SECRETION OF CUMULUS EXPANSION-ENABLING FACTOR (CEEF) IN PORCINE FOLLICLES

The objective of this study was to find out whether porcine cumulus an d mural granulosa cells can secrete cumulus expansion-enabling factor (CEEF). Culture drops of M-199 medium were conditioned with denuded po rcine oocytes (1 oocyte/mu l), cumulus cells from oocytectomized compl exes (1 OOX/mu l), pieces of mural granulosa isolated from preantral t o preovulatory follicles (1000 cells/mu l), or oviductal cells (1000 c ells/mu l) for 24 hr. The production of CEEF was assessed by the addit ion of mouse OOX and follicle-stimulating hormone (FSH) (1 mu g/ml) to microdrops of the conditioned medium. After 16-18 hr, expansion of th e mouse OOX was scored on a scale of 0 to 4 by morphologic criteria. M ouse OOX did not expand in nonconditioned FSH-supplemented medium. Imm ature porcine oocytes produced +3 to +4 expansion of the mouse OOX. Gr anulosa cells isolated from preantral and early antral follicles and c umulus cells isolated from all stages of follicle development constitu tively secreted CEEF under in vitro conditions. Mural granulosa cells of small, medium, and preovulatory (PMSG) follicles also secreted CEEF in vitro; however, FSH or leutenizing hormone (LH) stimulation was es sential for this secretion. Hormonally induced secretion of CEEF was a ccompanied by expansion of the mural granulosa itself. Granulosa cells isolated from follicles of gilts 20 hr after PMSG and human chorionic gonadotropin (hCG) administration did not produce CEEF and did not ex pand in response to FSH and LH in vitro. CEEF activity also was found in the follicular fluid of small antral follicles, was reduced in medi um follicles, and was not detectable in PMSG-stimulated follicles. How ever, CEEF activity was reestablished in the follicular fluid of preov ulatory follicles by hCG injection, conceivably due to increased produ ction of CEEF by cumulus cells. We conclude that (1) porcine cumulus a nd mural granulosa cells are capable of CEEF production in vitro and ( 2) autocrine secretion of CEEF by cumulus cells is involved in regulat ion of porcine cumulus expansion both in vitro and in vivo. (C) 1998 W iley-Liss, Inc.