scholarly journals Signal transducer and activator of transcription (STAT) 1 and STAT3 are expressed in the human ovary and have Janus kinase 1-independent functions in the COV434 human granulosa cell line

2020 ◽  
Vol 32 (12) ◽  
pp. 1027
Author(s):  
E. R. Frost ◽  
E. A. Ford ◽  
A. E. Peters ◽  
N. L. Reed ◽  
E. A. McLaughlin ◽  
...  
Keyword(s):  
Cell Line ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Cell Function ◽  
Janus Kinase ◽  
Ovary Development ◽  
Signal Transducer ◽  
Human Ovary ◽  
Human Granulosa Cell ◽  
Independent Functions

Ovarian granulosa cells are fundamental for oocyte maintenance and maturation. Recent studies have demonstrated the importance of members of the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signalling pathway in the granulosa cell population of mouse and horse ovaries, with perturbation of JAK1 signalling in the mouse shown to impair oocyte maintenance and accelerate primordial follicle activation. The presence and role of the JAK/STAT pathway in human granulosa cells has yet to be elucidated. In this study, expression of JAK1, STAT1 and STAT3 was detected in oocytes and granulosa cells of human ovarian sections from fetal (40 weeks gestation) and premenopausal ovaries (34–41 years of age; n=3). To determine the effects of JAK1 signalling in granulosa cells, the human granulosa-like cell line COV434 was used, with JAK1 inhibition using ruxolitinib. Chemical inhibition of JAK1 in COV434 cells with 100nM ruxolitinib for 72h resulted in significant increases in STAT3 mRNA (P=0.034) and p-Y701-STAT1 protein (P=0.0117), demonstrating a role for JAK1 in modulating STAT in granulosa cells. This study implicates a conserved role for JAK/STAT signalling in human ovary development, warranting further investigation of this pathway in human granulosa cell function.

Phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) and Janus kinase/signal transducer and activator of transcription (JAK/STAT) follicular signalling is conserved in the mare ovary

2018 ◽  
Vol 30 (4) ◽  
pp. 624 ◽  
Author(s):  
Sally E. Hall ◽  
Rose M. O. Upton ◽  
Eileen A. McLaughlin ◽  
Jessie M. Sutherland
Keyword(s):  
Protein Kinase ◽  
Granulosa Cells ◽  
Protein Kinase B ◽  
Quantitative Polymerase Chain Reaction ◽  
Janus Kinase ◽  
Signalling Pathways ◽  
Ovary Development ◽  
Signal Transducer ◽  
Adult Ovary ◽  
Phosphoinositide 3 Kinase

The mare ovary is unique in its anatomical structure; however, the signalling pathways responsible for physiological processes, such as follicular activation, remain uncharacterised. This provided us with the impetus to explore whether signalling molecules from important folliculogenesis pathways, phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) and Janus kinase/signal transducer and activator of transcription (JAK/STAT), are conserved in the mare ovary. Messenger RNA expression of six genes important in follicle development was measured using quantitative polymerase chain reaction and protein localisation of key pathway members (PI3K, AKT1, phosphatase and tensin homologue (PTEN), JAK1, STAT3 and suppressor of cytokine signalling 4 (SOCS4)) was compared in tissue from fetal and adult mare ovaries. Tissue from adult ovaries exhibited significantly increased levels of mRNA expression of PI3K, AKT1, PTEN, JAK1, STAT3 and SOCS4 compared with tissue from fetal ovaries. PI3K, AKT1, JAK1 and STAT3 demonstrated redistributed localisation, from pregranulosa cells in fetal development, to both the oocyte and granulosa cells of follicles in the adult ovary, whilst negative feedback molecules PTEN and SOCS4 were only localised to the granulosa cells in the adult ovary. These findings suggest that the PI3K/AKT and JAK/STAT signalling pathways are utilised during folliculogenesis in the mare, similarly to previously studied mammalian species, and may serve as useful biomarkers for assessment of ovary development in the horse.

scholarly journals The role of IGFs in the regulation of ovarian follicular growth in the brushtail possum (Trichosurus vulpecula)

Reproduction ◽  
2010 ◽  
Vol 140 (2) ◽  
pp. 295-303 ◽  
Author(s):  
Jennifer L Juengel ◽  
Lisa J Haydon ◽  
Brigitta Mester ◽  
Brian P Thomson ◽  
Michael Beaumont ◽  
...  
Keyword(s):  
Granulosa Cell ◽  
Granulosa Cells ◽  
Cell Function ◽  
Antral Follicle ◽  
Brushtail Possum ◽  
Follicular Growth ◽  
Theca Cells ◽  
Ovarian Follicular Growth

IGFs are known to be key regulators of ovarian follicular growth in eutherian mammals, but little is known regarding their role in marsupials. To better understand the potential role of IGFs in the regulation of follicular growth in marsupials, expression of mRNAs encoding IGF1, IGF2, IGF1R, IGF-binding protein 2 (IGFBP2), IGFBP4 and IGFBP5 was localized by in situ hybridization in developing ovarian follicles of the brushtail possum. In addition, the effects of IGF1 and IGF2 on granulosa cell function were tested in vitro. Both granulosa and theca cells synthesize IGF mRNAs, with the theca expressing IGF1 mRNA and granulosa cell expressing IGF2 mRNA. Oocytes and granulosa cells express IGF1R. Granulosa and theca cells expressed IGFBP mRNAs, although the pattern of expression differed between the BPs. IGFBP5 mRNA was differentially expressed as the follicles developed with granulosa cells of antral follicles no longer expressing IGFBP5 mRNA, suggesting an increased IGF bioavailability in the antral follicle. The IGFBP protease, PAPPA mRNA, was also expressed in granulosa cells of growing follicles. Both IGF1 and IGF2 stimulated thymidine incorporation but had no effect on progesterone production. Thus, IGF may be an important regulator of ovarian follicular development in marsupials as has been shown in eutherian mammals.

P–229 Oxidative stress and Metformin; an in-vitro study on serum and primary human granulosa cell cultures

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
F Alam ◽  
R Rehman ◽  
N Farooqui ◽  
F Jehan ◽  
S H Abidi
Keyword(s):  
Oxidative Stress ◽  
Sample Size ◽  
Granulosa Cell ◽  
Cell Cultures ◽  
Granulosa Cells ◽  
Metformin Treatment ◽  
Serum Samples ◽  
Human Granulosa Cells ◽  
Human Granulosa Cell ◽  
Infertile Patients

Abstract Study question What is the effect of administration of Metformin on the oxidative stress (OS) levels in serum and primary human granulosa cell cultures of infertile females? Summary answer Metformin suppresses oxidative stress in serum and human granulosa cells and increases the expression of SIRT1 in OS induced environment. What is known already Oxidative stress (OS) is a resultant of mitochondrial dysfunction when it either fails to fight against the oxidants or the expression of the antioxidants is not sufficient. Cellular damage including DNA damage is a common resultant of oxidative stress. OS effects the oocyte maturation and moreover, the cleavage phase in the early embryonic stage. The raised levels of OS makers are hypothesized to compromise the nuclear maturation and the mitotic spindles of the maturing oocytes. Metformin seemed to decrease oxidative stress and improve insulin resistance, dyslipidaemia and endothelial dysfunction in PCOS patients Study design, size, duration This cross-sectional study was conducted from August 2017 – July 2019, at Aga Khan Hospital in collaboration with Australian Concept Infertility Medical Centre (ACIMC) on ten infertile patients undergoing egg retrieval after ethical approval from of Aga Khan Hospital (AKU-ERC–2018–0557–601). Participants/materials, setting, methods Serum samples were obtained and analysed. Follicular fluid of these subjects was collected for establishment of primary cell culture model of normal human granulosa cells (hGCs). Serum and hGC cultures were grouped as; a) control: treatment, b) Test1: H2O2 induced OS, and c) Test2: H2O2 induced OS treated with metformin. OS was estimated in all groups by Mishra method. The two Test groups were assessed for SIRT1 levels using quantitative PCR employing SIRT1 specific primers Main results and the role of chance With mean age of 32.04 ± 2.29 years the mean BMI was 27.61 ± 2.15 kg/m2. OS was induced and measured by an increase in optical density (OD) in hGC Test samples which showed 0.28 (0.16–0.40) OD when compared with control hGC samples 0.153 (0.09–0.23). There was a significant reduction in ODs after metformin treatment in the stress induced cells 0.182 (0.05–0.30). A similar pattern was observed in the serum samples in ODs; control: 0.105 (0.09–0.15), stress induced samples: 0.199 (0.19–0.20). and stress induced serum sample with metformin treatment: 0.1415 (0.06–0.18). The Ct values obtained to express the effect of metformin on SIRT1 levels, for OS induced (Test1) and OS induced metformin treated (Test2) cells were found to be 29.12 and 26.42, respectively. We also observed a significant (85%) difference in the fold change of SIRT1 expression between metformin treated and untreated cells. Limitations, reasons for caution Small sample size is the limitation of this study. The impact of metformin on cell cultures due to different causes of infertility could not be ascertained Wider implications of the findings: Metformin suppresses oxidative stress in serum and human granulosa cells and increases the expression of SIRT1 in OS induced environment, therefore, metformin may be considered as a treatment of oxidative stress in infertile patients. Randomized control trial with large sample size is recommended to confirm the cause and effect relationship. Trial registration number Not applicable

scholarly journals Conjugated linoleic acids attenuate FSH- and IGF1-stimulated cell proliferation; IGF1, GATA4, and aromatase expression; and estradiol-17β production in buffalo granulosa cells involving PPARγ, PTEN, and PI3K/Akt

Reproduction ◽  
2012 ◽  
Vol 144 (3) ◽  
pp. 373-383 ◽  
Author(s):  
Isha Sharma ◽  
Dheer Singh
Keyword(s):  
Cell Proliferation ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Cell Function ◽  
Endocrine System ◽  
Peroxisome Proliferator ◽  
Peroxisome Proliferator Activated Receptor ◽  
Beneficial Effects ◽  
Tensin Homolog ◽  
Estradiol 17Β

Conjugated linoleic acid (CLA) has drawn much interest in last two decades in the area ranging from anticancer activity to obesity. A number of research papers have been published recently with regard to CLA's additional biological functions as reproductive benefits. However, not much is known how this mixture of isomeric compounds mediates its beneficial effects particularly on fertility. In this study, we demonstrated the cross talk between downstream signaling of CLA and important hormone regulators of endocrine system, i.e. FSH and IGF1, on buffalo granulosa cell function (proliferation and steroidogenesis). Experiments were performed in primary serum-free buffalo granulosa cell culture, where cells were incubated with CLA in combination with FSH (25 ng/ml) and IGF1 (50 ng/ml). Results showed that 10 μM CLA inhibits FSH- and IGF1-induced granulosa cell proliferation; aromatase,GATA4, andIGF1mRNA; and estradiol-17β production. Western blot analysis of total cell lysates revealed that CLA intervenes the IGF1 signaling by decreasing p-Akt. In addition, CLA was found to upregulate peroxisome proliferator-activated receptor-gamma (PPARG) and phosphatase and tensin homolog (PTEN) level in granulosa cells. Further study using PPARG- and PTEN-specific inhibitors supports the potential role of CLA in granulosa cell proliferation and steroidogenesis involving PPARG, PTEN, and PI3K/Akt pathway.

scholarly journals Establishment of a Human Nonluteinized Granulosa Cell Line that Transitions from the Gonadotropin-Independent to the Gonadotropin-Dependent Status

Endocrinology ◽  
2012 ◽  
Vol 153 (6) ◽  
pp. 2851-2860 ◽  
Author(s):  
Bayasula ◽  
Akira Iwase ◽  
Tohru Kiyono ◽  
Sachiko Takikawa ◽  
Maki Goto ◽  
...  
Keyword(s):  
Cell Line ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Early Stage ◽  
Regulatory Protein ◽  
Cell Types ◽  
Mrna Levels ◽  
Steroidogenic Cell ◽  
Morphogenetic Protein ◽  
Steroidogenic Acute Regulatory

The ovary is a complex endocrine organ responsible for steroidogenesis and folliculogenesis. Follicles consist of oocytes and two primary steroidogenic cell types, the granulosa cells, and the theca cells. Immortalized human granulosa cells are essential for researching the mechanism of steroidogenesis and folliculogenesis. We obtained granulosa cells from a 35-yr-old female and immortalized them by lentivirus-mediated transfer of several genes so as to establish a human nonluteinized granulosa cell line (HGrC1). We subsequently characterized HGrC1 and investigated its steroidogenic performance. HGrC1 expressed enzymes related to steroidogenesis, such as steroidogenic acute regulatory protein, CYP11A, aromatase, and gonadotropin receptors. Stimulation with FSH increased the mRNA levels of aromatase, which consequently induced the aromatization of androstenedione to estradiol. Activin A increased the mRNA levels of the FSH receptor, which were synergistically up-regulated with FSH stimulation. HGrC1 also expressed a series of ligands and receptors belonging to the TGF-β superfamily. A Western blot analysis showed that bone morphogenetic protein (BMP)-4, BMP-6, and BMP-7 phosphorylated small mother against decapentaplegic (Smad)1/5/8, whereas growth differentiation factor-9 phosphorylated Smad2/3. BMP-15 and anti-Müllerian hormone phosphorylated Smad1/5/8 while also weakly phosphorylating Smad2/3. These results indicate that HGrC1 may possess the characteristics of granulosa cells belonging to follicles in the early stage. HGrC1 might also be capable of displaying the growth transition from a gonadotropin-independent status to gonadotropin-dependent one.

scholarly journals Studies of Human FSH Receptor Mutant K376A Which Does Not Bind APPL1 when Expressed in Human Granulosa Cell Line KGN.

2008 ◽  
Vol 78 (Suppl_1) ◽  
pp. 171-172 ◽  
Author(s):  
Richard M. Thomas ◽  
Cheryl A. Nechamen ◽  
Alfredo Ulloa-Aguirre ◽  
Joseph E. Mazurkiewicz ◽  
Kerri Kluetzman ◽  
...  
Keyword(s):  
Cell Line ◽  
Granulosa Cell ◽  
Fsh Receptor ◽  
Human Granulosa Cell ◽  
Receptor Mutant

scholarly journals Modulation of granulosa cell function via CRISPR-Cas fuelled editing of BMPR-IB gene in goats (Capra hircus)

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Sai Kumar ◽  
Meeti Punetha ◽  
Bosco Jose ◽  
Jaya Bharati ◽  
Shivani Khanna ◽  
...  
Keyword(s):  
Cell Viability ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Cell Function ◽  
Biological Effects ◽  
Capra Hircus ◽  
Gene Modulation ◽  
High Fecundity ◽  
Smad Signaling ◽  
Sheep Breeds

AbstractBMPs are multifunctional growth factors implicated in regulating the ovarian function as key intra-ovarian factors. Biological effects of BMPs are mediated through binding with membrane bound receptors like BMPR-IB and initiating downstream Smad signaling pathway. FecB mutation, regarded as a loss of function mutation in the BMPR-IB gene was identified in certain sheep breeds having high fecundity. Similar type of fecundity genes in goats have not been discovered so far. Hence, the current study was designed to investigate the effects of BMPR-IB gene modulation on granulosa cell function in goats. The BMPR-IB gene was knocked out using CRISPR-Cas technology in granulosa cells and cultured in vitro with BMP-4 stimulation for three different durations In addition, the FecB mutation was introduced in the BMPR-IB gene applying Easi-CRISPR followed by BMP-4/7 stimulation for 72 h. Steroidogenesis and cell viability were studied to explore the granulosa cell function on BMPR-IB gene modulation. BMPRs were found to be expressed stage specifically in granulosa cells of goats. Higher transcriptional abundance of R-Smads, LHR and FSHR indicating sensitisation of Smad signaling and increased gonadotropin sensitivity along with a significant reduction in the cell proliferation and viability was observed in granulosa cells upon BMPR-IB modulation. The inhibitory action of BMP-4/7 on P4 secretion was abolished in both KO and KI cells. Altogether, the study has revealed an altered Smad signaling, steroidogenesis and cell viability upon modulation of BMPR-IB gene in granulosa cells similar to that are documented in sheep breeds carrying the FecB mutation.

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