A comparison between oocyte growth in coculture with granulosa cells and oocytes with granulosa cell-oocyte junctional contact maintained in vitro

1979 ◽  
Vol 209 (2) ◽  
pp. 345-353 ◽  
Author(s):  
John J. Eppig
Keyword(s):  
Granulosa Cell ◽  
Granulosa Cells ◽  
Oocyte Growth

Addition of granulosa cell mass to the culture medium of oocytes derived from early antral follicles increases oocyte growth, ATP content, and acetylation of H4K12

Zygote ◽  
2016 ◽  
Vol 24 (6) ◽  
pp. 848-856 ◽  
Author(s):  
Miyako Sugiyama ◽  
Mei Sumiya ◽  
Koumei Shirasuna ◽  
Takehito Kuwayama ◽  
Hisataka Iwata
Keyword(s):  
Granulosa Cell ◽  
Granulosa Cells ◽  
Culture Medium ◽  
Cell Mass ◽  
Fertilization Rate ◽  
Atp Content ◽  
Oocyte Growth ◽  
Acetylation Level ◽  
Antral Follicles

SummaryThe main aim of the present study was to examine the hypothesis that an increase in the number of granulosa cells surrounding developing bovine oocytes results in both high ATP levels and an increase in the acetylation level of H4K12 in oocytes grown in vitro. Oocyte–granulosa cell complexes (OGCs) were collected from early antral follicles (EAFs, 0.4–0.7 mm in diameter), and individually cultured on 96-well plates with or without additional granulosa cell mass that had been prepared from other OGCs. After 16 days of culture, we examined: (i) the rate of antrum formation of the OGCs; (ii) the diameter, maturation, and fertilization rate of the oocytes; and (iii) the ATP content and acetylation level of H4K12 in the oocytes grown in vitro. Granulosa cell mass added to the culture medium contributed to the development of OGCs with a higher rate of antrum formation and oocyte growth. Furthermore, the addition of granulosa cells increased the ATP content and acetylation level of H4K12 in oocytes grown in vitro compared with those developed without addition of granulosa cells. In addition, there was a positive correlation between the ATP content in oocytes grown in vitro and the number of granulosa cells in the corresponding OGCs. The results suggest that granulosa cells play a role not only in the development of OGCs and the growth of oocytes, but also in the determination of ATP content and the acetylation of H4K12 in the oocytes developed in vitro.

scholarly journals R-spondin2 signaling is required for oocyte-driven intercellular communication and follicular growth

2020 ◽  
Vol 27 (10) ◽  
pp. 2856-2871 ◽  
Author(s):  
Marie-Cécile De Cian ◽  
Elodie P. Gregoire ◽  
Morgane Le Rolle ◽  
Simon Lachambre ◽  
Magali Mondin ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Cell Cycle Progression ◽  
Ovarian Function ◽  
Follicular Growth ◽  
Oocyte Growth

Abstract R-spondin2 (RSPO2) is a member of the R-spondin family, which are secreted activators of the WNT/β-catenin (CTNNB1) signaling pathway. In the mouse postnatal ovary, WNT/CTNNB1 signaling is active in the oocyte and in the neighboring supporting cells, the granulosa cells. Although the role of Rspo2 has been previously studied using in vitro experiments, the results are conflicting and the in vivo ovarian function of Rspo2 remains unclear. In the present study, we found that RSPO2/Rspo2 expression is restricted to the oocyte of developing follicles in both human and mouse ovaries from the beginning of the follicular growth. In mice, genetic deletion of Rspo2 does not impair oocyte growth, but instead prevents cell cycle progression of neighboring granulosa cells, thus resulting in an arrest of follicular growth. We further show this cell cycle arrest to be independent of growth promoting GDF9 signaling, but rather associated with a downregulation of WNT/CTNNB1 signaling in granulosa cells. To confirm the contribution of WNT/CTNNB1 signaling in granulosa cell proliferation, we induced cell type specific deletion of Ctnnb1 postnatally. Strikingly, follicles lacking Ctnnb1 failed to develop beyond the primary stage. These results show that RSPO2 acts in a paracrine manner to sustain granulosa cell proliferation in early developing follicles. Taken together, our data demonstrate that the activation of WNT/CTNNB1 signaling by RSPO2 is essential for oocyte-granulosa cell interactions that drive maturation of the ovarian follicles and eventually female fertility.

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.

scholarly journals AKT is involved in granulosa cell autophagy regulation via mTOR signaling during rat follicular development and atresia

Reproduction ◽  
2014 ◽  
Vol 147 (1) ◽  
pp. 73-80 ◽  
Author(s):  
JongYeob Choi ◽  
MinWha Jo ◽  
EunYoung Lee ◽  
DooSeok Choi
Keyword(s):  
Cell Death ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Apoptotic Cell ◽  
Follicular Development ◽  
Negative Regulator ◽  
Metabolic Clearance ◽  
Akt Inhibitor ◽  
Western Blots

In this study, we examined whether granulosa cell autophagy during follicular development and atresia was regulated by the class I phosphoinositide-3 kinase/protein kinase B (AKT) pathway, which is known to control the activity of mammalian target of rapamycin (mTOR), a major negative regulator of autophagy. Ovaries and granulosa cells were obtained using an established gonadotropin-primed immature rat model that induces follicular development and atresia. Autophagy was evaluated by measuring the expression level of microtubule-associated protein light chain 3-II (LC3-II) using western blots and immunohistochemistry. The activity of AKT and mTOR was also examined by observing the phosphorylation of AKT and ribosomal protein S6 kinase (S6K) respectively. After gonadotropin injection, LC3-II expression was suppressed and phosphorylation of AKT and S6K increased in rat granulosa cells. By contrast, gonadotropin withdrawal by metabolic clearance promoted LC3-II expression and decreased phosphorylation of AKT and S6K. In addition,in-vitroFSH treatment of rat granulosa cells also indicated inhibition of LC3-II expression accompanied by a marked increase in phosphorylation of AKT and S6K. Inhibition of AKT phosphorylation using AKT inhibitor VIII suppressed FSH-mediated phosphorylation of S6K, followed by an increase in LC3-II expression. Furthermore, co-treatment with FSH and AKT inhibitor increased the levels of apoptosis and cell death of granulosa cells compared with the single treatment with FSH. Taken together, our findings indicated that AKT-mediated activation of mTOR suppresses granulosa cell autophagy during follicular development and is involved in the regulation of apoptotic cell death.

scholarly journals MiRNA-143 mediates the proliferative signaling pathway of FSH and regulates estradiol production

2017 ◽  
Vol 234 (1) ◽  
pp. 1-14 ◽  
Author(s):  
Li Zhang ◽  
XiaoXin Zhang ◽  
Xuejing Zhang ◽  
Yu Lu ◽  
Lei Li ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Loss Of Function ◽  
Cellular Processes ◽  
Cell Functions ◽  
Genes Expression ◽  
Underlying Mechanisms

MicroRNAs (MiRNAs) play important regulatory roles in many cellular processes. MiR-143 is highly enriched in the mouse ovary, but its roles and underlying mechanisms are not well understood. In the current study, we show that miR-143 is located in granulosa cells of primary, secondary and antral follicles. To explore the specific functions of miR-143, we transfected miR-143 inhibitor into primary cultured granulosa cells to study the loss of function of miR-143 and the results showed that miR-143 silencing significantly increased estradiol production and steroidogenesis-related gene expression. Moreover, our in vivo and in vitro studies showed that follicular stimulating hormone (FSH) significantly decreased miR-143 expression. This function of miR-143 is accomplished by its binding to the 3’-UTR of KRAS mRNA. Furthermore, our results demonstrated that miR-143 acts as a negative regulating molecule mediating the signaling pathway of FSH and affecting estradiol production by targeting KRAS. MiR-143 also negatively acts in regulating granulosa cells proliferation and cell cycle-related genes expression. These findings indicate that miR-143 plays vital roles in FSH-induced estradiol production and granulosa cell proliferation, providing a novel mechanism that involves miRNA in regulating granulosa cell functions.

scholarly journals Inhibition of oocyte growth factors in vivo modulates ovarian folliculogenesis in neonatal and immature mice

Reproduction ◽  
2010 ◽  
Vol 139 (3) ◽  
pp. 587-598 ◽  
Author(s):  
Samu Myllymaa ◽  
Arja Pasternack ◽  
David G Mottershead ◽  
Matti Poutanen ◽  
Minna M Pulkki ◽  
...  
Keyword(s):  
Growth Factors ◽  
Granulosa Cells ◽  
Microscopic Analysis ◽  
Corpora Lutea ◽  
Electron Microscopic ◽  
Oocyte Growth ◽  
Long Term Study ◽  
Ovarian Folliculogenesis

Growth differentiation factor-9 (GDF9) and bone morphogenetic protein-15 (BMP15) are among the key regulators transmitting the signaling between the oocyte and the surrounding granulosa cells. Previously, it has been shown that a recombinant BMP type II receptor ectodomain–Fc fusion protein (BMPR2ecd–Fc) is able to inhibit the actions of GDF9 and BMP15 in vitro. Here, we have produced bioactive BMPR2ecd–Fc, which was injected i.p. into neonatal mice. Early folliculogenesis was first studied by injecting mice five times with various doses of BMPR2ecd–Fc during the postnatal days 4–12. Folliculogenesis was affected dose dependently, as evidenced by a decreased mitogenesis of granulosa cells of the growing follicles. Furthermore, we also noticed a decrease in the number of secondary and tertiary follicles as well as an increase in the oocyte size. Electron microscopic analysis revealed that the ultrastructure of the granulosa cells of the primary follicles was not affected by the BMPR2ecd–Fc treatment. A second study was conducted to investigate whether a longer treatment with 12 injections during postnatal days 4–28 would inhibit folliculogenesis. Similar effects were observed in the two studies on the early follicular developmental stages. However, in the long-term study, later stages of folliculogenesis were not blocked but rather increased numbers of antral follicles, preovulatory follicles, and corpora lutea were found. We conclude that BMPR2ecd–Fc is a potent modulator of ovarian folliculogenesis in vivo, and thus, is a valuable tool for studying the physiology and downstream effects of oocyte-derived growth factors in vivo.

scholarly journals SMAD7 antagonizes key TGFβ superfamily signaling in mouse granulosa cells in vitro

Reproduction ◽  
2013 ◽  
Vol 146 (1) ◽  
pp. 1-11 ◽  
Author(s):  
Yang Gao ◽  
Haixia Wen ◽  
Chao Wang ◽  
Qinglei Li
Keyword(s):  
Granulosa Cell ◽  
Granulosa Cells ◽  
Transforming Growth Factor ◽  
Negative Regulator ◽  
Fine Tuning ◽  
Female Reproduction ◽  
Tgfβ Signaling ◽  
Tgfβ Superfamily

Transforming growth factor β (TGFβ) superfamily signaling is essential for female reproduction. Dysregulation of the TGFβ signaling pathway can cause reproductive diseases. SMA and MAD (mothers against decapentaplegic) (SMAD) proteins are downstream signaling transducers of the TGFβ superfamily. SMAD7 is an inhibitory SMAD that regulates TGFβ signalingin vitro. However, the function of SMAD7 in the ovary remains poorly defined. To determine the signaling preference and potential role of SMAD7 in the ovary, we herein examined the expression, regulation, and function of SMAD7 in mouse granulosa cells. We showed that SMAD7 was expressed in granulosa cells and subject to regulation by intraovarian growth factors from the TGFβ superfamily. TGFB1 (TGFβ1), bone morphogenetic protein 4, and oocyte-derived growth differentiation factor 9 (GDF9) were capable of inducingSmad7expression, suggesting a modulatory role of SMAD7 in a negative feedback loop. Using a small interfering RNA approach, we further demonstrated that SMAD7 was a negative regulator of TGFB1. Moreover, we revealed a link between SMAD7 and GDF9-mediated oocyte paracrine signaling, an essential component of oocyte–granulosa cell communication and folliculogenesis. Collectively, our results suggest that SMAD7 may function during follicular development via preferentially antagonizing and/or fine-tuning essential TGFβ superfamily signaling, which is involved in the regulation of oocyte–somatic cell interaction and granulosa cell function.

Prolactin, LH, and estradiol-17β in utilization of lipoprotein substrate by porcine granulosa cells in vitro

1988 ◽  
Vol 66 (5) ◽  
pp. 561-566 ◽  
Author(s):  
K. Rajkumar ◽  
P. Klingshorn ◽  
P. J. Chedrese ◽  
B. D. Murphy
Keyword(s):  
Granulosa Cell ◽  
Cell Cultures ◽  
Granulosa Cells ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Low Density ◽  
Porcine Granulosa Cells ◽  
Progesterone Synthesis ◽  
Serum Free Conditions

Porcine granulosa cells cultured under serum free conditions responded by increased progesterone secretion to the addition of the leuteotropic hormones, LH, prolactin, and estradiol. Provision of extracellular substrate for steroidogenesis in the form of porcine high density lipoprotein or low density lipoprotein enhanced progesterone accumulation by granulosa cell cultures. Estradiol, LH, and prolactin all greatly increased progesterone accumulation in the presence of either high or low density lipoproteins. Increases in progesterone accumulation following addition of prolactin or LH in combination with estradiol suggested the presence of a synergistic interaction among leuteotropins. Pre-exposure of granulosa cell cultures to estradiol increased the subsequent stimulatory effect of prolactin on lipoprotein utilization. It is concluded that all three leuteotropins function to enhance and may interact in the utilization of extracellular lipoprotein substrate for progesterone synthesis.

Effect of in vitro copper supplementation on granulosa cell estradiol synthesis and associated genes

2018 ◽  
Author(s):  
Ravi, P.S.P. Gupta, S. Nandi, S. Mondal, Kumar Soni­ ◽  
P.S.P. Gupta ◽  
S. Nandi ◽  
S. Mondal, J.R. Ippala, Avantika Mor, A Mondal ◽  
J.R. Ippala ◽  
...  
Keyword(s):  
Cell Survival ◽  
Mrna Expression ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Culture Medium ◽  
Estrus Synchronization ◽  
Ovarian Granulosa Cells ◽  
Effect Of Copper ◽  
Estradiol Synthesis

The study was conducted by supplementing cupric chloride dihydrate to modulate the estradiol synthesis in granulosa cells with a hypothesis of possible use of copper to potentiate or partially replace the hormones for estrus induction / estrus synchronization in future studies. In present study copper at three doses (0.1, 0.5 and 1 mM level in culture medium) were tested to deserve see their effects on in vitro granulosa cell survival, estradiol synthesis and their associated genes of ovarian granulosa cells of goat.There was no effect of copper on the ovarian granulosa cell survival rate. There was a considerable increase in the estradiol level per ml culture medium basis by 6th day of in vitro culture with the second dose of copper i.e. 0.5 mM, but the increase was non-significant (P greator than 0.05). There was no significant effect of copper on estradiol synthesis when expressed on per 30000 cell basis. Effect of copper (0.1 mM and 0.5 mM) on the mRNA expression of genes of aromatase (CYP19A1) and cyclin D2 (CCND2) was estimated. Copper had significantly (P less than 0.05) increased the mRNA expression of CCND2 and CYP19A1in ovarian granulosa cells with only one of the two doses tested i.e. 0.5 mM. Hence, copper can be considered as a potential mineral to supplement along with hormones in estrus induction or estrus synchronization protocols to minimize the use of hormones.

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