210 EXPRESSION OF mRNA ENCODING STEROIDOGENIC ENZYMES IN THE DEVELOPING BOVINE OVARY

Abstract

Recent findings suggest a role for estradiol in the regulation of early folliculogenesis. Estradiol production is greatest in the fetal ovary during early gestation in cattle, and both estradiol and progesterone inhibit primordial follicle activation (Yang MY and Fortune JE 2008 Biol. Reprod. 78 (Suppl 6), 1153-1161). Aromatase expression is detected in early stages of bovine pregnancy (Garverick HA et al. 2009 Anim. Reprod. Sci. in press). The mechanisms controlling steroidogenesis in the bovine fetal ovary remain to be fully elucidated. The objective of this study was to assess mRNA expression patterns of enzymes involved in steroid production [steroidogenic acute regulatory protein (STAR), side-chain cleavage P450 (CYP11), cytochrome P450 17 alpha-hydroxylase (CYP17A1), 3 beta-hydroxysteroid dehydrogenase (3fi-HSD), aromatase cytochrome P450 (CYP19), and 17 beta-hydroxysteroid dehydrogenase (17fi-HSD)] in bovine fetal ovaries during gestation. Bovine fetal ovaries were obtained in a local slaughterhouse, fetal age was estimated by the crown-rump length, and samples were grouped according with days of gestation as follows: 60 (n = 5), 75 (n = 8), 90 (n = 6), 120 (n = 7), 150 (n = 7), and 210 (n = 6). Expression of mRNA encoding steroidogenic enzymes was determined by semiquantitative real-time RT-PCR using bovine-specific primers and cyclophilin A as endogenous control. Reverse transcription was performed with SuperScriptIII® (Invitrogen, Carlsbad, CA, USA) and PCR with Power SYBR green master mix (Applied Biosystems, Foster City, CA, USA) in an ABI Prism® 7500 (Applied Biosystems). Gene expression values were determined by the Pfaffl equation and effect of day of gestation on gene expression was analyzed with Fisher’s protected test, except when data were not normally distributed and nonparametric analysis was performed. Expression of mRNA encoding all steroidogenic enzymes was detected throughout gestation. The mRNA abundance of CYP17A1 and CYP19 was highest at 60 days of gestation and decreased thereafter (P < 0.05). Expression of all other genes did not significantly vary with time of gestation. In conclusion, all major enzymes required for steroidogenesis were expressed in the bovine fetal ovary. Expression of CYP17A1 and CYP19 was suppressed after 60 days of gestation, suggesting that these enzymes may be involved in the mechanisms controlling estradiol production and follicle formation in the bovine fetal ovary. Supported by CAPES and FAPESP.