Interferon-tau infusion into the ovine corpus luteum delays luteolysis†

Abstract

Conceptus-derived interferon-tau (IFNT) initiates maternal recognition of pregnancy in ewes by paracrine actions on the endometrium and endocrine action on the corpus luteum (CL). To examine the effect of IFNT on the CL without inducing IFN-stimulated genes (ISGs) in the endometrium, recombinant ovine IFNT (roIFNT) or bovine serum albumin was delivered directly into CLs via osmotic pumps at a rate of 10, 50, or 100 ng/h from days 9 to 12 of the estrous cycle. Endometrial and CL samples were collected on day 12. 50 ng/h of roIFNT induced ISG15 in the CL on day 12 without affecting endometrial ISG15 concentrations. In a second experiment, roIFNT (50 ng/h) was infused into the CL from days 10 to 17 of the estrous cycle and serum samples were collected daily. Serum progesterone concentrations were significantly higher from days 15 to 17 in roIFNT-infused ewes compared to controls. Levels of LHCGR, STAR, CYP11A1, HSL, OPA1, and protein kinase A mRNA and proteins were higher in the roIFNT-infused CLs compared to the controls. Levels of ISG15 and MX1 mRNA increased in the CLs of roIFNT-infused ewes but not in the endometrium. Endometrial ESR1 mRNA and protein concentrations were higher in the controls compared to roIFNT-infused ewes. In conclusion, intra-luteal delivery of roIFNT induced ISGs, stabilized steroidogenesis in the CL, and delayed luteolysis without inducing endometrial IFN-stimulated genes. Inhibition of ESR1 in the endometrium of roIFNT-infused ewes was observed suggesting that direct delivery of IFNT to the CL has an additional anti-luteolytic effect on the endometrium.

Keywords: corpus luteum; interferon-tau; ovine; pregnancy; steroidogenesis.

Graphical Abstract

Figure 1

Graphical demonstration of IFNT-infusing osmotic pump installation to the CL.

Figure 2

Levels of ISG15 mRNA in the luteal (A) and endometrial (B) tissues following the direct delivery of BSA (50 ng/h) (n = 6) or roIFNT 10 (n = 3), 50 (n = 4), or 100 ng/h (n = 3) by the osmotic pumps directly into the targeted CL from days 9 to 12 in Experiment 1.

Figure 3

Representative images of immunohistochemical localization of ISG15 (A) and western blot of free and conjugated ISG15 proteins (B) in ovine CL on day 17 following the intra-luteal delivery (50 ng/h) of BSA or IFNT from days 10 to 17 in Experiment 2. (Micron bar represents 1 mm, 40x and inset is 10x). (C), (D), and (E) show luteal ISG15 mRNA, free, and conjugated ISG15 protein concentrations on day 17 following the infusions, ^*P < 0.05, ^**P < 0.01. RU, densitometry of target adjusted for densitometry of ACTB (β-actin).

Figure 4

(A) Serum progesterone concentrations following intra-luteal infusion (50 ng/h) of BSA (n = 6) or roIFNT (n = 10) from day 10 (surgery) until day 17 (necropsy) in Experiment 2. ^*P < 0.05, ^**  P < 0.01. (B) AUC of the percent change in serum progesterone (P < 0.01). Serum progesterone concentrations are expressed as a percentage of the initial progesterone concentration on day 13 when the blood collection began.

Figure 5

Levels of LHCCR (A), STAR (B), CYP11A1 (C), PTGFR (D), and PTGS2 (E) mRNA in the luteal tissues on day 17 of the estrous cycle following the infusions in Experiment 2. ^**P < 0.01, ^***P < 0.001.

Figure 6

Protein concentrations of steroidogenic regulators: LHCGR (A), STAR (B), CYP11A1 (C), HSL (D), OPA1 (E), and PKA substrate (F) in the luteal tissues on day 17 of the estrous cycle following the infusion in Experiment 2. ^*P < 0.05, ^**P < 0.01, ^***P < 0.001. RU, densitometry of target adjusted for densitometry of ACTB (β-actin).

Figure 7

Levels of ESR1 mRNA (A) and protein (B) in the endometrial tissues on day 17 of the estrous cycle following the infusion in Experiment 2. ^**P < 0.01. RU, densitometry of target adjusted for densitometry of ACTB (β-actin).