scholarly journals Cellular heterogeneity of the LH receptor and its significance for cyclic GMP signaling in mouse preovulatory follicles

2020 ◽  
Author(s):  
Valentina Baena ◽  
Corie M. Owen ◽  
Tracy F. Uliasz ◽  
Katie M. Lowther ◽  
Siu-Pok Yee ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Cyclic Gmp ◽  
Cellular Localization ◽  
Cumulus Cells ◽  
Luteinizing Hormone Receptor ◽  
Cellular Heterogeneity ◽  
Signaling Proteins ◽  
Meiotic Arrest ◽  
Lh Receptor ◽  
Preovulatory Follicles

AbstractMeiotic arrest and resumption in mammalian oocytes are regulated by two opposing signaling proteins in the cells of the surrounding follicle: the guanylyl cyclase NPR2, and the luteinizing hormone receptor (LHR). NPR2 maintains a meiosis-inhibitory level of cyclic GMP (cGMP) until LHR signaling causes dephosphorylation of NPR2, reducing NPR2 activity, lowering cGMP to a level that releases meiotic arrest. However, the signaling pathway between LHR activation and NPR2 dephosphorylation remains incompletely understood, due in part to imprecise information about the cellular localization of these two proteins. To investigate their localization, we generated mouse lines in which HA epitope tags were added to the endogenous LHR and NPR2 proteins, and used immunofluorescence and immunogold microscopy to localize these proteins with high resolution. The results showed that the LHR protein is absent from the cumulus cells and inner mural granulosa cells, and is present in only 13-48% of the outer mural granulosa cells. In contrast, NPR2 is present throughout the follicle, and is more concentrated in the cumulus cells. Less than 20% of the NPR2 is in the same cells that express the LHR. These results suggest that to account for the LH-induced inactivation of NPR2, LHR-expressing cells send a signal that inactivates NPR2 in neighboring cells that do not express the LHR. An inhibitor of gap junction permeability attenuates the LH-induced cGMP decrease in the outer mural granulosa cells, consistent with this mechanism contributing to how NPR2 is inactivated in cells that do not express the LHR.

scholarly journals Cellular Heterogeneity of the Luteinizing Hormone Receptor and Its Significance for Cyclic GMP Signaling in Mouse Preovulatory Follicles

Endocrinology ◽  
2020 ◽  
Vol 161 (7) ◽  
Author(s):  
Valentina Baena ◽  
Corie M Owen ◽  
Tracy F Uliasz ◽  
Katie M Lowther ◽  
Siu-Pok Yee ◽  
...  
Keyword(s):  
Luteinizing Hormone ◽  
Granulosa Cells ◽  
Hormone Receptor ◽  
Cellular Localization ◽  
Cumulus Cells ◽  
Luteinizing Hormone Receptor ◽  
Cellular Heterogeneity ◽  
Signaling Proteins ◽  
Meiotic Arrest ◽  
Preovulatory Follicles

Abstract Meiotic arrest and resumption in mammalian oocytes are regulated by 2 opposing signaling proteins in the cells of the surrounding follicle: the guanylyl cyclase natriuretic peptide receptor 2 (NPR2), and the luteinizing hormone receptor (LHR). NPR2 maintains a meiosis-inhibitory level of cyclic guanosine 5′-monophosphate (cGMP) until LHR signaling causes dephosphorylation of NPR2, reducing NPR2 activity, lowering cGMP to a level that releases meiotic arrest. However, the signaling pathway between LHR activation and NPR2 dephosphorylation remains incompletely understood, due in part to imprecise information about the cellular localization of these 2 proteins. To investigate their localization, we generated mouse lines in which hemagglutinin epitope tags were added to the endogenous LHR and NPR2 proteins, and used immunofluorescence and immunogold microscopy to localize these proteins with high resolution. The results showed that the LHR protein is absent from the cumulus cells and inner mural granulosa cells, and is present in only 13% to 48% of the outer mural granulosa cells. In contrast, NPR2 is present throughout the follicle, and is more concentrated in the cumulus cells. Less than 20% of the NPR2 is in the same cells that express the LHR. These results suggest that to account for the LH-induced inactivation of NPR2, LHR-expressing cells send a signal that inactivates NPR2 in neighboring cells that do not express the LHR. An inhibitor of gap junction permeability attenuates the LH-induced cGMP decrease in the outer mural granulosa cells, consistent with this mechanism contributing to how NPR2 is inactivated in cells that do not express the LHR.

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 LH-Receptor Gene Expression in Human Granulosa and Cumulus Cells from Antral and Preovulatory Follicles

2012 ◽  
Vol 97 (8) ◽  
pp. E1524-E1531 ◽  
Author(s):  
Janni Vikkelsø Jeppesen ◽  
Stine Gry Kristensen ◽  
Maria Eilsø Nielsen ◽  
Peter Humaidan ◽  
Maria Dal Canto ◽  
...  
Keyword(s):  
Gene Expression ◽  
Receptor Gene ◽  
Cumulus Cells ◽  
Lh Receptor ◽  
Preovulatory Follicles ◽  
Receptor Gene Expression

scholarly journals Intercellular signaling via cyclic GMP diffusion through gap junctions restarts meiosis in mouse ovarian follicles

2015 ◽  
Vol 112 (17) ◽  
pp. 5527-5532 ◽  
Author(s):  
Leia C. Shuhaibar ◽  
Jeremy R. Egbert ◽  
Rachael P. Norris ◽  
Paul D. Lampe ◽  
Viacheslav O. Nikolaev ◽  
...  
Keyword(s):  
Gap Junctions ◽  
Granulosa Cells ◽  
Cyclic Gmp ◽  
Ovarian Follicle ◽  
General Mechanism ◽  
Intercellular Signaling ◽  
Meiotic Arrest ◽  
Physiological Signal ◽  
Direct Demonstration ◽  
Cell Layers

Meiosis in mammalian oocytes is paused until luteinizing hormone (LH) activates receptors in the mural granulosa cells of the ovarian follicle. Prior work has established the central role of cyclic GMP (cGMP) from the granulosa cells in maintaining meiotic arrest, but it is not clear how binding of LH to receptors that are located up to 10 cell layers away from the oocyte lowers oocyte cGMP and restarts meiosis. Here, by visualizing intercellular trafficking of cGMP in real-time in live follicles from mice expressing a FRET sensor, we show that diffusion of cGMP through gap junctions is responsible not only for maintaining meiotic arrest, but also for rapid transmission of the signal that reinitiates meiosis from the follicle surface to the oocyte. Before LH exposure, the cGMP concentration throughout the follicle is at a uniformly high level of ∼2–4 μM. Then, within 1 min of LH application, cGMP begins to decrease in the peripheral granulosa cells. As a consequence, cGMP from the oocyte diffuses into the sink provided by the large granulosa cell volume, such that by 20 min the cGMP concentration in the follicle is uniformly low, ∼100 nM. The decrease in cGMP in the oocyte relieves the inhibition of the meiotic cell cycle. This direct demonstration that a physiological signal initiated by a stimulus in one region of an intact tissue can travel across many layers of cells via cyclic nucleotide diffusion through gap junctions could provide a general mechanism for diverse cellular processes.

scholarly journals Ovulation Involves the Luteinizing Hormone-Dependent Activation of Gq/11 in Granulosa Cells

2013 ◽  
Vol 27 (9) ◽  
pp. 1483-1491 ◽  
Author(s):  
Shawn M. Breen ◽  
Nebojsa Andric ◽  
Tai Ping ◽  
Fang Xie ◽  
Stefan Offermans ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Target Genes ◽  
Inositol Phosphate ◽  
Conclusive Evidence ◽  
Corpora Lutea ◽  
Heterotrimeric G Proteins ◽  
Lh Receptor ◽  
Physiological Importance ◽  
Preovulatory Follicles ◽  
Inositol Phosphate Accumulation

The LH receptor (LHR) activates several families of heterotrimeric G proteins, but only the activation of Gs and subsequent generation of cAMP are universally accepted as important mediators of LH actions. To examine the involvement of the Gq/11 family on the actions of LH, we crossed Cyp19Cre and Gαqf/f;Gα11−/− mice to generate mice with a granulosa cell-specific deletion of Gαq in the context of a global deletion of Gα11. Granulosa cells from Gαqf/f;Gα11−/−;Cre+ mice have barely detectable levels of Gαq/11, have a normal complement of LHR, and respond to LHR activation with a transient increase in cAMP accumulation, but they fail to respond with increased inositol phosphate accumulation, an index of the activation of Gαq/11. The LHR-provoked resumption of meiosis, cumulus expansion, and luteinization are normal. However, the Gαqf/f;Gα11−/−;Cre+ mice display severe subfertility because many of the oocytes destined for ovulation become entrapped in preovulatory follicles or corpora lutea. Because follicular rupture is known to be dependent on the expression of the progesterone receptor (Pgr), we examined the LHR-induced expression of Pgr and 4 of its target genes (Adamts-1, Ctsl1, Edn2, and Prkg2). These actions of the LHR were impaired in the ovaries of the Gαqf/f;Gα11−/−;Cre+ mice. We conclude that the defect in follicular rupture is secondary to the failure of the LHR to fully induce the expression of the Pgr. This is the first conclusive evidence for the physiological importance of the activation of Gq/11 by the LHR and for the involvement of Gαq/11 in ovulation.

scholarly journals Luteinizing Hormone Receptor mRNA Down-Regulation Is Mediated through ERK-Dependent Induction of RNA Binding Protein

2011 ◽  
Vol 25 (2) ◽  
pp. 282-290 ◽  
Author(s):  
Bindu Menon ◽  
Megan Franzo-Romain ◽  
Shadi Damanpour ◽  
K. M. J. Menon
Keyword(s):  
Granulosa Cells ◽  
Rna Binding ◽  
Binding Protein ◽  
Primary Cultures ◽  
Binding Activity ◽  
Luteinizing Hormone Receptor ◽  
Down Regulation ◽  
Lh Receptor ◽  
Receptor Mrna ◽  
Mrna Binding

Abstract The ligand-induced down-regulation of LH receptor (LHR) expression in the ovaries, at least in part, is regulated by a posttranscriptional process mediated by a specific LH receptor mRNA binding protein (LRBP). The LH-mediated signaling pathways involved in this process were examined in primary cultures of human granulosa cells. Treatment with 10 IU human chorionic gonadotropin (hCG) for 12 h resulted in the down-regulation of LHR mRNA expression while producing an increase in LHR mRNA binding to LRBP as well as a 2-fold increase in LRBP levels. The activation of ERK1/2 pathway in LH-mediated LHR mRNA down-regulation was also established by demonstrating the translocation of ERK1/2 from the cytosol to the nucleus using confocal microcopy. Inhibition of protein kinase A using H-89 or ERK1/2 by U0126 abolished the LH-induced LHR mRNA down-regulation. These treatments also abrogated both the increases in LRBP levels as well as the LHR mRNA binding activity. The abolishment of the hCG-induced increase in LRBP levels and LHR mRNA binding activity was further confirmed by transfecting granulosa cells with ERK1/2 specific small interfering RNA. This treatment also reversed the hCG-induced down-regulation of LHR mRNA. These data show that LH-regulated ERK1/2 signaling is required for the LRBP-mediated down-regulation of LHR mRNA.

Molecular control of oocyte meiotic arrest and resumption

2013 ◽  
Vol 25 (3) ◽  
pp. 463 ◽  
Author(s):  
Lei Liu ◽  
Nana Kong ◽  
Guoliang Xia ◽  
Meijia Zhang
Keyword(s):  
Growth Factors ◽  
Granulosa Cells ◽  
Cumulus Cells ◽  
Follicle Cells ◽  
Egf Receptors ◽  
Meiotic Arrest ◽  
Molecular Control ◽  
Peptide Precursor ◽  
Camp Hydrolysis ◽  
Regulatory Functions

Mammalian oocytes within Graafian follicles are arrested at prophase I by factors from surrounding follicle cells, and resume meiosis after an LH surge from the pituitary. The maintenance of meiotic arrest requires high levels of cAMP, resulting from G-protein-coupled receptor (GPR) 3 and/or GPR12 activation of adenylyl cyclase within the oocyte. Recent studies indicate that natriuretic peptide precursor C (NPPC), acting via its cognate receptor NPR2, increases cGMP levels in granulosa cells; the cGMP then diffuses into oocytes and inhibits phosphodiesterase 3A activity and cAMP hydrolysis. Meiotic resumption is induced by LH via the generation of epidermal growth factor (EGF)-like growth factors in mural granulosa cells that activate EGF receptors in cumulus cells. However, the exact mechanisms underlying the actions of these growth factors on oocyte maturation are unclear. Herein we summarise the regulatory functions of NPPC and NPR2 in maintaining oocyte meiotic arrest and discuss the possibility that LH could stimulate meiotic resumption by decreasing NPPC content and NPR2 activity.

scholarly journals Cellular localization of cyclic AMP and cyclic GMP in rat bone marrow

1979 ◽  
Vol 29 ◽  
pp. 85
Author(s):  
Nobuyoshi Yoshida ◽  
Kohtaro Taniyama ◽  
Chikako Tanaka
Keyword(s):  
Bone Marrow ◽  
Cyclic Amp ◽  
Cyclic Gmp ◽  
Cellular Localization ◽  
Rat Bone

scholarly journals Human Granulosa Cells—Stemness Properties, Molecular Cross-Talk and Follicular Angiogenesis

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1396
Author(s):  
Claudia Dompe ◽  
Magdalena Kulus ◽  
Katarzyna Stefańska ◽  
Wiesława Kranc ◽  
Błażej Chermuła ◽  
...  
Keyword(s):  
Stem Cells ◽  
Granulosa Cells ◽  
Polycystic Ovary ◽  
Ovarian Follicle ◽  
Cumulus Cells ◽  
Ovary Syndrome ◽  
Different Types ◽  
Metabolite Exchange ◽  
Reproductive Techniques ◽  
New Strategies

The ovarian follicle is the basic functional unit of the ovary, comprising theca cells and granulosa cells (GCs). Two different types of GCs, mural GCs and cumulus cells (CCs), serve different functions during folliculogenesis. Mural GCs produce oestrogen during the follicular phase and progesterone after ovulation, while CCs surround the oocyte tightly and form the cumulus oophurus and corona radiata inner cell layer. CCs are also engaged in bi-directional metabolite exchange with the oocyte, as they form gap-junctions, which are crucial for both the oocyte’s proper maturation and GC proliferation. However, the function of both GCs and CCs is dependent on proper follicular angiogenesis. Aside from participating in complex molecular interplay with the oocyte, the ovarian follicular cells exhibit stem-like properties, characteristic of mesenchymal stem cells (MSCs). Both GCs and CCs remain under the influence of various miRNAs, and some of them may contribute to polycystic ovary syndrome (PCOS) or premature ovarian insufficiency (POI) occurrence. Considering increasing female fertility problems worldwide, it is of interest to develop new strategies enhancing assisted reproductive techniques. Therefore, it is important to carefully consider GCs as ovarian stem cells in terms of the cellular features and molecular pathways involved in their development and interactions as well as outline their possible application in translational medicine.

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