δ⁴ and δ⁵ pathway steroidogenesis by human granulosa lutein cells (GLC) in long-term cultures (6–8 days) was investigated under basal (10% serum + medium + GLC) and stimulated (human chorionic gonadotropins (hCG), 100 mIU/ml) conditions. In the δ⁴ pathway, 17-hydroxyprogesterone (17-OHP) secretion increased from 160-fold at 48 hours of culture to 360-fold at 144 hours under basal conditions, when compared with the respective controls (concentrations in the 10% serum added to the medium) (p 0.005). HCG further augmented 17-OHP production significantly at 96, 144 and 192 hours of culture. Progesterone (P) secretion behaved similarly, and increased from 150-fold at 48 hours to 560-fold at 96 hours when compared with controls, being further stimulated by hCG. In contrast, androstenedione (A) secretion throughout the entire culture period increased only slightly (3–5-fold) under both basal and stimulated conditions, when compared with the respective controls. In the δ⁵ pathway, the secretory pattern of 17-hydroxypregnenolone (17-OHPregn) and dehydroepiandro-sterone (DHEA) was similar to that observed with A, and both steroids increased only slightly under basal and stimulated conditions, when compared with their respective controls.

In conclusion, 17-OHP and P are secreted in very significant amounts during the entire culture period while 17-OHPregn, DHEA and A are secreted in extremely small amounts. These results demonstrate that the δ⁵ pathway is inactive in long-term human GLC cultures while the δ⁴ pathway is active in certain portions only. There is evidence of marked 17α-hydroxylase activity but a deficiency in C_17–20 lyase, therefore A production is extremely low. Human GLC in long-term cultures are unable to secrete androgens de novo to serve as precursors for aromatization and estradiol (E₂) production.