Abstract
Arsenic is highly toxic to the human bladder. In the present study, we established a human bladder epithelial cell line that closely mimics normal human bladder epithelial cells by immortalizing primary uroplakin 1B-positive human bladder epithelial cells with human telomerase reverse transcriptase (HBladEC-T). The uroplakin 1B-positive human bladder epithelial cell line was then used to evaluate the toxicity of seven arsenicals (iAsV, iAsIII, MMAV, MMAIII, DMAV, DMAIII, and DMMTAV). The cellular uptake and metabolism of each arsenical was different. Trivalent arsenicals and DMMTAV exhibited higher cellular uptake than pentavalent arsenicals. Except for MMAV, arsenicals were transported into cells by aquaglyceroporin 9 (AQP9). In addition to AQP9, DMAIII and DMMTAV were also taken up by glucose transporter 5. Microarray analysis demonstrated that arsenical treatment commonly activated the NRF2-mediated oxidative stress response pathway. ROS production increased with all arsenicals, except for MMAV. The activating transcription factor 3 (ATF3) was commonly upregulated in response to oxidative stress in HBladEC-T cells: ATF3 is an important regulator of necroptosis, which is crucial in arsenical-induced bladder carcinogenesis. Inorganic arsenics induced apoptosis while MMAV and DMAIII induced necroptosis. MMAIII, DMAV, and DMMTAV induced both cell death pathways. In summary, MMAIII exhibited the strongest cytotoxicity, followed by DMMTAV, iAsIII, DMAIII, iAsV, DMAV, and MMAV. The cytotoxicity of the tested arsenicals on HBladEC-T cells correlated with their cellular uptake and ROS generation. The ROS/NRF2/ATF3/CHOP signaling pathway emerged as a common mechanism mediating the cytotoxicity and carcinogenicity of arsenicals in HBladEC-T cells.
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The datasets generated and/or analyzed during the current study are available from the corresponding author on reasonable request.
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Acknowledgements
This work was supported by a grant from the Food Safety Commission, Cabinet Office, Government of Japan (Research Program for Risk Assessment Study on Food Safety, No. JPCAFSC20212102), Health and Labour Sciences Research Grants from the Ministry of Health, Labour and Welfare of Japan (No. 22KD1003), and grants from Japan Society for the Promotion of Science (No. 23K09652). The authors gratefully acknowledge the technical assistance of Rie Onodera, Keiko Sakata, Yuko Hisabayashi, and Yukiko Iura (Department of Molecular Pathology, Osaka Metropolitan University Graduate School of Medicine, Osaka, Japan), and Yoriko Yabunaka, and Yukimi Kira (Research Support Platform, Osaka Metropolitan University Graduate School of Medicine, Osaka, Japan). We would like to thank Dr. Hiroyuki Miyoshi and RIKEN Bioresource center for providing the lentivirus and packaging plasmids.
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Vachiraarunwong, A., Gi, M., Kiyono, T. et al. Characterizing the toxicological responses to inorganic arsenicals and their metabolites in immortalized human bladder epithelial cells. Arch Toxicol (2024). https://doi.org/10.1007/s00204-024-03750-1
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DOI: https://doi.org/10.1007/s00204-024-03750-1