Abstract:
Background Benzoapyrene (BaP) can cause respiratory damage in people exposed to it. Previous studies indicate that a 24-hour exposure to BaP and its active metabolites could cause abnormal cell energy metabolism and oxidative damage, leading to reduced cell survival.
Objective This experiment observes the changes of gene expression profile of human bronchial epithelial cells (16HBE) caused by short-term exposure to benzoapyrene diol epoxide (BPDE), an active metabolite of BaP, and to provide evidence for the potential mechanism of acute toxicity of BPDE.
Methods This experiment included four BPDE exposure groups (0.5, 1.0, 2.0, and 4.0 μmol·L-1, respectively), a solvent control group (dimethyl sulfoxide), and a blank control group (normal saline). CCK8 method was used to measure cell survival rate after 24 h treatment. According to the cell survival rate, an exposure group and a solvent control group were selected for transcriptional sequencing (RNA-Seq) using the BGISEQ platform. The DEseq2 package in R software was used to detect the differentially expressed genes (DEGs) in different samples. The phyper function in R software was used to perform Gene Ontology (GO) functional annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of the selected DEGs.
Results The survival rates of 16HBE cells decreased with the increase of BPDE concentration (Z=-7.79, Ptrend < 0.01). The cell survival rates of the 0.5, 1.0, 2.0, and 4.0 μmol·L-1 BPDE groups (94.56%±1.74%, 84.80%±3.19%, 80.08%±1.72%, and 62.72%±1.95%, respectively) were significantly reduced compared with the solvent control group (98.61%±0.61%) and the blank control group (100.00%±0.00%) (P < 0.05). The cells in the 2.0 μmol·L-1 BPDE group and the solvent control group were selected for sequencing, and 191 DEGs satisfying both|log2FC|>1 and P < 0.05 were detected in the exposure group, including 87 genes up-regulated and 104 genes down-regulated. The results of gene co-expression network found that chemokines CXCL6, CXCL1, CXCL3, and CXCL8 had strong correlations with colony stimulating factor CSF2. The GO analysis results showed that these DEGs were mainly involved in the biological process of cell proliferation, and the KEGG pathway analysis results showed that these genes were mainly enriched in the interleukin (IL)-17 signaling pathway and the inflammatory chemokine signaling pathway. Further mRNA analysis results revealed that 670 mRNAs were up-regulated and 878 mRNAs were downregulated. The differentially expressed mRNAs were mainly involved in negative regulation of execution phase of apoptosis, negative regulation of extracellular signal transduction, regulation of signal receptor activity, etc., affecting ferroptosis, regulation of stem cells' pluripotency, lysine degradation, and other signaling pathways.
Conclusion Short-term exposure to BPDE can cause changes in gene expression profile of 16HBE cells, and bioinformatics analysis results suggest that IL-17 signaling pathway and ferroptosis pathway might be involved in the BPDE associated cell injury.