An in vitro assessment of the toxicity of 2,4‐dichloroaniline using rainbow trout primary epidermal cell cultures

Abstract

Abstract2,4‐Dichloroaniline (2,4‐DCA) is a common pollutant of freshwaters. Information on the effects of this chemical on the aquatic environment is lacking. In this work, we present a technique using primary cell culture of rainbow trout (Oncorhynchus mykiss) epidermal cells as a model for the evaluation of the potential ecotoxicologic hazards of 2,4‐DCA. The effects of a wide dose range of the chemical on survival and function of the cells were studied. 2,4‐Dichloroaniline was found to show an increasingly toxic effect over the dose range of 100 to 1,000 μM. The toxicity threshold was observed to occur at approximately 400 μM in serum‐containing media and 200 μM in serum‐free media. The LC50 in serum‐containing media at 24, 48, 72, and 96 h of exposure ranged from 650 to 740 μM. In serum‐free media the LC50 ranged from 340 to 500 μM over a similar time period. Therefore, the toxicity of 2,4‐DCA to the primary cells increased in the absence of serum in the culture medium. As the 2,4‐DCA concentration increased the number of goblet (mucus) cells present in the cultures decreased, as did the normal healthy epidermal cells. Cell death was predominantly necrotic as opposed to apoptotic. This study is the first demonstration of an in vitro technique using fish primary epidermal cultures as a toxicology assessment tool. The major advantage of using primary cultures as opposed to immortalized cell lines is in the ability of these cultures to maintain their in vivo characteristics for approximately 10 d postexposure, allowing the study of the acute effects of aquatic contaminants. The characteristics of established cells deviate substantially from those of normal cells.