scholarly journals Influence of Estradiol-17beta on Progesterone and Estrogen Receptor mRNA Expression in Porcine Follicular Granulosa Cells during Short-Term, In Vitro Real-Time Cell Proliferation

2016 ◽  
Vol 2016 ◽  
pp. 1-8 ◽  
Author(s):  
Sylwia Ciesiółka ◽  
Joanna Budna ◽  
Karol Jopek ◽  
Artur Bryja ◽  
Wiesława Kranc ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Estrogen Receptor ◽  
Progesterone Receptor ◽  
Real Time ◽  
Granulosa Cells ◽  
Housekeeping Genes ◽  
Cumulus Cells ◽  
Short Term ◽  
Estrogen Receptor Mrna

Progesterone (P4) and estradiol (E2) play a significant role in mammalian reproduction. Our study demonstrated that separated porcine cumulus cells (CCs) and/or granulosa cells (GCs) might proliferate in vitro during short-term, real-time primary culture. The GCs were analyzed according to gene expression of the progesterone receptor (nuclear form) (pgr), progesterone receptor membrane component 1 (pgrmc1), and estrogen-related receptor beta 3 (esrrb3) in relation to two housekeeping genes: actb and pbgd. GCs were cultivated in medium with the E2. Both pgr/actb and pgr/pbgd revealed higher expression between 24 and 168 h of IVC of prolonged E2 treatment and at 48 h of IVC after acute E2 administration. The pgrmc1/actb and pgrmc1/pbgd displayed increased expression after prolonged E2 treatment between 24 and 120 h of IVC. The highest level of esrrb3/actb at 120 and 144 h, as well as esrrb3/pbgd at 120 h, in untreated controls as compared to the hormone-stimulated group, was observed. We suggest that E2 significantly influences the upregulation of pgr, pgrmc1, and esrrb3 expression in porcine GCs during real-time cell proliferation. Since esrrb3 expression is stimulated by E2 in both an acute and prolonged manner, estradiol may be recognized as a potential estrogen receptor agonist in GCs.

scholarly journals Time- and Dose-Dependent Effects of 17 Beta-Estradiol on Short-Term, Real-Time Proliferation and Gene Expression in Porcine Granulosa Cells

2017 ◽  
Vol 2017 ◽  
pp. 1-9 ◽  
Author(s):  
Sylwia Ciesiółka ◽  
Joanna Budna ◽  
Karol Jopek ◽  
Artur Bryja ◽  
Wiesława Kranc ◽  
...  
Keyword(s):  
Real Time ◽  
Granulosa Cells ◽  
Hormone Receptor ◽  
Proliferative Activity ◽  
Cell Transformation ◽  
Luteinizing Hormone Receptor ◽  
Luteal Cell ◽  
Ovarian Activity ◽  
Short Term

The key mechanisms responsible for achievement of full reproductive and developmental capability in mammals are the differentiation and transformation of granulosa cells (GCs) during folliculogenesis, oogenesis, and oocyte maturation. Although the role of 17 beta-estradiol (E2) in ovarian activity is widely known, its effect on proliferative capacity, gap junction connection (GJC) formation, and GCs-luteal cells transformation requires further research. Therefore, the goal of this study was to assess the real-time proliferative activity of porcine GCs in vitro in relation to connexin (Cx), luteinizing hormone receptor (LHR), follicle stimulating hormone receptor (FSHR), and aromatase (CYP19A1) expression during short-term (168 h) primary culture. The cultured GCs were exposed to acute (at 96 h of culture) and/or prolonged (between 0 and 168 h of culture) administration of 1.8 and 3.6 μM E2. The relative abundance of Cx36, Cx37, Cx40, Cx43, LHR, FSHR, and CYP19A1 mRNA was measured. We conclude that the proliferation capability of GCs in vitro is substantially associated with expression of Cxs, LHR, FSHR, and CYP19A1. Furthermore, the GC-luteal cell transformation in vitro may be significantly accompanied by the proliferative activity of GCs in pigs.

scholarly journals The Differential Metabolomes in Cumulus and Mural Granulosa Cells from Human Preovulatory Follicles

2021 ◽  
Author(s):  
Er-Meng Gao ◽  
Bongkoch Turathum ◽  
Ling Wang ◽  
Di Zhang ◽  
Yu-Bing Liu ◽  
...  
Keyword(s):  
Oocyte Maturation ◽  
Granulosa Cells ◽  
Quantitative Polymerase Chain Reaction ◽  
Cumulus Cells ◽  
Cholesterol Transport ◽  
Vitro Fertilization ◽  
Expression Of Genes ◽  
Preovulatory Follicles ◽  
Morphological Differences

AbstractThis study evaluated the differences in metabolites between cumulus cells (CCs) and mural granulosa cells (MGCs) from human preovulatory follicles to understand the mechanism of oocyte maturation involving CCs and MGCs. CCs and MGCs were collected from women who were undergoing in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) treatment. The differences in morphology were determined by immunofluorescence. The metabolomics of CCs and MGCs was measured by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) followed by quantitative polymerase chain reaction (qPCR) and western blot analysis to further confirm the genes and proteins involved in oocyte maturation. CCs and MGCs were cultured for 48 h in vitro, and the medium was collected for detection of hormone levels. There were minor morphological differences between CCs and MGCs. LC-MS/MS analysis showed that there were differences in 101 metabolites between CCs and MGCs: 7 metabolites were upregulated in CCs, and 94 metabolites were upregulated in MGCs. The metabolites related to cholesterol transport and estradiol production were enriched in CCs, while metabolites related to antiapoptosis were enriched in MGCs. The expression of genes and proteins involved in cholesterol transport (ABCA1, LDLR, and SCARB1) and estradiol production (SULT2B1 and CYP19A1) was significantly higher in CCs, and the expression of genes and proteins involved in antiapoptosis (CRLS1, LPCAT3, and PLA2G4A) was significantly higher in MGCs. The level of estrogen in CCs was significantly higher than that in MGCs, while the progesterone level showed no significant differences. There are differences between the metabolomes of CCs and MGCs. These differences may be involved in the regulation of oocyte maturation.

scholarly journals NRF2-mediated signaling is a master regulator of transcription factors in bovine granulosa cells under oxidative stress condition

2021 ◽  
Author(s):  
Mohamed Omar Taqi ◽  
Mohammed Saeed-Zidane ◽  
Samuel Gebremedhn ◽  
Dessie Salilew-Wondim ◽  
Ernst Tholen ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Cell Proliferation ◽  
Endoplasmic Reticulum ◽  
Transcription Factors ◽  
Endoplasmic Reticulum Stress ◽  
Granulosa Cells ◽  
Mitochondrial Activity ◽  
Small Interference Rna ◽  
Nrf2 Signaling Pathway

AbstractTranscription factors (TFs) are known to be involved in regulating the expression of several classes of genes during folliculogenesis. However, the regulatory role of TFs during oxidative stress (OS) is not fully understood. The current study was aimed to investigate the regulation of the TFs in bovine granulosa cells (bGCs) during exposure to OS induced by H2O2 in vitro. For this, bGCs derived from ovarian follicles were cultured in vitro till their confluency and then treated with H2O2 for 40 min. Twenty-four hours later, cells were subjected to various phenotypic and gene expression analyses for genes related to TFs, endoplasmic reticulum stress, apoptosis, cell proliferation, and differentiation markers. The bGCs exhibited higher reactive oxygen species accumulation, DNA fragmentation, and endoplasmic reticulum stress accompanied by reduction of mitochondrial activity after exposure to OS. In addition, higher lipid accumulation and lower cell proliferation were noticed in H2O2-challenged cells. The mRNA level of TFs including NRF2, E2F1, KLF6, KLF9, FOS, SREBF1, SREBF2, and NOTCH1 was increased in H2O2-treated cells compared with non-treated controls. However, the expression level of KLF4 and its downstream gene, CCNB1, were downregulated in the H2O2-challenged group. Moreover, targeted inhibition of NRF2 using small interference RNA resulted in reduced expression of KLF9, FOS, SREBF2, and NOTCH1 genes, while the expression of KLF4 was upregulated. Taken together, bovine granulosa cells exposed to OS exhibited differential expression of various transcription factors, which are mediated by the NRF2 signaling pathway.

Activation of chromosomal vitellogenin genes in Xenopus oocytes by pure estrogen receptor and independent activation of albumin genes

1990 ◽  
Vol 10 (12) ◽  
pp. 6674-6682
Author(s):  
E A McKenzie ◽  
N A Cridland ◽  
J Knowland
Keyword(s):  
Estrogen Receptor ◽  
Xenopus Oocytes ◽  
Liver Cells ◽  
Receptor Protein ◽  
Estrogen Receptor Protein ◽  
Receptor Mrna ◽  
Estrogen Receptor Mrna ◽  
Vitellogenin Gene ◽  
Vitellogenin Genes

We generate pure estrogen receptor protein in Xenopus oocytes by injecting them with estrogen receptor mRNA synthesized in vitro. A chromosomal vitellogenin gene, which normally responds to estrogen only in liver cells, is activated. Primer extension shows that initiation is accurate, and ribonuclease mapping shows that the first exon is correctly spliced out of the initial transcript. Long transcripts are produced, one being equal in length to poly(A)- vitellogenin mRNA. Immunochemical estimates of receptor levels in the oocyte nuclei suggest that pure receptor, acting alone, cannot activate oocyte vitellogenin genes unless unusually large amounts are present. However, when a receptor-free extract from liver cells is also injected, the amount of receptor required is reduced. Such an extract, but not pure receptor, can also activate albumin genes in oocytes.

scholarly journals Investigating the Intrafollicular Factors Affecting Oocyte Developmental Competency

2021 ◽  
Author(s):  
◽  
Zaramasina Clark
Keyword(s):  
Gene Expression ◽  
Oocyte Maturation ◽  
Granulosa Cells ◽  
Embryo Quality ◽  
Cumulus Cells ◽  
Significant Finding ◽  
High Quality ◽  
Final Maturation

<p>The number of cycles of assisted reproductive technologies (ART) performed increased by ~9.5 % globally between 2008 and 2010. In spite of this, the success rate in terms of delivery was only ~19.0 % (Dyer et al., 2016). This discrepancy between the demand for, and success of, these technologies necessitates the development of tools to improve ART efficiency. To facilitate this, a better understanding of how the microenvironment changes within the developing follicle to culminate in a mature, developmentally-competent oocyte is required. This study employed an in vivo and in vitro ovine model to investigate the relationship between the surrounding microenvironment and oocyte maturation, and in particular, the attainment of oocyte developmental competency and high-quality embryos.  The first objective of this PhD study was to comprehensively investigate the changing microenvironment of in vivo matured, presumptive preovulatory (PPOV) follicles from wild-type (++) and high ovulation rate (OR; I+B+) ewes. The high OR ewes were heterozygous carriers of mutations in BMP15 (I+) and BMPRIB (B+). Functional differences in follicular somatic (granulosa and cumulus) cells between these genotypes, including differential gonadotropin responsiveness of granulosa cells, composition of follicular fluid and gene expression profiles in cumulus cells were evident. These differences emerged as part of a compensatory mechanism by which oocytes from smaller follicles, containing fewer granulosa cells, achieved developmental competency in I+B+ ewes.  The second objective of this PhD study was to develop new approaches for improving current in vitro maturation (IVM) strategies. The first approach utilised in this study focused on developing biomarkers that could be used to improve prediction of developmental competency in oocytes and in vitro produced embryos. This involved interrogating the hypothesis that a combination of molecular and morphokinetic biomarkers would better predict the developmental competency of oocytes and embryos compared to using these biomarkers alone. The second approach utilised in this PhD study tested the effects of modulating IVM conditions to better mimic the follicular microenvironment of a high, compared to a low, OR species on oocyte developmental competency and embryo quality. This involved supplementing IVM media with different ratios of two oocyte-secreted growth factors, i.e. GDF9:BMP15, that were representative of low or high OR species. These approaches demonstrated significant potential and warrant further investigation.  The most significant finding of this study was that despite variances in the surrounding microenvironment during in vivo and in vitro oocyte maturation that culminated in differential gene expression patterns in cumulus cells, and divergent gonadotropin-responsiveness of granulosa cells, the gene expression signatures of developmentally-competent oocytes and the morphokinetics of high-quality embryos were unaltered. This confirms the value of developing such biomarkers for oocyte development competency and embryo quality that remain unaltered despite a changing surrounding environment. Interestingly, simulating the ratio of GDF9:BMP15 that oocytes from high OR species are exposed to during maturation improved developmental competency in oocytes as demonstrated by increased blastocyst rates. Furthermore, this study has demonstrated that combinations of molecular (cumulus cell gene expression) and morphokinetic biomarkers improved the ability to predict developmental competency in oocytes and embryos. Overall, this study revealed novel information regarding the follicular microenvironment during final maturation and identified several novel approaches to improving the efficiency of ART.</p>

scholarly journals Activation of chromosomal vitellogenin genes in Xenopus oocytes by pure estrogen receptor and independent activation of albumin genes.

1990 ◽  
Vol 10 (12) ◽  
pp. 6674-6682 ◽  
Author(s):  
E A McKenzie ◽  
N A Cridland ◽  
J Knowland
Keyword(s):  
Estrogen Receptor ◽  
Xenopus Oocytes ◽  
Liver Cells ◽  
Receptor Protein ◽  
Estrogen Receptor Protein ◽  
Receptor Mrna ◽  
Estrogen Receptor Mrna ◽  
Vitellogenin Gene ◽  
Vitellogenin Genes

We generate pure estrogen receptor protein in Xenopus oocytes by injecting them with estrogen receptor mRNA synthesized in vitro. A chromosomal vitellogenin gene, which normally responds to estrogen only in liver cells, is activated. Primer extension shows that initiation is accurate, and ribonuclease mapping shows that the first exon is correctly spliced out of the initial transcript. Long transcripts are produced, one being equal in length to poly(A)- vitellogenin mRNA. Immunochemical estimates of receptor levels in the oocyte nuclei suggest that pure receptor, acting alone, cannot activate oocyte vitellogenin genes unless unusually large amounts are present. However, when a receptor-free extract from liver cells is also injected, the amount of receptor required is reduced. Such an extract, but not pure receptor, can also activate albumin genes in oocytes.

Relative importance of estrogen and progesterone receptor assays as prognostic indicators in primary breast cancer: a short-term study.

1989 ◽  
Vol 35 (2) ◽  
pp. 238-240 ◽  
Author(s):  
D Brocklehurst ◽  
C E Wilde ◽  
J A Finbow ◽  
R Brett ◽  
A E Champion ◽  
...  
Keyword(s):  
Estrogen Receptor ◽  
Progesterone Receptor ◽  
Prognostic Indicators ◽  
Lymph Node Status ◽  
Relative Importance ◽  
Short Term ◽  
Term Study ◽  
Estrogen Receptor Positivity ◽  
Short Term Study ◽  
Grade Iii

Abstract This short-term study of the relative importance of estrogen and progesterone receptors shows that progesterone receptor correlates better than estrogen receptor with tumor recurrence regardless of lymph-node status. Life-table analysis has effectively identified only two groups of patients that may be classified by progesterone receptor status alone. Progesterone-receptor negativity correlated well with tumors of histological Grade III; estrogen-receptor positivity correlated with Grade I and II tumors. The earlier recurrence of Grade III breast tumors may explain why progesterone receptor is a better prognostic indicator than estrogen receptor in short-term studies.

70 NON-PLATED GRANULOSA AND CUMULUS CELLS AND FIRST PASSAGE FIBROBLASTS AS NUCLEUS DONOR FOR GOAT CLONING

2006 ◽  
Vol 18 (2) ◽  
pp. 143
Author(s):  
D. Salamone ◽  
M. Catala ◽  
A. Gibbons ◽  
F. Pereyra Bonnet ◽  
M. Cueto
Keyword(s):  
Granulosa Cells ◽  
Room Temperature ◽  
Uv Light ◽  
Cumulus Cells ◽  
Cleavage Rate ◽  
First Passage ◽  
Chi Square ◽  
Donor Nucleus ◽  
Chi Square Test

Different types of somatic cells have been used as nucleus donors for cloning. Most of them were previously cultured in vitro as a monolayer through several plate passages. The experiment reported here was conducted to study the potential usages of granulosa and cumulus cells for cloning without previous culture as a monolayer. A first-plate-passage fibroblast was also used. Oocytes were aspirated by laparoscopy from Criolla goats and matured in TCM-199 + 5% FCS at 39°C for 24 h. Matured oocytes were denuded by vortexing for 3 min in TL HEPES with 1 mg/mL bovine testis hyaluronidase. Metaphases were assessed and oocytes were enucleated by visualization with Hoechst 33342 (5 μg/mL) under UV light (<6 s). Granulosa and cumulus cells were also recovered by laparoscopy and maintained in maturation medium in cryotube for 20 h at room temperature or 39°C, respectively. Goat adult ear fibroblasts were cultured for 1 or 2 weeks and used 2 days after confluence. All types of donor cells were transferred to the perivitlline space of enucleated oocytes and fused by an electrical pulse. After 2 h, activation was induced by incubation in TL-HEPES with 5 µM ionomycin for 4 min and 2 mM 6-DMAP for 3 h. The oocytes were then washed with TL-HEPES and cultured in SOF medium and atmosphere of 5% CO2 + 5% O2 + 90% N2. Cleavage (Day 2) and development to blastocysts (Day 6) were recorded and analyzed by chi-square test. The cleavage rate for non-plated granulosa cells was higher than for the other treatment goups; cumulus cells had a lower rate of development to blastocysts (Table 1). These results suggest that granulosa cells collected and maintained for 24 h at room temperature could be used to produce cloned blastocysts. Table 1. Effect of non-plated granulosa and cumulus cells and first passage fibroblasts as donor nucleus oocytes in goat cloning

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