Inhibition of iodoacetamide and t-butylhydroperoxide toxicity in LLC-PK1 cells by antioxidants: a role for lipid peroxidation in alkylation induced cytotoxicity.
Previously we reported that thiol depletion and lipid peroxidation were associated with the cytotoxicity of nephrotoxic cysteine S-conjugates, a group of toxins which kill LLC-PK1 cells after metabolic activation and covalent binding. To determine if this is a general mechanism of cytotoxicity in these cells, we compared the effect of antioxidants, an iron chelator, and a thiol reducing agent on the toxicity of an alkylating agent, iodoacetamide (IDAM), and an organic peroxidant, t-butylhydroperoxide (TBHP). IDAM or TBHP toxicity was concentration (0.01 to 1.0 mM) and time (1 to 6 h) dependent. Both toxins caused lipid peroxidation which occurred prior to cell death as determined by leakage of lactate dehydrogenase (LDH). The alkylating agent IDAM bound to cellular macromolecules and depleted cellular non-protein thiols almost completely by 1 h, while LDH release occurred first at 2 to 3 h. The toxicity of IDAM and TBHP was inhibited by the antioxidants DPPD, BHA, BHQ, PGA, and BHT and the iron chelator deferoxamine. However, DPPD blocked TBHP- and IDAM-induced lipid peroxidation and toxicity without affecting binding and depletion of cellular nonprotein thiols. Furthermore, the thiol reducing agent dithiothreitol was able to block lipid peroxidation and toxicity. Therefore it is possible that with an alkylating agent, depletion of cellular nonprotein thiols cooperates with covalent binding and contributes to lipid peroxidation and cell death. There appear to be common elements in the toxicity of alkylating agents and organic peroxidants in LLC-PK1 cells.
Chen
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W. Alton Jones Cell Science Center, Lake Placid, New York 12946.
Stevens
J L
JL
D000477:Alkylating Agents / Q000633:toxicity*; D000818:Animals; D000975:Antioxidants; D002470:Cell Survival; D003676:Deferoxamine / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism; D007460:Iodoacetamide / Q000633:toxicity*; D007668:Kidney / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D015227:Lipid Peroxidation; D010545:Peroxides / Q000633:toxicity*; D013552:Swine; D020122:tert-Butylhydroperoxide
D000477:Alkylating Agents; D000975:Antioxidants; D010545:Peroxides; D020122:tert-Butylhydroperoxide; D007770:L-Lactate Dehydrogenase; D003676:Deferoxamine; D004229:Dithiothreitol; D007460:Iodoacetamide