IN VIVO EFFECT OF GONADOTROPIN RELEASING HORMONE ANALOGS ON RAT OVARIAN FOLLICLE APOPTOSIS

PARBORELL, FERNANDA; ABRAMOVICH DALHIA; TESONE, MARTA

Quebec, Canadá

Congreso; 38 th Annual Meeting of Society for the Study of Reproduction (SSR); 2005

Society for the Study of Reproduction (SSR)

Gonadotropin-releasing hormone (GnRH) analogs: agonists (GnRH-a) or antagonists (GnRH-ant) have been widely used to inhibit gonadotropin pituitary release, though studies have also shown that GnRH has extrapituitary effects, particularly on the rat and human ovary. In previous works from our laboratory, we showed an apoptotic follicular action of GnRH-a correlated with an imbalance in the ratio of antiapoptotic:proapoptotic proteins (Bcl-xL/Bcl-xS), reducing the stability of the Bcl-xL. In addition, GnRH-ant inhibited the apoptosis in preovulatory follicles (POF) through a decrease of Bax traslocation to mitochondria. To elucidate whether these changes were mediated by effector caspases, the localization and the levels of caspase-3 were determined. Prepubertal rats superovulated with eCG (Control group) were injected during 48 h, each 12 h with GnRH-a (Leuprolide Acetate, 1µg/rat/day, LA group) and/or GnRH-ant (Antide, 10 ug/rat/day; LA+Ant group or Ant group). Healthy POF (>400 mm in diameter) from ovaries were dissected microscopically and analyzed to study the expression of the caspase-3 (inactive/active) by western blot using an anticaspase-3 antibody, which recognizes procaspase-3 and p17/cleavage, the active subunit of caspase-3. LA caused an significant increase in the level of the p17/cleavage product compared to the control group (DENSITOMETRIA). The co-injection of Ant interfered with this LA effect (DENSITOMETRIA) and Ant alone significantly decreased caspase-3 cleavage (DENSITOMETRIA). The treatments did not change the level of procaspase-3. In addition, it appeared an 20 kDa immunoreactive band which represents an intermediate cleavage product of caspase-3. Immunohistochemistry studies showed that immunostaining for caspase-3 was absent or weak in granulosa cells of preantral follicles (PF) and antral follicles (AF) at all treatments. The theca cells (TC) in PF exhibited a moderate signal for caspase-3 only in control and LA group. The TC from AF  showed immunoreactivity for caspase-3 in all groups, appearing more intense in LA group, and in addition in this group the staining was more evident in the nucleus of TC. We suggest that LA enhances the apoptosis in POF through an increase in caspase-3 expression, whereas Ant decreased it, and these changes mainly occur in TC.