Department of Chemistry, University of North Carolina at Chapel Hill, 125 South Road, Chapel Hill, North Carolina 27599-3290, United States.
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, 450 West Drive, Chapel Hill, North Carolina 27599-7295, United States.
Anal Chem. 2023 Feb 7;95(5):3054-3061. doi: 10.1021/acs.analchem.2c05201. Epub 2023 Jan 26.
Cellular invasion is the gateway to metastasis, with cells moving from a primary tumor into neighboring regions of healthy tissue. Invasion assays provide a tractable experimental platform to quantitatively assess cellular movement in the presence of potential chemokines or inhibitors. Many such assays involve cellular movement from high cell densities to cell-free regions. To improve the physiological relevance of such assays, we developed an assay format to track cellular movement throughout a uniform density of cells. This assay format imparts diffusion-dominated environments along the channel, resulting in oxygen and nutrient gradients found in spheroids or poorly vascularized tumors. By incorporating oxygen- and pH-sensing films, we quantified spatial and temporal changes in the extracellular environment while simultaneously tracking the movement of a subset of cells engineered to express fluorescent proteins constitutively. Our results show the successful invasion into neighboring tissues likely arises from a small population with a highly invasive phenotype. These highly invasive cells continued to move throughout the 48 h experiment, suggesting they have stem-like or persister properties. Surprisingly, the distance these persister cells invaded was unaffected by the density of cells in the channel or the presence or absence of an oxygen gradient. While these datasets cannot determine if the invasive cells are inherent to the population or if diffusion-dominated environments promote them, they highlight the need for further study.
细胞侵袭是转移的门户，细胞从原发性肿瘤移动到健康组织的邻近区域。侵袭测定提供了一个可行的实验平台，可以定量评估在潜在趋化因子或抑制剂存在的情况下细胞的运动。许多这样的测定涉及细胞从高细胞密度向无细胞区域的运动。为了提高这些测定的生理相关性，我们开发了一种测定格式来跟踪整个细胞均匀密度中的细胞运动。这种测定格式沿着通道赋予扩散主导的环境，导致在球体或血管不良的肿瘤中发现的氧和营养物质梯度。通过结合氧和 pH 感应膜，我们量化了细胞外环境的时空变化，同时同时跟踪表达荧光蛋白的一组细胞的运动，这些细胞被设计为持续表达。我们的结果表明，侵入邻近组织的成功可能源于具有高度侵袭表型的一小部分细胞。这些高侵袭性细胞在 48 小时的实验中持续移动，表明它们具有干细胞样或持久细胞特性。令人惊讶的是，这些持久性细胞侵入的距离不受通道中细胞密度或氧梯度的存在与否的影响。虽然这些数据集不能确定侵袭性细胞是否是群体固有的，或者扩散主导的环境是否促进了它们，但它们强调了进一步研究的必要性。
