A study of enzymatic activity in cell cultures via the analysis of volatile biomarkers
A study of enzymatic activity in cell cultures via the analysis of volatile biomarkers
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DOI：
10.1039/c2an35815h
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发表时间：
2012-01-01
期刊：
影响因子：
4.2
通讯作者：
Smith, David
中科院分区：
文献类型：
--
作者：
Chippendale, Thomas W. E.;Hu, Bin;Smith, David
醛脱氢酶（ALDH）负责醛类（包括乙醛（AA））的代谢，并与疾病有关。我们描述了一种方法来研究ALDH活性在细胞培养物中涉及AA浓度的测量在气体/蒸汽相。这已经实现了使用选择离子流管质谱法（SIFT-MS），开发用于快速定量的微量气体在潮湿的介质。hepG 2肝细胞癌细胞系的人细胞和原代骨髓源性间充质干细胞（hMSC）从培养基中耗尽AA，但ALDH抑制剂二乙氨基苯甲醛（DEAB）和双硫仑（DSF）的应用抑制了这种耗尽或在某些情况下导致AA浓度升高。此外，细胞显示出将二甲亚砜（DMSO）还原成二甲硫醚，这是由甲硫氨酸亚砜还原酶A（MsrA）酶介导的。有趣的是，这个过程也被DEAB和DSF抑制。本研究的结果表明，SIFT-MS气相分析可以应用于细胞内酶反应的挥发性代谢产物的研究，这在疾病研究和药物发现具有潜在的效用。
Aldehyde dehydrogenase (ALDH) enzymes are responsible for the metabolism of aldehydes, including acetaldehyde (AA), and are linked to disease. We describe a method to study ALDH activity in cell cultures involving the measurement of AA concentrations in the gas/vapour phase. This has been achieved using selected ion flow tube mass spectrometry (SIFT-MS), developed for the rapid quantification of trace gases in humid media. Human cells of the hepG2 hepatocellular carcinoma cell line and primary bone marrow-derived mesenchymal stem cells (hMSCs) depleted AA from the culture media, but the application of ALDH inhibitors diethylaminobenzaldehyde (DEAB) and disulfiram (DSF), suppressed this depletion or in some cases resulted in elevated AA concentrations. Further, the cells were shown to reduce the dimethyl sulphoxide (DMSO) to dimethyl sulphide, which is mediated by methionine sulfoxide reductase A (MsrA) enzymes. Interestingly, this process was also inhibited by DEAB and DSF. The results of this study indicate that SIFT-MS gas phase analysis could be applied to the study of volatile metabolites of intracellular enzyme reactions, this having potential utility in disease research and drug discovery.