Identification of Human Lung Epithelial Metabolic Responses to Acute Benzo(a)pyrene Exposure

Abstract

Polycyclic aromatic hydrocarbons (PAH) are a family of compounds commonly produced in the burning of wood, fuels, or grilled meats. Several PAH members, such as benzo(a)pyrene (BaP) have been classified as carcinogenic; however, other risk factors include both asthma and inflammation. The concentration of BaP in plasma has been found to be in nanomolar to micromolar range based on exposure. Previous studies by our laboratory found alterations to the following metabolic pathways in the sera of individuals with elevated benzo(a)pyrene: linoleate, drug, among others. The purpose of this study was to evaluate the effects of the PAH’s, benzo(a)pyrene (BaP) on metabolism and cellular function in A549 lung epithelial cells. The A549 cells were grown until 90% confluency in complete culture media (10% FBS) and then incubated in low serum media (0.5% FBS) overnight before treatment with vehicle or 3 µM BaP for 24h. Treatment at these concentrations were calculated to be approximately 3 times higher than the highest concentration observed in human plasma. The cells were then lysed in 150 μL extraction buffer including 14 internal standards and extracted samples were analyzed by a Q-Exactive mass spectrometer by following the workflow for high-resolution metabolomics as previously reported. The results show that of 8532 m/z features that were detected, 621 metabolic features were significantly different (p ≤ 0.05) after BaP treatment compared to control. Using pathway enrichment analyses, 22 metabolic pathways were significantly affected by BaP, including amino acid, urea cycle, and fatty acid metabolisms, several of which were observed in the human sera samples. Subsequent time course studies (0-24h) were performed using the median physiological concentration of BaP (14.2 nM) detected from the human study were performed as described above. Significant alterations to amino acid metabolism, such as changes to tyrosine, glutamine and methionine were found using high resolution metabolomics analysis. Taken together, these results suggest a novel mechanism of acute BaP exposure to A549 cells by eliciting metabolic perturbations on critical metabolic pathways for proper lung physiology and function.