Effects of Culture Conditions on Estrogen-mediated Hepatic In Vitro Gene Expression and Correlation to In Vivo Responses.

Fong, C.J., Burgoon, L.D., and Zacharewski, T.R.

Abstract

Development of in vitro systems for predictive toxicology is important in order to develop high throughput early toxicity screening assays and to minimize animal testing studies. This study assesses the ability of mouse Hepa-1c1c7 hepatoma cell model under differing culture conditions to predict in vivo estrogen-induced hepatic gene expression changes. Custom mouse cDNA microarrays, were used to compare Hepa-1c1c7 temporal gene expression profiles treated with 10 nM 17?-estradiol (E2) in serum-free and stripped-serum supplemented media at 1, 2, 4, 8, 12 and 24 hrs. Stripped serum conditions increase the number gene expression changes and overall responsiveness likely due to the presence of serum factors supporting proliferation and mitochondrial activity. Data from both experiments were compared to a gene expression time course study examining the effects of 100 ?g/kg 17?-ethynylestradiol (EE) in C57BL/6 mouse liver at 2, 4, 8, 12, 18 and 24 hrs. Only 18 genes overlapped between the serum-free and in vivo studies, whereas 238 genes were in common between Hepa-1c1c7 cells in stripped serum data and C57BL/6 liver samples. Stripped-serum cultured cells exhibited E2-elicited gene expression changes associated with proliferation, changes in cytoskeletal organization, increased cholesterol uptake and synthesis, increased fatty acid ?-oxidation and responses to oxidative stress which correlates well with reported in vivo hepatic responses. These results demonstrate that the importance of serum in cell responsiveness and that Hepa-1c1c7 cells are predictive of estrogen elicited in vivo hepatic responses for selected pathways.

Supplementary Data

Supplementary Table S1 2.06MB: [MS Excel]

Supplementary Table S2 2.06MB: [MS Excel]

Supplementary Table S3 4.24MB: [MS Excel]

Supplementary Table S4 4.24MB: [MS Excel]

Supplementary Table S5 1.71MB: [MS Excel]

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