PARENT SESSION
OVARY - C

Wednesday, August 4, 2004
10:30 AM–12:30 PM
Buchanan Courtyard

(775) GRANULOSA CELL CYCLE PROGRESSION DURING FOLLICLE GROWTH: A DECREASE IN S PHASE PRIOR TO AN OVULATORY STIMULUS.

Cannon, Jennifer¹, Swan, Tara¹, Chaffin, Charles¹, ¹ Medical College of Georgia, Augusta, GA

ABSTRACT- Follicular development and subsequent corpus luteum formation involves complex orchestration of granulosa cell proliferation, growth, and differentiation. The goal of the present study was to elucidate the proliferative profile of granulosa cells during follicle growth, prior to an ovulatory stimulus. Immature rats were given a single injection of PMSG to initiate follicle growth, and granulosa cells collected before (0 h) and up to 48 h after PMSG. FACS analysis was used to determine the proportion of cells in S phase of the cell cycle. Prior to PMSG (0 h), 8.0±1.4% of granulosa cells were in S phase, while the proportion of S phase cells increased significantly between 6 and 12 h (8.8±0.6 to 13.9±1.1, p<0.05). This increase was maintained 24 h post-PMSG (13.4±0.9) but was transiently reduced at 30 h to pre-PMSG levels (9.2±0.8, p<0.05 v. 24 h). There was a tendency for the proportion of S phase cells to increase 36 h after PMSG relative to 30 h (11.7±0.9, p=0.08). 48 h post-PMSG, the percentage of granulosa cells in S phase was not different than 0 h (8.0±1.4). Analysis of BrdU uptake by immunohistochemistry showed a decline in the number of positive cells between 24 and 48 h after administration of PMSG. No significant changes were found in the mRNA levels of cyclin D2, cyclin E, cdk 2, 4, or 6 as measured by real-time RT-PCR. However, Cdk 2 protein was increased significantly 24 h after PMSG (4-fold, p<0.05) and returned to pre-PMSG levels at 48 h. The expression of c-myc mRNA did not change during follicle growth, although mRNA levels of the myc antagonist mad 1 were increased significantly 42 and 48 h after PMSG (p<0.05). Results from this study suggest that the rate of granulosa cell proliferation peaks between 12-24 h after PMSG and begins to decrease prior to an ovulatory stimulus by mechanisms that involve suppressed cdk 2 and mad 1. These data lend further support to the hypothesis that the regulation of granulosa cell proliferation is independent of luteal development. (Supported in part by a National Science Foundation Graduate Research Fellowship to JC.)

KEY WORDS: proliferation, follicle growth, granulosa cell, ovary