Lizardo Michael M, Sorensen Poul H
Pediatric Oncology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health; BC Cancer Agency, Provincial Health Services Authority.
BC Cancer Agency, Provincial Health Services Authority; Department of Pathology and Laboratory Medicine, University of British Columbia;
J Vis Exp. 2018 Mar 12(133):56332. doi: 10.3791/56332.
The pulmonary metastasis assay (PuMA) is an ex vivo lung explant and closed cell culture system that permits researchers to study the biology of lung colonization in osteosarcoma (OS) by fluorescence microscopy. This article provides a detailed description of the protocol, and discusses examples of obtaining image data on metastatic growth using widefield or confocal fluorescence microscopy platforms. The flexibility of the PuMA model permits researchers to study not only the growth of OS cells in the lung microenvironment, but also to assess the effects of anti-metastatic therapeutics over time. Confocal microscopy allows for unprecedented, high-resolution imaging of OS cell interactions with the lung parenchyma. Moreover, when the PuMA model is combined with fluorescent dyes or fluorescent protein genetic reporters, researchers can study the lung microenvironment, cellular and subcellular structures, gene function, and promoter activity in metastatic OS cells. The PuMA model provides a new tool for osteosarcoma researchers to discover new metastasis biology and assess the activity of novel anti-metastatic, targeted therapies.
肺转移分析（PuMA）是一种离体肺外植体和封闭细胞培养系统，它使研究人员能够通过荧光显微镜研究骨肉瘤（OS）肺定植的生物学特性。本文详细描述了该方案，并讨论了使用宽视野或共聚焦荧光显微镜平台获取转移生长图像数据的示例。PuMA模型的灵活性使研究人员不仅能够研究OS细胞在肺微环境中的生长，还能够评估抗转移治疗随时间的效果。共聚焦显微镜能够对OS细胞与肺实质的相互作用进行前所未有的高分辨率成像。此外，当PuMA模型与荧光染料或荧光蛋白基因报告基因结合时，研究人员可以研究转移OS细胞中的肺微环境、细胞和亚细胞结构、基因功能以及启动子活性。PuMA模型为骨肉瘤研究人员发现新的转移生物学特性和评估新型抗转移靶向治疗的活性提供了一种新工具。
