M. Fielden, R. Halgren, C. Fong, K. Chou, T. Zacharewski. Department of Biochemistry & Molecular Biology, National Food Safety & Toxicology Center, Department of Animal Science, Institute for Environmental Toxicology, Michigan State University, East Lansing, MI, USA, 48824.

To explore the molecular events underlying the adverse effects on sperm quality following developmental exposure to estrogenic chemicals, we have constructed cDNA microarrays to examine testicular gene expression in B6D2F1 offspring of mice exposed by gavage to 10 µg/kg diethylstilbestrol (DES) from gestational day 12 to postnatal day 21. Replicate gene expression profiles were examined in male offspring at 3, 15 and 45 weeks of age to determine if the effects on gene expression paralleled the long term decreases in testis weight, sperm count, and sperm fertilizing ability, as previously described. A mouse cDNA microarray containing approximately 1948 genes was constructed and used to compare testicular gene expression in the offspring of DES-exposed mice to that of vehicle-exposed mice. An independent reference design coupled with paired t-tests was used to identify genes significantly (p<0.05) altered in expression by DES. Based on adjusted p values, there was only one gene significantly altered at 3 weeks of age (a component of the 20S proteasome). At 15 weeks of age, there were 46 genes significantly altered in expression. There were no significant changes at 45 weeks of age. Raw p values and functional annotation were used to prioritize the selection of genes for verification. Using real-time PCR, both age and dose-dependent changes in gene expression were examined. There was a significant decrease in the expression of ER alpha mRNA at 3 weeks of age (p<0.01), while mRNA expression was below the limits of detection in 15 and 45 week old mice. Other genes selected for verification include inhibin, the orphan receptor TR2, and Xmr, a component of the synaptonemal complex. These results suggest that the adverse effects on sperm fertilizing ability may be due to altered expression of ER alpha, and possibly other genes, in the testis.