Immunohistochemical localization of steroidogenic enzymes and comparison with hormone production during follicle development in the pig

Abstract

The steroidogenic enzymes, P450 aromatase (P450 _arom ) and P450 17a-hydroxylase (P450 _17a ), were precisely located within the healthy porcine follicle by immunohistochemistry. Enzyme distribution was examined throughout follicular development during natural oestrous cycles (n = 14 gilts) and was compared with steroid production by healthy whole and theca-only follicles. All follicles 2 mm or more in diameter were either fixed for immunohistochemistry (n = 380 of which 197 were assessed as healthy) or incubated as whole (n = 110) or theca-only (n = 110) follicles to measure steroidogenesis. P450 _17a was confined to the theca layer. The number of positive cells and staining intensity increased with follicle size. P450 _arom was consistently detected in the granulosa layer of follicles measuring 6 mm or more in diameter and those cells furthest from the antrum were most strongly stained. P450_arom was also detected in the theca layer of these large follicles. Whole and theca-only follicles produced oestradiol and androstenedione, and the levels of both hormones increased with follicle size (P<0.001). Whole follicles produced more oestradiol (P<0.001), but less androstenedione (P = 0.01) than theca-only follicles of the same size. Although granulosa cells contained P450 _arom and synthesized oestradiol, only theca cells contained P450 _17a. Theca cells therefore provided granulosa cells with androgen substrate. In addition, theca cells possessed P450 _arom , making them capable of independent oestradiol production, which may be required to trigger the LH surge. This study confirms the vital role of theca cells in follicular steroidogenesis in the pig.