Ellis James Keith, Athersuch Toby James, Cavill Rachel, Radford Robert, Slattery Craig, Jennings Paul, McMorrow Tara, Ryan Michael P, Ebbels Timothy Mark David, Keun Hector Charles
Department of Surgery and Cancer, Faculty of Medicine, Imperial College London, Sir Alexander Fleming Building, South Kensington, London SW7 2AZ, UK.
Mol Biosyst. 2011 Jan;7(1):247-57. doi: 10.1039/c0mb00146e. Epub 2010 Nov 19.
Toxicity testing is vital to protect human health from exposure to toxic chemicals in the environment. Furthermore, combining novel cellular models with molecular profiling technologies, such as metabolomics can add new insight into the molecular basis of toxicity and provide a rich source of biomarkers that are urgently required in a 21st Century approach to toxicology. We have used an NMR-based metabolic profiling approach to characterise for the first time the metabolome of the RPTEC/TERT1 cell line, an immortalised non-tumour human renal epithelial cell line that recapitulates phenotypic characteristics that are absent in other in vitro renal cell models. RPTEC/TERT1 cells were cultured with either the dosing vehicle (DMSO) or with exposure to one of six compounds (nifedipine, potassium bromate, monuron, D-mannitol, ochratoxin A and sodium diclofenac), several of which are known to cause renal effects. Aqueous intracellular and culture media metabolites were profiled by (1)H NMR spectroscopy at 6, 24 and 72 hours of exposure to a low effect dose (IC(10)). We defined the metabolome of the RPTEC/TERT1 cell line and used a principal component analysis approach to derive a panel of key metabolites, which were altered by chemical exposure. By considering only major changes (±1.5 fold change from control) across this metabolite panel we were able to show specific alterations to cellular processes associated with chemical treatment. Our findings suggest that metabolic profiling of RPTEC/TERT1 cells can report on the effect of chemical exposure on multiple cellular pathways at low-level exposure, producing different response profiles for the different compounds tested with a greater number of major metabolic effects observed in the toxin treated cells. Importantly, compounds with established links to chronic renal toxicity produced more diverse and severe perturbations to the cellular metabolome than non-toxic compounds in this model. As these changes can be rationalised with the different pharmacological and toxicity profiles of the chemicals it is suggested that metabolic profiling in the RPTEC/TERT1 model would be useful in investigating the mechanism of action of toxins at a low dose.
毒性测试对于保护人类健康免受环境中有毒化学物质的暴露至关重要。此外，将新型细胞模型与分子谱分析技术（如代谢组学）相结合，可以为毒性的分子基础提供新的见解，并提供21世纪毒理学方法中迫切需要的丰富生物标志物来源。我们首次使用基于核磁共振的代谢谱分析方法来表征RPTEC/TERT1细胞系的代谢组，该细胞系是一种永生化的非肿瘤人肾上皮细胞系，概括了其他体外肾细胞模型中不存在的表型特征。RPTEC/TERT1细胞用给药载体（二甲基亚砜）培养，或暴露于六种化合物（硝苯地平、溴酸钾、灭草隆、D-甘露醇、赭曲霉毒素A和双氯芬酸钠）之一，其中几种已知会对肾脏产生影响。在暴露于低效应剂量（IC(10)）6、24和72小时时，通过(1)H核磁共振光谱对细胞内和培养基中的水性代谢物进行谱分析。我们定义了RPTEC/TERT1细胞系的代谢组，并使用主成分分析方法得出一组关键代谢物，这些代谢物因化学暴露而发生改变。通过仅考虑该代谢物组中的主要变化（与对照相比变化±1.5倍），我们能够显示与化学处理相关的细胞过程的特定改变。我们的研究结果表明，RPTEC/TERT1细胞的代谢谱分析可以报告低水平暴露时化学暴露对多种细胞途径的影响，对不同测试化合物产生不同的反应谱，在毒素处理的细胞中观察到更多的主要代谢效应。重要的是，在该模型中，与慢性肾毒性有既定联系的化合物对细胞代谢组产生的扰动比无毒化合物更多样化和更严重。由于这些变化可以根据化学物质的不同药理和毒性谱进行合理解释，因此建议在RPTEC/TERT1模型中进行代谢谱分析有助于研究低剂量毒素的作用机制。
