Submitted to: Biology of Reproduction
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: June 19, 1996
Publication Date: N/A
Interpretive Summary: Reproductive efficiency in the pig is heavily dependent upon the number and fertilizability of follicular oocytes. The number of follicles forming after the current ovulations may determine the number that ovulate in the next cycle. Whether follicle cells proliferate or die is expression of specific genes. Steroids are produced by action of specific enzymes and steroids regulate gene expression by binding to specific receptors that are transcription factors. In this experiment, we tested a new histochemical procedure for identifying atretic (dying)follicles by measuring cells dying by apoptosis (programmed cell death). We then measured the amounts of two different enzyme proteins and one receptor/transcription factor, measured by immunohistochemistry procedures in frozen tissue sections, in growing and dying follicles. The two enzymes, aromatase and 17alpha-hydroxylase/C17-20 lyase (17alpha),and the androgen receptor transcription factor (AR) were less in dying than growing follicles. In fact, aromatase was completely absent in dying follicles showing that these follicles could not produce estradiol. The results of this basic research will be used by scientists as a basis for studies on the molecular regulation of follicular growth and atresia in swine. These studies will provide knowledge required better regulate ovulation rate.
Technical Abstract: The purpose of this study was to assess the expression of Androgen Receptors (AR), aromatase (Arom), and 17alpha-hydroxylase/C17-20 lyase (17alpha) by immunohistochemistry (IHC) in follicles developed on Days 3, 5 and 7 following the onset of estrous. In addition, apoptosis of follicular cells was assessed by in situ 3' end labeling as a marker of follicular atresia. Staining intensity of the four variables tested was assigned a numeric value (0-2 for AR, Arom and 3' end labeling, and 0-3 for 17alpha). Follicles assigned a value of 2 for 3' end labeling were considered atretic. Follicles were further classified according to size as small (<3 mm), medium (3-5 mm) or large (>5 mm) Analysis of Variance of rank transformed data revealed that staining for AR, Arom and 17alpha was significantly less (P=0.0001) in atretic follicles (AF) compared to non-atretic follicles (NAF). Expression of Arom was undetectable in AF of any size. Regression analysis revealed a decrease in AR expression (P<0.01), and an increase in Arom and 17alpha expression (P<0.01) in NAF with increasing follicle size. In AF, the expression of 17alpha was greater in medium and large follicles than in small follicles (P<0.01). In non-atretic small follicles, the expression of Arom and 17alpha decreased significantly (P<0.01) between days 3 and 7. These data suggest that atresia of antral follicles is associated with a reduction in the expression of AR, Arom and 17alpha. These data also suggest that loss of Arom precedes loss of 17alpha in AF.