Altered gene expression in immunotoxicology screening in vitro: Comparison with ex vivo analysis

We have proposed in vitro immunotoxicology screening systems utilizing cDNA probes to monitor alterations in cytokine expression in defined cell populations. In this study we compared responses observed following in vitro exposure with responses in cells obtained ex vivo. Peritoneal macrophages and splenic lymphocytes were cultured from C3H mice or PVG rats and exposed to graded concentrations of the immunomodulators, biostim, cyclosporin A, azathioprine or tributyltin oxide (TBTO). Biostim (1 mug/ml) induced the expression of cytokine mRNAs in peritoneal macrophage cultures and induced a similar spectrum of cytokine expression in macrophages following administration to mice (100 mug/kg ip). Cyclosporin A (1 mug/ml) totally inhibited the expression of IL-2 mRNA in murine mixed lymphocyte cultures and induced a 40% inhibition of IL-2 mRNA in splenocytes isolated from mice repetitively exposed (5 x 100 mg/kg over 10 days). Azathioprine (10 mug/ml) inhibited the expression of all mRNAs in rat lymphocyte cultures and inhibited both IL-2 mRNA and actin expression in thymocytes isolated from PVG rats (2 mg/kg/day for 28 days). TBTO (10 nm) induced a 25% inhibition of an early peak of IL-2 receptor mRNA expression within mixed lymphocyte cultures and induced a 10% inhibition of expression of IL-2 receptor mRNA within mixed lymphocyte cultures from rats exposed to 50 mg/kg/day for 28 days. Our results demonstrate that appropriate batteries of in vitro tests can identify the four immunomodulatory chemicals described here. Observations made in vitro using molecular biological analysis have been shown to correlate with parallel ex vivo analysis.