Role of Valence State and Solubility of Chromium Compounds on Induction of Cytotoxicity, Mutagenesis, and Anchorage Independence in Diploid Human Fibroblasts¹

We previously showed that carcinogenic nickel, arsenic, and chromium(VI) compounds induced anchorage independence (AI) in diploid human fibroblastic cells (HFC) derived from foreskins (K. A. Biedermann and J. R. Landolph, Cancer Res., 47: 3815–3823, 1987). To elucidate the role of the valence state of chromium and solubility of chromium compounds in inducing AI, we studied the ability of soluble and insoluble hexavalent [chromium(VI)] and trivalent [chromium(III)] chromium compounds to induce mutation and AI in HFC. Chromium(VI) compounds (PbCrO₄, CaCrO₄, Na₂CrO₄, and CrO₃) were 1000-fold more cytotoxic to HFC (average 50% lethal dose 0.5 µm) than chromium(III) compounds (CrCl₃, Cr₂O₃, Cr₂S₃; average 50% lethal dose 500 µm). However, equal concentrations (0.1–10.0 µm) of soluble or insoluble chromium compounds in either +6 or +3 valence states induced similar increases in frequencies of AI in HFC (100–200/10⁵). Chromium(VI)- and chromium(III)-induced AI was a stable phenotype.

All soluble chromium(VI) and insoluble chromium(III) compounds studied induced mutation to 6-thioguanine resistance at cytotoxic concentrations in HFC. Insoluble PbCr(VI)O₄ and a soluble form of Cr(III)Cl₃ were inactive in this assay. Mutation induction by chromium(III) compounds only occurred at cytotoxic concentrations (100–1000 µm) 1000-fold greater than those concentrations of chromium(VI) compounds (0.25–1 µm) which were cytotoxic, mutagenic, and induced AI. Soluble hexavalent Na₂⁵¹CrO₄ was taken up facilely by cells at concentrations that induced cytotoxicity, mutation, and AI. At concentrations of 0.25–1.0 µm, which induced AI but were not cytotoxic or mutagenic, or concentrations of 1–1000 µm, which were cytotoxic and mutagenic, soluble trivalent ⁵¹CrCl₃ was not taken up by cells. An insoluble form of CrCl₃ was not taken up intracellularly but did avidly associate with cells over the concentration range 1 to 100 µm which induced AI, cytotoxicity, and mutagenicity. Therefore, both chromium(VI) and chromium(III) compounds induced genotoxic effects in human fibroblasts. Cellular uptake, cytotoxicity, mutagenicity, and AI induced by soluble chromium(VI) compounds all occurred at the low concentrations of 0.2 to 1.0 µm; hence mutagenicity and induction of AI may be coupled for soluble chromium(VI) compounds but not for insoluble PbCrO₄, which induced AI but was not mutagenic. Cytotoxicity and mutagenicity of insoluble chromium(III) occurred at concentrations of 10–100 µm, but induction of AI occurred at concentrations of 0.1–10 µm, indicating that inductions of mutagenicity and AI were not coupled for chromium(III) compounds.

We postulate that induction of AI by insoluble PbCr(VI)O₄ and by insoluble and soluble Cr(III) compounds occurs by a mechanism not involving base substitution or frameshift mutations. Since there was no significant cellular uptake of Cr(III) compounds, induction of AI by chromium(III) compounds might involve Cr(III)-induced membrane damage, followed by generation of chemically reactive membrane components, which could act intracellularly to induce stable AI.