Gene expression in ovarian granulosa and theca cells in cattle selected for double ovulations and twin births

Abstract

Scope and Method of Study: Components of the IGF system and gonadotropin receptors (FSHR and LHR) might be involved in the control of the development of multiple dominant follicles leading to twinning. Gene expression of FSHR, LHR, IGF2R and the hedgehog system (Ihh and Ptch1) gene expression in granulosa and/or theca cells from cattle not selected (Control) or selected for multiple ovulations (Twinner) were quantitated at two different stages (Day 3, D3; Day 5, D5) of the follicular cycle and in the largest 3 follicles. The effect of IGF2 in small (1-5 mm) and large (8-22 mm) bovine granulosa cell cultures on steroid production, cell proliferation, and gene expression were also investigated.

Findings and Conclusions: Granulosa FSHR mRNA was greater (P < 0.05) at D3 versus D5 in healthy estrogen-active (E-A) follicles and in Control versus Twinner cows at D3. Granulosa IGF2R mRNA was lower (P < 0.05) in E-A versus atretic estrogen-inactive (E-I) follicles, and lower (P < 0.05) at D5. Granulosa IGF2R mRNA was greater (P < 0.05) in Control than Twinner cows at D3 and D5; Control F2 had greater (P<0.05) thecal IGF2R mRNA than F1, F2 and F3 of Twinner cows. IGF2 increased granulosa estradiol and progesterone production, stimulated aromatase mRNA and increased proliferation of granulosa cells. IGF1R antibodies reduced the stimulatory effect of IGF2 and IGF1 on estradiol production and cell proliferation. Granulosa Ihh mRNA was two-fold greater (P < 0.05) in healthy than atretic follicles and in D3 than D5; thecal Ptch1 mRNA was lower (P < 0.05) at D3 than D5, and in Twinner versus Control cows. In cultured granulosa cells, IGF1 decreased (P < 0.001) Ihh mRNA abundance, and both IGF1 and IGF2 decreased (P < 0.01) thecal Ptch1 mRNA abundance. Decreased IGF2R in granulosa and theca cells of Twinner cows likely increases bioavailable IGF2, which in turn enhances follicular development of two, rather than one follicle and further supports the idea that IGF2 and its receptor (IGF2R) may play a role in follicular development. For the first time, we demonstrate an interaction between the ovarian IGF and hedgehog systems.