Mechanisms of cytotoxicity induced by halobenzoquinone water disinfection byproducts

• Author / Creator

  Li, Jinhua

• Drinking water disinfection effectively kills pathogens and prevents waterborne diseases, but it also generates a large number of water disinfection byproducts (DBPs). Epidemiological studies have consistently linked long-term consumption of chlorinated water with an increased risk of developing bladder cancer; however, the responsible DBP or DBPs have not been identified. Halobenzoquinones (HBQs) are an emerging class of DBPs that are predicted to be potent bladder carcinogens. However, little is known about the toxicological effects of HBQs. This thesis investigated the cytotoxicity and potential mechanism of toxicity induced by HBQs. The cytotoxicity of HBQs was demonstrated in several cell lines: T24 human bladder carcinoma cells, CHO-K1 Chinese hamster ovary cells, and SV-HUC-1 human uroepithelial cells. At μM levels HBQs induced significantly higher toxicity than the regulated DBPs. HBQs generated significant intracellular reactive oxygen species (ROS), weakened the glutathione (GSH) antioxidant system, and caused severe oxidative DNA and protein damage (i.e., increased cellular level of 8-hydroxydeoxyguanosine (8-OHdG) and protein carbonyls). Additionally, toxicogenomic analysis showed that HBQs activate the nuclear factor E2-related factor 2 (Nrf2) antioxidant pathway; the major transcripts differentially up-regulated at both early and late exposure times were HMOX1, followed by NQO1, PTGS2 and TXNRD1. These results support oxidative stress as a key mechanism of HBQ-induced cytotoxicity. Structure–toxicity relationship analysis demonstrated that formation of ROS, cytotoxicity, and genotoxicity were strongly affected by the structure of HBQs (i.e., position, type, and number of substitutions of halogens on the benzoquinone ring). Halogen substitution groups, isomeric structure, and electron distribution were shown to affect toxicity. The cytotoxicity of di-halogenated HBQs followed the trend: iodo- > bromo- > chloro-HBQs. Additionally, 2,5-HBQs induced greater toxicity than their corresponding 2,6-HBQ isomers. Furthermore, HBQ toxicity was influenced by two major structural descriptors: the lowest unoccupied molecular orbital energy (ELUMO) and dipole moment. An established human embryonic kidney (HEK293) cell model, expressing specific multidrug resistance proteins (MRPs), was used to indirectly examine the role of MRPs on the cellular export of HBQs. Compared to the HEK-empty vector, HEK-MRP4 cells significantly decreased HBQ cytotoxicity and cellular ROS generation, supporting the role of MRP4 in the cellular detoxification of HBQs. The detoxification of HBQs by MRP4 was dependent on GSH. This research provides several original contributions to the field. 1) HBQ DBPs can cause high toxicity in multiple human cell lines, and show significantly higher toxicity than regulated DBPs in CHO cells. 2) The structure–toxicity analysis of HBQs identified ELUMO and dipole moment as a key parameter of HBQ toxicity. 3) Toxicogenomic analysis supports oxidative stress pathways as one of the key mechanisms of toxicity of HBQ DBPs. 4) The first study of MRPs and the toxicity of DBPs demonstrated the role of MRP4 in detoxifying HBQs. These results provide useful insight into the mechanisms of toxic effects of HBQs in in vitro systems.

• Subjects / Keywords

  • halobenzoquinone
  • water disinfection byproducts
  • oxidative stress
  • Structure–toxicity relationship analysis
  • multidrug resistance proteins
  • cytotoxicity

• Graduation date

  Fall 2017

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/R3Q815824

• License

  This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.