CeO2 nanoparticles induce pulmonary fibrosis via activating S1P pathway as revealed by metabolomics

CeO 2 nanoparticles (NPs) have been shown to cause lung fibrosis , however, the underlying molecular mechanisms are not well understood. In this study, we have conducted a mass spectrometry-based global metabolomic analysis of human bronchial epithelial BEAS-2B cells treated by CeO 2 NPs with different aspect ratios and assessed their toxicity on the bronchial epithelial cells by various cell-based functional assays. Although CeO 2 NPs at doses ranging from 12.5 μg/mL to 25 μg/mL displayed low cytotoxicity on the bronchial epithelial cells, the metabolomic analysis revealed a number of metabolites in the cellular metabolic pathways of sphingosine-1-phosphate, fatty acid oxidation , inflammation, etc. were significantly altered by CeO 2 NPs, especially those with high aspect ratios. The robustness of metabolomics findings was further validated in mouse models upon acute and chronic exposures to CeO 2 NPs. Mechanistically, CeO 2 NPs upregulated transforming growth factor beta-1 (TGF-β1) levels in BEAS-2B cells in an aspect ratio-dependent manner through enhancing the expression of early growth response protein 1 (EGR-1). In addition, both in vitro and in vivo studies demonstrated that CeO 2 NPs significantly induced the expression of sphingosine kinase 1 (SHPK1), phosphorylated Smad2/3 and lung fibrosis markers. Moreover, targeting SPHK1, TGFβ receptor or Smad3 phosphorylation significantly attenuated the fibrosis-promoting effects of CeO 2 NPs, and SPHK1-S1P pathway exerted a greater effect on the TGF-β1-mediated lung fibrosis compared to the conventional Smad2/3 pathway. Collectively, our studies have identified the metabolomic changes in BEAS-2B cells exposed to CeO 2 NPs with different aspect ratios and revealed the subtle changes in metabolic activities that traditional functional assays might have missed. More importantly, we have discovered a previously unknown molecular mechanism underlying CeO 2 NP-induced lung fibrosis with different aspect ratios, shedding new insights on the environmental hazard potential of CeO 2 NPs. • LC-MS-based metabolomics is a sensitive method for the assessment of nanotoxicity. • In vitro metabolomic analysis may be used for predicting nanotoxicity in vivo . • CeO 2 NPs alter the metabolomes of human BEAS2B cells and promote mouse lung fibrosis in an aspect ratio dependent manner. • CeO 2 NPs promote lung fibrosis through the TGF-β1/SPHK1/S1P pathway in addition to the conventional TGF-β1-SMAD pathway.