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Reproduction (2004) 127 117-124
DOI: 10.1530/rep.1.00071
Copyright © 2004 Society for Reproduction and Fertility
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RESEARCH

Local interaction of prostaglandin F2{alpha} with endothelin-1 and tumor necrosis factor-{alpha} on the release of progesterone and oxytocin in ovine corpora lutea in vivo: a possible implication for a luteolytic cascade

M Ohtani1, S Takase2, M P B Wijayagunawardane3, M Tetsuka2 and A Miyamoto2

1 The Field Center of Animal Science and Agriculture and 2 Department of Agriculture and Life Science, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Japan and 3 Department of Animal Science, University of Peradeniya, Peradeniya, Sri Lanka

Correspondence should be addressed to A Miyamoto; Email: akiomiya{at}obihiro.ac.jp


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Acknowledgements
 References
 
Endothelin-1 (ET-1) and tumor necrosis factor-{alpha} (TNF{alpha}) participate in the cascade of luteolysis. Thus, in the present study the interactions of ET-1 and TNF{alpha} with prostaglandin F2{alpha} (PGF2{alpha}) on the release of progesterone and oxytocin (OT) within the corpus luteum (CL) were investigated. A microdialysis system (MDS) was surgically implanted in ovine CL (one MDS line/CL; 5–10 lines/ewe) formed after super-ovulation. A 4-h perfusion with PGF2{alpha} (0.01–1 µmol l -1) induced no clear effect on progesterone release, but acutely stimulated OT release in a dose-dependent manner. A perfusion of PGF2{alpha} (1 µmol l -1) increased ET-1 release over a period of 12 h. Two perfusions of ET-1 (0.1 µmol l-1) or a perfusion of ET-1 followed by TNF{alpha} (200 ng ml-1) decreased progesterone release (56–64% at 36–48 h). When the CL were pre-perfused with PGF2{alpha} (1 µmol l-1), two consecutive perfusions of ET-1 decreased progesterone release more rapidly. Similarly, a pre-perfusion with PGF2{alpha} followed by consecutive perfusions of ET-1 and then TNF{alpha} rapidly decreased progesterone release, with the inhibition most pronounced (35%) at 36–48 h. The simultaneous infusion of ET-1 with PGF2{alpha} induced a rapid decrease in progesterone release (36% at 36–48 h). In a further study, the possible second messenger systems involved in PGF2{alpha} action on the release of progesterone, OT and ET-1 were investigated. A perfusion with 12-O-tetradecanoyl-phorbol-13-acetate (TPA; 10 µmol l-1), A23187 [GenBank] (10 µmol l-1), or PGF2{alpha} + A23187 [GenBank] increased progesterone release during infusion, but decreased it after perfusion. All treatments induced a massive release of OT during infusion, and increased ET-1 release after infusion. These results show that ET-1 is capable of suppressing progesterone release in the PGF2{alpha}-primed ovine CL in vivo and thus ET-1 works as a local luteolysin together with PGF2{alpha} during the process of functional luteolysis. During structural luteolysis, TNF{alpha} may interact with PGF2{alpha} and ET-1 to cause a rapid drop in progesterone release and accelerate the process of luteolysis. This result supports the contention that ET-1 and TNF{alpha} interact with PGF2{alpha} as local luteolytic mediators in the ewe as previously suggested.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Acknowledgements
 References
 
It is well known that a local counter-current transfer of uterine prostaglandin F2{alpha} (PGF2{alpha}) primes the luteolytic cascade of bovine corpora lutea (CL). Recent evidence in the cow suggests that endothelin-1 (ET-1), a vaso-constrictive 21-amino acid peptide, interacts with PGF2{alpha} in the control of functional luteolysis (Girsh et al. 1996a,b, Miyamoto et al. 1997, Ohtani et al. 1998, Meidan et al. 1999). This hypothesis has been further supported by in vivo studies in the ewe. A single intra-luteal injection of BQ123, a highly specific antagonist of the endothelin type A receptor (ETA-R), at the mid luteal phase mitigated the luteolytic effect of PGF2{alpha} (Hinckley & Milvae 2001). This study also demonstrated that a systemic injection of ET-1 15 min after an injection of a sub-luteolytic dose of PGF2{alpha} potentiated the decrease in plasma progesterone concentration, which resulted in the return of estrus (Hinckley & Milvae 2001). Moreover, the administration of a luteolytic dose of PGF2{alpha} rapidly stimulated gene expression for ET-1 in ovine CL collected at mid-cycle (Hinckley & Milvae 2001). It is likely, therefore, that ET-1 released from micro-vascular endothelial cells in the CL and luteal cells (Levy et al. 2001, Schams et al. 2003) cooperates with PGF2{alpha} to initiate luteolysis by both directly inhibiting progesterone release (Girsh et al. 1996a, Miyamoto et al. 1997) via ETA-R (Girsh et al. 1996a, Hinckley & Milvae 2001) and probably by vasoconstriction of arterioles that results in an acute drop in progesterone release.

The presence of tumor necrosis factor-{alpha} (TNF{alpha}) and its receptors in the pig and bovine CL was observed by Okuda et al. (1999), Sakumoto et al. (2000) and Miyamoto et al. (2002) and suggested that TNF{alpha} plays a significant role in the process of luteolysis (Murdoch et al. 1988, Benyo & Pate 1992, Shaw & Britt 1995, Wuttke et al. 1998). Elevated local secretion of TNF{alpha} has been found in the regressing ovine (Ji et al. 1991) and bovine (Shaw & Britt 1995) CL. Moreover, it was reported that TNF{alpha} acts synergistically with PGF2{alpha} during the process of luteolysis in pig (Wuttke et al. 1997, 1998). However, the possible synergism among PGF2{alpha}, ET-1 and TNF{alpha} during luteolysis (Meidan et al. 1999) in the ewe is not well established. Thus, the present study was designed to examine the direct local effects and interaction of ET-1 and TNF{alpha} with PGF2{alpha} on the release of progesterone and oxytocin (OT) within the CL, by using an in vivo microdialysis system (MDS) implanted into the CL of ewes (Miyamoto et al. 1998a,b). This system allows cells to maintain the integrity of CL structures, and thus enables observations to be made of real-time local changes of different substances in the CL that may play a role in cell-to-cell communication. The study was extended further to examine the possible second messenger systems involved in PGF2{alpha} action on the release of progesterone, OT and ET-1 in the CL in vivo.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Acknowledgements
 References
 
Experimental design and animals
The experiment was carried out at the Field Centre of Animal Science and Agriculture, Obihiro University and the experimental procedures complied with the Guide for Care and Use of Agriculture Animals of Obihiro University.

The study used a multiple CL model to implant the MDS, as the CL formed after super-ovulation were shown to regress in response to a luteolytic injection of PGF2{alpha} in a way that was very similar to that of intact CL (Miyamoto et al. 1998a). This model enables us to examine in parallel several experimental infusions into the MDS implanted in the different CL (one MDS line/CL) within a ewe (Miyamoto et al. 1998a,b). The study comprised three experiments. The first experiment (n = 3 ewes) observed the effect of PGF2{alpha} (Sigma Chemical Co., St Louis, MO, USA) infused at concentrations of 0.01 µmol l-1, 0.1 µmol l-1, or 1 µmol l-1 into the MDS on the local release of progesterone, OT and ET-1. The second experiment (n = 5 ewes) examined the effects of infusions with PGF2{alpha} (1 µmol l-1), ET-1 (0.1 µmol l-1; Peptide Institute Inc., Osaka, Japan), and/or TNF{alpha} (200 ng ml-1, recombinant human TNF{alpha}; 2.55 x 106 JRU mg-1 protein, generously provided by Dainippon Pharm. Co. Ltd, Osaka, Japan) on the release of progesterone and OT. The third experiment (n = 4 ewes) compared the pharmacological effect PGF2{alpha} (10 µmol l -1), 12-O-tetradecanoyl-phorbol-13-acetate (TPA; 10 µmol l-1), ionophore A23187 [GenBank] (10 µmol l-1), and PGF2{alpha} + A23187 [GenBank] (all substances from Sigma Chemical Co.) on the release of progesterone, OT and ET-1.

The experiments were conducted in October to December, during the breeding season. Mature Suffolk ewes were kept in a paddock under natural day length and temperature, and were fed a diet of concentrates and hay with water available ad libitum. All animals had at least two normal estrous cycles (16 or 17 days) before being used. The estrous cycles were based on progesterone concentrations in plasma taken every 3 days. In order to induce super-ovulation, ewes were pretreated according to the regimen of Miyamoto et al. (1998a,b) with an intra-vaginal sponge containing 60 mg medroxyprogesterone acetate (MPA, Repromap: The Upjohn Co. International Ltd, NSW, Australia) for 12 days. Pregnant mare’s serum gonadotropin (200 IU) (Serotropin; Teikokuzoki Co., Tokyo, Japan) was injected intramuscularly on the evening of the 9th day after the insertion of sponges, together with the first follicle-stimulating hormone (FSH) (6 mg Antorin 10; FSH of porcine origin, Denka Chemical Co., Kawasaki, Japan) injection. The FSH treatment comprised six intramuscular injections given at approximately 12-h intervals in a decreasing dose regimen (6, 6, 4, 4, 2, 2 mg) to give a total dose of 24 mg. The MPA sponges were withdrawn on the morning of the 12th day after the insertion. An injection of 100 µg synthetic gonadotropin releasing hormone (GnRH) (Conceral; Takeda Chemical Co., Osaka, Japan) was intramuscularly administered 24 h after the removal of the MPA sponges. The administration time of the GnRH injection was considered to be day 0. Daily blood samples (5 ml) for progesterone analysis in all experiments were collected through a jugular venous catheter over the whole period of the experiment, and the plasma samples were stored at - 30 °C until assay.

Implantation of MDS into the CL
On day 7 after GnRH injection, several lines of MDS were surgically implanted into CL (one MDS line/CL) in both ovaries of animals as detailed earlier (Miyamoto et al. 1998a,b). The CL formed after super-ovulation (5–10 CL/ewe) were selected so as to have similar macroscopic characteristics (size, color and consistency). Each CL was penetrated with one capillary dialysis membrane (Fresenius SPS 900 Hollow Fibers, cut-off Mr 1000 kDa, 0.2 mm diameter, 3 mm long; Fresenius AG, St Wendel, Germany). The capillary was affixed to the surface of the CL by Histoacryl Blue (B Braun-Dexon GmbH, Spangenberg, Germany). Both ends of the capillary were glued to 20-cm-long pieces of silastic tubing (i.d. 0.3 mm) that were connected to pieces of Teflon tubing leading to the outside of the abdomen. The exteriorized bundle of afferent and efferent Teflon tubes was taped to the body from the abdomen to the back with an adhesive bandage (Benefix; Japan Sigmax, Tokyo, Japan). The animals were brought to the pen where they were chained immediately after surgery. For perfusion, one end of the tube was connected to a peristaltic pump and the other was routed to a fraction collector. The CL were perfused with Ringer’s solution at a flow rate of 1.5-ml/30 min/fraction throughout the experiments. The MDS lines of each animal were used for perfusion with substances on days 9 to 10. At least one line was used for Ringer’s solution only (control) in each animal.

After 30-h perfusion, fractions of the perfusate were collected every 30 min for 32 to 48 h on days 9–11. After a period of 8 h for observing the spontaneous release of progesterone (baseline), further fractions were collected according to the experimental design. The transfer capacity of the microdialysis membrane was about 0.1% of the concentration infused as determined for OT, ET-1 and TNF{alpha}, and about 1% for progesterone and PGF2{alpha} under the conditions used (Miyamoto et al. 1997, 1998a). At the end of the experiments, the ewes were ovariectomized, and the CL were fixed in Bouin’s solution, dehydrated in a graded series of ethanol, cleared in xylene and embedded in paraffin wax, and then processed for histology after hematoxylin–eosin staining. In most cases, little damage and no connective tissues such as fibroblasts were observed around the microdialysis capillary membrane.

Hormone determination
Progesterone, OT and ET-1 concentrations in the perfusate fractions from the MDS, and progesterone concentrations in plasma were determined with second-antibody enzyme immunoassays (EIAs) that were based on a competitive assay using a horseradish peroxidase-labeled progesterone (Miyamoto et al. 1992) or biotin-labeled peptides as tracers (Miyamoto et al. 1997). Progesterone concentrations in plasma samples were assayed after extraction by diethyl ether, but those in the MDS fractions were assayed directly. The standard curve ranged from 0.05 to 50 ng ml-1 and the ED50 of the assay was 1.8 ng ml -1. The intra- and interassay coefficients of variation were on average 6.2% and 9.3% respectively.

For peptide extraction, 1-ml samples were taken from every fraction, and every 8 consecutive 1-ml samples (corresponding to a 4-h period) were pooled. BSA was added to each of the pooled samples to a final concentration of 1 mg ml-1, and the solution was adjusted to pH 2.5 with 1 M acetic acid. The samples were then applied to a Sep-Pak C18 cartridge (Waters, Milford, MA, USA) according to the established method (Miyamoto et al. 1997). The MDS fractions were concentrated 40-fold as a result of the process. The recoveries of synthetic OT and ET-1 added to Ringer’s solution were 73% and 62% respectively (n = 60). The EIAs for OT and ET-1 were conducted as described previously (Miyamoto et al. 1997). The standard curve for OT ranged from 1.6 to 200 pg ml-1 and the ED50 of the assay was 21 pg ml-1. The intra- and interassay coefficients of variation of the OT assay were on average 6.2% and 8.6% respectively. The standard curve for ET-1 ranged from 9.7 to 5000 pg ml-1 and the ED50 of the assay was 450 pg ml -1. The intra- and interassay coefficients of variation of the ET-1 assay were on average 8.7% and 12.6% respectively.

Statistical analyses
The mean hormone concentrations in the first 8 h (progesterone was based on each 0.5-h fraction, and OT and ET-1 were based on each 4-h pooled fraction) were used to calculate the individual baseline, because of the large variation among individuals in the basal concentrations of each hormone released into the MDS. All hormone concentrations in the fractions were then expressed as a proportion of this individual baseline. This treatment enables an evaluation of the relative changes of hormonal values between the CL of different animals. The change in hormonal release after substance infusions was tested based on individual time points throughout the experiment as compared with the baseline. Means were analyzed by time-dependent repeated measures ANOVA followed by Student’s t-test. Where a comparison among several treatment groups during the same time period was needed, means were analyzed by ANOVA followed by Tukey–Kramer test. For the figures showing MDS data, all hormonal concentrations were expressed as a percentage of the baseline with an 8 h basis. The absolute concentrations of the hormones in the MDS fractions (means±S.E.M.) are given in the figure legends.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Acknowledgements
 References
 
The super-ovulation treatment induced multiple CL (16.6 ± 1.8; mean±S.E.M., n = 12). There was no significant change in plasma progesterone levels during the MDS experiments, but the concentrations depended on the number of CL formed in each animal (8.2–27.8 ng ml-1).

Intra-luteal release of progesterone, OT and ET-1 in response to PGF2{alpha} infusion
The release of hormones into the MDS from days 9 to 10 after GnRH was relatively constant over the experimental period. A 4-h perfusion with PGF2{alpha} at 0.01 µmol l-1, 0.1 µmol l-1, or 1 µmol l-1 via the MDS induced no clear effect on progesterone release, but acutely stimulated OT release in a dose-dependent manner (P < 0.05; Fig. 1Go). The increase in OT was followed by a sustained drop after the stimulation. A 4-h perfusion with PGF2{alpha} at 1 µmol l-1 increased ET release over a period of 12 h (P < 0.05; Fig. 2Go), but the lower doses of PGF2{alpha} at 0.01 µmol l -1 or 0.1 µmol l-1 did not affect ET-1 release.



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  		Figure 1 Effects of 4-h intraluteal application of PGF2{alpha} (0.01 µmol l -1, 0.1 µmol l-1 and 1 µmol l-1) at 8–12 h on the local release of progesterone and OT from microdialyzed CL in the ewe (n = 3). The data are expressed as a percentage of the baseline (0–8 h) value, and are the means ± S.E.M. of 6–7 CL/group. The baseline (100%) values of each hormone were 2.2 ± 0.2 ng ml-1 for progesterone and 0.74 ± 0.19 pg ml-1 for OT. *P < 0.05, ***P < 0.001 compared with baseline.

 


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  		Figure 2 Effects of 4-h intra-luteal application of PGF2{alpha} (1 µmol l -1) at 8–12 h on the local release of ET-1 from microdialyzed CL in the ewe (n = 3). The data are expressed as a percentage of the baseline (0–8 h) value, and are the means± S.E.M. of 5 CL/group. The baseline (100%) value of ET-1 release was 3.8 ± 0.8 pg ml-1. *P < 0.05 compared with baseline.

 
Effect of pre- and simultaneous exposure with PGF2{alpha} on the suppressing activity of ET-1 and TNF{alpha}
Two 4-h perfusions of ET-1 (0.1 µmol l-1) between 12 and 16 h and 24 and 28 h decreased progesterone release between 24 and 48 h (78.2 ± 2.8%; P < 0.05; Fig. 3Go upper panel). Likewise, a 4-h perfusion of ET-1 between 12 and 16 h and of TNF{alpha} (200 ng ml-1) between 24 and 28 h decreased progesterone release between 24 and 48 h (76.2. ± 8.4%; P < 0.05; Fig. 3Go upper panel). A 4-h perfusion with PGF2{alpha} (1 µmol l-1) between 8 and 12 h slightly decreased progesterone release at 36 to 48 h (79.5 ± 6.4%; P < 0.05; Fig. 3Go lower panel). When the CL were pre-perfused with PGF2{alpha} for 4 h between 8 and 12 h, two consecutive perfusions of ET-1 between 12 and 16 h and 24 and 28 h rapidly decreased progesterone release between 16 and 48 h (74.6 ± 5.9%; P < 0.05; Fig. 3Go lower panel); this decrease occurred 8 h earlier than in the CL treated with ET-1 alone. Similarly, a pre-perfusion with PGF2{alpha} for 4 h between 8 and 12 h, followed by consecutive perfusions of ET-1 between 12 and 16 h and of TNF{alpha} between 24 and 28 h, rapidly decreased progesterone release at 16 to 48 h (67.5 ± 7.0%; P < 0.05; Fig. 3Go lower panel), but the inhibitory effect between 36 and 48 h was more pronounced than it was in the above-described treatments (35.1 ± 3.3%; P < 0.05; Fig. 3Go lower panel). The simultaneous infusion of ET-1 and PGF2{alpha} between 8 and 12 h induced a rapid and pronounced decrease in progesterone release (65.0 ± 6.5%; P < 0.05; Fig. 3Go lower panel). ET-1 alone slightly stimulated OT release (403.7 ± 72.6%; P < 0.05; Fig. 4Go upper panel). An infusion of PGF2{alpha} between 8 and 12 h induced an acute release of OT (628.2 ± 100.4%; P < 0.05; Fig. 4Go lower panel). The simultaneous infusion of ET-1 and PGF2{alpha} induced the highest stimulation in OT release (897.7 ± 70.7%; P < 0.05; Fig. 4Go lower panel), followed by a drop for the next 12 h (52.9 ± 16.0%; P < 0.05). ET-1 or TNF{alpha} after PGF2{alpha} also stimulated OT release (666.9 ± 123.4; P < 0.05; Fig. 4Go lower panel).



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  		Figure 3 Effects of intra-luteal application of ET-1 (0.1 µmol l-1) and TNF{alpha} (200 ng ml-1) with (lower panel) or without (upper panel) PGF2{alpha} (0.1 µmol l-1) on the local release of progesterone from microdialyzed CL in the ewe (n = 5). The data are expressed as a percentage of the baseline (0–8 h) value, and are the means± S.E.M. of 5–8 CL/group. The baseline (100%) value of progesterone release was 2.8 ± 0.3 ng ml-1. *P < 0.05, **P < 0.01, ***P < 0.001 compared with baseline.

 


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  		Figure 4 Effects of intra-luteal application of ET-1 (0.1 µmol l-1) and TNF{alpha} (200 ng ml-1) with (lower panel) or without (upper panel) PGF2{alpha} (0.1 µmol l-1) on the local release of OT from microdialyzed CL in the ewe (n = 5). The data are expressed as a percentage of the baseline (0–8 h) value, and are the means± S.E.M. of 5–7 CL/group. The baseline (100%) value of OT release was 0.67 ± 0.08 pg ml-1. *P < 0.05, ** P < 0.01, ***P < 0.001 compared with baseline.

 
Comparison of the effects of PGF2{alpha}, TPA and A23187
A 6-h perfusion with PGF2{alpha} (10 µmol l - 1) between 8 and 14 h slightly increased progesterone release during infusion (P < 0.05), but did not affect it during the 16-h period after infusion (14–30 h). A 6-h perfusion with TPA (10 µmol l-1), A23187 [GenBank] (10 µmol l-1), or PGF2{alpha} together with A23187 [GenBank] between 8 and 14 h increased progesterone release during infusion more than with PGF2{alpha} alone (P < 0.05), but induced a decrease in progesterone release after infusion, in the order A23187 [GenBank] (89%), TPA (67%), and PGF2{alpha} + A23187 [GenBank] (52%; Table 1Go). All three values were significantly different from each other (P < 0.05). All treatments induced a massive release of OT during infusion (P < 0.05), whereas only A23187 [GenBank] and PGF2{alpha} + A23187 [GenBank] depressed OT release after infusion (P < 0.05). All treatments similarly increased ET-1 release after infusion (P < 0.05; Table 1Go).


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  		Table 1 Comparison of secretion rates of progesterone, OT, and ET-1 into the MDS implanted in the CL formed after super-ovulation among 5 treatment groups during (8–14 h) and after (14–30 h) stimulant infusion at 10 µmol l-1 in the ewe (n = 4). Values are expressed as a percentage (means± S.E.M.) of the respective baseline (0–8 h). The baseline (100%) of progesterone release was 5.13 ± 0.14 ng ml-1, of OT release was 0.89 ± 0.17 pg ml-1, and of ET-1 release was 3.1 ± 0.9 pg ml-1.
 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Acknowledgements
 References
 
The results of the present study indicate that PGF2{alpha} acts cooperatively with ET-1 and TNF{alpha} to suppress progesterone secretion in the ovine CL in vivo. Substances infused through the MDS act in the local microenvironment around the capillary membrane. Thus, the model used here does not have any major effect on the constriction of blood vessels in the whole CL, and hence excludes a direct effect on the vasoconstriction that was shown to be a crucial process for luteolysis (Niswender et al. 1976, Azmi & O-Shea 1984, Knickerbocker et al. 1988, Acosta et al. 2002). In this context, the present in vivo data may give basic and important information on the interaction of PGF2{alpha} with ET-1 and TNF{alpha} in the cascade of functional luteolysis.

It was observed in the present study that a single infusion of PGF2{alpha} for 4 h did not change progesterone release in the CL up to 20 h after the infusion; notwithstanding the results of our similar in vivo investigation that a PGF2{alpha} infusion to the bovine mid-cycle CL via MDS stimulated, but did not inhibit progesterone secretion (Ohtani et al. 1999). These findings suggest that the countercurrent transfer of uterine or exogenous PGF2{alpha} to the ovary plays a crucial role in inducing a rapid drop in progesterone release from the CL during luteolysis. On the other hand, the present study showed that a high dose of PGF2{alpha} (1 µmol l-1) could increase ET-1 release from the CL. This fact further supports the concept that a vasoconstriction of arterioles in the CL (Niswender et al. 1976, Azmi & O-Shea 1984, Knickerbocker et al. 1988) at the beginning of luteolysis may be attributed to the strong effect of ET-1 as well as of PGF2{alpha}. It was observed in the present study that the simultaneous exposure of CL to PGF2{alpha} and ET-1 was much more effective in reducing progesterone release than was the sequential exposure to these two compounds. Indeed, PGF2{alpha} stimulates ET-1 release from ovine luteal cells (Hinckley & Milvae 2001) and it is this elevated local ET-1, stimulated by PGF2{alpha}, together with the infused ET-1 which collectively act on the luteal cells, rather than a direct action of PGF2{alpha}. This might result in a greater progesterone suppression when CL are exposured simultaneously to PGF2{alpha} and ET-1. Our recent report revealed that PGF2{alpha} and ET-1 cooperatively induce functional luteolysis of mid-cycle CL in the cow (Miyamoto et al. 2001). We recently proposed that another vasoconstrictive peptide, angiotensin II (Ang II), might have a similar role. As in the case for ET-1, PGF2{alpha} increases luteal Ang II release, and Ang II acts with PGF2{alpha} as a suppressor of progesterone release in the bovine CL (Hayashi & Miyamoto 1999, Hayashi et al. 2002). In both vasoactive peptides, a concomitant perfusion of CL with PGF2{alpha} had the greatest effect on progesterone suppression as observed in the present in vivo study, suggesting that these peptides are important luteal factors in inducing functional luteolysis.

Interestingly, PGF2{alpha} did not suppress progesterone release in the present MDS model, but TPA, a stimulator of protein kinase C (PKC) clearly suppressed it. Moreover, TPA but not PGF2{alpha} inhibited basal and low density lipoprotein-supported steroidogenesis in cultured regressing porcine luteal cells (Brannian et al. 1995). Thus, TPA and PGF2{alpha} have different effects on the CL. Stimulation of PKC in ovine large luteal cells was shown to be a dominant intracellular mechanism involved in progesterone suppression (Wiltbank et al. 1990). This was confirmed later using the in vivo model in which an infusion of TPA into the ovarian artery also decreased plasma progesterone concentrations (McGuire et al. 1994). Our present in vivo data further support this concept. It appears that TPA infusion into the CL affects luteal progesterone release but differently in cows and ewes: TPA inhibited progesterone release in the ewe (the present result), while it did not affect progesterone release in the cow (Ohtani et al. 1999). On the other hand, an infusion of A23187 [GenBank] slightly inhibited progesterone release in the ewe (the present result), but strongly inhibited it in the cow (Ohtani et al. 1999). PGF2{alpha} has been shown to increase the intracellular concentration of Ca2+ in the luteal cells in both species (Davis et al. 1987, Wiltbank et al. 1989, Wegner et al. 1991). Thus, there might be some difference between ewes and cows in cell sensitivity to PGF2{alpha} at the second messenger level. This may be especially true in large luteal cells, in which a PKC pathway is dominant in the ewe, whereas an increase in Ca2+ is a crucial function in the cow.

As a result of pharmacological stimulation of PKC and a calcium influx in the CL cells in vivo, a local release of OT was acutely stimulated during substance infusion while the ET-1 release was gradually stimulated after infusion. These data support the concept that Ca2+ and PKC are responsible for the release of ET-1 and OT (Cosola-Smith et al. 1990, Masaki 1993, Miyamoto et al. 1993). These data also support the concept that OT is released from large luteal cells by granule exocytosis (Cosola-Smith et al. 1990, Wathes & Denning-Kendall 1992), while ET-1 secretion is a result of transcriptional activation of prepro-ET-1 followed by cleavage steps from big-ET-1 to ET-1 in endothelial cells (Masaki 1993).

Our previous observation of a direct local effect of TNF{alpha} using the MDS implanted in the intact mid-cycle CL in the ewe showed that an infusion of TNF{alpha} alone induced a slight increase in the release of progesterone and PGF2{alpha}, but it did not inhibit progesterone release (Miyamoto et al. 1995). The effect of TNF{alpha} infusion was more or less similar when TNF{alpha} was perfused into the MDS implanted in the regressing CL in the same model (multiple CL) as the present study (Miyamoto et al. 1998a). However, the present results clearly indicated that TNF{alpha} is capable of decreasing progesterone release in the CL in vivo, if the CL is pre-treated by PGF2{alpha} and ET-1. As an increase in local secretion of TNF{alpha} has been found in the regressing CL only after the completion of functional luteolysis in the ovine CL (Ji et al. 1991) and bovine CL (Shaw & Britt 1995), the timing of the exposure of CL to TNF{alpha} may have a physiological meaning; TNF{alpha} may indeed be an effective suppressor of progesterone release when the luteal cells are stimulated by PGF2{alpha} and ET-1 in vivo. In fact, a similar phenomenon was observed in an MDS study in pigs in which TNF{alpha} and PGF2{alpha} synergistically inhibited steroidogenesis (Wuttke et al. 1997, 1998). Therefore, a lack of up-regulation of TNF{alpha} mRNA in the regressing CL (Penny et al. 1999, Petroff et al. 1999) may not exclude the possibility of such a synergism among PGF2{alpha}, ET-1 and TNF{alpha}. Consequently, we speculate that during structural luteolysis, TNF{alpha} inhibits progesterone release, and at the same time it further stimulates a local release of ET-1, since endothelial cells are identified as target cells of cytokines and as cytokine-producing cells (Martin & Resch 1988). ET-1, in turn, may stimulate TNF{alpha} secretion (Cunningham et al. 1997, Ruetten & Thiemermann 1997), thereby establishing a local positive feedback, which would accelerate the cascade of luteolysis. Indeed, the intra-luteal ET-1 concentration was shown to be maintained at a high level over the whole period of luteolysis in the cow (Ohtani et al. 1998). In this regard, Meidan et al. (1999) proposed the same hypothesis on the basis of their results showing that ET-1 induces secretion of TNF{alpha} by bovine macrophages, and that luteal cells express the p55 type receptor for TNF{alpha}. Moreover, mRNA for TNF{alpha} was detected in luteal tissue around natural and induced luteolysis (Penny et al. 1999) and this increase was associated with an influx of macrophages into the CL around the time of the second half of luteolysis, possibly in response to monocyte chemoattractant protein (Webb et al. 2002). Thus, the TNF{alpha} secreted by luteal cells and macrophages during structural luteolysis may interact with PGF2{alpha} and ET-1, resulting in a rapid drop in progesterone release and accelerating the process of luteolysis.

Collectively, the present results provide further evidence that ET-1 is capable of suppressing progesterone release in the PGF2{alpha}-primed ovine CL in vivo. This observation supports the previous finding that the systemic administration of ET-1 potentiates the progesterone-suppressing activity of PGF2{alpha} in the ewe (Hinckley & Milvae 2001). The in vivo data obtained in the ewe (Hinckley & Milvae 2001, present study), as well as the in vitro data obtained from the cow (Miyamoto et al. 1997) suggest that ET-1 alone cannot promote functional luteolysis. ET-1 works as a local luteolysin together with PGF2{alpha}, and the CL needs to be triggered by PGF2{alpha} to start the cascade of luteolysis in which ET-1 acts in response to the first stimuli of PGF2{alpha}, causing a rapid drop in progesterone release. In conclusion, ET-1 and TNF{alpha} together with PGF2{alpha} are capable of directly suppressing the local progesterone release in the ovine CL in vivo.


   Acknowledgements
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Acknowledgements
References
 
This study was supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science, Sport and Culture of Japan, the 21st Century COE Program (A-1), Ministry of Education, Culture, Science and Technology, Japan and the Mishima Kaiun Foundation for the Promotion of Science. The authors wish to thank Dr K Okuda, Okayama University, for the progesterone antiserum, Dr D Schams, Technical University of Munich, Germany, for the ET antiserum, Dr T Higuchi, Kochi University of Medicine, Japan, for the OT antiserum, and Fresenius AG, St Wendel, Germany, for the microdialysis capillary membrane. Dr M P B Wijayagunawardane is a postdoctoral fellow supported by Japan Society for the Promotion of Science (JSPS).


   Footnotes
 
Received 30 June 2003
First decision 28 August 2003
Accepted 1 October 2003


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Acknowledgements
 References
 

Acosta TJ, Yoshizawa N, Ohtani M & Miyamoto A 2002 Local changes in blood flow within the early and midcycle corpus luteum after prostaglandin F2{alpha} injection in the cow. Biology of Reproduction 66 651–658.[Abstract/Free Full Text]

Azmi TI & O-Shea JD 1984 Mechanism of deletion of endothelial cells during regression of the corpus luteum. Laboratory Investigation 51 206–217.[ISI][Medline]

Benyo DF & Pate JL 1992 Tumor necrosis factor-{alpha} alters bovine luteal cell synthetic capacity and viability. Endocrinology 130 854–860.[Abstract]

Brannian JD, Christianson H, Flynn S & Kurz SG 1995 Loss of low-density lipoprotein utilization by regressing porcine luteal cells: effects of protein kinase C activation. Biology of Reproduction 52 793–797.[Abstract]

Cosola-Smith CA, Appell LH, Abdelgadir SE & Stormshak F 1990 Stimulation of bovine luteal oxytocin secretion in vitro by a phorbol ester and calcium ionophore. Journal of Animal Science 68 2465–2470.[Abstract]

Cunningham ME, Huribal M, Bala RJ & McMillen MA 1997 Endothelin-1 and endothelin-4 stimulate monocyte production of cytokines. Critical Care Medicine 25 958–964.[CrossRef][ISI][Medline]

Davis JS, Weakland LL, Weiland DA, Farese RV & West LA 1987 Prostaglandin F2{alpha} stimulates phosphatidylinositol 4,5-bisphosphate hydrolysis and mobilizes intracellular Ca2+ in bovine luteal cells. PNAS 84 3728–3732.[Abstract/Free Full Text]

Girsh E, Milvae RA, Wang W & Meidan R 1996a Effect of endothelin-1 on bovine luteal cell function: role in prostaglandin F2{alpha}-induced antisteroidogenic action. Endocrinology 137 1306–1312.[Abstract]

Girsh E, Wang W, Mamluk R, Arditi F, Friedman A, Milvae RA & Meidan R 1996b Regulation of endothelin-1 expression in the bovine corpus luteum: elevation by prostaglandin F2{alpha}. Endocrinology 137 5191–5196.[Abstract]

Hayashi K & Miyamoto A 1999 Angiotensin II interacts with prostaglandin F2{alpha} and endothelin-1 as a local luteolytic factor in the bovine corpus luteum in vitro. Biology of Reproduction 60 1104–1109.[Abstract/Free Full Text]

Hayashi K, Tanaka J, Hayashi KG, Hayashi M, Ohtani M & Miyamoto A 2002 The cooperative action of angiotensin II with subluteolytic administration of PGF2{alpha} in inducing luteolysis and oestrus in the cow. Reproduction 124 311–315.[Abstract]

Hinckley ST & Milvae RA 2001 Endothelin-1 mediates prostaglandin F2{alpha}-induced luteal regression in the ewe. Biology of Reproduction 64 1619–1623.[Abstract/Free Full Text]

Ji I, Slaughter RG, Ellis JA, Ji TH & Murdoch WJ 1991 Analyses of ovine corpora lutea for tumor necrosis factor mRNA and bioactivity during prostaglandin-induced luteolysis. Molecular and Cellular Endocrinology 81 77–80.[CrossRef][ISI][Medline]

Knickerbocker JJ, Wiltbank MC & Niswender GD 1988 Mechanisms of luteolysis in domestic livestock. Domestic Animal Endocrinology 5 91–107.[CrossRef][ISI][Medline]

Levy N, Gordin M, Mamluk R, Yanagisawa M, Smith MF, Hampton JH & Meidan R 2001 Distinct cellular localization and regulation of endothelin-1 and endothelin-converting enzyme-1 expression in the bovine corpus luteum: implications for luteolysis. Endocrinology 142 5254–5260.[Abstract/Free Full Text]

McGuire WJ, Juengel JL & Niswender GD 1994 Protein kinase C second messenger system mediates the antisteroidogenic effects of prostaglandin F2{alpha} in the ovine corpus luteum in vivo. Biology of Reproduction 51 800–806.[Abstract]

Martin M & Resch K 1988 Interleukin 1: more than a mediator between leukocytes. Trends in Pharmacological Sciences 9 171–177.[CrossRef][Medline]

Masaki T 1993 Endothelins: homeostatic and compensatory actions in the circulatory and endocrine systems. Endocrine Reviews 14 256–268.[CrossRef][ISI][Medline]

Meidan R, Milvae RA, Weiss S, Levy N & Friedman A 1999 Intraovarian regulation of luteolysis. Journal of Reproduction and Fertility (Supplement) 54 217–228.[CrossRef]

Miyamoto A, Okuda K, Schweigert F & Schams D 1992 Effects of basic fibroblast growth factor, transforming growth factor-ß and nerve growth factor on the secretory function of the bovine corpus luteum in vitro. Journal of Endocrinology 135 103–114.[Abstract]

Miyamoto A, von Lutzow H & Schams D 1993 Acute actions of prostaglandin F2{alpha}, E2, and I2 in microdialyzed bovine corpus luteum in vitro. Biology of Reproduction 49 423–430.[Abstract]

Miyamoto A, Tashiro Y, Nakatsuka T, Meyer H, Taguchi K, Abe N & Fukui Y 1995 Effect of tumor necrosis factor-{alpha} and interleukin-1 on local release of progesterone, prostaglandin F2{alpha} and oxytocin in microdialyzed ovine corpus luteum in vitro. Assisted Reproductive Technology/Andrology 8 21–32.

Miyamoto A, Kobayashi S, Arata S, Ohtani M, Fukui Y & Schams D 1997 Prostaglandin F2{alpha} promotes the inhibitory action of endothelin-1 on the bovine luteal function in vitro. Journal of Endocrinology 152 R7–R11.[Abstract]

Miyamoto A, Nakatsuka T, Ohtani M & Fukui Y 1998a Intraluteal release of progesterone and prostaglandins during PGF2{alpha}-induced luteolysis in ewes: local effects of tumor necrosis factor-{alpha}. Journal of Reproduction and Development 44 385–391.[CrossRef]

Miyamoto A, Takemoto K, Acosta T, Ohtani M, Yamada J & Fukui Y 1998b Comparative activities of growth hormone and luteinizing hormone in the direct stimulation of local release of progesterone from microdialyzed ovine corpora lutea in vivo. Journal of Reproduction and Development 44 273–280.[CrossRef]

Miyamoto A, Oki N & Ohtani M 2001 Effect of endothelin-1 on functional luteolysis in the cow. Biology of Reproduction 64 Supplement 1 197.[Abstract/Free Full Text]

Miyamoto Y, Sakumoto R, Sakabe Y, Miyake M, Okano A & Okuda K 2002 Tumour necrosis factor-{alpha} receptors are present in the corpus luteum throughout the oestrous cycle and during the early gestation period in pigs. Reproduction Domestic Animals 37 105–110.[CrossRef]

Murdoch WJ, Steadman LE & Belden EL 1988 Immunoregulation of luteolysis. Medical Hypotheses 27 197–199.[CrossRef][ISI][Medline]

Niswender GD, Reimers TJ, Diekman MA & Nett TM 1976 Blood flow: a mediator of ovarian function. Biology of Reproduction 14 64–81.[Abstract]

Ohtani M, Kobayashi S, Miyamoto A, Hayashi K & Fukui Y 1998 Real-time relationships between intraluteal and plasma concentrations of endothelin, oxytocin, and progesterone during prostaglandin F2{alpha}-induced luteolysis in the cow. Biology of Reproduction 58 103–108.[Abstract/Free Full Text]

Ohtani M, Kobayashi S & Miyamoto A 1999 Direct effect of PGF2{alpha}, TPA and ionophore A23187 [GenBank] on progesterone release from the microdialyzed corpus luteum in cows. Journal of Reproduction and Development 45 233–238.[CrossRef]

Okuda K, Sakumoto R, Uenoyama Y, Berisha B, Miyamoto A & Schams D 1999 Tumor necrosis factor {alpha} receptors in microvascular endothelial cells from bovine corpus luteum. Biology of Reproduction 61 1017–1022.[Abstract/Free Full Text]

Penny LA, Armstrong D, Bramley TA, Webb R, Collins RA & Watson ED 1999 Immune cells and cytokine production in the bovine corpus luteum throughout the oestrous cycle and after induced luteolysis. Journal of Reproduction and Fertility 115 87–96.[Abstract]

Petroff MG, Petroff BK & Pate JL 1999 Expression of cytokine messenger ribonucleic acids in the bovine corpus luteum. Endocrinology 140 1018–1021.[Abstract/Free Full Text]

Ruetten H & Thiemermann C 1997 Endothelin-1 stimulates the biosynthesis of tumour necrosis factor in macrophages: ET-receptors, signal transduction and inhibition by dexamethasone. Journal of Physiological Pharmacology 48 675–688.

Sakumoto R, Berisha B, Kawate N, Schams D & Okuda K 2000 Tumor necrosis factor-{alpha} and its receptor in bovine corpus luteum throughout the estrous cycle. Biology of Reproduction 62 192–199.[Abstract/Free Full Text]

Schams D, Berisha B, Neuvians T, Amselgruber W & Kraetzl WD 2003 Real-time changes of the local vasoactive peptide systems (angiotensin, endothelin) in the bovine corpus luteum after induced luteal regression. Molecular Reproduction and Development 65 57–66.[CrossRef][ISI][Medline]

Shaw DW & Britt JH 1995 Concentrations of tumor necrosis factor {alpha} and progesterone within the bovine corpus luteum sampled by continuous-flow microdialysis during luteolysis in vivo. Biology of Reproduction 53 847–854.[Abstract]

Wathes DC & Denning-Kendall PA 1992 Control of synthesis and secretion of ovarian oxytocin in ruminants. Journal of Reproduction and Fertility (Supplement) 45 39–52.

Webb R, Woad KJ & Armstrong DG 2002 Corpus luteum (CL) function: local control mechanisms. Domestic Animal Endocrinology 23 277–285.[CrossRef][ISI][Medline]

Wegner JA, Martinez-Zaguilan R, Gillies RJ & Hoyer PB 1991 Prostaglandin F2{alpha}-induced calcium transient in ovine large luteal cells. II Modulation of the transient and resting cytosolic free calcium alters progesterone secretion. Endocrinology 128 929–936.[Abstract]

Wiltbank MC, Guthrie PB, Mattson MP, Kater SB & Niswender GD 1989 Hormonal regulation of free intracellular calcium concentrations in small and large ovine luteal cells. Biology of Reproduction 41 771–778.[Abstract]

Wiltbank MC, Diskin MG, Flores JA & Niswender GD 1990 Regulation of the corpus luteum by protein kinase C. II Inhibition of lipoprotein-stimulated steroidogenesis by prostaglandin F2{alpha}. Biology of Reproduction 42 239–245.[Abstract]

Wuttke W, Pitzel L, Knoke I, Theiling K & Jarry H 1997 Immune-endocrine interactions affecting luteal function in pigs. Journal of Reproduction and Fertility 52 (Suppl) 19–29.

Wuttke W, Spiess S, Knoke I, Pitzel L, Leonhardt S & Jarry H 1998 Synergistic effects of prostaglandin F2{alpha} and tumor necrosis factor to induce luteolysis in the pig. Biology of Reproduction 58 1310–1315.[Abstract/Free Full Text]


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