EXPRESSION OF MESSENGER RIBONUCLEIC-ACIDS OF INSULIN-LIKE GROWTH-FACTOR BINDING PROTEIN-2, PROTEIN-4, AND PROTEIN-5 IN THE OVINE OVARY - LOCALIZATION AND CHANGES DURING GROWTH AND ATRESIA OF ANTRAL FOLLICLES

in the sheep as in many mammalian species, growth and atresia of antra l follicles are characterized, respectively, by a decrease and a high increase in the intrafollicular levels of insulinlike growth factor bi nding proteins of less than 40 kDa (IGFBPs < 40 kDa), mainly IGFBP-2, -4, and -5. The objective of this study was to investigate whether suc h changes are associated with changes in follicular expression of the corresponding mRNA. For this purpose, ovaries were recovered from ewes slaughtered at the end of follicular phase (i.e., 30 h after progesta gen sponge removal; control ewes) or at 24 h, 36 h, or 72 h after hypo physectomy (hyper) performed 30 h after sponge removal. The expression of mRNA of IGFBPs of less than 40 kDa (IGFBPs < 40 kDA mRNA) was stud ied in ovine antral follicles from control and hyper ewes by in situ h ybridization using [S-35]-labeled human IGFBP-2, -4, and -5 cRNA as pr obes. In control ewes, IGFBP-2 mRNA was mainly expressed in granulosa as a gradient in healthy follicles, the expression being higher in gra nulosa cells close to the basal membrane than in granulosa cells borde ring the antrum and within the cumulus. The level of IGFBP-2 mRNA was lower both in granulosa cells close to the basal membrane and in those bordering the antrum from small follicles than in the corresponding c ompartments of granulosa cells from large healthy follicles (p < 0.05) . In healthy follicles, IGFBP-4 and -5 mRNA were mainly expressed in t hecal cells. No change in level of IGFBP-4 mRNA was observed between s mall and large follicles, whereas the level of IGFBP-5 mRNA tended to be lower in thecal cells from large compared to small follicles (p = 0 .055). In atretic follicles, expression of IGFBPs < 40 kDa mRNA strong ly increased in granulosa (IGFBP-2 and -5, p < 0.01) and in thecal cel ls (IGFBP-2 and -4, p < 0.01). In hyper ewes, the chronology of change s in expression of follicular IGFBPs < 40 kDa mRNA and in intrafollicu lar levels of the corresponding proteins was studied during atresia of large antral follicles. Early atresia of large follicles was associat ed with a strong decrease in intrafollicular estradiol levels (p < 0.0 01); an increase in intrafollicular levels of IGFBP-2, -4, and -5 (p < 0.001); an increase in both IGFBP-2 (p < 0.001) and -5 (p < 0.01) mRN A expression in granulosa and thecal cells; but no change in IGFBP-4 m RNA expression. Late atresia of large follicles was associated with a further decrease in intrafollicular estradiol levels (p < 0.001); a fu rther increase in intrafollicular levels of IGFBP-2, -4, and -5 (p < 0 .001); an increase in IGFBP-4 (p < 0.01) and -5 (p < 0.05) mRNA expres sion in theca and granulosa, respectively; a decrease in IGFBP-5 mRNA expression in theca (p < 0.05); but no further increase in IGFBP-2 mRN A expression. Overall, these data suggest that the decrease and the in crease in expression of mRNA of follicular IGFBPs < 40 kDa during foll icular growth and atresia, respectively, are involved in the decrease and the increase in intrafollicular levels of the corresponding protei ns. Moreover, the increase in expression of follicular IGFBPs < 40 kDa during atresia of large follicles in hypophysectomized ewes followed a specific time course, the increase in IGFBP-2 and -5 mRNA expression being earlier than the increase in IGFBP-4 mRNA expression.