Department of Chemistry, Kenan and Caudill Laboratories , 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. 2018 Oct 16;90(20):11981-11988. doi: 10.1021/acs.analchem.8b02486. Epub 2018 Oct 4.
The health risks associated with acute and prolonged exposure to estrogen receptor (ER) modulators has led to a concerted effort to identify and prioritize potential disruptors present in the environment. ER agonists and antagonists are identified with end-point assays, quantifying changes in cellular proliferation or gene transactivation in monolayers of estrogen receptor alpha expressing (ER+) cells upon exposure. While these monolayer cultures can be prepared, dosed, and analyzed in a highly parallelized manner, they are unable to predict the potencies of ER modulators in vivo accurately. Physiologically relevant model systems that better predict tissue- or organ-level responses are needed. To address this need, we describe here a screening platform capable of quantitatively assessing ER modulators in 96 chemically isolated 3D cultures. These cultures are supported in wax-patterned paper scaffolds whose design has improved performance and throughput over previously described paper-based setups. To highlight the potential of paper-based cultures for toxicity screens, we measured the potency of known ER modulators with a luciferase-based reporter assay. We also quantified the proliferation and invasion of two ER+ cell lines in the presence of estradiol. Despite the inability of the current setup to better predict in vivo potencies of ER modulators than monolayer cultures, the results demonstrate the potential of this platform to support increasingly complex and physiologically relevant tissue-like structures for environmental chemical risk assessment.
与急性和长期暴露于雌激素受体 (ER) 调节剂相关的健康风险促使人们共同努力，以确定和优先考虑环境中存在的潜在干扰物。通过终点测定法识别雌激素受体激动剂和拮抗剂，定量测定暴露于表达雌激素受体 alpha (ER+)细胞单层时细胞增殖或基因转录激活的变化。虽然可以以高度平行的方式制备、给药和分析这些单层培养物，但它们无法准确预测体内 ER 调节剂的效力。需要更能准确预测组织或器官水平反应的生理相关模型系统。为了解决这一需求，我们在这里描述了一种能够在 96 种化学分离的 3D 培养物中定量评估 ER 调节剂的筛选平台。这些培养物在蜡图案化的纸支架上得到支持，其设计提高了以前描述的基于纸的设置的性能和通量。为了突出基于纸的培养物用于毒性筛选的潜力，我们使用基于荧光素酶的报告基因测定法测量了已知 ER 调节剂的效力。我们还定量测量了两种 ER+细胞系在雌二醇存在下的增殖和侵袭。尽管当前设置无法比单层培养物更好地预测 ER 调节剂的体内效力，但结果表明该平台有潜力支持越来越复杂和生理相关的组织样结构，用于环境化学风险评估。
