Antioxidants inhibit the enhancement of malignant cell transformation induced by 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin

The mechanisms of the tumor promoting activity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) were studied using as in vitro model the enhancement ('promotion') of malignant transformation of C3H/M2 mouse fibroblasts induced by N-methyl-N'-nitro-N-nitrosoguanidine or 3-methylcholanthrene. In this assay, the promoting effect of TCDD was maximal at a very low concentration of 1.5 pM and was comparable to the effect of the reference tumor promoter, 12-O-tetradecanoylphorbol-13-acetate (TPA, 0.25 microg/ml). The role of reactive oxygen species in the promoting action was investigated: mannitol, a scavenger of hydroxyl radicals, or antioxidants, i.e. ascorbic acid plus alpha-tocopherol, abolished the in vitro promoting effects of TPA and TCDD. Furthermore, the involvement of protein kinase C (PKC) activation was studied: the protein kinase inhibitor H-7 markedly reduced the in vitro promoting activity of TPA but did not affect the promotion by TCDD. In accord with these results, TPA, but not TCDD, enhanced the PKC activity in C3H/M2 fibroblasts. Since the TPA-mediated activation of PKC was not affected by ascorbate plus alpha-tocopherol, it is concluded that the antioxidants interfere with tumor promotion at a step beyond PKC activation. Thus, the results suggest that the enhancement of malignant cell transformation by TPA and TCDD is dependent on a common mechanism, possibly induced by oxygen radicals, and, in addition, on further mechanisms that may involve agent-specific signalling pathways (e.g. PKC activation by TPA).