scholarly journals Human Chorionic Gonadotropin-Dependent Up-Regulation of Genes Responsible for Estrogen Sulfoconjugation and Export in Granulosa Cells of Luteinizing Preovulatory Follicles

Endocrinology ◽  
2006 ◽  
Vol 147 (9) ◽  
pp. 4222-4233 ◽  
Author(s):  
Kristy A. Brown ◽  
Monique Doré ◽  
Jacques G. Lussier ◽  
Jean Sirois
Keyword(s):  
Chorionic Gonadotropin ◽  
Granulosa Cells ◽  
Human Chorionic Gonadotropin ◽  
Cell Layer ◽  
Rt Pcr ◽  
Expression Of Genes ◽  
Preovulatory Follicles ◽  
Steady State Levels ◽  
17Β Estradiol ◽  
Theca Interna

Estrogen sulfotransferase (EST) is responsible for the sulfoconjugation of estrogens, thereby changing their physical properties and preventing their action via the estrogen receptors. These sulfoconjugated steroids no longer diffuse freely across the lipid bilayer; instead, they are exported by members of the ATP-binding cassette family, such as ABCC1. The objective of this study was to investigate the regulation of EST and ABCC1 during human chorionic gonadotropin (hCG)-induced ovulation/luteinization. The transcripts for EST and ABCC1 were cloned by RT-PCR, and the regulation of their mRNAs was studied in preovulatory follicles obtained during estrus at 0, 12, 24, 30, 33, 36, and 39 h after hCG. Results obtained from RT-PCR/Southern blot analyses showed significant changes in steady-state levels of both EST and ABCC1 mRNA after hCG treatment (P < 0.05). In granulosa cells, a significant increase in EST transcript was observed 30–39 h after hCG. Similarly, ABCC1 transcript levels were induced in granulosa cells 12–39 h after hCG. In contrast, no significant changes in either EST or ABCC1 were detected in theca interna samples after hCG. The increase in EST and ABCC1 transcripts observed in granulosa cells was reflected in preparations of intact follicle walls, suggesting that the granulosa cell layer contributes the majority of EST and ABCC1 expression in preovulatory follicles. The present study demonstrates that follicular luteinization is accompanied not only by a decrease in 17β-estradiol biosynthesis but also by an increase in expression of genes responsible for estrogen inactivation and elimination from granulosa cells, such as EST and ABCC1, respectively.

scholarly journals Human Chorionic Gonadotropin-Dependent Regulation of 17β-Hydroxysteroid Dehydrogenase Type 4 in Preovulatory Follicles and Its Potential Role in Follicular Luteinization

Endocrinology ◽  
2004 ◽  
Vol 145 (4) ◽  
pp. 1906-1915 ◽  
Author(s):  
Kristy A. Brown ◽  
Derek Boerboom ◽  
Nadine Bouchard ◽  
Monique Doré ◽  
Jacques G. Lussier ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Chorionic Gonadotropin ◽  
Human Chorionic Gonadotropin ◽  
Hydroxysteroid Dehydrogenase ◽  
Mrna Induction ◽  
Multidomain Structure ◽  
Acid Protein ◽  
Preovulatory Follicles ◽  
17Β Estradiol ◽  
Theca Interna

Abstract 17β-Hydroxysteroid dehydrogenase type 4 (17βHSD4) has a unique multidomain structure, with one domain involved in 17β-estradiol inactivation. The objective of the study was to investigate the regulation of 17βHSD4 during human chorionic gonadotropin (hCG)-induced ovulation/luteinization. The equine 17βHSD4 cDNA was cloned and was shown to encode a 735-amino acid protein that is highly conserved (81–87% identity) compared with other mammalian orthologs. RT-PCR/Southern blot analyses were performed to study the regulation of 17βHSD4 transcripts in equine preovulatory follicles isolated between 0–39 h after hCG treatment. Results showed the presence of basal 17βHSD4 mRNA expression before hCG treatment, but an increase was observed in follicles obtained 24 h after hCG (P < 0.05). Analyses of isolated preparations of granulosa and theca interna cells identified basal mRNA expression in both layers, but granulosa cells appeared as the predominant site of follicular 17βHSD4 mRNA induction. A specific polyclonal antibody was raised against a fragment of the equine protein and used to study regulation of the 17βHSD4 protein. Immunoblots showed an increase in full-length 17βHSD4 protein in follicles 24 h after hCG (P < 0.05), in keeping with mRNA results. Immunohistochemical data confirmed the induction of the enzyme in follicular cells after hCG treatment. Collectively, these results demonstrate that the gonadotropin-dependent induction of follicular luteinization is accompanied by an increase in 17βHSD4 expression. Considering the estrogen-inactivating function of 17βHSD4, its regulated expression in luteinizing preovulatory follicles appears as a potential complementary mechanism to reduce circulating levels of 17β-estradiol after the LH surge.

scholarly journals Regulation of transcripts encoding ADAMTS-1 (a disintegrin and metalloproteinase with thrombospondin-like motifs-1) and progesterone receptor by human chorionic gonadotropin in equine preovulatory follicles

2003 ◽  
Vol 31 (3) ◽  
pp. 473-485 ◽  
Author(s):  
D Boerboom ◽  
DL Russell ◽  
JS Richards ◽  
J Sirois
Keyword(s):  
Progesterone Receptor ◽  
Chorionic Gonadotropin ◽  
Granulosa Cells ◽  
Human Chorionic Gonadotropin ◽  
Reproductive Tract ◽  
Pcr Analysis ◽  
Preovulatory Follicles ◽  
Theca Interna ◽  
A Disintegrin And Metalloproteinase ◽  
Ovulatory Process

One member of a new family of metalloproteinases, a disintegrin and metalloproteinase with thrombospondin-like motifs-1 (ADAMTS-1), has been found to be expressed and hormonally induced in granulosa cells of ovulating rodent follicles. Furthermore, the targeted disruption of the ADAMTS-1 gene resulted in ovarian defects associated with severely impaired fertility. While these data demonstrate the importance of ADAMTS-1 in rodent ovarian physiology, the potential role of ADAMTS-1 in the ovulatory process of monoovulatory species remains unknown. The objectives of this study were to clone the equine ADAMTS-1 primary transcript and to study its regulation during human chorionic gonadotropin (hCG)-induced ovulation. A 3573 bp follicular cDNA library clone was isolated and found to encode a nearly complete, highly conserved ADAMTS-1 homologue. Real-time RT-PCR analysis detected this transcript in diverse tIssues, including previously unreported sites of ADAMTS-1 expression such as the male reproductive tract, the follicular theca interna and the mature corpus luteum. The tIssue distribution of the progesterone receptor (PR), a known regulator of ADAMTS-1 expression in rodent preovulatory follicles, was found to overlap that of ADAMTS-1 in some tIssues. A study of the regulation of follicular ADAMTS-1 and PR mRNAs during the hCG-induced ovulatory process revealed distinct patterns of regulation in granulosa cells and in theca interna. In granulosa cells, ADAMTS-1 mRNA was found to be induced at 12 h post-hCG (P<0.05), followed by a return to basal levels by 30 h and a re-increase at 33-39 h (P<0.05). A concomitant increase in PR mRNA (P<0.05) was observed at 12 h post-hCG. In theca interna, abundant ADAMTS-1 mRNA was detected at all timepoints, and levels increased transiently at 33 h post-hCG (P<0.05), whereas no significant change was observed in PR mRNA. Together, these data demonstrate for the first time the hormonally regulated ovarian expression of ADAMTS-1 in a monoovulatory species, and identify a novel biphasic regulation of ADAMTS-1 in granulosa cells and a regulated expression in theca interna that were not previously observed in rodents.

Expression and function of brain-derived neurotrophin factor and its receptor, TrkB, in ovarian follicles from the domestic hen (Gallus gallus domesticus)

2001 ◽  
Vol 204 (12) ◽  
pp. 2087-2095 ◽  
Author(s):  
T. Jensen ◽  
A. L. Johnson
Keyword(s):  
Granulosa Cell ◽  
Granulosa Cells ◽  
Cell Layer ◽  
Preliminary Evidence ◽  
Neurotrophin Receptor ◽  
Bdnf Mrna ◽  
Preovulatory Follicles ◽  
Layer Increase ◽  
Theca Interna ◽  
And Function

SUMMARY This report summarizes patterns of mRNA expression for the brain-derived neurotrophic factor (BDNF) together with its high-affinity neurotrophin receptor trkB within the hen ovary during follicle development, describes hormonal mechanisms for the regulation of trkB gene expression and provides preliminary evidence for a novel function for BDNF-mediated TrkB signaling within the granulosa layer. Levels of BDNF mRNA in the thecal layer and of trkB mRNA within the granulosa cell layer increase coincident with entrance of the follicle into the preovulatory hierarchy. Localization of the BDNF mRNA transcript correlates with expression of BDNF protein within the theca interna of preovulatory follicles, while localization of trkB mRNA and protein occurs extensively within the granulosa cell layer of preovulatory follicles. This pattern of expression suggests a paracrine relationship between theca and granulosa cells for BDNF signaling via TrkB. Vasoactive intestinal peptide and gonadotropin treatments stimulate increases in levels of trkB mRNA within cultured granulosa cells derived from both prehierarchal and preovulatory follicles, and this response is increased by co-treatment with 3-isobutyl-1-methylxanthine. Finally, BDNF treatment of cultured granulosa cells from preovulatory follicles results in a modest, but significant, reduction in basal progesterone production, whereas this effect was reversed by k252a, an inhibitor of Trk kinase activity. These results support the proposals that BDNF functions as a paracrine signal in hen granulosa cells and that its physiological functions may include the modulation of steroidogenesis.

scholarly journals Induction of Hyaluronan Synthase 2 by Human Chorionic Gonadotropin in Mural Granulosa Cells of Equine Preovulatory Follicles

Endocrinology ◽  
2002 ◽  
Vol 143 (11) ◽  
pp. 4375-4384 ◽  
Author(s):  
Angelika E. Stock ◽  
Nadine Bouchard ◽  
Kristy Brown ◽  
Andrew P. Spicer ◽  
Charles B. Underhill ◽  
...  
Keyword(s):  
Chorionic Gonadotropin ◽  
Granulosa Cells ◽  
Human Chorionic Gonadotropin ◽  
Hyaluronan Synthase ◽  
Preovulatory Follicles ◽  
Hyaluronan Synthase 2

scholarly journals SIRT3 Positively Regulates the Expression of Folliculogenesis- and Luteinization-Related Genes and Progesterone Secretion by Manipulating Oxidative Stress in Human Luteinized Granulosa Cells

Endocrinology ◽  
2014 ◽  
Vol 155 (8) ◽  
pp. 3079-3087 ◽  
Author(s):  
Houju Fu ◽  
Osamu Wada-Hiraike ◽  
Mana Hirano ◽  
Yumiko Kawamura ◽  
Ayako Sakurabashi ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Chorionic Gonadotropin ◽  
Granulosa Cells ◽  
Human Chorionic Gonadotropin ◽  
Hydroxysteroid Dehydrogenase ◽  
Human Reproduction ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Progesterone Secretion ◽  
Sirt3 Expression

SIRT3 is a member of the sirtuin family and has recently emerged as a vital molecule in controlling the generation of reactive oxygen species (ROS) in oocytes. Appropriate levels of ROS play pivotal roles in human reproductive medicine. The aim of the present study was to investigate SIRT3 expression and analyze the SIRT3-mediated oxidative response in human luteinized granulosa cells (GCs). Human ovarian tissues were subjected to immunohistochemical analysis to localize SIRT3 expression. Hydrogen peroxide and human chorionic gonadotropin were used to analyze the relationship between ROS and SIRT3 by quantitative RT-PCR and Western blotting. Intracellular levels of ROS were investigated by fluorescence after small interfering RNA–mediated knockdown of SIRT3 in human GCs. To uncover the role of SIRT3 in folliculogenesis and luteinization, mRNA levels of related genes and the progesterone concentration were analyzed by quantitative RT-PCR and immunoassays, respectively. We detected the expression of SIRT3 in the GCs of the human ovary. The mRNA levels of SIRT3, catalase, and superoxide dismutase 1 were up-regulated by hydrogen peroxide in both COV434 cells and human GCs and down-regulated by human chorionic gonadotropin. Knockdown of SIRT3 markedly elevated ROS generation in human GCs. In addition, SIRT3 depletion resulted in decreased mRNA expression of aromatase, 17β-hydroxysteroid dehydrogenase 1, steroidogenic acute regulatory protein, cholesterol side-chain cleavage enzyme, and 3β-hydroxysteroid dehydrogenase in GCs and thus resulted in decreased progesterone secretion. These results have the important clinical implication that SIRT3 might play a positive role in the folliculogenesis and luteinization processes in GCs, possibly by sensing and regulating the generation of ROS. Activation of SIRT3 function might help to sustain human reproduction by maintaining GCs as well as oocytes.

scholarly journals Human chorionic gonadotropin-dependent induction of an equine aldo-keto reductase (AKR1C23) with 20α-hydroxysteroid dehydrogenase activity during follicular luteinization in vivo

2006 ◽  
Vol 36 (3) ◽  
pp. 449-461 ◽  
Author(s):  
K A Brown ◽  
D Boerboom ◽  
N Bouchard ◽  
M Doré ◽  
J G Lussier ◽  
...  
Keyword(s):  
Chorionic Gonadotropin ◽  
Human Chorionic Gonadotropin ◽  
Cell Types ◽  
Hydroxysteroid Dehydrogenase ◽  
Biochemical Basis ◽  
Mrna Induction ◽  
Cell Extracts ◽  
Preovulatory Follicles ◽  
Theca Interna

Aldo–keto reductases (AKRs) are multifunctional enzymes capable of acting on a wide variety of substrates, including sex steroids. AKRs having 20α-hydroxysteroid dehydrogenase (20α-HSD) activity can reduce progesterone to 20α-hydroxy-4-pregnen-3-one (20α-DHP), a metabolite with lower affinity for the progesterone receptor. The objective of this study was to investigate the regulation of equine AKR1C23 during human chorionic gonadotropin (hCG)-induced ovulation/luteinization. The equine AKR1C23 cDNA was cloned and shown to encode a 322 amino acid protein that is conserved (71–81% identity) when compared with mammalian orthologs. RT-PCR/Southern blotting analyses were performed to study the regulation of AKR1C23 transcripts in equine preovulatory follicles isolated between 0 and 39 h after hCG treatment (ovulation occurring 39–42 h post-hCG). Results showed the presence of low AKR1C23 expression before hCG treatment, but a marked increase was observed in follicles obtained 12 h after hCG (P<0.05). Analyses of isolated preparations of granulosa and theca interna cells identified low mRNA expression in both cell types prior to hCG treatment, with granulosa cells clearly being the predominant site of follicular AKR1C23 mRNA induction. A specific polyclonal antibody was raised against a fragment of the equine protein and immunoblotting analyses showed an increase in AKR1C23 protein in granulosa cell extracts when comparing follicles isolated at 36 h post-hCG vs those collected prior to treatment, in keeping with mRNA results. Immunohistochemical data confirmed the induction of the enzyme in follicular cells after hCG treatment. The enzyme was tested for 20α-HSD activity and was shown to exhibit a KM of 3.12 μM, and a Vmax of 0.86 pmol/min per 10 μg protein towards progesterone. The levels of 20α-DHP measured in follicular fluid reflected this activity. Collectively, these results demonstrate for the first time that the gonadotropin-dependent induction of follicular luteinization is accompanied by an increase in AKR1C23 expression. Considering the 20α-HSD activity of AKR1C23, its regulated expression in luteinizing preovulatory follicles may provide a biochemical basis for the increase in ovarian 20α-DHP observed during gonadotropin-induced luteinization/ovulation. (The nucleotide sequence reported in this paper has been submitted to GenBank with accession number AY955082.)

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.

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