Protein alkylation, transcriptional responses and cytochrome c release during acrolein toxicity in A549 cells: influence of nucleophilic culture media constituents.
Acrolein is a toxic combustion product that elicits apoptotic and/or necrotic cell death depending on the conditions under which exposure occurs. As a strong electrophile, side-reactions with nucleophilic media constituents seem likely to accompany study of its toxicity in vitro, but these reactions are poorly characterized. We have thus examined the effect of media composition on the toxicity of acrolein in A549 cells. Cells were exposed to acrolein in either Dulbecco's buffered saline (DBS) or F12 supplemented with various concentrations of fetal bovine serum. Cell viability was assessed using the MTT assay, while heme oxygenase-1 (HO-1) and cytoplasmic cytochrome c were measured as respective markers of transcriptional response and apoptosis. Protein damage was evaluated using the protein carbonyl assay. Compared to F12 media (with or without serum), maximal cell death as evaluated using the MTT assay, as well as adduction of intracellular proteins, occurred when cells were exposed to acrolein in DBS. In contrast, cytochrome c release was maximal in cells exposed to acrolein in serum-containing F12, conditions which inhibited protein modification and overt cell death. These findings highlight the need for careful attention to experimental conditions when conducting in vitro toxicological studies of reactive substances.
Thompson
Colin A
CA
Pharmacology and Anaesthesiology Unit, School of Medicine and Pharmacology, The University of Western Australia, QEII Medical Centre, M Block, Nedlands, Perth WA 6009, Australia. colin.thompson@uwa.edu.au <colin.thompson@uwa.edu.au>
Burcham
Philip C
PC
D000171:Acrolein / Q000633:toxicity*; D000230:Adenocarcinoma / Q000378:metabolism / Q000473:pathology; D000478:Alkylation / Q000187:drug effects; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002417:Cattle; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D003470:Culture Media / Q000737:chemistry*; D045304:Cytochromes c / Q000187:drug effects* / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D051547:Heme Oxygenase-1 / Q000378:metabolism; D006801:Humans; D050050:Protein Carbonylation / Q000187:drug effects; D044967:Serum / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects*
D003470:Culture Media; D000171:Acrolein; D045304:Cytochromes c; D051547:Heme Oxygenase-1