An untargeted multi-technique metabolomics approach to studying intracellular metabolites of HepG2 cells exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin

Abstract

BackgroundIn vitro cell systems together with omics methods represent promising alternatives to conventional animal models for toxicity testing. Transcriptomic and proteomic approaches have been widely applied in vitro but relatively few studies have used metabolomics. Therefore, the goal of the present study was to develop an untargeted methodology for performing reproducible metabolomics on in vitro systems. The human liver cell line HepG2, and the well-known hepatotoxic and non-genotoxic carcinogen 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), were used as the in vitro model system and model toxicant, respectively.ResultsThe study focused on the analysis of intracellular metabolites using NMR, LC-MS and GC-MS, with emphasis on the reproducibility and repeatability of the data. State of the art pre-processing and alignment tools and multivariate statistics were used to detect significantly altered levels of metabolites after exposing HepG2 cells to TCDD. Several metabolites identified using databases, literature and LC-nanomate-Orbitrap analysis were affected by the treatment. The observed changes in metabolite levels are discussed in relation to the reported effects of TCDD.ConclusionsUntargeted profiling of the polar and apolar metabolites of in vitro cultured HepG2 cells is a valid approach to studying the effects of TCDD on the cell metabolome. The approach described in this research demonstrates that highly reproducible experiments and correct normalization of the datasets are essential for obtaining reliable results. The effects of TCDD on HepG2 cells reported herein are in agreement with previous studies and serve to validate the procedures used in the present work.

Cytotoxic effects and degradation products of three mycotoxins: Alternariol, 3-acetyl-deoxynivalenol and 15-acetyl-deoxynivalenol in liver hepatocellular carcinoma cells

“…HepG2 cells were chosen for experiments, derive from a human hepatocarcinoma and have preserved the activities of several phase I and phase II enzymes but with activity lower than in normal liver (Knasmüller et al, 1998;Ruiz-Aracama et al, 2011;Levsen et al, 2005). Certain mechanisms which cannot be detected in tests with metabolic incompetent cells can be identified properly in experiments with the HepG2 cells (Knasmüller et al, 1998).…”

The effects of early postnatal exposure to a low dose of decabromodiphenyl ether (BDE-209) on serum metabolites in male mice

“…Decrease in long-chain acylcarnitines was observed following dioxin exposure in an in vitro study (Ruiz-Aracama et al, 2011). A human cohort study found that concentration of long-chain acylcarnitines correlated positively, while short chain acylcarnitines correlated negatively, with serum free thyroxine (FT 4 ) levels (Jourdan et al, 2014).…”

Cross-platform metabolic profiling: application to the aquatic model organism Lymnaea stagnalis

“…However, the application of only one platform would imply the inevitable loss of information in terms of detected metabolites and biological information. As a consequence, cross-platform approaches in metabolomics have been applied and the recent trend is to move toward the combination of more than one analytical platform [25,26]. Nevertheless, the scientific literature is limited in studies that compare cross-platform methodologies.…”