Effect of prolactin and cyclic AMP on 125I-labelled low density lipoprotein uptake and metabolism by luteinized porcine granulosa cells in culture

Abstract

The present study examines the effect of prolactin (PRL) and N6-21-O-dibutyryladenosine 3′5′-cyclic monophosphate (cAMP) on low density lipoprotein (LDL) uptake and metabolism by luteinized porcine granulosa cells in culture. Granulosa cells from preovulatory follicles were plated with 1% serum and 1 μg/mL of insulin for the first 48 h. Following plating (day 3) the cells were cultured in serum-free media with the same dose of insulin. The next day the medium was replaced with serum- and insulin-free medium, and to some cultures 1.23 IU/mL of human chorionic gonadotropin (hCG) was added. On day 5 the medium was again replaced and graded amounts of PRL (0, 0.03, 0.3, and 3 μg/mL) were added. Following 48 h of incubation with PRL, 20 μg/mL of 125I-labelled LDL was added to cultures. Surface-bound, internalized, and degraded LDLs were quantitated at 12 h following addition of LDL. To examine the effect of cAMP on LDL metabolism, the cells were exposed for 24 h to cAMP (3 mM) on day 6 of culture. PRL had a stimulatory effect on LDL degradation by luteinized granulosa cells. Pre-exposure of cells to hCG augmented the stimulatory effect of PRL. Addition of cAMP also enhanced LDL degradation by luteinized granulosa cells. Both PRL and cAMP increased surface binding of LDL in cells pre-exposed to hCG, but there was no effect on internalization. The increase in cell surface binding of LDL with PRL and cAMP was less than their effect on LDL degradation. It is concluded that PRL and cAMP enhance LDL metabolism in luteinized porcine granulosa cells by increasing surface binding of LDL and by a post-receptor mechanism. PRL may play a role in the supply of LDL-carried cholesterol for steroidogenesis by luteinized granulosa cells.