Visualizing and Quantifying mRNA Localization at the Invasive Front of 3D Cancer Spheroids.
Visualizing and Quantifying mRNA Localization at the Invasive Front of 3D Cancer Spheroids.
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可视化和定量mRNA定位在3D癌症球体的侵袭性前沿。
DOI：
10.1007/978-1-0716-2887-4_16
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发表时间：
2023
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mRNA在迁移细胞前部的定位是一种广泛使用的机制，其在功能上支持有效的细胞运动。在二维表面上的单细胞中以及在多细胞三维（3D）结构和体内组织中观察到。3D多细胞培养可以揭示细胞外基质和细胞-细胞接触的拓扑结构如何影响亚细胞mRNA分布。在这里，我们描述了一种方法的mRNA成像在一个诱导系统的集体癌细胞入侵。将MDA-MB-231癌细胞球状体包埋在Matrigel中，诱导其侵入，并处理以使具有单分子灵敏度的mRNA成像。使用分析算法来定量和比较在侵入性前导细胞前部的mRNA分布。该方法可以很容易地适应和应用于分析RNA分布在额外的设置，其中细胞沿沿着一个线性轴。
Localization of mRNAs at the front of migrating cells is a widely used mechanism that functionally supports efficient cell movement. It is observed in single cells on two-dimensional surfaces, as well as in multicellular three-dimensional (3D) structures and in tissue in vivo. 3D multicellular cultures can reveal how the topology of the extracellular matrix and cell-cell contacts influence subcellular mRNA distributions. Here we describe a method for mRNA imaging in an inducible system of collective cancer cell invasion. MDA-MB-231 cancer cell spheroids are embedded in Matrigel, induced to invade, and processed to image mRNAs with single-molecule sensitivity. An analysis algorithm is used to quantify and compare mRNA distributions at the front of invasive leader cells. The approach can be easily adapted and applied to analyze RNA distributions in additional settings where cells polarize along a linear axis.