Protection from cytotoxic effects induced by the nitrogen mustard mechlorethamine on human bronchial epithelial cells in vitro.
The present study was undertaken to find potent molecules against the toxicity of nitrogen mustard mechlorethamine (HN2) on respiratory epithelial cells, using a human bronchial epithelial cell line (16HBE14o-) as an in vitro model. The compounds examined included inhibitors of poly(ADP-ribose) polymerase (PARP), sulfhydryl-group donors as nucleophiles, and iron chelators and inhibitors of lipid peroxidation as antioxidants. Their effectiveness was determined upon observance of metabolic dysfunction induced by HN2 following a 4-h exposure, using (3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) reduction and ATP-level assays as indicators. Moreover, the fluorescent probe, monobromobimane (mBBr), and 2',7'-dichlorofluorescin-diacetate (H2DCF-DA) were used to assess intracellular sulfhydryl and peroxide level modifications by flow cytometry, respectively, following a 3-h exposure. At last, cell death was assessed by flow cytometry using the propidium iodide (PI)-dye-exclusion assay following 24-h exposure. PARP inhibitors (niacinamide, 3-aminobenzamide, 6(5H)-phenanthridinone), and two sulfhydryl-group donors (N-acetylcysteine, WR-1065) were found to be effective in preventing HN2-induced metabolic dysfunction when added in immediate or delayed treatment with HN2. Only N-acetylcysteine, however, was found to prevent cell death induced by HN2, though it must be present at the time of the HN2 challenge. Flow cytometric measurements of intracellular sulfhydryl levels strongly suggested that N-acetylcysteine and WR-1065 are preventive in alkylation of cellular compounds, mainly by direct extracellular interaction with HN2. PARP inhibitors prevent secondary deleterious effects induced by HN2, considering metabolism dysfunction as the endpoint. Elsewhere, the oxidative stress appears to be a side effect in HN2 toxicity only upon considering the inefficiency of several antioxidants.
Rappeneau
S
S
Laboratoire de Cytophysiologie et Toxicologie Cellulaire, Université Paris VII-Denis Diderot, France. rappeneau@paris7.jussieu.fr
Baeza-Squiban
A
A
Jeulin
C
C
Marano
F
F
D000255:Adenosine Triphosphate / Q000378:metabolism; D000970:Antineoplastic Agents / Q000633:toxicity*; D001980:Bronchi / Q000166:cytology* / Q000187:drug effects; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects*; D005434:Flow Cytometry; D006801:Humans; D007502:Iron Chelating Agents / Q000494:pharmacology; D015227:Lipid Peroxidation / Q000187:drug effects; D008466:Mechlorethamine / Q000031:analogs & derivatives* / Q000633:toxicity; D018384:Oxidative Stress; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011355:Prodrugs / Q000633:toxicity*; D013438:Sulfhydryl Compounds / Q000378:metabolism; D013439:Sulfhydryl Reagents / Q000633:toxicity; D013454:Sulfoxides / Q000633:toxicity*; D013778:Tetrazolium Salts; D013844:Thiazoles
D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D007502:Iron Chelating Agents; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011355:Prodrugs; D013438:Sulfhydryl Compounds; D013439:Sulfhydryl Reagents; D013454:Sulfoxides; D013778:Tetrazolium Salts; D013844:Thiazoles; C074926:4-(methylsulfinyl)phenyl nitrogen mustard; D008466:Mechlorethamine; D000255:Adenosine Triphosphate; C022616:thiazolyl blue