Ensemble Analysis of Angiogenic Growth in Three-Dimensional Microfluidic Cell Cultures
Ensemble Analysis of Angiogenic Growth in Three-Dimensional Microfluidic Cell Cultures
复制标题
DOI：
10.1371/journal.pone.0037333
复制
发表时间：
2012-05-25
期刊：
影响因子：
3.7
通讯作者：
Asada, H. Harry
中科院分区：
文献类型：
--
作者：
Farahat, Waleed A.;Wood, Levi B.;Asada, H. Harry
我们展示了合奏三维细胞培养和定量分析的血管生成生长均匀的内皮细胞单层。我们的方法结合了两个关键要素：微流体检测，可以在常见细胞外条件下并行检测血管生成情况，以及自动图像采集和处理方案，可以高通量、无偏见地量化血管生成生长。由于与现有的三维形态发生测定相比，测定的通量增加，因此可以可靠地估计血管生成生长的统计特性。我们使用该试验来评估血管内皮生长因子（VEGF）和信号脂质鞘氨醇-1-磷酸（S1 P）的联合作用。我们的研究结果表明，S1 P在VEGF梯度存在下放大血管生成反应的重要性。此外，应用S1 P与VEGF梯度导致血管生成芽具有更高的纵横比S1 P与背景水平的VEGF，尽管总迁移活性降低。这意味着生长因子之间在促进血管生成活性方面存在协同效应。最后，发现所计算的血管生成度量的方差（如通过集合标准差测量的）随着集合平均值线性增加。这一发现是一致的随机代理为基础的数学模型的血管生成，代表血管生成的生长作为一系列独立的随机细胞水平的决定。
We demonstrate ensemble three-dimensional cell cultures and quantitative analysis of angiogenic growth from uniform endothelial monolayers. Our approach combines two key elements: a micro-fluidic assay that enables parallelized angiogenic growth instances subject to common extracellular conditions, and an automated image acquisition and processing scheme enabling high-throughput, unbiased quantification of angiogenic growth. Because of the increased throughput of the assay in comparison to existing three-dimensional morphogenic assays, statistical properties of angiogenic growth can be reliably estimated. We used the assay to evaluate the combined effects of vascular endothelial growth factor (VEGF) and the signaling lipid sphingoshine-1-phosphate (S1P). Our results show the importance of S1P in amplifying the angiogenic response in the presence of VEGF gradients. Furthermore, the application of S1P with VEGF gradients resulted in angiogenic sprouts with higher aspect ratio than S1P with background levels of VEGF, despite reduced total migratory activity. This implies a synergistic effect between the growth factors in promoting angiogenic activity. Finally, the variance in the computed angiogenic metrics (as measured by ensemble standard deviation) was found to increase linearly with the ensemble mean. This finding is consistent with stochastic agent-based mathematical models of angiogenesis that represent angiogenic growth as a series of independent stochastic cell-level decisions.