病，称为溶酶体贮积症（LSD）。尽管储存物质的积累对这些疾病的病理至关重要，但对于大多数 LSD 来说，监测溶酶体中货物积累的方法仍然缺乏。我们使用被动粒子跟踪纳米流变学和光漂白后荧光恢复，报告在几种 LSD 模型中，随着货物的积累，溶酶体的粘度显著增加。在尼曼-匹克 C 的哺乳动物细胞培养模型中，溶酶体的粘度与积累的胆固醇水平直接相关。我们还在秀丽隐杆线虫的各种 LSD 模型中观察到了粘度的增加，这表明溶酶体粘度是一种强大的报告器，可以监测 LSD 中底物的积累，用于新的诊断或检测治疗效果。
