title: Comparison of three different cell viability assays for evaluation of vanadyl sulphate cytotoxicity in a Chinese hamster ovary K1 cell line.
abstract: Previously, evaluation of sodium metavanadate (NaVO3) cytotoxicity after 24 h exposure of Chinese hamster ovary K1 (CHO-K1) cells revealed different sensitivity of the in vitro assays used starting from the neutral red (NR, 3-amino-7-dimethylamino-2-methylphenazine hydrochloride) test (detecting lysosomal and possibly the Golgi apparatus damage) as the most sensitive followed by the 2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxyanilide inner salt (XTT) and resazurin (7-hydroxy-3H-phenoxazin-3-one-10-oxide) tests (mitochondrial disruption). The trypan blue (TB) staining (plasma membrane permeability) showed cytotoxicity of NaVO3 at a much higher NaVO3 concentration than the above-mentioned assays. In the current study, using the same experimental approach, we have assessed the toxicity of vanadyl sulphate (VOSO4) and compared the obtained results with NaVO3 action. Unlike metavanadate, VOSO4 treatment at 24 h resulted in similar sensitivity of the NR and resazurin tests. Nevertheless, following the 48-h incubation with VOSO4, the NR test showed markedly higher sensitivity than the resazurin test when comparing the half maximal inhibitory concentration values (61 and 110 µM for the NR and resazurin test, respectively, p < 0.05). The TB staining method was the least susceptible for detecting vanadyl cytotoxicity at each exposure time point. In summary, both the NR and resazurin tests can be advocated as similarly sensitive in detection of VOSO4-induced cytotoxicity in the CHO-K1 cell line at 24 h. However, the longer incubation time with VOSO4 showed that the NR test is more sensitive than the resazurin assay. The differences in the results between the cytotoxicity tests employed probably arise from dissimilar susceptibility of the endpoints (targets) measured with these tests to the damage by vanadium. Considering this, the current and the previous studies highlight the role of lysosomes (and possibly the Golgi apparatus) apart from mitochondria in the toxicity mechanism induced by inorganic vanadium in mammalian cells.
authors.affiliation: Department of Cell Biology, Institute of Environmental Protection, The John Paul II Catholic University of Lublin, Poland iglina@kul.lublin.pl.
authors.forename: Iwona
authors.initials: I
authors.lastname: Zwolak
mesh_terms: D000818:Animals; D001681:Biological Assay; D016466:CHO Cells; D002470:Cell Survival / Q000187:drug effects*; D006224:Cricetinae; D003412:Cricetulus; D006056:Golgi Apparatus / Q000187:drug effects; D020128:Inhibitory Concentration 50; D008247:Lysosomes / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects; D009499:Neutral Red / Q000737:chemistry; D010078:Oxazines / Q000633:toxicity; D012680:Sensitivity and Specificity; D013778:Tetrazolium Salts / Q000633:toxicity; D018675:Toxicity Tests / Q000379:methods*; D014638:Vanadates / Q000633:toxicity; D017968:Vanadium Compounds / Q000633:toxicity*; D014966:Xanthenes / Q000633:toxicity
chemical_list: D010078:Oxazines; D013778:Tetrazolium Salts; D017968:Vanadium Compounds; D014966:Xanthenes; C059087:2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-((phenylamino)carbonyl)-2H-tetrazolium hydroxide; C005843:resazurin; D009499:Neutral Red; D014638:Vanadates; C034028:vanadyl sulfate
keywords: CHO-K1 cells; neutral red; resazurin; toxicity mechanism; trypan blue; vanadium