In Vitro Setting of Dose–effect Relationships of 32 Metal Compounds in the Balb/3T3 Cell Line, as a Basis for Predicting their Carcinogenic Potential

Abstract

The results are reported of the second stage in a programme for a systematic in vitro study on the carcinogenic potential of metal compounds with Balb/3T3 clone A31-1-1 mouse fibroblasts. Nineteen metal compounds that exhibited a strong cytotoxic effect during a previous screening run with a 100µM fixed dose were tested with a 72-hour exposure over a wide range of concentrations (from 0.1µM to 1000µM), to produce dose-effect curves to permit extrapolation of the 50% inhibition concentration (IC50) values for each metal compound. This allows the establishment of a suitable range of doses for individual metal species, for use in the subsequent Balb/3T3 assay based on a two-stage concurrent cytotoxicity and morphological transformation protocol. Another 13 metal compounds were also tested, to determine whether the Balb/3T3 cell transformation assay really is a valuable in vitro model in relation to the problem of metal speciation. Of the metal compounds assayed, 26 showed a dose related cytotoxic response with calculated IC50 values ranging from 0.25µM (CH 3 HgCl) to 140µM [(C 5 H 5) 2 TiCl 2 ], whereas six metal compounds, namely (NH 4) 6 Mo 7 O 24 •4H 2 O, CH 3 AsO(OH) 2 , (CH 3) 2 AsNaO 2 ⋅3H 2 O, KBr, CrCl 3 •6H 2 O and (NH 4) 2 [TiO(C 2 O 4) 2 ]⋅H 2 O, displayed no observable cytotoxicity or low cytotoxicity at all the doses tested. The determination of IC50 values permits a ranking of the cytotoxicity responses of metal compounds with the highest cytotoxicities. Dose-effect curves and IC50 values of different chemical forms of individual metal compounds of As, Br, Cr, Hg, Ir, Pt, Te, Ti and V (cationic/anionic inorganic or organometallic species) showed clearly how the chemical nature of the metal strongly influences the toxic response. This confirms that the Balb/3T3 cell line is a valuable in vitro model with respect to the problem of metal speciation. This is a fundamental aspect to be considered when incorporating the results from in vitro cell transformation assays of the carcinogenic potential of metal compounds into regulatory testing schemes. In this context, the choice of test metal species for the development and validation of such assays cannot disregard the possibility that humans will be exposed to specific chemical forms of individual metal compounds (different oxidation states, and inorganic or organometallic natures) that can profoundly affect their toxicity.

Mutagenicity, Carcinogenicity, and Teratogenicity

“…Recently, in-vitro transformation of SHE cells into a neoplastic state was shown to be associated with overexpression of c-myc and c-Ha-ras oncogenes . The Balb3T3 mouse cell line which has also been a classical model for cell transformation has recently been used for the assessment of cytotoxicity of a large series of metal compounds in order to predict their carcinogenic potential (Mazzotti et al 2001(Mazzotti et al , 2002.…”

The Mutagenicity and Carcinogenicity of Inorganic Manganese Compounds: A Synthesis of The Evidence

“…Assessment of mechanistic data on the neurotoxic effects of Mn has implicated mitochondrial disruption, cytotoxicity, and oxidative stress as leading to neurodegeneration in Mazzotti et al 2002;Pascal and Tessier 2004;Takano et al 2002;Tessier and Pascal 2006;Tsui et al 2006). While not directly related to genotoxicity, this information provides useful insight into the molecular interactions of Mn in relation to the observed nongenotoxic effects and thus may offer some support to a mechanism involving similar molecular interactions underlying the weak mutagenic and genotoxic effects of Mn.…”

Chromium(III) and iron(III) inhibits replication of DNA and RNA viruses

“…The concentrations of chromium chloride and iron chloride for these studies were chosen on the basis of other reports (Mazzotti et al 2001, 2002) and our earlier investigations. Our previous experiments have shown that both of them slightly stimulated cell proliferation (Terpiłowska and Siwicki 2009, 2010).…”

“…The aim of this study was to examine the effect of chromium(III) and iron(III) used separately and simultaneous treating of chromium(III) and iron(III) on HSV-1 an BVDV replication. The concentrations of chromium chloride and iron chloride for these studies were chosen on the basis of other reports (Mazzotti et al 2001, 2002) and our earlier investigations (Terpiłowska and Siwicki 2009, 2010, 2012). …”