Abstract: The purpose of this experiment was to study the effects of PGF2α injection at different luteal stages on luteal tissue morphology, expression of PGF2α receptors and programmed necroptosis-associated genes in Hu sheep. Forty eight healthy ewes with similar body weight were used in this study, which were in good condition and have undergone estrus synchronization. They were randomly divided into 6 groups, namely the early-luteal phase experiment group and control group, the mid-luteal phase experiment group and control group, and the late-luteal phase experiment group and control group. The ewes in 3 experimental groups were injected with 1 mL PGF2α (0.1 mg) on day 6 (early-luteal phase), day 11 (mid-luteal phase) and day 16 (late-luteal phase), respectively. The ewes in 3 control groups were injected with 1 mL normal saline at the same time. The corpus luteum tissue was collected at 3 h after each injection for gene expression detection and HE staining for the detection of morphological changes of corpus luteum. The results showed that, after the injection of PGF2α, the morphology of corpus luteum changed, with inflammatory cell infiltration and apoptosis, especially in the mid-luteal phase, with a higher degree of degeneration. For ewes that were not injected with PGF2α, the mRNA expression level of FPA in the corpus luteum in the early-luteal phase was significantly higher in the early-luteal and mid-luteal phases compared to the late-luteal phase (P<0.05); the FPB expression level in the corpus luteum in the early-luteal phase was significantly higher than that in the late-luteal phase (P<0.05), and there was no significant difference from the mid-luteal phase (P>0.05); the expression level of TNFR1 in the mid-luteal phase was significantly higher than that in the early-luteal and late-luteal phases (P<0.05); The expression level of RIPK3 in the mid-luteal phase was significantly higher than that in the late-luteal phase (P<0.05), and both of them was no significant difference to the early-luteal phase (P>0.05); TNF-α, RIPK1 and MLKL expression levels did not significantly change throughout the luteal phase (P>0.05). In the early-luteal phase, injection of PGF2α significantly reduced the expression of FPA (P<0.05), and significantly increased the expression of FPB, RIPK1, MLKL (P<0.05), and had no significant effect on the expression of TNF-α, TNFR1, and RIPK3 (P>0.05).In the mid-luteal phase, injection of PGF2α could significantly up-regulate the expression of FPB, TNF-α, RIPK1 and MLKL (P<0.05), but had no significant effect on the expression of FPA, TNFR1 and RIPK3 (P>0.05); In the late-luteal phase, injection of PGF2α could significantly up-regulate the expression of RIPK1 (P<0.05)), had no effect on the expression of FPA, FPB, TNF-α, RIPK3 and MLKL (P>0.05). The mRNA expression level of FPA was positively correlated with RIPK3 and TNFR1 (P<0.05), and the mRNA expression level of FPB was positively correlated with MLKL and TNF-α (P<0.05). The results show that PGF2α may initiate necroptosis through the TNF-α/TNFR1 pathway after binding to its receptors FPA and FPB, and promote the degeneration of the corpus luteum. FPB plays a major role in mediating PGF2α-induced luteal degeneration and necroptosis, and PGF2α has a better effect on inducing luteal degeneration in the mid-luteal phase.

Key words: ewe, prostaglandin F2α, luteal phase, necroptosis, prostaglandin F receptor