Analysis of the combined osteolathyritic effects of β-aminopropionitrile and diethyldithiocarbamate on Xenopus development

Abstract

In order to examine the mechanistic basis between combined effects and mechanisms of action, two osteolathyrogens, β-aminopropionitrile (βAPN) and diethyldithiocarbamate (DTC), were tested together on Xenopus embryos. In a separate test, DTC was also tested with copper sulfate to determine the importance of copper in DTC-induced osteolathyrism. Frog embryos (Xenopus laevis) were exposed for 96 h, with daily solution removal and replacement. Preserved tadpoles were evaluated for osteolathyritic lesions. For the βAPN:DTC test, a 1.2-factor matrix design was used, producing two single chemical and seven mixture-response curves. The χ2 goodness-of-fit test was used to compare the experimental mixture-response curves with theoretical effects for two combined effects models, dose-addition and independence. All seven mixture curves were consistent with expected results for dose-addition, but the correlations were generally not high. For the DTC:copper test, the three mixture-response curves generated showed that added copper increased the DTC-alone EC50, but there was no corresponding right shift at the top of the response curves, as observed previously with βAPN and copper. In the βAPN:DTC and DTC:copper tests, DTC alone showed a biphasic concentration-osteolathyrism curve, and the slope of the response curve for DTC alone in each test was statistically different than the slope for the βAPN alone response curve. Taken together, the results suggest the potential for a second osteolathyritic effect of DTC that affected the combined toxicity enough to produce a dose-addition correlation without the chemicals necessarily having the same mechanism.