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2-Methoxyethanol (ME) and its metabolite, methoxyacetic acid (MAA), produce testicular lesions characterized by pachytene spermatocyte degeneration. To understand the molecular basis of this action on meiotic prophase cells, mRNA differential display was used to identify gene expression changes in control and treated cells. When pachytene spermatocytes were cultured with 5 mM ME or 5 mM MAA for 24 h, two complementary DNAs (cDNAs), of 557 nucleotides (clone 5) and 388 nucleotides (clone 6), were up-regulated; and a cDNA of 648 nucleotides (clone 1) was down-regulated. The altered expression pattern shown by differential display was confirmed by Northern blotting. Sequence analyses indicate that clones 1 and 6 have 83% and 79% homology at the nucleotide level to a polo-like kinase and a thiol-specific antioxidant, respectively. Clone 5 shows no homology to any known gene in the database. Messenger RNAs (mRNAs) encoding the thiol-specific antioxidant and clone 5 are up-regulated within 30 min of the addition of MAA, whereas the polo-like kinase mRNA decreased to undetectable levels after 6 h. Changes in Sertoli cell gene expression were also detected when Sertoli cells were cultured with 5 mM ME or MAA for 24 h. Two cDNAs, of 367 nucleotides (clone 2) and 676 nucleotides (clone 3), were up-regulated; and a cDNA of 538 nucleotides (clone 4) was down-regulated. Homology searches revealed that clones 3 and 4 have 90 and 91% homology at the nucleotide level to an oxidative stress protein and a phosphodiesterase (PDE), respectively. Northern blotting confirmed the differential display expression pattern for the PDE and oxidative stress protein. mRNAs for the latter were induced within 30 min, and PDE mRNAs were down-regulated within one h, after the addition of MAA. To determine whether the changes in gene expression seen with cells in culture also occur in vivo, rats were given a single oral dose of 250 mg/kg ME or MAA. After 24 h, total testis RNAs from control and treated rats were purified and hybridized. The expression patterns seen in vivo for the differentially expressed cDNAs were identical to those seen in vitro. We conclude that, although pachytene spermatocytes seem to be selectively affected by ME and MAA, changes in gene expression are also detected in Sertoli cells, suggesting that the action(s) of ME or MAA on pachytene spermatocytes could be mediated through Sertoli cells.

endocrinology

Rat Pachytene Spermatocytes Down-Regulate a Polo-Like Kinase and Up-Regulate a Thiol-Specific Antioxidant Protein, Whereas Sertoli Cells Down-Regulate a Phosphodiesterase and Up-Regulate an Oxidative Stress Protein after Exposure to Methoxyethanol and Methoxyacetic Acid<sup>1</sup>

Rat Pachytene Spermatocytes Down-Regulate a Polo-Like Kinase and Up-Regulate a Thiol-Specific Antioxidant Protein, Whereas Sertoli Cells Down-Regulate a Phosphodiesterase and Up-Regulate an Oxidative Stress Protein after Exposure to Methoxyethanol and Methoxyacetic Acid1

Rat Pachytene Spermatocytes Down-Regulate a Polo-Like Kinase and Up-Regulate a Thiol-Specific Antioxidant Protein, Whereas Sertoli Cells Down-Regulate a Phosphodiesterase and Up-Regulate an Oxidative Stress Protein after Exposure to Methoxyethanol and Methoxyacetic Acid¹