Abstract
Aflatoxin B1 (AFB1) is widely distributed in nature and is confirmed to be the most toxic of all the aflatoxins, whose predominant metabolism site is the liver. As a well-studied and vital mode of epigenetic modifications, aberrant methylation of the promoters in eukaryotic cells may cause the silence of essential genes, affecting their related transcriptional pathways and ultimately leading to the development of disease and cancers. This study investigated the mechanisms of AFB1-induced hepatotoxicity in S phase-arrested L02 cells using single-cell RNA-seq and single-cell reduced representation bisulfite sequencing (RRBS). AFB1 induced apoptosis and cell cycle S phase arrest, reduced mitochondrial membrane potential (ΔΨm), and increased reactive oxygen species (ROS) generation, as well as the DNA methylation level. Hepatotoxicity mechanism patterns induced by AFB1 in S phase-arrested L02 cells were revealed by combining single-cell RNA-seq with single-cell RRBS analysis, in which DNA methylation played a role via regulating the gonadotropin-releasing hormone receptor pathway, the Wnt signaling pathway, and the TGF-beta signaling pathway. Moreover, a novel strategy for precision toxicology exploration was obtained, including the selection of target cells, multi-group non-directional sequencing, and pathway analysis.
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Acknowledgments
We sincerely thank Geek Gene Beijing for their technical support. Thank you to Fei Wang from the National Center for Protein Sciences at Peking University for assistance with the cell sorting.
Funding
This work was supported by the National GMO Cultivation Major Project of New Varieties (2018ZX0801102B).
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WX conceived and designed the experiments. BZ and YD performed the experiments and wrote the paper. LZ provided help during experiments. XH and KH provided help during data analysis.
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Zhang, B., Dai, Y., Zhu, L. et al. Single-cell sequencing reveals novel mechanisms of Aflatoxin B1-induced hepatotoxicity in S phase-arrested L02 cells. Cell Biol Toxicol 36, 603–608 (2020). https://doi.org/10.1007/s10565-020-09547-z
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DOI: https://doi.org/10.1007/s10565-020-09547-z