Abstract
We recently demonstrated that caspase-3 is important for apoptosis during spontaneous involution of the corpus luteum (CL). These studies tested if prostaglandin F2α (PGF2α) or FAS regulated luteal regression, utilize a caspase-3 dependent pathway to execute luteal cell apoptosis, and if the two receptors work via independent or potentially shared intracellular signaling components/pathways to activate caspase-3. Wild-type (WT) or caspase-3 deficient female mice, 25–26 days old, were given 10 IU equine chorionic gonadotropin (eCG) intraperitoneally (IP) followed by 10 IU human chorionic gonadotropin (hCG) IP 46 h later to synchronize ovulation. The animals were then injected with IgG (2 micrograms, i.v.), the FAS-activating antibody Jo2 (2 micrograms, i.v.), or PGF2α (10 micrograms, i.p.) at 24 or 48 h post-ovulation. Ovaries from each group were collected 8 h later for assessment of active caspase-3 enzyme and apoptosis (measured by the TUNEL assay) in the CL. Regardless of genotype or treatment, CL in ovaries collected from mice injected 24 h after ovulation showed no evidence of active caspase-3 or apoptosis. However, PGF2α or Jo2 at 48 h post-ovulation and collected 8 h later induced caspase-3 activation in 13.2 ± 1.8% and 13.7 ± 2.2 % of the cells, respectively and resulted in 16.35 ± 0.7% (PGF2α) and 14.3 ± 2.5% TUNEL-positive cells when compared to 1.48 ± 0.8% of cells CL in IgG treated controls. In contrast, CL in ovaries collected from caspase-3 deficient mice whether treated with PGF2α , Jo2, or control IgG at 48 h post-ovulation showed little evidence of active caspase-3 or apoptosis. CL of WT mice treated with Jo2 at 48 h post-ovulation had an 8-fold increase in the activity of caspase-8, an activator of caspase-3 that is coupled to the FAS death receptor. Somewhat unexpectedly, however, treatment of WT mice with PGF2α at 48 h post-ovulation resulted in a 22-fold increase in caspase-8 activity in the CL, despite the fact that the receptor for PGF2α has not been shown to be directly coupled to caspase-8 recruitment and activation. We hypothesize that PGF2α initiates luteolysis in vivo, at least in part, by increasing the bioactivity or bioavailability of cytokines, such as FasL and that multiple endocrine factors work in concert to activate caspase-3-driven apoptosis during luteolysis.
Highlights
Prostaglandin F2α (PGF2α) has been implicated as a luteolysin in a number of mammalian species [1,2,3]
In striking contrast to the results with WT mice, there was no significant increase in the percentage of transferase-mediated dUTP nick end-labeling (TUNEL)-positive cells in corpus luteum (CL) derived from caspase-3 deficient female mice treated at 48 h post-ovulation with Jo2 (Fig. 1D; 0.32 ± 0.17) or PGF2α (Fig. 1F; 0.22 ± 0.08) when compared with IgG-treated controls (Fig. 1B; 0.06 ± .003)
There was no evidence of CM1-positive cells in CL derived from caspase3 deficient female mice treated at 48 h post-ovulation with IgG (Fig. 2B), Jo2 (Fig. 2D) or PGF2α (Fig. 2F)
Summary
Prostaglandin F2α (PGF2α) has been implicated as a luteolysin in a number of mammalian species [1,2,3]. FAS immunostaining is observed in human granulosa-lutein cells during the early luteal phase, and progressively intensifies during the mid-luteal phase through the late luteal phase [17]. This expression pattern is observed in the CL of mice [18] and rats [19,20]. In keeping with the proposal that FAS plays a role in luteolysis, in vitro studies have shown that FasL or FAS-activating antibodies induce luteal cell death in the human [10], mouse [5,18], rat [19,20] and cow [21]. Limited in vivo work has demonstrated that intravenous or intraperitoneal administration of FAS-activating antibody causes luteolysis in the mouse [18]
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