The role of bone morphogenetic proteins 2, 4, 6 and 7 during ovarian follicular development in sheep: contrast to rat

The intraovarian roles of BMP family members such as BMP2, 4, 6 and 7 are not well understood, particularly in species with low ovulation rates such as sheep. Therefore, the objectives of these experiments were to determine the expression patterns of mRNAs encoding BMP2, 4, 6 and 7 during ovarian follicular development in sheep, and to determine the effects of these growth factors on ovine granulosa cell functions in vitro. For comparative purposes, the effects of these BMPs were also determined in rat granulosa cells since these factors have been most widely studied in this poly-ovulatory species. As assessed by in situ hybridization, non-atretic ovine follicles expressed mRNA for BMP6 but not 2, 4 or 7. Furthermore, expression of BMP6 was limited to the oocyte of primordial as well as primary, pre-antral and antral follicles. Reverse transcription-PCR of granulosa cell mRNA detected low levels of all the BMPs in some pools of cells. BMP2, 4, 6 and 7 each inhibited progesterone production from ovine granulosa cells without affecting cellular proliferation/survival. Similarly, these BMPs inhibited progesterone production from rat granulosa cells. However, they also stimulated cellular proliferation/survival of the rat granulosa cells highlighting a species-specific difference for these growth factors. In conclusion, in sheep, BMP2, 4, 6 and 7 inhibit granulosa cell differentiation without affecting proliferation. However, as BMP2, 4 and 7 were not detectable by in situ hybridization in any cells of non-atretic ovarian follicles, it seems unlikely that these proteins would have an important intra-ovarian role in regulating follicular development in sheep. In contrast, localization of BMP6 mRNA in the oocyte suggests that this BMP family member may have a paracrine and/or autocrine role in regulating follicular growth in sheep, as has been shown for two other oocyte derived from members of the transforming growth factor superfamily, BMP15 and growth differentiation factor 9.

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IGF1 induces up-regulation of steroidogenic and apoptotic regulatory genes via activation of phosphatidylinositol-dependent kinase/AKT in bovine granulosa cells

Reproduction ◽

10.1530/rep-09-0050 ◽

2010 ◽

Vol 139 (1) ◽

pp. 139-151 ◽

Cited By ~ 89

Author(s):

Arul Murugan Mani ◽

Mark A Fenwick ◽

Zhangrui Cheng ◽

Mohan K Sharma ◽

Dheer Singh ◽

...

Keyword(s):

Mrna Expression ◽

Granulosa Cell ◽

Granulosa Cells ◽

Cellular Proliferation ◽

Mapk Pathway ◽

Regulatory Gene ◽

Follicular Development ◽

Pi3k Pathway ◽

Cell Number ◽

Regulatory Genes

IGF1, a potent stimulator of cellular proliferation, differentiation and development, regulates granulosa cell steroidogenesis and apoptosis during follicular development. Depending upon species and stage of follicular growth, IGF1 acts on granulosa cell steroidogenesis either alone or together with FSH. We examined the mechanism of action of IGF1 in bovine granulosa cells in serum-free culture without insulin to determine its potential role in the regulation of steroidogenic and apoptotic regulatory gene expression and to investigate the interaction of FSH with IGF1 on this mechanism. Bovine granulosa cells treated with IGF1 demonstrated a significant increase in 17β-oestradiol (OE2) production, cell number and in mRNA expression ofCYP11A1,HSD3B1,CYP19A1,BAX, type 1 IGF receptor (IGF1R) andFSHR, while FSH alone had no significant effects. IGF1 or FSH alone or both together had no effect onBCL2expression. IGF1 with FSH resulted in a synergistic increase in granulosa cell number and in mRNA expression ofCYP19A1andIGF1Rwithout altering OE2production. IGF1 stimulated the phosphoinositide 3′-OH kinase (PI3K) but not the MAPK pathway in granulosa cells, as evidenced by increased phosphorylation of AKT but not extracellular-regulated kinase 1/2. Addition of the PI3K pathway inhibitor LY294002 (but not the MAPK pathway inhibitor PD98059) abrogated the increased expression of genes induced by IGF1. IGF1 therefore up-regulates the steroidogenic and apoptotic regulatory genes via activation of PI3K/AKT in bovine granulosa cells. The synergistic action of IGF1 with FSH is of likely key importance for the development of small antral follicles before selection; subsequently, other factors such as LH may also become necessary for continued cell survival.

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The conflict between hierarchical ovarian follicular development and superovulation treatment

Reproduction ◽

10.1530/rep-10-0165 ◽

2010 ◽

Vol 140 (2) ◽

pp. 287-294 ◽

Cited By ~ 11

Author(s):

Kenneth P McNatty ◽

Derek A Heath ◽

Norma L Hudson ◽

Karen L Reader ◽

Laurel Quirke ◽

...

Keyword(s):

Granulosa Cell ◽

Granulosa Cells ◽

Follicular Development ◽

Follicular Phase ◽

Ovulation Rate ◽

Cell Populations ◽

Antral Follicles ◽

Functional States ◽

Ovarian Follicular Development

In mammals with a low ovulation rate phenotype, ovarian follicular development is thought to be hierarchical with few, if any, antral follicles at similar stages of development. The hypothesis being tested herein was that if most follicles are in a functionally different state, then the application of exogenous hormones to increase ovulation rate will not overcome the hierarchical nature of follicular development. Using sheep as the experimental model, the functional states of all non-atretic antral follicles ≥2 mm diameter were assessed in individual ewes (N=10/group) during anoestrus with or without pregnant mare's serum gonadotrophin (PMSG) treatment, or after a standard superovulation regimen, or during the follicular phase of the oestrous cycle. The functional states of these follicles were assessed by measuring the FSH- or human chorionic gonadotrophin (hCG)-induced cAMP responses of granulosa cellsin vitro. There were significant overall effects across the treatment groups on the responses of granulosa cells to either FSH or LH (bothP<0.001). It was concluded that for anoestrous ewes with or without PMSG treatment, and ewes during the follicular phase, granulosa cell populations of many follicles (≥2 mm diameter) did not share a similar cAMP response to FSH (∼50% of follicles) or hCG (>90% of follicles) either on a per cell or total cell basis. After superovulation, ≤30 and 10% respectively of the granulosa cell populations shared similar responses to FSH and LH with regard to follicular diameter and cAMP output. Thus, exogenous hormone treatments used routinely for increasing oocyte yield do not effectively override the hierarchical pattern of ovarian follicular development during the follicular phase.

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Effect of hypothyroidism on ovarian follicular development, granulosa cell proliferation and peripheral hormone levels in the prepubertal rat

Acta Endocrinologica ◽

10.1530/eje.0.1340649 ◽

1996 ◽

Vol 134 (5) ◽

pp. 649-654 ◽

Cited By ~ 54

Author(s):

Grietje Dijkstra ◽

Dirk G de Rooij ◽

Frank H de Jong ◽

Robert van den Hurk

Keyword(s):

Cell Proliferation ◽

Granulosa Cell ◽

Granulosa Cells ◽

Ovarian Development ◽

Follicular Development ◽

Corpora Lutea ◽

Hormone Levels ◽

Antral Follicles ◽

Ovarian Follicular Development ◽

Serum Tsh

Dijkstra G, de Rooij DG, de Jong FH, van den Hurk R. Effect of hypothyroidism on ovarian follicular development, granulosa cell proliferation and peripheral hormone levels in the prepubertal rat. Eur J Endocrinol 1996;134:649–54. ISSN 0804–4643 The aim of this study was to examine the effects of prepubertal hypothyroidism on ovarian development in rats. Therefore, from birth up to day 40 postpartum, rats were given 6-propyl-2-thiouracil (PTU) via the drinking water of mothers and pups. At ages ranging from 12 to 40 days, ovarian weights were measured and serum was collected to estimate thyrotrophin (TSH), folliclestimulating hormone (FSH) and inhibin levels. Two hours before sacrifice the animals received an injection of bromodeoxyuridine (BrdU) to estimate the proliferative activity of the follicular granulosa cells. Ovaries were fixed in Carnoy's fluid and follicle counts were performed on sections stained with anti-BrdU and with haematoxylin and eosin. The PTU treatment resulted in increased serum TSH levels, indicative of hypothyroidism, and markedly lower body and ovarian weights, whereas serum FSH and inhibin levels were hardly affected. At day 40, ovaries of PTU-treated animals contained relatively more secondary and less antral follicles, smaller non-atretic antral follicles and more atretic follicles when compared with untreated rats, while corpora lutea were absent. It is concluded that this disturbed folliculogenesis is due to inadequate thyroid hormone supply, which hampers the differentiation and not the proliferation of granulosa cells because diameters of antral follicles were significantly smaller whereas the BrdU-labelling index had not changed. Robert van den Hurk, Department of Functional Morphology, Faculty of Veterinary Medicine, PO Box 80.157, 3508 TD Utrecht, The Netherlands

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Oocyte control of ovarian follicular development and function in mammals

Reproduction ◽

10.1530/rep.0.1220829 ◽

2001 ◽

pp. 829-838 ◽

Cited By ~ 547

Author(s):

JJ Eppig

Keyword(s):

Granulosa Cell ◽

Granulosa Cells ◽

Follicular Development ◽

Primordial Follicle ◽

Dominant Factor ◽

Ovarian Follicular Development ◽

New Perspective ◽

Regulatory Loop ◽

Sphere Of Influence ◽

And Function

A new perspective on ovarian follicular development has emerged over the last decade. Whereas the oocyte was previously considered only a passive recipient of developmental signals from oocyte-associated granulosa cells, it is now clear that communication between oocytes and granulosa cells is bidirectional. A complex interplay of regulatory factors governs the development of both types of cell. This interplay is essential not only for oocyte development but also for follicular development, beginning with the initial assembly of the primordial follicle and continuing throughout ovulation. The existence of an oocyte-granulosa cell regulatory loop, essential for normal follicular differentiation as well as for the production of an oocyte competent to undergo fertilization and embryogenesis, is proposed. Although gonadotrophins are essential for driving the differentiation of granulosa cell phenotypes, within its sphere of influence, the oocyte is probably the dominant factor determining the direction of differentiation and the function of the granulosa cells associated with it.

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rno-miR-128-3p promotes apoptosis in rat granulosa cells (GCs) induced by norepinephrine through Wilms tumor 1 (WT1)

In Vitro Cellular & Developmental Biology - Animal ◽

10.1007/s11626-021-00609-y ◽

2021 ◽

Author(s):

Ming Li ◽

Ling Xue ◽

Weibin Xu ◽

Pingping Liu ◽

Feng Li

Keyword(s):

Granulosa Cell ◽

Granulosa Cells ◽

Ovarian Function ◽

Wilms Tumor ◽

Sympathetic Innervation ◽

Follicular Development ◽

Wilms Tumor 1 ◽

Ovarian Follicular Development ◽

Induced Apoptosis

AbstractThe mechanism related to ovarian follicular is complex, which has not been fully elucidated. Abundant reports have confirmed that the ovarian function development is closely related to sympathetic innervation. As one of the major neurotransmitters, norepinephrine (NE) is considered an effective regulator of ovarian functions like granulosa cell (GC) apoptosis. However, the mechanism between NE and GC apoptosis in rat is still unclear. In our study, GCs were isolated and cultured in vitro with NE treatment. The apoptosis of GCs was facilitated by NE. Wilms tumor 1 (WT1) was found to be significantly downregulated in GCs after NE treatment, and overexpression of WT1 repressed apoptosis in rat GCs induced by NE. rno-miR-128-3p was found to be significantly enhanced by NE in GCs, and inhibition of rno-miR-128-3p repressed apoptosis in rat GCs induced by NE. Mechanistically, rno-miR-128-3p interacted with WT1 and repressed its expression. In summary, inhibition of rno-miR-128-3p may enhance WT1 expression, and then repress NE-induced apoptosis in rat GCs. Our research may provide a new insight for the improvement of ovarian follicular development.

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MicroRNA Mediating Networks in Granulosa Cells Associated with Ovarian Follicular Development

BioMed Research International ◽

10.1155/2017/4585213 ◽

2017 ◽

Vol 2017 ◽

pp. 1-18 ◽

Cited By ~ 4

Author(s):

Baoyun Zhang ◽

Long Chen ◽

Guangde Feng ◽

Wei Xiang ◽

Ke Zhang ◽

...

Keyword(s):

Transcription Factors ◽

Granulosa Cells ◽

Messenger Rna ◽

Expression Patterns ◽

Follicular Development ◽

Functional Networks ◽

Signal Pathways ◽

Differentially Expressed Mirnas ◽

Selection Of

Ovaries, which provide a place for follicular development and oocyte maturation, are important organs in female mammals. Follicular development is complicated physiological progress mediated by various regulatory factors including microRNAs (miRNAs). To demonstrate the role of miRNAs in follicular development, this study analyzed the expression patterns of miRNAs in granulosa cells through investigating three previous datasets generated by Illumina miRNA deep sequencing. Furthermore, via bioinformatic analyses, we dissected the associated functional networks of the observed significant miRNAs, in terms of interacting with signal pathways and transcription factors. During the growth and selection of dominant follicles, 15 dysregulated miRNAs and 139 associated pathways were screened out. In comparison of different styles of follicles, 7 commonly abundant miRNAs and 195 pathways, as well as 10 differentially expressed miRNAs and 117 pathways in dominant follicles in comparison with subordinate follicles, were collected. Furthermore, SMAD2 was identified as a hub factor in regulating follicular development. The regulation of miR-26a/b onsmad2messenger RNA has been further testified by real time PCR. In conclusion, we established functional networks which play critical roles in follicular development including pivotal miRNAs, pathways, and transcription factors, which contributed to the further investigation about miRNAs associated with mammalian follicular development.

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

Reproduction ◽

10.1530/rep-13-0386 ◽

2014 ◽

Vol 147 (1) ◽

pp. 73-80 ◽

Cited By ~ 43

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.

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

Journal of Endocrinology ◽

10.1530/joe-16-0488 ◽

2017 ◽

Vol 234 (1) ◽

pp. 1-14 ◽

Cited By ~ 19

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.

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The mouse has a Polycomb-like chromobox gene

Development ◽

10.1242/dev.114.4.921 ◽

1992 ◽

Vol 114 (4) ◽

pp. 921-929 ◽

Cited By ~ 11

Author(s):

J.J. Pearce ◽

P.B. Singh ◽

S.J. Gaunt

Keyword(s):

Cellular Proliferation ◽

Expression Patterns ◽

Northern Analysis ◽

Heterochromatin Protein 1 ◽

Heterochromatin Protein ◽

Drosophila Gene ◽

Acid Protein ◽

Temporal And Spatial ◽

Amino Acid Protein

The Drosophila gene Polycomb (Pc) has been implicated in the clonal inheritance of determined states and is a trans-regulator of the Antennapedia-like homeobox genes. Pc shares a region of homology (the chromobox) with the Drosophila gene Heterochromatin Protein 1 (HP1), a component of heterochromatin. The Pc chromobox has been used to isolate a mouse chromobox gene, M33, which encodes a predicted 519 amino acid protein. The M33 chromodomain is more similar to that in the Pc protein, than that in the HP1 protein. In addition to the chromodomain, the M33 and Pc proteins also share a region of homology at their C termini. The temporal and spatial expression patterns of M33 have been studied by in situ hybridization and northern analysis. During the final 10 days of embryonic development, M33 expression mirrors that of the cell-cycle-specific cyclin B gene. It is therefore suggested that the rate of cellular proliferation controls M33 expression. From comparisons of the characteristics of M33 with those of Pc it is proposed that M33 is a Pc-like chromobox gene. The roles of M33 and Pc in models of cellular memory are examined and implications of the memory models addressed.

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Pentachloronitrobenzene alters progesterone production and primordial follicle recruitment in cultured granulosa cells and rat ovary†

Biology of Reproduction ◽

10.1093/biolre/ioz195 ◽

2019 ◽

Vol 102 (2) ◽

pp. 511-520

Author(s):

Yanrong Kuai ◽

Xiaobo Gao ◽

Huixia Yang ◽

Haiyan Luo ◽

Yang Xu ◽

...

Keyword(s):

Protein Kinase ◽

Granulosa Cells ◽

Crop Production ◽

Follicular Development ◽

Mitogen Activated Protein Kinase ◽

Dependent Manner ◽

Serum Progesterone ◽

Primordial Follicles ◽

Progesterone Production

Abstract Pentachloronitrobenzene (PCNB) is an organochlorine fungicide widely used for crop production and has become an environmental concern. Little is known about the effect of PCNB on ovarian steroidogenesis and follicular development. We found that PCNB stimulated Star expression and progesterone production in cultured rat granulosa cells in a dose-dependent manner. PCNB activated mitogen-activated protein kinase (MAPK3/1) extracellulat regulated kinase (ERK1/2), thus inhibition of either protein kinase A (PKA) or MAPK3/1 signaling pathway significantly attenuated progesterone biosynthesis caused by PCNB, suggesting that PCNB induced progesterone production by activating the cyclic adenosine monophosphate (cAMP/PKA) and MAPK3/1 signaling pathways. Further investigation demonstrated that PCNB induced Star expression and altered MAPK3/1 signaling in ovary tissues of immature SD rats treated with PCNB at the dose of 100, 200, or 300 mg/kg by daily gavage for 7 days, while serum progesterone level was dose-dependently decreased. We demonstrated that PCNB exposure accelerated the recruitment of primordial follicles into the growing follicle pool in ovary tissues, accompanied by increased levels of anti-Mullerian hormone (AMH) in both ovary tissues and serum. Taken together, our data demonstrate for the first time that PCNB stimulated Star expression, altered MAPK3/1 signaling and progesterone production in vivo and in vitro, and accelerated follicular development with a concomitant increase in AMH in ovary tissues and serum. Our findings provide novel insight into the toxicity of PCNB to animal ovary function.

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