Monoethylhexyl phthalate (MEHP) is one of the main active metabolites of the plasticizer di(2-ethylhexyl) phthalate. It has been known that MEHP has an impact on lipolysis; however, its mechanism on the cellular lipid metabolism remains largely unclear. Here, we first utilized global lipid profiling to fully characterize the lipid synthesis and degradation pathways upon MEHP treatment on hepatic cells. Meanwhile, we further identified the possible MEHP-targeted proteins in living cells using the cellular thermal shift assay (CETSA) method. The lipidomics results showed that there was a significant accumulation of fatty acids and other lipids in the cell. The CETSA identified 18 proteins and fatty acid β-oxidation inhibition pathways that were significantly perturbed. MEHP's binding with selected proteins HADH and HSD17B10 was further evaluated using molecule docking, and results showed that MEHP has higher affinities as compared to endogenous substrates, which was further experimentally confirmed in the surface plasma resonance interaction assay. In summary, we found a novel mechanism for MEHP-induced lipid accumulation, which was probably due to its inhibitive effects on the enzymes in fatty acid β-oxidation. This mechanism substantiates the public concerns on the high exposure level to plasticizers and their possible role as an obesogen.
Chemical
proteomics methods have been used as effective tools to
identify novel protein targets for small molecules. These methods
have great potential to be applied as environmental toxicants to figure
out their mode of action. However, these assays usually generate dozens
of possible targets, making it challenging to validate the most important
one. In this study, we have integrated the cellular thermal shift
assay (CETSA), quantitative proteomics, metabolomics, computer-assisted
docking, and target validation methods to uncover the protein targets
of monoethylhexyl phthalate (MEHP). Using the mass spectrometry implementation
of CETSA (MS-CETSA), we have identified 74 possible protein targets
of MEHP. The Gene Ontology (GO) enrichment integration was further
conducted for the target proteins, the cellular dysregulated proteins,
and the metabolites, showing that cell cycle dysregulation could
be one primary change due to the MEHP-induced toxicity. Flow cytometry
analysis confirmed that hepatocytes were arrested at the G1 stage
due to the treatment with MEHP. Subsequently, the potential protein
targets were ranked by their binding energy calculated from the computer-assisted
docking with MEHP. In summary, we have demonstrated the development
of interactomics workflow to simplify the redundant information from
multiomics data and identified novel cell cycle regulatory protein
targets (CPEB4, ANAPC5, and SPOUT1) for MEHP.
Liquid crystal monomers (LCMs) are a large family of artificial ingredients that have been widely used in global liquid crystal display (LCD) industries. As a major constituent in LCDs as well as the end products of e-waste dismantling, LCMs are of growing research interest with regard to their environmental occurrences and biochemical consequences. Many studies have analyzed LCMs in multiple environmental matrices, yet limited research has investigated the toxic effects upon exposure to them. In this study, we combined in silico simulation and in vitro assay validation along with omics integration analysis to achieve a comprehensive toxicity elucidation as well as a systematic mechanism interpretation of LCMs for the first time. Briefly, the high-throughput virtual screen and reporter gene assay revealed that peroxisome proliferatoractivated receptor gamma (PPARγ) was significantly antagonized by certain LCMs. Besides, LCMs induced global metabolome and transcriptome dysregulation in HK2 cells. Notably, fatty acid β-oxidation was conspicuously dysregulated, which might be mediated through multiple pathways (IL-17, TNF, and NF-kB), whereas the activation of AMPK and ligand-dependent PPARγ antagonism may play particularly important parts. This study illustrated LCMs as a potential PPARγ antagonist and explored their toxicological mode of action on the trans-omics level, which provided an insightful overview in future chemical risk assessment.
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