Effects of pregnant mare serum gonadotropin (eCG) on follicle development and granulosa-cell apoptosis in the pig

Abstract

The effect of eCG on follicular development and granulosa-cell apoptosis in sexually mature and immature gilts and on granulosa-cell apoptosis in vitro were studied. The sexually mature gilts were treated with eCG on Day 11 of the estrous cycle, and effects were analyzed at different times after treatment with untreated animals at corresponding stages of the cycle as controls. Apoptosis was determined by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL), hematoxylin and eosin staining, and DNA ladder. The proportion of apoptotic cells in atretic follicles (39%) was significantly higher ( P<0.01) than that in healthy follicles (9%). At 24 h after eCG treatment in mature gilts, the total number of follicles visible on the ovarian surface (57 per ovary), the number of small (<3 mm) follicles (31.5 per ovary) and the number of medium-sized (3–5 mm) follicles (23 per ovary) were significantly higher ( P<0.05) than those of control animals (28, 20 and 6.5 per ovary, respectively), and declined gradually thereafter to below the level of control animals. The number of large (≥5 mm) follicles began to show a marked increase at 72 h after eCG (8.5 versus 2.5, P<0.05). At 24 h after eCG treatment, the proportions of apoptotic cells in small (7.2%) and medium-sized follicles (7.4%) were markedly lower ( P<0.01) than those in controls (21.5 and 21%, respectively) and increased gradually thereafter to approach the level in controls. The percentage of apoptotic cells in large follicles (10% at 24 h post-eCG) did not change significantly. Before eCG treatment, there were markedly fewer follicles of all types on ovaries of immature gilts than of mature gilts (9 versus 25 per ovary) and the proportion of apoptotic cells in small and medium follicles was high (25 and 34%, respectively). After eCG treatment, the changes in follicle number and proportion of apoptotic cells in the immature gilts followed a similar pattern to that of the mature gilts. Equine chorion gonadotropin inhibited apoptosis of granulosa cells cultured either in vitro or in intact follicles in a dose-dependent manner. Thus, follicular atresia in the pig, as in other animals, was characterized by apoptosis of large numbers of granulosa cells, and eCG promoted follicular development by inhibition of granulosa-cell apoptosis.