scholarly journals Genomic and non-genomic effects of progesterone on prostaglandin (PG) F2α and PGE2 production in the bovine endometrium

2016 ◽  
Vol 28 (10) ◽  
pp. 1588 ◽  
Author(s):  
Mariko Kuse ◽  
Ryosuke Sakumoto ◽  
Kiyoshi Okuda
Keyword(s):  
Epithelial Cells ◽  
Mrna Expression ◽  
Stromal Cells ◽  
Follicular Phase ◽  
Pge2 Production ◽  
Cell Type ◽  
Short Term ◽  
Endometrial Cells ◽  
Receptor Mrna ◽  
Short Time

Progesterone (P4) acts through different actuating pathways called genomic and non-genomic pathways. Here we investigated whether P4 regulates prostaglandin (PG) F2α (PGF) and PGE2 production in bovine endometrium through different pathways. Cultured endometrial cells were exposed to P4 for a short time (5–20 min) or bovine serum albumin (BSA)-conjugated P4 (P4-BSA) for 24 h. Progesterone treatment for 24 h stimulated PGE2 production in epithelial cells, but suppressed both PGF and PGE2 production and the expression of PG-metabolising enzymes including phospholipase A2 (PLA2) and cyclooxygenase-2 (COX2) in stromal cells. Short-term (5–20 min) P4 treatment did not affect PLA2 or COX2 transcript levels in either cell type. P4-BSA increased PGF and PGE2 production only in epithelial cells. Nuclear P4 receptor mRNA expression in endometrium was higher at the follicular phase than at the early- to mid-luteal stages, whereas membrane P4 receptor mRNA expression did not change throughout the oestrous cycle. The overall results suggest that P4 controls PG production by inhibiting enzymes via a genomic pathway and by stimulating signal transduction via a non-genomic pathway. Consequently, P4 may protect the corpus luteum by attenuating PGF production in stromal cells and by increasing PGE2 secretion from epithelial cells.

P–369 Three-dimensional co-culture of human endometrial cells with aneuploid and euploid embryos

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
S Amiri ◽  
F Amjadi ◽  
M Ashrafi ◽  
R Aflatoonian ◽  
A Akbar. Sene ◽  
...  
Keyword(s):  
Cell Culture ◽  
Epithelial Cells ◽  
Pregnancy Rate ◽  
Stromal Cells ◽  
Phase Contrast ◽  
Three Dimensional ◽  
3D Culture ◽  
Cell Type ◽  
Endometrial Cells ◽  
Culture Systems

Abstract Study question Is implantation different in euploid and aneuploid embryos? Summary answer By simulating the human endometrium using a three-dimensional scaffold, aneuploid embryos were unable to attach to the endometrial cells, while euploid embryos attached. What is known already Although embryo selection for transfer is usually based on morphology, 70% of embryos with high morphological quality have chromosomal abnormalities. The results of implantation and pregnancy rate assessments following Preimplantation Genetic Screening (PGS) are controversial. There is still no in vitro study to compare the implantation of human euploid and aneuploid embryos. Study design, size, duration After informed consent, 10 endometrial biopsies were taken from fertile women. For scaffolding, the stromal cells were resided within the matrix, after 24 hours, the epithelial cells were seeded on the scaffold. Cell culture continued for 5 days to reach the appropriate confluence. The embryos were also examined by performing PGS following CGH Array. 10 euploid and 10 aneuploid blastocysts were selected and co-cultured for 72 hours with the 3D structure of human endometrial cells. Participants/materials, setting, methods Endometrial cells were isolated and expanded in 2D cultures to achieve enough cells. The fibrin-agarose scaffold was made and stromal and epithelial cells were cultured into and on the scaffold, respectively. Then, cell proliferation was assessed by MTT assay. The simulated endometrial construct was confirmed by H&E and IHC. Partial hatching of blastocysts was performed using a laser system. The blastocyst’s attachment to the endometrial-like structure was examined under a phase-contrast microscope and SEM. Main results and the role of chance The MTT OD of scaffolds increased during 5 days of cell culture (P < 0.05). The histological evaluation of the co-culture systems was done under light microscopy by H&E staining. On the top of the 3D culture system, epithelial cells shaped a constricted cell monolayer. Stromal cells combined with the fibrin-agarose scaffold got lengthened and expanded, displaying that the 3D culture systems supplied a suitable environment for the growth of endometrial cells. In the 3D culture, the origins and locations of epithelial and stromal cells were defined by cytokeratin and vimentin immunostaining, respectively. IHC for cytokeratin was only positive for epithelial cells in the surface epithelium. IHC for the vimentin was positive for the stromal cells located in the 3D matrix. These results showed that fibrin-agarose scaffold could simulate the human endometrial structure. Using SEM and phase-contrast microscopy, it was found that only euploid embryos were able to attach to the endometrial construct while aneuploid embryos weren’t. Limitations, reasons for caution Since the co-culture does not contain a unique cell type, and the MTT OD standard curve against cell number is specified for cell type, the number of growing cells in the co-culture cannot be calculated; therefore it is reported as OD. Wider implications of the findings: Our findings determined that PGS allows us to transfer top-quality embryos with higher implantation potential. It improves implantation and pregnancy rate during ART cycles, especially in patients with recurrent implantation failure. Trial registration number Not applicable

Lysophosphatidic Acid Stimulates Prostaglandin E2 Production in Cultured Stromal Endometrial Cells Through LPA1 Receptor

2009 ◽  
Vol 234 (8) ◽  
pp. 986-993 ◽  
Author(s):  
Izabela Woclawek-Potocka ◽  
Katarzyna Kondraciuk ◽  
Dariusz Jan Skarzynski
Keyword(s):  
Epithelial Cells ◽  
Intracellular Calcium ◽  
Mrna Expression ◽  
Lysophosphatidic Acid ◽  
Stromal Cells ◽  
Calcium Ion ◽  
Cell Types ◽  
Endometrial Cells ◽  
Intracellular Calcium Ion

Lysophosphatidic acid (LPA) has been shown to be a potent modulator of prostaglandin (PG) secretion during the luteal phase of the estrous cycle in the bovine endometrium in vivo. The aims of the present study were to determine the cell types of the bovine endometrium (epithelial or stromal cells) responsible for the secretion of PGs in response to LPA, the cellular, receptor, intracellular, and enzymatic mechanisms of LPA action. Cultured bovine epithelial and stromal cells were exposed to LPA (10−5–10−9 M), tumor necrosis factor α (TNFα; 10 ng/mL) or oxytocin (OT; 10−7 M) for 24 h. LPA treatment resulted in a dose-dependent increase of PGE2 production in stromal cells, but not in epithelial cells. LPA did not influence PGF2α production in stromal or epithelial cells. To examine which type of LPA G-protein–coupled receptor (LP-GPCR; LPA1, LPA2, or LPA3) is responsible for LPA action, stromal cells were preincubated with three selected blockers of LPA receptors: NAEPA, DGPP, and Ki16425 for 0.5 h, and then stimulated with LPA. Only Ki16425 inhibited the stimulatory effect of LPA on PGE2 production and cell proliferation in the stromal cells. LPA-induced intracellular calcium ion mobilization was also inhibited only by Ki16425. Finally, we examined whether LPA-induced PGE2 synthesis in stromal cells is via the influence on mRNA expression for the enzymes responsible for PGE2 synthesis— PGE 2 synthase ( PGES) and PG-endoperoxide synthase 2 ( PTGS2). We demonstrated that the stimulatory effect of LPA on PGE2 production in stromal cells is via the stimulation of PTGS2 and PGES mRNA expression in the cells. The overall results indicate that LPA stimulates PGE2 production, cell viability, and intracellular calcium ion mobilization in cultured stromal endometrial cells via Ki16425-sensitive LPA1 receptors. Moreover, LPA exerts a stimulatory effect on PGE2 production in stromal cells via the induction of PTGS2 and PGES mRNA expression.

scholarly journals Bovine herpesvirus 4 is tropic for bovine endometrial cells and modulates endocrine function

Reproduction ◽  
2007 ◽  
Vol 134 (1) ◽  
pp. 183-197 ◽  
Author(s):  
Gaetano Donofrio ◽  
Shan Herath ◽  
Chiara Sartori ◽  
Sandro Cavirani ◽  
Cesidio Filippo Flammini ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Stromal Cells ◽  
Bovine Herpesvirus ◽  
Endocrine Function ◽  
Uterine Disease ◽  
Endometrial Stromal Cells ◽  
Persistently Infected ◽  
Endometrial Cells ◽  
Bovine Herpesvirus 4 ◽  
Herpesvirus 4

Bovinepostpartumuterine disease, metritis, affects about 40% of animals and is widely considered to have a bacterial aetiology. Although the γ-herpesvirus bovine herpesvirus 4 (BoHV-4) has been isolated from several outbreaks of metritis or abortion, the role of viruses in endometrial pathology and the mechanisms of viral infection of uterine cells are often ignored. The objectives of the present study were to explore the interaction, tropism and outcomes of BoHV-4 challenge of endometrial stromal and epithelial cells. Endometrial stromal and epithelial cells were purified and infected with a recombinant BoHV-4 carrying an enhanced green fluorescent protein (EGFP) expression cassette to monitor the establishment of infection. BoHV-4 efficiently infected both stromal and epithelial cells, causing a strong non-apoptotic cytopathic effect, associated with robust viral replication. The crucial step for the BoHV-4 endometriotropism appeared to be after viral entry as there was enhanced transactivation of the BoHV-4 immediate early 2 gene promoter following transient transfection into the endometrial cells. Infection with BoHV-4 increased cyclooxygenase 2 protein expression and prostaglandin estradiol secretion in endometrial stromal cells, but not epithelial cells. Bovine macrophages are persistently infected with BoHV-4, and co-culture with endometrial stromal cells reactivated BoHV-4 replication in the persistently infected macrophages, suggesting a symbiotic relationship between the cells and virus. In conclusion, the present study provides evidence of cellular and molecular mechanisms, supporting the concept that BoHV-4 is a pathogen associated with uterine disease.

107 PRODUCTION OF EPIDERMAL GROWTH FACTOR BY BOVINE ENDOMETRIAL CELLS AND ITS EFFECTS ON EMBRYONIC INTERFERONT AND PROSTAGLANDIN

2015 ◽  
Vol 27 (1) ◽  
pp. 146 ◽  
Author(s):  
T. J. Acosta ◽  
K. Takatsu
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Epithelial Cells ◽  
Stromal Cells ◽  
Pge2 Production ◽  
Bioactive Molecules ◽  
Significant Difference ◽  
Epidermal Growth ◽  
Maternal Recognition Of Pregnancy ◽  
Cells Cultured

A dynamic interaction between bioactive products of the embryo (blastocyst) and the endometrium is crucial for the successful establishment of pregnancy. In ruminants, the principal signal for maternal recognition of pregnancy is interferon-τ (IFNT) secreted by the trophoectoderm between Days 8 and 20 post-fertilization. Epidermal growth factor (EGF) produced by the endometrium acting through EGF receptors (EGFR) present in the blastocyst seems to regulate embryonic production of IFNT. Epidermal growth factor and IFNT have been shown to play crucial roles in controlling luteolytic prostaglandin (PG) F2α (PGF) and luteotropic PGE2 production by bovine endometrium. However, it is unknown how these bioactive molecules regulate uterine function during maternal recognition of pregnancy. To clarify the main source of EGF in bovine endometrium and the mechanisms regulating the interaction between the hatched blastocyst and maternal uterine environment, the production of EGF by cultured endometrial epithelial and stromal cells and the effects of EGF on embryonic IFNT and PG were investigated. In addition, the effects of EGF on PGE2 and PGF production by cultured epithelial or stromal cells were examined. Endometrial epithelial and stromal cells were enzymatically isolated on the day of ovulation, seeded at a density of 100 000 viable cells mL–1, and cultured at 38°C in a humidified atmosphere of 5% CO2 in air. After the cells reached 90% confluence, they were cultured in the presence or absence of EGF (0.1, 1.0, 10, and 100 ng mL–1) for 24 h. Cells cultured in the absence of EGF and their cultured media were collected separately for protein analysis. Hatched bovine blastocysts (Days 8–10) were also cultured and exposed to EGF (1, 10, and 100 ng mL–1) for 24 h. Protein concentrations of EGF and IFNT in the cultured media were determined by commercial enzyme immunoassay kit. Hormonal concentrations were analysed by ANOVA followed by Fisher's protected least-significant difference procedure (PLSD) as a multiple comparison test by StatView (Abacus Concepts Inc., Berkeley, CA, USA). The concentration of EGF in the culture media of epithelial cells cultured in the absence of EGF was significantly (P < 0.05) higher than in the cultured media of endometrial stromal cells. Epidermal growth factor (10 and 100 ng mL–1) increased embryonic production of IFNT and luteotropic PGE2 production but not luteolytic PGF by hatched blastocyst. EGF (100 ng mL–1) increased both PGE2 and PGF production (P < 0.05) by cultured endometrial epithelial and stromal cells. The overall results suggest that endometrial epithelial cells rather than stromal cells are the main source of EGF. Epidermal growth factor produced by epithelial cells stimulates the production of IFNT by bovine trophoblasts. The capacity of conceptus to increase IFNT and luteotropic PGE2 production rather than luteolytic PGF in response to EGF stimulation may be essential for the establishment of pregnancy in cattle.

scholarly journals Increased expression of CYP1A1 and CYP1B1 in ovarian/peritoneal endometriotic lesions

Reproduction ◽  
2016 ◽  
Vol 151 (6) ◽  
pp. 683-692 ◽  
Author(s):  
Carla A Piccinato ◽  
Rosa M Neme ◽  
Natália Torres ◽  
Lívia Renta Sanches ◽  
Priscilla Bento Mattos Cruz Derogis ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Stromal Cells ◽  
Premenopausal Women ◽  
Estrogen Metabolism ◽  
Cross Sectional ◽  
Eutopic Endometrium ◽  
Endometrial Cells ◽  
Specific Regulation ◽  
A Site

Abstract Endometriosis is an estrogen-dependent disease affecting up to 10% of all premenopausal women. There is evidence that different endometriosis sites show distinct local estrogen concentration, which, in turn, might be due to a unique local estrogen metabolism. We aimed to investigate whether there was a site-specific regulation of selected enzymes responsible for the oxidative metabolism of estrogens in biopsy samples and endometrial and endometriotic stromal cells. Cytochrome P450 (CYP) 1A1 and CYP1B1 mRNA and protein expressions in deep-infiltrating (rectal, retossigmoidal, and uterossacral) lesions, superficial (ovarian and peritoneal) lesions, and eutopic and healthy (control) endometrium were evaluated by real-time PCR and western blot. Using a cross-sectional study design with 58 premenopausal women who were not under hormonal treatment, we were able to identify an overall increased CYP1A1 and CYP1B1 mRNA expression in superficial lesions compared with the healthy endometrium. CYP1A1 mRNA expression in superficial lesions was also greater than in the eutopic endometrium. Interestingly, we found a similar pattern of CYP1A1 and CYP1B1 expression in in vitro stromal cells isolated from ovarian lesions (n=3) when compared with stromal cells isolated from either rectum lesions or eutopic endometrium. In contradiction, there was an increased half-life of estradiol (measured by HPLC-MS-MS) in ovarian endometriotic stromal cells compared with paired eutopic stromal endometrial cells. Our results indicate that there is a site-dependent regulation of CYP1A1 and CYP1B1 in ovarian/peritoneal lesions and ovarian endometriotic stromal cells, whereas a slower metabolism is taking place in these cells.

scholarly journals Decorin induced by progesterone plays a crucial role in suppressing endometriosis

2014 ◽  
Vol 223 (2) ◽  
pp. 203-216 ◽  
Author(s):  
Yoshihiro Joshua Ono ◽  
Yoshito Terai ◽  
Akiko Tanabe ◽  
Atsushi Hayashi ◽  
Masami Hayashi ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Mrna Expression ◽  
Cell Lines ◽  
Stromal Cells ◽  
Crucial Role ◽  
Dependent Manner ◽  
Endometrial Stromal Cells ◽  
Endometrial Cells ◽  
Cycle Arrest ◽  
Dose Dependent

Dienogest, a synthetic progestin, has been shown to be effective against endometriosis, although it is still unclear as to how it affects the ectopic endometrial cells. Decorin has been shown to be a powerful endogenous tumor repressor acting in a paracrine fashion to limit tumor growth. Our objectives were to examine the direct effects of progesterone and dienogest on the in vitro proliferation of the human ectopic endometrial epithelial and stromal cell lines, and evaluate as to how decorin contributes to this effect. We also examined DCN mRNA expression in 50 endometriosis patients. The growth of both cell lines was inhibited in a dose-dependent manner by both decorin and dienogest. Using a chromatin immunoprecipitation assay, it was noted that progesterone and dienogest directly induced the binding of the decorin promoter in the EMOsis cc/TERT cells (immortalized human ovarian epithelial cells) and CRL-4003 cells (immortalized human endometrial stromal cells). Progesterone and dienogest also led to significant induced cell cycle arrest via decorin by promoting production of p21 in both cell lines in a dose-dependent manner. Decorin also suppressed the expression of MET in both cell lines. We confirmed that DCN mRNA expression in patients treated with dienogest was higher than that in the control group. In conclusion, decorin induced by dienogest appears to play a crucial role in suppressing endometriosis by exerting anti-proliferative effects and inducing cell cycle arrest via the production of p21 human ectopic endometrial cells and eutopic endometrial stromal cells.

scholarly journals Ovarian steroids affect prostaglandin production in equine endometrial cells in vitro

2014 ◽  
Vol 220 (3) ◽  
pp. 263-276 ◽  
Author(s):  
Anna Z Szóstek ◽  
António M Galvão ◽  
Graça M Ferreira-Dias ◽  
Dariusz J Skarzynski
Keyword(s):  
Cell Proliferation ◽  
Epithelial Cells ◽  
Protein Expression ◽  
Stromal Cells ◽  
Positive Control ◽  
Dependent Manner ◽  
Ovarian Steroids ◽  
Endometrial Cells ◽  
Prostaglandin Endoperoxide Synthase

This study aimed to evaluate the influence of ovarian steroids on equine endometrial epithelial and stromal cells, specifically i) prostaglandin (PG) production in a time-dependent manner, ii) specific PG synthases mRNA transcription and protein expression, and iii) cell proliferation. After passage I, cells were exposed to vehicle, oxytocin (OT, positive control, 10−7M), progesterone (P4, 10−7M), 17β estradiol (E2, 10−9M), or P4+E2for 12, 24, 48, or 72 h. Following treatment, PG concentration was determined using the direct enzyme immunoassay (EIA) method. Alterations inPGsynthases mRNA transcriptions,PGsynthases protein expression, and cell proliferation in response to the treatments were determined after 24 h using real-time PCR, western blot, or 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide respectively. After 24 h, E2and P4+E2increased PGE2and PGF2αsecretion as well as specific prostaglandin-endoperoxide synthase-2 (PTGS2), PGE2synthases (PGES), and PGF2αsynthases (PGFS) expression in the epithelial cells (P<0.05). Additionally, E2and P4+E2increased PTGS2 expression in stromal cells after 24 h (P<0.05). In stromal cells, P4+E2increased PGE2production as well as PGES expression after 24 h (P<0.05). Both E2and P4+E2increased PGF2αproduction by stromal cells after 24 h (P<0.05). Ovarian steroids affected proliferation of stromal and epithelial cells during the 24-h incubation period (P<0.05). We provide evidence that ovarian steroids affect PG production in equine endometrial cells, upregulating PTGS2, PGES, and PGFS expression. Ovarian steroid-stimulated PG production could be an important mechanism occurring in the equine endometrium that is involved in the regulation of the estrous cycle and early pregnancy.

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