Keratinization of mouse teratoma cell line XB produced by 2,3,7,8-tetrachlorodibenzo-p-dioxin: An in vitro model of toxicity

Abstract

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), an extremely potent toxin, serves as the prototype of a large group of halogenated aromatic hydrocarbons, all of which produce a similar and characteristic pattern of toxic lesions, which includes hyperkeratosis and squamous metaplasia in the skin; induce a battery of coordinately expressed enzymes, the most studied of these being aryl hydrocarbon hydroxylase (AHH) activity; and appear to exert both their toxic effects and enzyme induction through stereospecific, reversible binding to a cytosolic receptor protein. Previous studies performed with over 30 cell types have failed to demonstrate any in vitro toxicity from TCDD. When XB cells derived from a mouse teratoma are cultured at high density (to prevent spontaneous differentiation) along with lethally irradiated 3T3 cells, the addition of TCDD produces a dose-related keratinization response as detected by red staining with Rhodanile blue. Maximal keratinization is produced by 5 × 10 −11 M TCDD. This is a direct effect of TCDD on XB cells, and will occur in the absence of the feeder cells if the teratoma cells are cultured in 3T3-conditioned media. The keratinization produced by TCDD was examined by electron microscopy and histologic staining, and appears to be similar to that which occurs spontaneously when XB cells are plated at low density. XB cells contain the cytosol receptor and respond to TCDD with a dose-related induction of AHH activity. The potencies of halogenated aromatic hydrocarbon congeners [dibenzo-p-dioxins, dibenzofurans, biphenyls and azo(xy)benzenes] to produce keratinization in the XB/3T3 system corresponds to their binding affinities for the cytosol receptor. Nonhalogenated aromatic hydrocarbons, which are agonists for the receptor [such as benz(a)anthracene and 5,6-benzoflavone], also produce a dose-related keratinization in XB/3T3 cultures, whereas unrelated toxins (such as alkylating agents and inhibitors of nucleic acid synthesis) do not. Our results suggest that the keratinization produced by TCDD and congeners in XB/3T3 cultures is mediated by the cytosolic receptor, and that this system provides an in vitro model for the in vivo toxic effects produced by halogenated aromatic hydrocarbons in the epidermis.