Expression of cytochrome P450 mRNA in rat germ cells was characterized by reverse transcription PCR with various primers located at the 3 -end of the coding region. At least two unusual isoforms (Ex10-S and INT) of P450 aromatase (P450arom) mRNA were expressed. Analysis of their sequences demonstrated that an alternative splicing event occurred first at the exon-intron boundary of the GT consensus sequence of the last coding exon, and second in the internal 5 donor inside exon 9 used as a minor cryptic splicing site. These isoforms lacked the last coding exon which contained the heme-binding domain; in addition, for the Ex10-S transcript, the catalytic domain was also absent because of a frameshift in the open reading frame. The deduced amino acid sequences led to truncated P450arom polypeptides without the heme-binding domain, which were probably unable to convert androgens into estrogens. Adult rat germ cells are able to express P450arom mRNA, which is then translated into a biologically active enzyme which is involved in estrogen production. Moreover, for the first time, we report the existence of alternative splicing events of P450arom mRNA in pachytene spermatocytes and round spermatids, which probably cannot encode functional aromatase molecules.

Increasing evidence suggests that cytokines may play a role in ovarian processes. The purpose of this study was to determine if interleukin-1 (IL-1) could modulate rabbit pre-ovulatory follicular function and to explore cellular sites of IL-1 biosynthesis in rabbit follicles. Development of rabbit pre-ovulatory follicles was induced by 200 mIU equine chorionic gonadotropin daily for 2 days. Seventy-two hours after the last injection, ovaries were excised and granulosa and theca cells isolated. The two types of cell were pre-incubated for 24 h in Minimum Essential Medium (MEM) with 5% fetal calf serum (FCS), and then incubated for 24 h in MEM with 2.5% FCS with appropriate stimulants. Results showed that rabbit granulosa and theca cell culture supernatants contain IL-1 bioactivity and that this bioactivity was diminished by gonadotropins, FSH and human chorionic gonadotropin, in a dose-dependent manner. Low doses of IL-1 (1 ng/ml) inhibited gonadotropin-induced progesterone production in both cell types and at the same time increased cell numbers. A study of the mechanism of IL-1 action demonstrated that it affects cAMP generation, and also steps distal to cAMP formation. We conclude that in our model gonadotropins, by inhibiting IL-1 production, could control IL-1 modulation of gonadotropin action on steroidogenesis.

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