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Evaluation of the Aryl Hydrocarbon Receptor Response in LMH 3D Spheroids.
Environmental Toxicology and Chemistry ( IF 3.6 ) Pub Date : 2020-05-25 , DOI: 10.1002/etc.4783 Tasnia Sharin 1, 2 , Doug Crump 1 , Jason M O'Brien 1
Environmental Toxicology and Chemistry ( IF 3.6 ) Pub Date : 2020-05-25 , DOI: 10.1002/etc.4783 Tasnia Sharin 1, 2 , Doug Crump 1 , Jason M O'Brien 1
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In the present study, we investigated whether the immortalized chicken hepatocellular carcinoma cell line, leghorn male hepatoma (LMH), had a comparable aryl hydrocarbon receptor (AhR) response to primary chicken embryonic hepatocytes (CEHs) when used in a well‐established assay for chemical screening and prioritization. The LMH cells were grown as 2‐dimensional (2D) confluent cells and 3D spheroids to determine the optimal cell culture states for chemical screening. Cytochrome P450 1A4 and 1A5 (CYP1A) activity and gene expression were compared between CEHs and LMH cells grown in 2 culture states following exposure to the dioxin‐like compound 3,3′,4,4′,5‐pentachlorobiphenyl (PCB‐126). The CYP1A activity was measured using the ethoxyresorufin‐O‐deethylase (EROD) assay, and changes in mRNA expression associated with the AhR pathway were determined using a custom‐designed polymerase chain reaction array. Among LMH cell culture states (i.e., 2D vs 3D), EROD induction was observed only in 3D LMH spheroids. Similarly, 3D spheroids had the greatest number of changes in AhR‐related genes compared with confluent cells. Overall, these results suggest that LMH cells grown as 3D spheroids have a metabolic and gene expression profile that is comparable to that of CEH, and may represent a suitable animal‐free alternative for in vitro screening of chemicals. Environ Toxicol Chem 2020;39:1693–1701. © 2020 SETAC
中文翻译：
在LMH 3D球体中评估芳烃受体的响应。
在本研究中，我们调查了永生化的鸡肝细胞癌细胞系来克霍恩男性肝癌（LMH）对原代鸡胚肝细胞（CEHs）是否具有可比的芳烃受体（AhR）反应。化学筛选和优先级排序。LMH细胞生长为二维（2D）融合细胞和3D球体，以确定用于化学筛选的最佳细胞培养状态。比较暴露于二恶英样化合物3,3'，4,4'，5-五氯联苯（PCB-126）后在2种培养状态下生长的CEH和LMH细胞的细胞色素P450 1A4和1A5（CYP1A）活性和基因表达。CYP1A活性是通过乙氧基异佛光素-O-脱乙基酶（EROD）测定的，并使用定制设计的聚合酶链反应阵列确定了与AhR途径相关的mRNA表达变化。在LMH细胞培养状态（即2D与3D）中，仅在3D LMH球体中观察到EROD诱导。同样，与融合细胞相比，3D球体在AhR相关基因中的变化最多。总体而言，这些结果表明，以3D球体形式生长的LMH细胞具有与CEH相当的代谢和基因表达谱，并且可能代表了一种适合动物的体外筛选化学替代品。2020年《环境毒理学》； 39：1693–1701。©2020 SETAC
更新日期：2020-05-25
中文翻译：
在LMH 3D球体中评估芳烃受体的响应。
在本研究中，我们调查了永生化的鸡肝细胞癌细胞系来克霍恩男性肝癌（LMH）对原代鸡胚肝细胞（CEHs）是否具有可比的芳烃受体（AhR）反应。化学筛选和优先级排序。LMH细胞生长为二维（2D）融合细胞和3D球体，以确定用于化学筛选的最佳细胞培养状态。比较暴露于二恶英样化合物3,3'，4,4'，5-五氯联苯（PCB-126）后在2种培养状态下生长的CEH和LMH细胞的细胞色素P450 1A4和1A5（CYP1A）活性和基因表达。CYP1A活性是通过乙氧基异佛光素-O-脱乙基酶（EROD）测定的，并使用定制设计的聚合酶链反应阵列确定了与AhR途径相关的mRNA表达变化。在LMH细胞培养状态（即2D与3D）中，仅在3D LMH球体中观察到EROD诱导。同样，与融合细胞相比，3D球体在AhR相关基因中的变化最多。总体而言，这些结果表明，以3D球体形式生长的LMH细胞具有与CEH相当的代谢和基因表达谱，并且可能代表了一种适合动物的体外筛选化学替代品。2020年《环境毒理学》； 39：1693–1701。©2020 SETAC
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