Corpus Luteum In Cattle Research Articles

The Role of Peroxisome Proliferator-Activated Receptors in PGF2α-Induced Luteolysis in the Bovine Corpus Luteum.

Simple SummaryThe corpus luteum (CL) is responsible for progesterone (P4) secretion. In the absence of pregnancy, luteolysis occurs, which leads to a reduction in P4 production, followed by the structural regression of the CL. In cows, prostaglandin F2α (PGF2α) is the main luteolytic factor. It is also an endogenous ligand for peroxisome proliferator-activated receptors (PPARs), which are important factors regulating mammalian reproductive function. However, the mechanisms of action of PPAR isoforms, i.e., PPARα, PPARδ and PPARγ, in the luteolytic pathways in cattle are still not fully understood. The aim of this in vitro study was to determine the expression of PPAR isoforms in the bovine CL throughout the estrous cycle, and their involvement in PGF2α-induced processes related to luteolysis. The obtained results indicate that the expression of PPARs changes in the bovine CL throughout the estrous cycle; moreover, PGF2α affects its expression. This study provides evidence that PPARγ, among all examined PPAR isoforms, could be involved in the regulation of PGF2α-induced luteolysis in cattle, and PPARs may affect CL regression at multiple sites. These results help to widen the knowledge of the mechanisms of luteal regression in the bovine CL.The participation of peroxisome proliferator-activated receptors (PPARs) in ovarian function in cattle is still not fully understood. The aim of this in vitro study was to determine: (i) the immunolocalization, mRNA expression and tissue concentration of PPARα, PPARδ and PPARγ in the bovine corpus luteum (CL) (n = 40) throughout the estrous cycle, and (ii) the involvement of PPAR in PGF2α-induced processes related to luteolysis. CL (n = 9) explants were cultured in the presence of PPAR antagonists (10−5 M) in combination with or without PGF2α receptor antagonist (10−5 M) and PGF2α (10−6 M). The mRNA and protein expression of PPARs was evaluated through qPCR, IHC, and ELISA, respectively. The results showed that PPAR mRNA and protein expression differed according to the luteal stages. PGF2α upregulated PPARδ and PPARγ mRNA expression in the bovine CL in vitro, whereas PPARγ increased the inhibitory effect of PGF2α by decreasing progesterone secretion and the mRNA expression of hydroxy-delta-5-steroid dehydrogenase, 3 β- and steroid delta-isomerase 1 (HSD3B1) in the CL explants; mRNA transcription of tumor necrosis factor α (TNFα) and inducible nitric oxide synthase (iNOS) was increased. The obtained results indicate that the mRNA and protein expression of PPARs changes in the bovine CL throughout the estrous cycle and under the influence of PGF2α. We suggest that isoform γ, among all examined PPARs, could be a factor involved in the regulation of PGF2α-induced processes related to luteolysis in the bovine CL. Further studies are needed to understand the role of PPAR in luteal regression in the CL of cattle.

Contribution of the immune system to follicle differentiation, ovulation and early corpus luteum formation.

Much of what we know about the involvement of the immune system in periovulatory follicle differentiation, ovulation and subsequent formation of the corpus luteum in cattle is drawn from the findings of studies in several mammalian livestock species. By integrating published histological data from cattle, sheep and pigs and referring back to the more comprehensive knowledge bank that exists for mouse and humans we can sketch out the key cells of the immune system and the cytokines and growth factors that they produce that are involved in follicle differentiation and luteinization, ovulation and early follicle development. These contributions are reviewed and the key findings, discussed.

Do differences in the endometrial transcriptome between uterine horns ipsilateral and contralateral to the corpus luteum influence conceptus growth to day 14 in cattle?

Embryo transfer to the uterine horn contralateral to the ovary containing the corpus luteum (CL) negatively impacts pregnancy establishment in cattle. Our aim was to compare the transcriptome and ability of the ipsilateral and contralateral uterine horns to support preimplantation conceptus survival and growth to day 14. In experiment 1, endometrial samples from both horns were collected from synchronized heifers slaughtered on day 5, 7, 13, or 16 post-estrus (n=5 per time) and subjected to RNA sequencing. In experiment 2, 10 day 7 in vitro produced blastocysts were transferred into the uterine horn ipsilateral (n=9) or contralateral to the CL (n=8) or into both horns (i.e., bilateral, n=9) of synchronized recipient heifers. Reproductive tracts were recovered at slaughter on day 14, and the number and dimensions of recovered conceptuses were recorded for each horn. A total of 217, 54, 14, and 18 differentially expressed genes (>2-fold change, FDR P<0.05) were detected between ipsilateral and contralateral horns on days 5, 7, 13, and 16, respectively, with signaling pathways regulating pluripotency of stem cells, ErbB signaling pathway, and mTOR signaling pathway amongst the top canonical pathways. Site of embryo transfer did not affect recovery rate (48.0%, 168/350) or length of conceptuses (mean±SE 2.85±0.27 mm). Although differences in gene expression exist between the endometrium of uterine horns ipsilateral and contralateral to the CL in cattle, they do not impact conceptus survival or length between day 7 and 14.

133 COMPARISON OF ENDOMETRIAL TRANSCRIPTOME CHANGES BETWEEN IPSI- AND CONTRALATERAL HORNS DURING DIESTROUS AND ITS RELATIONSHIP WITH THE ABILITY TO SUPPORT CONCEPTUS ELONGATION IN CATTLE

Data from embryo transfer studies indicate that transfer to the uterine horn contralateral to the ovary containing the corpus luteum (CL) reduces pregnancy rate. The aim of this study was to compare the ipsi- v. contralateral horns in terms of (1) endometrial gene expression and (2) the ability to support conceptus growth to Day 14. In Experiment 1, endometrial samples from the ipsi- and contralateral horns were collected from synchronized nonpregnant heifers slaughtered on Day 5, 7, 13 or 16 post-oestrus (n = 5 per time point) and snap frozen for subsequent analysis. In Experiment 2, fresh Day 7 in vitro-produced blastocysts were transferred to the uterine horn(s) ipsilateral (Group I, n = 9 recipients) or contralateral to the CL (Group C, n = 8) or to both horns (i.e. bilateral, Group B, n = 9) of synchronized recipients (10 embryos transferred per horn per recipient). All recipients were slaughtered on Day 14. Reproductive tracts were recovered and each horn was flushed independently. The number and dimensions (length and width) of recovered conceptuses were recorded independently for each horn. Embryo data were analysed using PROC MIXED in SAS (SAS Institute Inc., Cary, NC, USA). The model had treatment as a fixed effect and heifer within treatment as a random effect. Experiment 1, RNA-Seq analysis of endometrium, detected 217, 54, 14, and 18 differentially expressed genes (&gt;2-fold change, false discovery rate P &lt; 0.05) between ipsi- and contralateral horns on Day 5, 7, 13, and 16 of the oestrous cycle, respectively. In Experiment 2, no differences were observed in the recovery rate of conceptuses on Day 14 from the 3 groups [53.3% (48/90), 48.7% (39/80), and 45% (81/180)], for the I, C, and B transfer groups, respectively. Conceptus length from the ipsi- (2.24 ± 0.35 mm) and contralateral (2.91 ± 0.38 mm) horns was not different (P &gt; 0.05). Thus, significant differences in endometrial gene expression exist between the uterine horns ipsi- and contralateral to the CL in cattle, but those differences do not affect conceptus elongation on Day 14. Research was supported by Science Foundation Ireland (13/IA/1983) and the Irish Department of Agriculture, Food and The Marine (13S528).

Expression of the Mir-183 cluster in the follicles and corpus luteum of cattle

In this study, the expression profiles of the miR-183 cluster were explored in bovine dominant and subordinate follicles and different luteal phases. This miRNA cluster showed higher expression levels in dominant bovine follicles than in subordinates. The expression levels of miR- 182 and miR-96 were reduced from the early phase to the regressed phase, whereas miR-183 showed lower expression levels in the mid and late CL phases than in the early and regressed CL phases.

The Accuracy of Transrectal Ultrasonography Diagnostic for Examination of Ovarian Structures in Cattle

The study aimed to determine the accuracy of ovarian follicle and corpus luteum measurements in vivo using ultrasonography diagnostic and in vitro after slicing of the ovaries. A total of 15 cows were examined using transrectal ultrasonography just before being slaughtered. Ultrasonographic examination was performed using 7.5 MHz transrectal tranducer on ovaries, measured dominant follicle and corpus luteum diameters. Follicleappeared echogenic, black with round shape. Corpus luteum, appeared as low echogenicity structure and grayish in color.Ovaries were taken out soon after animals slaughtered, soaked in 10% buffered formalin for 12 hours,then cut in thin slices and measured follicle and corpus luteum diameters. Results of follicle and corpus luteum diameter measurements in vitro and in vivo compared using two sample or paired t-test, correlation and regression analyses. The two methods of follicle and corpus luteum measurements had linear orrelation (follicle measurement, R2 = 0.933 , r = 0.966, n = 15, P &lt; 0.05) (corpus luteum, R2 = 0.912, r = 0.955, n = 15, P &lt; 0.05).Measurements of dominant follicle diameter and corpus luteum size manifested the real size of the two ovarian structures. Transrectal ultrasonography diagnostic on ovaries proved to be having high accuracy and reliability for identification, measurement of ovarian structure diameters, follicle and corpus luteum in cattle.

Cattle and sheep use different promoters to direct the expression of the aromatase cytochrome P450 encoding gene, Cyp19, during pregnancy

Gestagens and oestrogens are important regulators of pregnancy and parturition. The aim of the present study was the comparative quantification of steroidogenic transcripts in placenta and corpus luteum of cattle and sheep during pregnancy and post partum. Cyp19 transcript variants, derived from different promoters, as well as transcripts of Hsd3b, Cyp11A1, and Cyp17, encoding the steroidogenic enzymes P450 arom, 3β-HSD, P450 SCC, and P450 C17, respectively, were quantified by newly developed real-time PCR assays. All steroidogenic transcripts were detected in ovine and bovine corpus luteum and placenta during pregnancy, however at a very different concentration. In both species Cyp11A1 and especially Hsd3b transcripts predominated in corpus luteum, outnumbering transcripts of Cyp17 and Cyp19 by more than two and three orders of magnitude, respectively. Cyp19 transcript were found at high concentration in the placenta and at a very low concentration in corpus luteum. Cyp17 transcripts had a relatively low concentration in both, placenta and corpus luteum, however showed a peak of expression in the ovine and bovine term placenta. Tissue- and species-specific Cyp19 transcripts derived from different promoters were detected. In order to map all promoters, the bovine Cyp19 locus was reconstructed by in silico analysis. In the placenta, transcripts were primarily derived from the proximal promoter P 1.5 in sheep, but from the distally located P 1.1 in cattle. Corpora lutea of both species predominantly expressed P 1.1 derived transcripts. Contrary to the bovine, the sheep corpus luteum also showed considerable P 1.5 derived expression. This demonstrates that cattle and sheep use different promoters to direct Cyp19 expression during pregnancy.

Identification and physical mapping of genes expressed in the corpus luteum in cattle1

A representational difference analysis was performed to identify genes expressed in the corpus luteum of cattle. The corpus luteum is an ovarian structure that is essential for the establishment and maintenance of pregnancy. Knowledge of gene expression and function of corpus luteum will be important to improve fertility in humans and domestic animals. Housekeeping genes were removed from the corpus luteum representation (tester) using skeletal muscle as the subtracting agent (driver). A total of 80 clones of the final subtraction product were analysed by sequencing and 11 new bovine gene sequences were identified (pBTCL1-11). The sequences were mapped to segments of 10 different chromosomes using a somatic cell hybrid panel and a radiation hybrid panel. With one exception the locations are in agreement with published comparative maps of cattle and man. Expression in corpus luteum was verified by RT-PCR for all the 11 clones.

Morphological and functional characteristics of the dominant follicle and corpus luteum in cattle and their influence on ovarian function

Predicting the functional activity of a dominant follicle (DF) and corpus luteum (CL) might be important before starting a superovulation regime or a synchronization program. The DF and CL were characterized morphologically by using ultrasonography and were characterized functionally by estimating the estradiol-17β/progesterone (E2/P4) ratio. Their influence on ovarian function was estimated through their ability to ovulate at different stages of development in response to PGF2α-application. A total of 47 Holstein Friesian (35 cows and 12 heifers) were used in two experiments. In Experiment 1, 25 animals were examined by daily transrectal palpation and ultrasonography to follow the morphological development of the DF. The status of the DF was categorized into 3 groups (A1, B1, C1). The A1 group (n=7) contained animals with DF in the growing phase or in early static growth phase for less than 3 days. Group B1 (n=13) included animals with DF in static growth phase for 3 to 4 days, while Group C1 (n=5) comprised animals with DF keeping a plateau for more than 4 days or animals with DF in the regression phase. The DF were aspirated transvaginally and the follicular fluid (FF) was analyzed for E2 and P4. In Experiment 2, 22 animals were included. As in Experiment 1, the animals were classified into three groups (A2, n=10; B2, n=5; C2, n=7). They were treated by a single dose of PGF2α (25 mg, im) between Days 8 and 12 of the cycle. Results showed that luteolyses occurred in all animals. The DF, which were in growing or in early static growth phase < 3 days were always E2-dominant (E2 > P4) and ovulated after PGF2α-application in 68 of cases and persisted in 2 (Group A2). The DF persisting > 4 days or that had been in regression were always P4-dominant. This type of DF regressed after PGF2α-application (Group C2). The DF in early static growth phase for 3 to 4 days in 513 cases were E2-dominant and in 813 cases were P4-dominant. This type of DF ovulated in 35 cases and regressed in 25 cases after PGF2α-application (Group B2). These results suggest that the DF is morphologically and functionally defined as long as the DF is in the growing or early static growth phase (A1, A2) for at least 2 days or if the DF is in regression (C1, C2). However, when the DF is in the static growth phase for 3 or 4 days (B1, B2), their morphological and functional characteristics are different. The CL controls ovulation in the A and C groups and plays an abettor's roll in the B-group.

Bovine interferon-tau stimulates the Janus kinase-signal transducer and activator of transcription pathway in bovine endometrial epithelial cells.

Trophoblastic bovine interferon-tau (bIFN-tau) suppresses luteolytic pulses of endometrial prostaglandin F(2alpha) (PGF(2alpha)) at the time of maternal recognition of pregnancy. This results in maintenance of the corpus luteum in cattle. The hypothesis that effects of bIFN-tau in the endometrium were through activation of the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway of signal transduction was tested. Whole cell, cytosolic, and nuclear extracts from bovine endometrial cells treated with bIFN-tau were analyzed by immunoprecipitation, immunoblotting, and electrophoretic mobility shift assays in a series of dose- and time-dependency experiments. Bovine IFN-tau stimulated tyrosine phosphorylation, homo- and heterodimer formation, nuclear translocation, and DNA binding of STAT proteins 1, 2, and 3. Moreover, bIFN-tau induced synthesis of interferon-regulatory factor. In conclusion, bIFN-tau stimulates the JAK-STAT pathway in the bovine endometrium. It is proposed that activation of the JAK-STAT pathway is involved in regulating the antiluteolytic effects of bIFN-tau.

Corpus luteum development and function in cattle with episodic release of luteinizing hormone pulses inhibited in the follicular and early luteal phases of the estrous cycle.

The influence of episodic LH pulses before and subsequent to ovulation on size and function of the corpus luteum (CL) in cattle was examined. Treatments were 1) control; 2) LHRH antagonist starting 2 days before the preovulatory LH surge (Antagonist [Ant] -2); 3) LHRH antagonist at initiation of the preovulatory LH surge (Ant 0); and 4) LHRH antagonist starting 2 days after the preovulatory LH surge (Ant 2). Treatments with an LHRH antagonist were continued until 7 days after the preovulatory surge. Diameter of the CL and concentrations of progesterone were monitored during the luteal phase that ensued after treatment. Maximum average diameters of CL were 9.5, 17.5, 21.6, and 28.8 mm for females from the Ant -2, Ant 0, Ant 2, and control groups, respectively (P < 0. 01). Compared with those in control animals, concentrations of progesterone in plasma were less (P < 0.01) in animals in which release of LH pulses was inhibited by treatment with antagonist. Arbitrary units under the curve for concentrations of progesterone during the luteal phase of the estrous cycle for Ant -2, Ant 0, Ant 2, and control groups were 19.6, 41.6, 43.6, and 142.2, respectively. There was no difference in circulating concentrations of progesterone (P > 0.1) among antagonist-treated groups. In conclusion, episodic release of LH pulses before, during, and after the time of the preovulatory surge of LH may stimulate development and function of the CL in cattle.

Effect of long-term infusion with recombinant growth hormone-releasing factor and recombinant bovine somatotropin on development and function of dominant follicles and corpora lutea in Holstein cows.

The objective of this study was to evaluate the effects of recombinant bovine growth hormone-releasing factor (rGRF) or recombinant bovine somatotropin (rbST) on growth and function of the first-wave dominant follicle and corpus luteum. Primiparous Holstein cows (117 d postpartum) were infused with 12 mg/d of rGRF (n = 10) or 29 mg/d of rbST (n = 10) for 63 d, and non-infused cows (n = 10) were controls. At slaughter on d 5 of an estrous cycle, blood and ovaries were collected and data from cows with a corpus luteum were analyzed (control, n = 8; rGRF, n = 5; rbST, n = 6). Treatment with rGRF or rbST increased somatotropin (ST) and IGF-I in serum similarly compared with controls. In contrast, rbST-treated cows had higher concentrations of ST in follicular fluid (FF) compared with rGRF-treated and control cows. In addition, rbST, but not rGRF, increased the number and decreased the size of estrogen-active follicles (EA; estradiol > progesterone concentrations in FF), increased the abundance of IGF binding proteins-2, -3, and -4 in FF from EA follicles, and increased the number but decreased the size of corpora lutea and decreased concentration of progesterone in serum compared with controls. Based on these results, we concluded that long-term infusion of rbST alters growth and function of the first-wave dominant follicle and the corpus luteum in cattle.

Transvaginal, ultrasound-guided biopsy of the corpus luteum in cattle

An ultrasound-guided transvaginal technique for corpus luteum biopsy was developed and tested in cattle. The biopsy needle set consisted of an inner needle (o.d. 1 mm) with a 20-mm long specimen notch, an outer cannula (o.d. 1.2 mm) with a cutting edge, and an automated spring-loaded handle with trigger. The biopsy needle set was inserted into the channel guide of the handle of a convex-array transvaginal ultrasound probe. The transducer was positioned in the vaginal fornix, and the ovary was manipulated transrectally against the vaginal wall and transducer face. During monitoring on the ultrasound screen, the inner needle was pushed through the vaginal wall into the corpus luteum, and the cutting cannula was fired, cutting and trapping luteal tissue in the specimen notch. Three luteal biopsies at each of Hours 0 and 4 were taken 10 d after ovulation in heifers; 6 other heifers served as controls. A biopsy core was obtained in 36 of 39 attempts (92%). The tissue specimens seemed normal based on gross evaluation. The effect of biopsy on luteal function was assessed by daily ultrasound monitoring of luteal area, by assay of progesterone concentrations in blood samples obtained daily, and by the length of the interval from biopsy to ovulation. No significant differences were found for postbiopsy function for any of the 3 end points. The results indicated repeated transvaginal, ultrasound-guided biopsy of the corpus luteum in cattle is a practical procedure and may be useful for experimental and diagnostic purposes,

Immunohistochemical distribution of follistatin in dominant and subordinate follicles and the corpus luteum of cattle.

The study was done to quantitatively characterize the distribution of follistatin in ovarian follicles and corpora lutea at specific stages of development. Transrectal ultrasonography was used to monitor the growth of individually identified follicles from 2 days before ovulation until the day of ovariectomy on Day 3 of wave 1 (n = 8), Day 6 of wave 1 (n = 6), Day 1 of wave 2 (n = 6), or after onset of proestrus, at least 17 days postovulation (n = 7). Days of ovariectomy represent the growing, early-static, late-static, and regressing phases of the dominant follicle of wave 1, respectively. Subordinate (n = 24), preselection (n = 15), and preovulatory (n = 6) follicles and corpora lutea (n = 31) were also analyzed. Follistatin was localized using immunohistochemical labeling of paraffin sections, and relative amounts were quantitated using densitometric analysis. Follistatin was distributed in the perinuclear cytoplasm of granulosa and luteal cells but not in theca cells. Dominant follicles contained more (p < 0.05) follistatin than corresponding subordinate follicles. The amount of follistatin was maximal during the mid-growing phase of the dominant follicle and decreased thereafter (p < 0.05). Among the corpora lutea, the maximal amount was detected at mid-diestrus (Day 11 postovulation). Less than half of luteal cells displayed the stain for follistatin; positively stained luteal cells were located in close proximity to blood capillaries. Follistatin was not detectable in the corpus luteum during metestrus (Day 3 postovulation) or proestrus (Day >/= 17 postovulation). In summary, the degree of immunohistochemical expression of follistatin was phase specific for both follicles and corpora lutea. The most intense staining in follicles was associated with the period of functional dominance and in corpora lutea was seen during the phase of maximal development. Significant phase-related differences in follistatin expression provide rationale for the hypothesis that follistatin is involved in the final stages of follicle and luteal gland development in cattle.

Unilateral relationship of embryos and the corpus luteum in cattle.

The unilateral relationship between embryo and corpus luteum was studied in two experiments in Holstein heifers. Two fertilized ova were inserted into one uterine horn of recipient heifers on Day 6 after estrus. Recipients were given progesterone from Day 13 through Day 23 and were necropsied on Day 24 (18 days after ova transfer). Corpora lutea were weighed and classified as maintained or regressed. In Exp. 1, the uterine horns were separated and the fertilized ova were inserted ipsilateral (group 1, 5 heifers) or contralateral (group 2, 5 heifers) to the CL. Nine of 10 heifers were pregnant at necropsy. Mean CL weight for group 1 (3725 mg) was greater (P<0.01) than that for group 2 (1300 mg). Results indicated that in heifers with a surgically isolated uterine horn a unilateral utero-ovarian mechanism was involved in maintenance of the CL during early pregnancy. Experiment 2 was designed to determine if embryos inserted into the uterine horn contralateral to the CL in uterine-intact heifers would maintain the CL and, if so, whether the luteotropic message passed from gravid to nongravid side through the uterine body and horns or through the intercornual area. Recipient heifers were randomized into 4 groups (n=4): 1) uterine-intact controls; 2) ipsilateral horn transected, intercornual ligament intact; 3) uterine horns intact, intercornual ligament severed; 4) ipsilateral horn and intercornual ligament severed. Mean CL weight was smaller (P<0.05) in group 1 (2900 mg), group 2 (1405 mg) and group 4 (578 mg) than in group 3 (4848 mg). There was a significant difference between groups 1 and 3 although the CL in all heifers in both groups were classified as maintained. Mean CL weight was not significantly different between groups 2 and 4. Results indicated that embryos transferred to the contralateral horn were capable of maintaining the CL if the uterine horns were not separated. The passage of the luteotropic message from gravid to nongravid side was exerted through the uterine body and horns rather than through the intercornual area and is most likely attributable to the growth of embryonic membranes into the nongravid horn.

Glycosidases in the corpus luteum.

SUMMARY 1. The corpus luteum in cattle, pigs and women is rich in α-l-fucosidase, β-d-fucosidase, β-d-galactosidase, α-d-mannosidase, β-d-glucuronidase and β-N-acetyl-d-glucosaminidase activity. In comparison, the activity of these enzymes in the follicular fluid is usually negligible. The remainder of the ovarian tissue approximates to the follicles in activity. 2. The activities found in the corpora lutea of pregnant and non-pregnant animals were the same.