Bovine Corpus Luteum Research Articles

Expression and Localization of Gap Junctional Connexins 26 and 43 in Bovine Periovulatory Follicles and in Corpus Luteum During Different Functional Stages of Oestrous Cycle and Pregnancy

The aim of this study was to characterize the regulation of connexins (Cx26 and Cx43) in the bovine ovary (experiment 1-3). Experiment 1: ovaries containing preovulatory follicles or corpora lutea (CL) were collected at 0, 4, 10, 20, 25 (follicles) and 60 h (CL) relative to injection of GnRH. Experiment 2: CL were assigned to the following stages: days 1-2, 3-4, 5-7, 8-12, 13-16, >18 (after regression) of oestrous cycle and of early and late pregnancy (<4 and >4 months). Experiment 3: induced luteolysis, cows on days 8-12 were injected with PGF2alpha analogue (Cloprostenol), and CL were collected by transvaginal ovariectomy before and 0.5, 2, 4, 12, 24, 48 and 64 h after PGF2alpha injection. Real-time RT-PCR was applied to investigate mRNA expression and immunofluorescence was utilized for protein localization. Cx26 mRNA increased rapidly 4 h after GnRH injection (during LH surge) and decreased afterwards during the whole experimental period. Cx43 mRNA expression decreased continuously after GnRH application. Cx26 mRNA in CL increased significantly in the second part of oestrous cycle and after regression. In contrast, the highest mRNA expression for Cx43 in CL was detected during the early luteal phase. After induced luteolysis the mRNA expression of Cx26 increased significantly at 24 h. As shown by immunofluorescence, Cx26 was predominantly localized in the connective tissue and blood vessels of bovine CL, whereas Cx43 was present in the luteal cells and blood vessels. This resulted in a strong increase of Cx26 expression during the late luteal phase and after luteal regression. Subsequently, Cx43 expression was distinctly decreased after luteal regression. These data suggest that Cx26 and Cx43 are involved in the local cellular mechanisms participating in tissue remodelling during the critical time around periovulation as well as during CL formation (angiogenesis), function and regression in the bovine ovary.

Differential Gene Expression in the Bovine Corpus Luteum During Transition from Early Phase to Midphase and Its Potential Role in Acquisition of Luteolytic Sensitivity to Prostaglandin F2 Alpha1

Prostaglandin F2 alpha (PGF(2alpha)) brings about regression of the bovine corpus luteum (CL). This luteolytic property of PGF(2alpha) is used in beef and dairy cattle to synchronize estrus. A limitation of this protocol is insensitivity of the early CL to luteolytic actions of PGF(2alpha). The mechanisms underlying this differential luteal sensitivity are poorly understood. The developing CL has a maximum number of PGF(2alpha) receptors; therefore, differences in signaling events may be responsible for luteal insensitivity. Hence, differential gene expression at two developmental stages of CL, Day 4 (D-4) and D-10 after estrus, might account for differences in signal transduction pathways associated with luteal sensitivity. This possibility was examined in these studies. Microarray analysis (n = 3 cows per stage) identified 167 genes that were differentially expressed (P < 0.05). These were categorized into genes involved in protein biosynthesis and modification (18.5%), transcription regulation and DNA biosynthesis (18.5%), miscellaneous (17.0%), cell signaling (12.0%), steroidogenesis and metabolism (10.2%), extracellular matrix and cytoskeletal proteins (9.5%), unknown functions (6.0%), protein degradation (5.3%), and antioxidant property (3.0%). Real-time PCR confirmed the differential expression of nine selected genes, including tyrosine 3-monooxygenase/tryptophan 5-monooxygense activation protein zeta polypeptide (YWHAZ) and regulator of G protein signaling 2 24-kDa (RGS2), observed in microarray. Furthermore, the in vivo effect of exogenous PGF(2alpha) (n = 3 cows per stage) on selected genes that were found to be differentially expressed during this developmental transition was examined. PGF(2alpha) increased the expression of a guanine nucleotide-binding protein (G protein) beta polypeptide 1 (GNB1) in D-4 CL and calcium/calmodulin-dependent kinase kinase 2 beta (CAMKK2) in D-10 CL. Therefore, GNB1, CAMKK2, YWHAZ, and RGS2 are candidate genes that may have a significant role in acquisition of luteal sensitivity to PGF(2alpha). Additional evidence supporting the significance of the microarray data was obtained from the observation that the amount of CAMKK2 paralleled the differential mRNA expression observed for this gene when examined by microarray analysis and by real-time RT-PCR. Furthermore, the two types of luteal steroidogenic cells known to be targets for PGF(2alpha) actions were demonstrated to be a cellular source for CAMKK2.

Survival Role of Locally Produced Acetylcholine in the Bovine Corpus Luteum1

The present study was conducted to explore the source of acetylcholine (ACH) in the corpus luteum (CL) and to test our hypothesis of an antiapoptotic role of ACH in the bovine CL and, further, to investigate whether nerve growth factor (NGF), insulin-like growth factor 1 (IGF1), and transforming growth factor beta1 (TGFB1) influence the expression of choline acetyltransferase (CHAT), the biosynthetic enzyme of ACH, in cultured bovine luteal cells. Protein expression and immunolocalization of CHAT were carried out at different stages throughout the luteal phase and in cultured luteal and endothelial cells. ACH was measured in luteal tissue at the different luteal stages and in luteal cells cultured for 8 and 24 h. Cell viability and TUNEL assays were performed on cultured midluteal cells treated with or without tumor necrosis factor alpha (TNF)/interferon gamma (IFNG) in the presence of ACH and its muscarinic (atropine) and nicotinic (mecamylamine) receptor antagonists. The CL was devoid of cholinergic nerve fibers. CHAT immunostaining was evident in luteal, endothelial, and stromal cells in luteal tissue sections and in cultured luteal and endothelial cells. CHAT protein was expressed throughout the cycle without any significant changes. ACH concentration in luteal tissue was not changed during the luteal stages but increased over time and with increased cell numbers in luteal cell cultures. ACH increased cell viability and prevented cell death induced by TNF/IFNG. Atropine significantly attenuated ACH action, whereas mecamylamine had no effect. TNF/IFNG treatment downregulated CHAT expression, whereas NGF, IGF1, and TGFB1 upregulated CHAT expression, in cultured luteal cells. The overall findings strongly suggest a nonneural source and antiapoptotic role of ACH in the bovine CL. Locally produced ACH appears to be regulated by NGF, IGF1, and TGFB1.

KIT variants in bovine ovarian cells and corpus luteum

We report the presence of KIT variants in granulosa and thecal cells of the follicle and endothelial and steroidogenic cells of the corpus luteum. Transcripts of both full-length splice variants, KIT and KITA, were ubiquitously detected in all cell types, in contrast to transcripts for truncated KIT. RT-PCR with exon-intron-specific primers suggested that KIT transcripts retained intron sequences. We used domain-specific KIT antibodies to identify truncated KIT proteins in cell conditioned media and lysates. These proteins represented soluble KIT and a so far disregarded intracellular KIT fragment, and were ubiquitously present. In contrast, glycosylated variants of full-length KIT were predominantly detected in thecal and endothelial cells. All KIT variants were encountered again in COS-7 cells transfected with a vector containing KITA. Phorbol 12-myristate-13-acetate treatment induced levels of truncated KITs, and this effect was repressed by the metalloproteinase inhibitor TAPI-1. Our findings show that ectodomain cleavage of full-length KIT generates an intracellular KIT. Our experiments suggest that replenishing full-length KIT differs among various ovarian cell types.

157. EXPRESSION PATTERNS OF EXTRACELLULAR MATRIX IN MICE OVARIES

Extracellular matrix (ECM) has been shown to have distinct expression patterns during bovine follicle and corpus luteum (CL) formation and regression. To date little is known about ECM patterns during follicle and CL formation in the mouse ovary. Twenty nine day old mice were treated with PMSG on experimental day 0 and 1 to induce follicle development, and subsequently with hCG on day 2 to induce ovulation. Ovaries were collected for immunohistochemistry on days 0, 2 and 5 (n = 10 per group). Another group was similarly treated but was additionally mated, and ovaries examined in the pregnant mice on experimental day 11 (n = 7). The follicular basal lamina (BM) of all developmental stages contained collagen IV α1 and α2, laminin α1, β1 and γ1 chains, nidogen 1 and 2, and perlecan. Collagen XVIII was only found in BMs of primordial, primary and some preantral follicles, whereas laminin α2 was only present in some preantral and antral follicles. BMs of atretic follicles showed similar composition to healthy follicles. A specialized matrix of the membrana granulosa (focimatrix) was detected in both healthy and atretic follicles. The focimatrix contained collagen IV α1 and α2, laminin α1, β1 and γ1 chains, nidogen 1 and 2, perlecan and collagen XVIII, but not laminin α2. The immunostaining in CL was restricted to capillary sub-endothelial basal laminas and contained collagen IV α1 and α2, laminin α1, β1 and γ1 chains, nidogen 1 and 2, perlecan and collagen XVIII. No parenchymal matrix, as observed in cow and human, was detected. In addition, laminin α4 and α5 were not immunolocalised to any structure in the mouse ovary. The composition of the BM of follicles in the mouse ovary is similar to cow and rat, but the appearance and composition of the focimatrix differs from bovine ovaries.

Changes in the Vasculature of Bovine Corpus Luteum During the Estrous Cycle and Prostaglandin F2&amp;alpha;-induced Luteolysis

To investigate the possible role of the vasculature in the local regulation of corpus luteum (CL) function, we determined the densities of capillaries and large blood vessels in the center of the bovine CL during the estrous cycle and following prostaglandin (PG) F2alpha-induced luteolysis. The CLs at the early (Days 2-3 post-ovulation), developing (Days 5-7), mid (Days 8-12), late (Days 15-17) and regressed (Days 19-21) stages were collected. In addition, the CLs were collected by transvaginal ovariectomy from 12 cows (Day 10 after ovulation), i.e., non-treated (n=3, 0 h, control), at 0.5 (n=3), 2 (n=3) and 12 h (n=3) after injection of a luteolytic dose of PGF2alpha. Immunohistochemical staining with von Willebrand Factor (specific for endothelial cells that are found in both types of blood vessels) revealed that the density of the luteal blood vessels was significantly higher at the developing and late luteal stages (P<0.05) than at the other stages, whereas the number of larger blood vessels (those stained with alpha-smooth muscle actin) was higher at the late and regressed luteal stages (P<0.05) than at the other stages. Furthermore, both the density of blood vessels and the number of blood vessels with smooth muscle were significantly higher in the CLs obtained at 2 h and 12 h after PGF2alpha administration (P<0.05) than in those without PGF2alpha treatment. These results suggest that the number of blood vessels with smooth muscle per unit area in the regressing CL increased as a result of losing steroidogenic cells and capillaries. The overall results demonstrate that the capillaries disappeared earlier than the large blood vessels during structural luteolysis and suggest that the loss of capillaries in the CL results in a reduced supply of nutrients and oxygen to luteal cells followed by cell death.

Interrelationships among morphology, echotexture, and function of the bovine corpus luteum during the estrous cycle

It has been suggested that ultrasound image attributes are a potential indicator of physiological and functional status of the corpus luteum (CL) in several species, including cattle. The aims of this study were to evaluate CL morphological, functional and echotextural characteristics, and also to investigate the hypothesis that those attributes are correlated and change similarly throughout an estrous cycle. Ovaries of crossbred ( Bos taurus taurus × Bos taurus indicus) heifers were evaluated using ultrasonography daily throughout an interestrus interval using a B-mode, real-time ultrasound machine equipped with a 5 MHz linear-array rectal transducer, during a natural estrous cycle (Experiment 1; n = 12) or during a shortened cycle, with luteolysis induction 10 d after estrus (Experiment 2; n = 6). Blood samples were collected for assay of plasma progesterone concentrations. Corpora lutea areas were measured and daily images of each CL were videotaped and digitized for computer-assisted analysis using custom-developed software. In Experiment 1, area of luteal tissue increased until a maximum value 10 d after estrus ( P < 0.001), followed by a plateau phase, and then a decline beginning 14 d after estrus. Luteal tissue area was highly correlated to plasma progesterone concentrations ( r = 0.86; P < 0.001). When luteolysis was induced in Experiment 2, loss of CL function (decrease in plasma progesterone concentrations to metestrous values) preceded tissue regression by 48 h (24 h compared with 72 h; P < 0.001). The mean pixel value of ultrasound images did not change in Experiment 1 ( P > 0.70), but a day effect on this attribute was observed in Experiment 2 ( P = 0.052). In contrast, mean pixel value was correlated to plasma progesterone concentrations in Experiment 1 ( r = −0.63; P < 0.05), but not in Experiment 2 ( r = −0.28; P > 0.10). In regard to CL heterogeneity, defined as the standard deviation of the mean pixel value of the luteal tissue, a time effect was observed following both natural (Experiment 1; P < 0.009) and luteolysis-induced (Experiment 2; P < 0.05) estrous cycles ( P < 0.05). Moreover, this variable was correlated with plasma progesterone concentrations ( r = −0.71 and −0.58 in Experiments 1 and 2, respectively; P < 0.01), indicating that CL images were more heterogeneous during metestrus and after luteolysis (functional regression). In summary, morphological and echotextural attributes were correlated with CL function and underwent similar changes during the estrous cycle. Luteal tissue heterogeneity, assessed by ultrasonography, is considered a potential indicator of CL functional status, because it is correlated to circulating progesterone concentrations.

Progesterone receptor membrane component 1 (PGRMC1) gene expression in corpus luteum during the estrous cycle in cows

The aim of the study was to investigate progesterone receptor membrane component 1 (PGRMC1) mRNA expression in bovine corpus luteum (CL) obtained from heifers or non-pregnant cows on the following days of the estrous cycle: 1-5, 6-10, 11-16 and 17-21 (n=4/each time period). The expression of PGRMC1 mRNA, analyzed by semiquantitative RT-PCR, was the highest on days 6-10 (p<0.01) and then it declined (p<0.05). The lowest expression was found on days 1-5 (p<0.05). A significant correlation (p<0.05) was also observed between luteal progesterone (P(4)) concentration and PGRMC1 mRNA expression. These data indicate that PGRMC1 mRNA is expressed in bovine CL and this expression varies throughout the luteal phase. It is assumed that PGRMC1 may be involved in a non-genomic effect of P(4) on luteal cells.

The influence of tumor necrosis factor α (TNF) on the secretory function of bovine corpus luteum: TNF and its receptors expression during the estrous cycle

Tumor necrosis factor alpha (TNF) inversely regulates the function of bovine corpus luteum (CL). Whereas the low doses of TNF induce luteolysis, the high doses prolong CL lifespan and prevent luteolysis in vivo. We suggest that the varying effects of TNF may be caused by its action exerted on CL via multiple signaling pathways involving two distinct receptors: TNFR-I (responsible for induction of the cell death) and TNFR-II (implicated in cell proliferation). In the study, we determined CL expressions of TNF, TNFR-I and TNFR-II mRNAs during the bovine estrous cycle using semi-quantitative RT-PCR. Specific transcripts for TNF, TNFR-I and TNFR-II were found in the CL with the highest (p<0.05) expression in the regressed CL. We also examined the TNF influence on the bovine CL function in vivo. On Day 15 of the estrous cycle, cows were infused (via aorta abdominalis) with saline, TNF (1 or 10 microg) or analogue of prostaglandin (PG)F(2alpha) (aPGF(2alpha) , 500 microg; a positive control). Four hours after infusions, CLs were collected by colpotomy and luteal contents of progesterone (P(4)), stable metabolites of nitric oxide (NO; nitrite/nitrate), leukotriene (LT)C(4), luteolytic PGF(2alpha),and luteotropic PGE(2) were determined. Moreover, caspase-3 activity was measured in the CLs as an indicator of apoptosis induction. The luteal content of P(4) decreased (p<0.05) after infusion of 1 microg of TNF. TNF inversely affected PGs content in CL: the low dose increased (p<0.01) the PGF(2alpha) level and the high dose increased (p<0.05) PGE(2) level. Contents of LTC(4) and nitrite/nitrate increased (p<0.01) after the low dose of TNF. Moreover, 1 microg of TNF induced apoptosis and increased (p<0.05) caspase-3 activity in the CLs collected during the late luteal phase. In conclusion, the high expressions of TNF and TNF receptors mRNAs were observed during or just after the luteolysis. A low concentration of TNF stimulated in vivo luteolytic factors such as PGF(2alpha), LTC(4) and NO as well as induced apoptosis; whereas the high concentration of TNF stimulated a survival pathway in the bovine CL increasing luteal content of P(4) and PGE(2).

Evidence for an inhibitory role of bone morphogenetic protein(s) in the follicular–luteal transition in cattle

Bone morphogenetic proteins (BMPs) and their receptors are expressed in ovarian theca cells (TC) and granulosa cells (GC) and BMPs have been implicated in the regulation of several aspects of follicle development including thecal androgen production and granulosal oestrogen production. Their potential involvement in luteal function has received less attention. In this study, we first compared relative abundance of mRNA transcripts for BMPs, activin-betaA and BMP/activin receptors in bovine corpus luteum (CL) and follicular theca and granulosa layers before undertaking functional in vitro experiments to test the effect of selected ligands (BMP6 and activin A) on luteinizing bovine TC and GC. Relative to beta-actin transcript abundance, CL tissue contained more BMP4 and -6 mRNA than granulosa, more BMP2 mRNA than theca but much less activin-betaA mRNA than both granulosa and theca. Transcripts for all seven BMP/activin receptors were readily detected in each tissue and two transcripts (BMPRII, ActRIIA) were more abundant in CL than either theca or granulosa, consistent with tissue responsiveness. In vitro luteinization of TC and GC from antral follicles (4-6 mm) was achieved by culturing with 5% serum for 6 days. Treatment with BMP6 (0, 2, 10, and 50 ng/ml) and activin A (0, 2, 10 and 50 ng/ml) under these conditions dose-dependently suppressed forskolin-induced progesterone (P4) secretion from both cell types without affecting cell number. BMP6 reduced forskolin-stimulated upregulation of STAR mRNA and raised 'basal' CYP17A1 mRNA level in theca-lutein cells without affecting expression of CYP11A1 or hydroxy-Delta-5-steroid dehydrogenase, 3 beta- and steroid Delta-isomerase 1 (HSD3B1). In granulosa-lutein cells, STAR transcript abundance was not affected by BMP6, whereas forskolin-induced expression of CYP11A1, HSD3B1, CYP19A1 and oxytocin transcripts was reduced. In both cell types, follistatin attenuated the suppressive effect of activin A and BMP6 on forskolin-induced P4 secretion but had no effect alone. Granulosa-lutein cells secreted low levels of endogenous activin A ( approximately 1 ng/ml); BMP6 reduced this, while raising follistatin secretion thus decreasing activin A:follistatin ratio. Collectively, these findings support inhibitory roles for BMP/activin signalling in luteinization and steroidogenesis in both TC and GC.

ウシ黄体における発情周期を通じた血管分布変化および中期黄体内血管分布に及ぼす PGF2α の影響

ウシ黄体における発情周期を通じた血管分布変化および中期黄体内血管分布に及ぼす PGF2α の影響

Luteolytic Effect of Prostaglandin F2α on Bovine Corpus Luteum Depends on Cell Composition and Contact

Prostaglandin F(2 alpha) (PGF(2 alpha)) is a main luteolytic factor in vivo; however, its direct luteolytic influence on steroidogenic cells of bovine corpus luteum (CL) is controversial and not fully understood. The aim of the study was to clarify PGF(2 alpha) action on bovine CL in different in vivo and in vitro conditions and to examine whether the contact among all main types of CL cells is necessary for luteolytic PGF(2 alpha) action. In experiment 1, the bovine CL (day 15 of the oestrous cycle) was perfused using in vivo microdialysis system with dinoprost (an analogue of PGF(2 alpha)) for 0.5 h. Dinoprost caused a short-time increase in progesterone (P4), whose concentration decreased thereafter (at 6-, 10-, 12- and 24-h after treatment). In experiment 2, the direct effect of PGF(2 alpha) on P4 accumulation in CL steroidogenic cells cultured in monolayer (day 15 of the cycle) was determined. PGF(2 alpha) after 24 h of incubation increased P4 accumulation in steroidogenic CL cells. In experiment 3 steroidogenic, endothelial CL and immune cells (day 15 of the cycle) were incubated with PGF(2 alpha) in cocultures for 24 h in glass tubes and the levels of P4, stable metabolites of nitric oxide (NO) and leukotriene (LT) C(4) were determined. Although PGF(2 alpha) treatment increased P4 secretion in homogeneous steroidogenic CL cell culture, the decrease in P4 secretion in cocultures of all types of CL cells was observed. The secretion of NO and LTC(4) increased after the treatment of PGF(2 alpha) both in pure cultures of CL cells and in cocultures. The interactions between endothelial and immune cells with steroidogenic CL cells are needed for luteolytic PGF(2 alpha) action within the bovine CL. Our results indicate that the cell coculture model, including the main types of CL cells, is the most approximate to study PGF(2 alpha) role in vitro.

Regulation of Luteal Function and Corpus Luteum Regression in Cows: Hormonal Control, Immune Mechanisms and Intercellular Communication

The main function of the corpus luteum (CL) is production of progesterone (P4). Adequate luteal function to secrete P4 is crucial for determining the physiological duration of the oestrous cycle and for achieving a successful pregnancy. The bovine CL grows very fast and regresses within a few days at luteolysis. Mechanisms controlling development and secretory function of the bovine CL may involve many factors that are produced both within and outside the CL. Some of these regulators seem to be prostaglandins (PGs), oxytocin, growth and adrenergic factors. Moreover, there is evidence that P4 acts within the CL as an autocrine or paracrine regulator. Each of these factors may act on the CL independently or may modify the actions of others. Although uterine PGF(2 alpha) is known to be a principal luteolytic factor, its direct action on the CL is mediated by local factors: cytokines, endothelin-1, nitric oxide. The changes in ovarian blood flow have also been suggested to have some role in regulation of CL development, maintenance and regression.

Regulatory changes of apoptotic factors in the bovine corpus luteum after induced luteolysis

The corpus luteum (CL) offers the opportunity to study not only proliferative, but also regressive processes. During luteolysis of the CL a sudden death of luteal and endothelial cells seems to be involved (apoptosis). The aim of this study was to examen the mRNA expression of factors known to be involved in apoptotic processes: monocyte chemoattractant protein-1 (MCP-1), factors of the extrinsic and intrinsic apoptotic pathways, caspase3, -6, -7 and interferone gamma (IFNgamma). Luteolysis was induced by injection of 500 microg Cloprostenol during mid-luteal phase. The CLs were collected at 0.5, 2, 4, 12, 24, 48, and 64 hr after PGF2alpha-injection. Control CLs (Days 8-12) were collected at the slaugtherhouse. Real-time RT-PCR determined the mRNA expressions. Western blot analysis of poly(ADP-ribose) polymerase (PARP-1) and IFNgamma as well as protein measurement of tumor necrosis factor alpha (TNFalpha) by EIA were performed. The mRNA levels of MCP-1, IFNgamma and most factors of the extrinsic pathway were significantly increased between 0.5 and 2 hr. The factors of the intrinsic pathway were mostly later up-regulated at 24-48 hr after PGF2alpha. Caspase6 and 3 revealed a significant increase from 2 and 12 hr, respectively, whereas caspase7 was significantly up-regulated after 24 hr. The protein level of TNFalpha increased significantly to a maximum level at 12 hr. The Western blot revealed an increasing level of an 89 kDa fragment of PARP-1 from 12 to 24 hr, which is specific for apoptosis. We assume that the extrinsic pathway is more important for the onset of luteolysis, because of its earlier and higher increase during induced luteolysis.

Vascular dynamics in relation to immunolocalisation of VEGF-A, VEGFR-2 and Ang-2 in the bovine corpus luteum

Vascular dynamics during development and regression of the bovine corpus luteum were investigated morphometrically in relation to immunolocalisation of vascular endothelial growth factor A (VEGF-A), vascular endothelial growth factor receptor 2 (VEGFR-2) and angiopoietin 2 (Ang-2) in the different cell types. Angiogenesis and remodelling of the capillary network between day 8 and 17 of the oestrous cycle was driven by luteal cells, which were highly immunopositive for VEGF-A. Thereafter, immunoreactivity for VEGF-A and VEGFR-2 was mainly found in vascular smooth muscle cells, the predominant cell type in late luteolysis. During early regression, apoptosis of luteal and endothelial cells was closely correlated, resulting in a significant decrease of capillarity. In late regression, an increase in capillary density was found, suggesting that regression and transformation into the corpus albicans requires adequate perfusion. In the phases of vascular remodelling, i.e. mature and late regressing corpus luteum, high scores of Ang-2-immunopositive endothelial and smooth muscle cells were found. Therefore, it may be hypothesised that Ang-2 supports the angiogenic effects of VEGF-A in these luteal stages. Results emphasise that precise staging and a differentiated view on the cellular populations is important in evaluating the controlled regression and transformation of the corpus luteum.

Anti-Apoptotic Roles of Prostaglandin E2 and F2alpha in Bovine Luteal Steroidogenic Cells1

Production of prostaglandins (PGs) and expression of their receptors have been demonstrated in bovine corpus luteum (CL). The aim of the present study was to determine whether PGE2 and PGF2alpha have roles in bovine luteal steroidogenic cell (LSC) apoptosis. Cultured bovine LSCs obtained at the midluteal stage (Days 8-12 of the cycle) were treated for 24 h with PGE2 (0.001-1 microM) and PGF2alpha (0.001-1 microM). Prostaglandin E2 (1 microM) and PGF2alpha (1 microM) significantly stimulated progesterone (P4) production and reduced the levels of cell death in the cells cultured with or without tumor necrosis factor alpha (TNF)/interferon gamma (IFNG), in the presence and absence of FAS ligand (P < 0.05). Furthermore, DNA fragmentation induced by TNF/IFNG was observed to be suppressed by PGE2 and PGF2alpha. Prostaglandin E2 and PGF2alpha also attenuated mRNA expression of caspase 3 and caspase 8, as well as caspase 3 activity (P < 0.05) in TNF/IFNG-treated cells. FAS mRNA and protein expression were decreased only by PGF2alpha (P < 0.05). A specific P4 receptor antagonist (onapristone) attenuated the apoptosis-inhibitory effects of PGE2 and PGF2alpha in the absence of TNF/IFNG (P < 0.05). A PG synthesis inhibitor (indomethacin) reduced cell viability in PGE2- and PGF2alpha-treated cells (P < 0.05). A specific inhibitor of cyclooxygenase (PTGS), PTGS2 (NS-398), also reduced cell viability, whereas an inhibitor of PTGS1 (FR122047) did not affect it. The overall results suggest that PGE2 and PGF2alpha locally play luteoprotective roles in bovine CL by suppressing apoptosis of LSCs.

Bovine Corpus Luteum as a Heterogeneous Endocrine Organ: Vascular Structure Regulates Site-Dependent Response to PGF2α.

Bovine Corpus Luteum as a Heterogeneous Endocrine Organ: Vascular Structure Regulates Site-Dependent Response to PGF2α.

Expression Profiles of Cytokine and Progesterone Receptor Messenger Ribonucleic Acids (mRNA) in Bovine Corpus Luteum During Pregnancy.

Expression Profiles of Cytokine and Progesterone Receptor Messenger Ribonucleic Acids (mRNA) in Bovine Corpus Luteum During Pregnancy.

Effect of Prostaglandin F2α (PGF2α) Administration on the Luteotropic and Angiogenic Factors During Functional Luteolysis in the Bovine Corpus Luteum.

The essential role of endometrium PGF2α in induction of the corpus luteum (CL) regression (luteolysis) is well documented in the cow. However, the following regulatory cascade of functional luteolysis is still not fully elucidated. The aim of the present study was therefore to determine the regulation patterns of local luteotropic factors (progesterone, oxytocin and insulin-like growth factor (IGF) family members) and angiogenic factors (vascular endothelial growth factor-A (VEGF) and angiopoietin (ANPT) family members) during functional luteolysis (first 12 h after PGF2α injection) in time-defined CL classes. Cows (n=4-5 per group) in the mid-luteal phase (days 8-12) were injected with the PGF2α-analogue (cloprostenol), and CLs were collected by transvaginal ovariectomy at 0 h, 0.5 h, 2 h, 4 h and 12 h after PGF2α injection. The mRNA expression was analyzed by a quantitative real-time PCR (Rotor-Gene 3000), and the protein concentration was evaluated by enzyme immunoassay (EIA) or radio immunoassay (RIA). Progesterone concentration in blood serum and in CL tissue as well as mRNA expression of progesterone receptor in CL tissue, were significantly downregulated at 12 h after PGF2α. Oxytocin and IGF-1 peptide concentrations rapidly declined after 0.5 h and remained at low level afterwards. The IGF binding protein (BP)-1 mRNA was strongly upregulated with the maximal level at 4 h after PGF2α. This may cause further reduction of bioactive free IGF-1 in tissue. The VEGF protein decreased also at 0.5 h, with a synchronous acute and temporal increase of angiopoietin (ANPT)-2 peptide concentrations (0.5 h and 2 h after PGF2α). The similar tendency of upregulation of ANPT-2 was found for mRNA expression level, whereas the level of ANPT-1 mRNA did not change. The results suggest that the acute decrease of the local luteotropic factors and of VEGF may play a key role during the early step of functional luteolysis. The increase of ANPT-2 after PGF2α suggests a change in vascular stability to enhance the cascade of functional luteolysis.

Potential Role of Secreted Phosphoprotein 1 in the Bovine Corpus Luteum.

Potential Role of Secreted Phosphoprotein 1 in the Bovine Corpus Luteum.