Modulation of Ras signaling alters the toxicity of hydroquinone, a benzene metabolite and component of cigarette smoke.

BACKGROUND: Benzene is an established human leukemogen, with a ubiquitous environmental presence leading to significant population exposure. In a genome-wide functional screen in the yeast Saccharomyces cerevisiae, inactivation of IRA2, a yeast ortholog of the human tumor suppressor gene NF1 (Neurofibromin), enhanced sensitivity to hydroquinone, an important benzene metabolite. Increased Ras signaling is implicated as a causal factor in the increased pre-disposition to leukemia of individuals with mutations in NF1. METHODS: Growth inhibition of yeast by hydroquinone was assessed in mutant strains exhibiting varying levels of Ras activity. Subsequently, effects of hydroquinone on both genotoxicity (measured by micronucleus formation) and proliferation of WT and Nf1 null murine hematopoietic precursors were assessed. RESULTS: Here we show that the Ras status of both yeast and mammalian cells modulates hydroquinone toxicity, indicating potential synergy between Ras signaling and benzene toxicity. Specifically, enhanced Ras signaling increases both hydroquinone-mediated growth inhibition in yeast and genotoxicity in mammalian hematopoetic precursors as measured by an in vitro erythroid micronucleus assay. Hydroquinone also increases proliferation of CFU-GM progenitor cells in mice with Nf1 null bone marrow relative to WT, the same cell type associated with benzene-associated leukemia. CONCLUSIONS: Together our findings show that hydroquinone toxicity is modulated by Ras signaling. Individuals with abnormal Ras signaling could be more vulnerable to developing myeloid diseases after exposure to benzene. We note that hydroquinone is used cosmetically as a skin-bleaching agent, including by individuals with cafe-au-lait spots (which may be present in individuals with neurofibromatosis who have a mutation in NF1), which could be unadvisable given our findings.

Formaldehyde induces micronuclei in mouse erythropoietic cells and suppresses the expansion of human erythroid progenitor cells.

Although formaldehyde (FA) has been classified as a human leukemogen, the mechanisms of leukemogenesis remain elusive. Previously, using colony-forming assays in semi-solid media, we showed that FA exposure in vivo and in vitro was toxic to human hematopoietic stem/progenitor cells. In the present study, we have applied new liquid in vitro erythroid expansion systems to further investigate the toxic effects of FA (0-150 µM) on cultured mouse and human hematopoietic stem/progenitor cells. We determined micronucleus (MN) levels in polychromatic erythrocytes (PCEs) differentiated from mouse bone marrow. We measured cell growth, cell cycle distribution, and chromosomal instability, in erythroid progenitor cells (EPCs) expanded from human peripheral blood mononuclear cells. FA significantly induced MN in mouse PCEs and suppressed human EPC expansion in a dose-dependent manner, compared with untreated controls. In the expanded human EPCs, FA slightly increased the proportion of cells in G2/M at 100 µM and aneuploidy frequency in chromosomes 7 and 8 at 50 µM. Our findings provide further evidence of the toxicity of FA to hematopoietic stem/progenitor cells and support the biological plausibility of FA-induced leukemogenesis.

Bone marrow genotoxicity of 2,5-dimethylfuran, a green biofuel candidate.

2,5-Dimethylfuran (DMF) is being considered as a potential green transportation biofuel, but there is limited information about its toxicity and safety. We examined DMF toxicity in the bone marrow using a murine in vitro erythropoietic micronucleus assay and found that exposure to DMF (0.1 mM, 1 hr) induced an increase in micronuclei frequency compared with controls. These data suggest that DMF may be genotoxic to hematopoietic cells and that more thorough toxicological studies on DMF should be conducted to ensure public and occupational safety before it is considered a viable biofuel and produced in mass quantities. As well as specific data on DMF, our study further validates an in vitro cell culture system that captures the essential features of the in vivo mammalian micronucleus genotoxicity assay, enabling increased throughput and controlled studies on hematopoietic DNA damage response, while reducing animal sacrifice. In vitro assays, such as the in vitro micronucleus assay, will be essential as international chemical policy is increasingly utilizing green chemistry principles that require more toxicological testing.