Original Research Article
Different expression of PGE synthase, PGF receptor, TNF, Fas and oxytocin in the bovine corpus luteum of the estrous cycle and pregnancy

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Abstract

Functional differences between the corpus luteum (CL) of pregnancy and CL of the cycle in cows were examined. Messenger RNA and protein levels of prostaglandin (PG) E synthase (PGES), PGF2α receptor (PGFR), tumor necrosis factor-α (TNF) and Fas were found to be higher in the CL of pregnancy than in CL of the cycle. Oxytocin (OT) mRNA and protein levels were lower in the CL of pregnancy. Messenger RNA levels of progesterone receptor (PR), luteinizing hormone receptor (LHR), PGE2 receptor (PGER), PGF synthase (PGFS), TNF receptor type I (TNFRI) and TNF receptor type II (TNFRII) did not differ between the cycle and pregnancy. PGE2 and PGF2α production by cultured bovine endometrial tissues was decreased by a supernatant derived from the homogenized CL of pregnancy but not by that of the CL of the cycle, suggesting that specific substances in the CL of pregnancy affect endometrial PG production in cows. Collectively, PGES, PGFR, TNF, Fas or OT may contribute to differences between the CL of pregnancy and CL of the estrous cycle in cows.

Introduction

The corpus luteum (CL) is a transient ovarian organ established by cells of the follicle following ovulation. The primary product of the CL, progesterone (P4), is required for the establishment and maintenance of pregnancy. If pregnancy does not occur, the CL degenerates. On the other hand, when pregnancy is established, the CL lifespan is prolonged and the CL continues to produce P4 during the gestation period [1]. The bovine CL produces not only P4, but also various other intraluteal factors, including prostaglandins (PG), oxytocin (OT) and cytokines [1]. In particular, PGE2 and PGF2α are well known as potent luteal P4 regulators in cows [2], [3], [4], [5]. Intraluteal OT is a key regulator that induces PGF2α production by the endometrium [6], and tumor necrosis factor-α (TNF) and Fas stimulate luteal PG production and induce apoptosis of luteal cells at the time of luteolysis [7]. This indicates that these intraluteal factors play important roles in regulating reproductive activity in an interaction-based manner.

Although the CL is required during the early phase of pregnancy in all mammals, in some species (e.g., cows, pigs and dogs) it is required for the entire gestation [8]. In other species (e.g., primates), the CL is not required for the entire gestation, because luteal P4 secretion can be replaced by placental P4 secretion [9]. Many differences have been observed between the CL of the estrous cycle and pregnancy. Corpora lutea of the cycle and pregnancy are morphologically different in cows [10], [11], sheep [12], [13], and pigs [14]. In pregnant sheep, volume densities of steroidogenic small and large luteal cells gradually increase, peaking between days 60 and 142 of pregnancy [12], [13]. The gene expression of PGE and PGF synthases in pigs is higher in the CL of pregnancy than in the CL of the cycle [15]. The expression of PGE and PGG and the activities of copper–zinc superoxide dismutase (Cu, Zn-SOD) are higher in the human CL of pregnancy than in the CL of the midcycle [16]. In the pig, the activity of glutathione peroxidase is higher and the activity of SOD is lower in the CL of pregnancy than in the CL of the cycle [17]. In sheep, the response of small luteal cells to luteinizing hormone (LH) was weaker at days 40–50 of pregnancy than at day 25 of pregnancy [13], [18].

In the cow, the CL of pregnancy produces more PGE2 [19] and less OT [20] than the CL of the cycle. Additionally, the binding sites of PGF2α [21] and TNF [22] have been demonstrated in the CL of pregnancy. However, a systematic approach to investigate the functional differences between the CL of pregnancy and the CL of the cycle has not yet been demonstrated. The objective of the present study was to determine possible differences in gene expression and protein concentration of major intraluteal factors between the CL of pregnancy and CL of the cycle. In addition, we examined whether supernatants derived from the homogenized CL of pregnancy and the CL of the cycle differentially affected PGE2 and PGF2α secretion by bovine endometrial tissues in vitro.

Section snippets

Collection of the bovine corpora lutea and uteri

Bovine ovaries containing corpora lutea (CLs) and uteri were obtained from Japanese-Black cows in the institute ranch within 10–30 min of exsanguination. Tissue samples were collected from cows on day 8–12 of the estrous cycle (cyclic), day 20–30 (early I), day 40–50 (early II), and day 150–165 (mid) of gestation (n = 4 animals/stage). The day of artificial insemination was designated as day 1 of gestation. The CLs were immediately separated from the ovaries and then cut into small pieces (<0.8 cm3

Results

Specific transcripts for all tested substances were detected in the bovine CL of the estrous cycle and each stage of pregnancy. PR, LHR, PGER and PGFS mRNA expression did not differ between cycle and pregnancy (Fig. 1A–C and F). Transcripts of PGFR and PGES were more abundant (p < 0.05) in the mid-stage (PGFR) and early I stage (PGES) of the CL of pregnancy than in the CL of the cycle (Fig. 1D and E). The expression of TNF mRNA was higher in the CL of pregnancy from the early I and mid-stage than

Discussion

The results of the current study demonstrated that in cows the gene expression and protein concentration of intraluteal factors in the CL of pregnancy differ from those in the CL of the cycle. Similarly, the effects of the supernatant derived from homogenized CL on PG production by endometrial tissues varied dependent on the CL origin (cycle or pregnancy). This suggests that in cows the CL of pregnancy has different properties than the CL of the cycle.

As expected, luteal gene expression of PR

Authors’ contributions

RS participated in the design of the study, collected the materials, carried out all experiments and drafted the manuscript. KGH and TT were responsible for all animal care, collected the materials and helped to carry out a part of experiments.

Conflict of interest

The authors declare that they have no competing interests.

Acknowledgments

This research was supported by Grants-in-Aid for Scientific Research (No. 25780276) from the Japanese Society for the Promotion of Science (JSPS), and Research Program on Innovative Technologies for Animal Breeding, Reproduction, and Vaccine Development (REP-1001) from the Ministry of Agriculture, Forestry and Fisheries of Japan.

The authors thank the staff of the National Institute of Livestock and Grassland Sciences, Japan, for their skilled technical assistance.

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    Present address: Laboratory of Theriogenology, Faculty of Agriculture, Iwate University, Iwate 020-8550, Japan.

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