Szatmary A C, Nossal R
Eunice Kennedy Shriver National Institute of Child Health, and Human Development, National Institutes of Health, Bethesda, Maryland 20892, USA; Department of Mechanical Engineering, Bucknell University, 1 Dent Drive, Lewisburg, PA 17837, USA.
Eunice Kennedy Shriver National Institute of Child Health, and Human Development, National Institutes of Health, Bethesda, Maryland 20892, USA.
J Theor Biol. 2017 Jul 21;425:103-112. doi: 10.1016/j.jtbi.2017.05.014. Epub 2017 May 10.
Chemotaxis, the motion of cells directed by a gradient of chemoattractant molecules, guides cells in immune response, development, wound healing, and cancer. Unfortunately, this process is difficult to distinguish from chemokinesis, i.e., stimulated random cell motion. Chemotaxis is frequently inferred by determining how many cells cross a boundary in a chemotaxis assay, for example how many cells crawl into a chemoattractant-infused filter, or how many cells enter a defined region in an under-agarose assay or agarose spot assay. To mitigate possible ambiguity in whether motion observed in these assays is directed by the chemoattractant gradient or by chemokinesis, we developed a mathematical model to determine when such methods indeed indicate directed motion of cells. In contrast to previous analyses of chemotaxis assays, we report not just the gradients that arise in the assays but also resulting cell motion. We applied the model to data obtained from rigorous measurements and show, as examples, that MDA-MB-231 breast-cancer cells are at least 20 times less sensitive to gradients of EGF or CXCL12 than neutrophils are to formyl peptides; we then used this information to determine the extent to which gradient sensing increases the rate of boundary crossing relative to a random-motility control. Results show, for example, that in the filter assay, 2-4 times as many neutrophils pass through the filter when exposed to a gradient as when the gradient is absent. However, in the other combinations of cells and assays we considered, only 10-20% more cells are counted as having migrated in a directed, rather than random, motility condition. We also discuss the design of appropriate controls for these assays, which is difficult for the under-agarose and agarose spot assays. Moreover, although straightforward to perform with the filter assay, reliable controls are often not done. Consequently, we infer that chemotaxis is frequently over-reported, especially for cells like MDA-MB-231 cells, which move slowly and are relatively insensitive to gradients. Such results provide insights into the use of chemotaxis assays, particularly if one wants to acquire and analyze quantitative data.
趋化作用，即细胞在趋化性吸引分子梯度的引导下运动，在免疫反应、发育、伤口愈合和癌症中引导细胞。不幸的是，这个过程很难与化学增活现象区分开来，即受刺激的随机细胞运动。趋化作用通常是通过在趋化性实验中确定有多少细胞穿过边界来推断的，例如有多少细胞爬入注入趋化性吸引剂的滤膜，或者有多少细胞在琼脂糖下层实验或琼脂糖斑点实验中进入一个确定的区域。为了减轻在这些实验中观察到的运动是由趋化性吸引剂梯度还是由化学增活现象所引导的可能的模糊性，我们开发了一个数学模型来确定这些方法何时确实表明细胞的定向运动。与之前对趋化性实验的分析不同，我们不仅报告了实验中出现的梯度，还报告了由此产生的细胞运动。我们将该模型应用于从严格测量中获得的数据，并举例表明，MDA-MB-231乳腺癌细胞对表皮生长因子（EGF）或CXC趋化因子配体12（CXCL12）梯度的敏感性至少比中性粒细胞对甲酰肽的敏感性低20倍；然后我们利用这些信息来确定相对于随机运动对照，梯度感知增加边界穿越速率的程度。结果表明，例如，在滤膜实验中，当暴露于梯度时，穿过滤膜的中性粒细胞数量是没有梯度时的2-4倍。然而，在我们考虑的细胞和实验的其他组合中，只有10-20%更多的细胞被计数为在定向而非随机运动条件下迁移。我们还讨论了这些实验的适当对照的设计，这对于琼脂糖下层实验和琼脂糖斑点实验来说是困难的。此外，尽管滤膜实验很容易进行，但可靠的对照往往没有做。因此，我们推断趋化作用经常被过度报告，特别是对于像MDA-MB-231细胞这样运动缓慢且对梯度相对不敏感的细胞。这些结果为趋化性实验的使用提供了见解，特别是如果有人想要获取和分析定量数据的话。
