Toxicity of smoke extracts towards A549 lung cells: role of acrolein and suppression by carbonyl scavengers.
The noxious 3-carbon electrophile acrolein forms on combustion of diverse organic matter including synthetic polymers such as polyethylene. While known to play a key role in smoke inhalation injury (SII), the molecular basis for the pulmonary toxicity of high dose acrolein-containing smoke is unclear. As a result, drug interventions in SII are poorly directed against pathogenetic smoke toxicants such as acrolein. The first aim of this study was to confirm a role for acrolein in the acute toxicity of smoke extracts towards A549 lung cells by monitoring adduction of known acrolein targets and the expression of acrolein-inducible genes. A second aim was to evaluate carbonyl scavengers for their abilities to protect cell targets and block smoke extract toxicity. Extracts were prepared by bubbling smoke released by smouldering polyethylene through a buffered saline-trap. Acrolein levels in the extracts were estimated via HPLC after derivatisation with 2,4-dinitrophenylhydrazine. Extracts were highly toxic towards A549 cells, eliciting greater ATP depletion than an equivalent concentration of acrolein alone. The toxicity was accompanied by pronounced carbonylation of several cytoskeletal targets, namely vimentin and keratins-7, -8 and -18. Western blotting revealed that polyethylene combustion products also upregulated several acrolein-responsive protein markers, including GADD45beta, NQO1, HMOX, Hsp70, Nur77 and Egr1. Several carbonyl scavengers (bisulfite, d-penicillamine, hydralazine and 1-hydrazinoisoquinoline) strongly attenuated smoke extract toxicity, with bisulfite suppressing both the adduction and cross-linking of intermediate filament targets. Bisulfite also suppressed the cytotoxicity of smoke extracts when detected using real-time monitoring of cellular impedance. These findings confirm a key role for acrolein in smoke cytotoxicity and suggest drugs that block acrolein toxicity deserve further investigation as possible interventions against SII.
Burcham
Philip C
PC
Pharmacology and Anaesthesiology Unit, School of Medicine and Pharmacology, The University of Western Australia, Nedlands, WA 6009, Australia. philip.burcham@uwa.edu.au
Raso
Albert
A
Thompson
Colin A
CA
D000171:Acrolein / Q000633:toxicity*; D000255:Adenosine Triphosphate / Q000378:metabolism; D045744:Cell Line, Tumor; D051766:Early Growth Response Protein 1 / Q000378:metabolism; D016166:Free Radical Scavengers / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D051547:Heme Oxygenase-1 / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D053538:Keratin-18 / Q000378:metabolism; D053552:Keratin-7 / Q000378:metabolism; D053533:Keratin-8 / Q000378:metabolism; D008175:Lung Neoplasms; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000378:metabolism; D057105:Nuclear Receptor Subfamily 4, Group A, Member 1 / Q000378:metabolism; D020959:Polyethylene / Q000633:toxicity; D050050:Protein Carbonylation / Q000187:drug effects; D012906:Smoke; D014746:Vimentin / Q000378:metabolism; D000097642:GADD45 Proteins
D000171:Acrolein; D000255:Adenosine Triphosphate; D051766:Early Growth Response Protein 1; D016166:Free Radical Scavengers; D018840:HSP70 Heat-Shock Proteins; D051547:Heme Oxygenase-1; D047908:Intracellular Signaling Peptides and Proteins; D053538:Keratin-18; D053552:Keratin-7; D053533:Keratin-8; D016660:NAD(P)H Dehydrogenase (Quinone); D057105:Nuclear Receptor Subfamily 4, Group A, Member 1; D020959:Polyethylene; D012906:Smoke; D014746:Vimentin; D000097642:GADD45 Proteins; C497325:EGR1 protein, human; C539821:NR4A1 protein, human; C447479:NQO1 protein, human