Transcription profile of the insulin-like growth factor signaling pathway during human ovarian follicular development
J Assist Reprod Genet. 2019 Mar 15. doi: 10.1007/s10815-019-01432-x. [Epub ahead of print]
Bøtkjær JA1, Pors SE2, Petersen TS2, Kristensen SG2, Jeppesen JV2, Oxvig C3, Andersen CY2. Author information Abstract PURPOSE: The IGF signaling cascade exerts important regulatory functions in human ovarian folliculogenesis. The scope of this study was to evaluate the transcription profile of insulin-like growth factor (IGF) genes during human ovarian follicle development and to analyze follicle fluid levels of key IGF proteins. METHODS: Gene expression profiling was performed with microarray gene analysis. The analysis was assessed from ovarian follicles and granulosa cells (GCs) obtained from isolated stage-specific human ovarian follicles, including preantral follicles, small antral follicles, and preovulatory follicles. Numerous genes involved in the IGF signaling pathway was evaluated and key genes were validated by qPCR from GCs. Protein levels of various IGF components of human follicular fluid (FF) were measured by ELISA and time-resolved immunofluorometric assays (TRIFMA). RESULTS: The gene expression levels of PAPPA, IGF2, IGF receptors and intracellular IGF-activated genes increased with increasing follicle size. This was especially prominent in the late preovulatory stage where IGF2 expression peaked. Protein levels of intact IGF binding protein-4 decreased significantly in FF from large preovulatory follicles compared with small antral follicles concomitant with higher protein levels of PAPP-A. The IGF modulators IGF-2 receptor, IGFBPs, stanniocalcins, and IGF-2 mRNA binding proteins were all observed to be expressed in the different follicle stages. CONCLUSIONS: This study confirms and highlights the importance of PAPP-A regulating bioactive IGF levels throughout folliculogenesis and especially for the high rate of granulosa cell proliferation and expression of key ovarian hormones important in the last part of the follicular phase of the menstrual cycle. KEYWORDS: Human ovarian follicles; IGF system; IGFBPs; PAPP-A; Stanniocalcins PMID: 30877600 DOI: 10.1007/s10815-019-01432-x
Inactivation of the LOX-1 pathway promotes the Golgi apparatus during cell differentiation of mural granulosa cells
J Cell Physiol. 2014 Dec;229(12):1946-51. doi: 10.1002/jcp.24644.
Weitzel JM1, Vernunft A, Krüger B, Plinski C, Viergutz T.
In female mammals, granulosa cells of the ovarian follicle differentiate into the corpus luteum after ovulation of the pregnable oocyte into the fallopian tube. During these differentiation processes several morphological alterations have to occur and the molecular basis is not fully understood. As an endpoint estradiol production from granulosa cells has to switch off in favor for progesterone production from the proceeding corpus luteum to sustain the developing embryo. Previously, we demonstrated that the multiligand receptor LOX-1 plays a critical role in steroid hormone synthesis of granulosa cells via intracellular calcium release from endoplasmic (ER)-dependent and ER-independent calcium pools. In the present study, we show that inhibition of LOX-1 leads to a rearrangement of ceramide from the basal membrane toward the Golgi apparatus. This activity is accomplished by a calcium-dependent phosphorylation of aromatase, the key step in estradiol production. Phosphorylated aromatase increased estradiol production in a dose-dependent manner. Our data indicate that the ceramide cascade is essential for proper granulosa cell function and ceramide redistribution serves as a first step in order to proceed with the prosperous differentiation into a corpus luteum. © 2014 Wiley Periodicals, Inc. PMID 24710763
Transcriptomic Diversification of Developing Cumulus and Mural Granulosa Cells in Mouse Ovarian Follicles
Biol Reprod. 2014 Nov 5. pii: biolreprod.114.121756.
Wigglesworth K, Lee KB, Emori C, Sugiura K, Eppig JJ.
Cumulus cells and mural granulosa cells (MGCs) have functionally distinct roles in antral follicles, and comparison of their transcriptomes at a global and systems level can propel future studies on mechanisms underlying their functional diversity. These cells were isolated from small and large antral follicles before and after stimulation of immature mice with gonadotropins, respectively. Both cell types underwent dramatic transcriptomic changes and differences between them increased with follicular growth. Although cumulus cells of both stages of follicular development are competent to undergo expansion in vitro, they were otherwise remarkably dissimilar with transcriptomic changes quantitatively equivalent to those of MGCs. Gene Ontology analysis revealed that cumulus cells of small follicles were enriched in transcripts generally associated with catalytic components of metabolic processes, while those from large follicles were involved in regulation of metabolism, cell differentiation, and adhesion. Contrast of cumulus cells versus MGCs revealed that cumulus cells were enriched in transcripts associated with metabolism and cell proliferation while MGCs were enriched for transcripts involved in cell signaling and differentiation. In-vitro and in-vivo models were used to test the hypothesis that higher levels of transcripts in cumulus cells versus MGCs is the result of stimulation by oocyte-derived paracrine factors (ODPFs). Surprisingly ~48% of transcripts higher in cumulus cells than MGCs were not stimulated by ODPFs. Those stimulated by ODPFs were mainly associated with cell division, mRNA processing, or the catalytic pathways of metabolism, while those not stimulated by ODPFs were associated with regulatory processes such as signaling, transcription, phosphorylation, or the regulation of metabolism. Copyright 2014 by The Society for the Study of Reproduction. KEYWORDS: Cumulus cells; Developmental biology; Follicular development; Gene expression; Granulosa cells
Wt1 functions in ovarian follicle development by regulating granulosa cell differentiation
Hum Mol Genet. 2014 Jan 15;23(2):333-41. doi: 10.1093/hmg/ddt423. Epub 2013 Sep 5.
Gao F1, Zhang J, Wang X, Yang J, Chen D, Huff V, Liu YX.
The Wt1 gene encodes a nuclear transcription factor that is specifically expressed in ovarian granulosa cells. However, the physiological significance of Wt1 in ovarian follicle development remains elusive. In this study, we found that Wt1(+/R394W) mice were grossly normal, however, the females displayed severe reproductive defects. Only ∼15% of the Wt1(+/R394W) females became pregnant after mating with wild-type males, compared with 88.2% of control females. Further study revealed that the subfertility of Wt1(+/R394W) females was caused by aberrant ovarian follicle development. Compared with control females, the ovary size and the number of developing follicles was significantly decreased in Wt1 mutant ovaries which was very similar to premature ovarian failure (POF) in human patients. The results of in vitro studies demonstrated that the expression of follicle stimulating hormone receptor (FSHR), 3β-hydroxysteroid dehydrogenase and Aromatase was inhibited by Wt1 in granulosa cells, and mutation of Wt1 resulted in the upregulation of these genes and in the premature differentiation of granulosa cells. We also found that Wt1 was likely involved in granulosa cell development via the regulation of E-cadherin and Par6b expression. Mutation in Wt1 caused defects in polarity establishment in granulosa cells, which also likely contributed to the observed aberrant follicle development. The results of this study provide new mechanisms for understanding the regulation of ovarian follicle development and potential pathological cause of POF in human patients.
Increased GDF9 and BMP15 mRNA levels in cumulus granulosa cells correlate with oocyte maturation, fertilization, and embryo quality in humans
Reprod Biol Endocrinol. 2014 Aug 20;12:81. doi: 10.1186/1477-7827-12-81.
Li Y, Li RQ, Ou SB, Zhang NF, Ren L, Wei LN1, Zhang QX, Yang DZ.
BACKGROUND: Oocyte secreted factors (OSFs), including growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15), play an important role in the process of follicular development and oocyte maturation. Since OSFs are expressed in oocytes and cumulus granulosa cells, the aim of the present study was to explore whether the expression levels of GDF9 and BMP15 mRNAs in cumulus granulosa cells can be used as molecular markers for predicting oocyte developmental potential. METHODS: Cumulus cells of 2426 cumulus-oocyte complexes were collected from 196 female patients who underwent intracytoplasmic sperm injection (ICSI) and were used for mRNA detection on the egg retrieval day. Pearson correlation analysis was used to analyze the correlation between OSF expression and general physiological parameters. Partial correlation analysis was used to analyze the correlation between OSF expression and oocyte developmental potential. Covariance analysis was used to compare OSF expression among different groups. Receiver operating characteristic curves were used to examine the diagnostic value of GDF9 and BMP15 mRNA for predicting pregnancy. RESULTS: The expression levels of GDF9 and BMP15 mRNAs were significantly associated with age, body mass index (BMI), oocyte maturation, normal fertilization, and cleavage rate (P < 0.05). The expression levels of GDF9 and BMP15 mRNAs in the group with high-quality embryos were significantly higher than those in the group without high-quality embryos (P < 0.05). The expression levels of GDF9 and BMP15 mRNAs in the pregnancy group were significantly higher than those in the nonpregnancy group (P < 0.05). The cut-off value of GDF9 mRNA for predicting pregnancy was 4.82, with a sensitivity of 82% and a specificity of 64%. The cut-off value of BMP15 mRNA for predicting pregnancy was 2.60, with a sensitivity of 78% and a specificity of 52%. CONCLUSIONS: The expression levels of GDF9 and BMP15 mRNAs were closely associated with oocyte maturation, fertilization, embryo quality, and pregnancy outcome; therefore, GDF9 and BMP15 mRNAs in cumulus granulosa cells may be considered as new molecular markers for predicting oocyte developmental potential. PMID 25139161
Somatic cells regulate maternal mRNA translation and developmental competence of mouse oocytes
Nat Cell Biol. 2013 Dec;15(12):1415-23. doi: 10.1038/ncb2873. Epub 2013 Nov 24.
Chen J1, Torcia S, Xie F, Lin CJ, Cakmak H, Franciosi F, Horner K, Onodera C, Song JS, Cedars MI, Ramalho-Santos M, Conti M.
Abstract Germ cells divide and differentiate in a unique local microenvironment under the control of somatic cells. Signals released in this niche instruct oocyte reentry into the meiotic cell cycle. Once initiated, the progression through meiosis and the associated programme of maternal messenger RNA translation are thought to be cell autonomous. Here we show that translation of a subset of maternal mRNAs critical for embryo development is under the control of somatic cell inputs. Translation of specific maternal transcripts increases in oocytes cultured in association with somatic cells and is sensitive to EGF-like growth factors that act only on the somatic compartment. In mice deficient in amphiregulin, decreased fecundity and oocyte developmental competence is associated with defective translation of a subset of maternal mRNAs. These somatic cell signals that affect translation require activation of the PI(3)K-AKT-mTOR pathway. Thus, mRNA translation depends on somatic cell cues that are essential to reprogramme the oocyte for embryo development. Comment in Somatic guidance for the oocyte. [Dev Cell. 2013] PMID 24270888
Comparative analysis of follicle morphology and oocyte diameter in four mammalian species (mouse, hamster, pig, and human)
J Exp Clin Assist Reprod. 2006 Mar 1;3:2.
Griffin J1, Emery BR, Huang I, Peterson CM, Carrell DT.
BACKGROUND: Laboratory animals are commonly used for evaluating the physiological properties of the mammalian ovarian follicle and the enclosed oocyte. The use of different species to determine the morphological relationship between the follicle and oocyte has led to a recognizable pattern of follicular stages, but differences in follicle size, oocyte diameter and granulosa cell proliferation are not consistent across the different species. In an effort to better understand how these differences are expressed across multiple species, this investigation evaluates oocyte and follicle diameters and granulosa cell proliferation in the mouse, hamster, pig, and human. METHODS: Histological sections of ovaries from the mouse, hamster, pig, and human were used to calculate the diameter of the oocyte and follicle and the number of granulosa cells present at pre-determined stages of follicular development. A statistical analysis of these data was performed to determine the relationship of follicular growth and development within and between the species tested. RESULTS: These data have revealed that the relationships of the features listed are tightly regulated within each species, but they vary between the species studied. CONCLUSION: This information may be useful for comparative studies conducted in different animal models and the human. PMID 16509981
Formation of mammalian oocytes and their growth, differentiation and maturation within ovarian follicles
Theriogenology. 2005 Apr 1;63(6):1717-51.
van den Hurk R1, Zhao J.
Abstract The limited knowledge on the regulation of oocyte formation, the different steps of folliculogenesis and the required conditions for oocytes to undergo proper growth, differentiation and maturation are major causes of the failure in obtaining viable offspring from in vitro cultured early oocytes from domestic animals and humans. This review highlights the factors that at present are known to be involved in the formation of mammalian oocytes and their growth, differentiation and maturation within ovarian follicles. PMID 15763114
Ovarian granulosa cell lines
Mol Cell Endocrinol. 2004 Dec 30;228(1-2):67-78.
Havelock JC1, Rainey WE, Carr BR.
The ovary is a complex endocrine gland responsible for production of sex steroids and is the source of fertilizable ova for reproduction. It also produces various growth factors, transcription factors and cytokines that assist in the complex signaling pathways of folliculogenesis. The ovary possesses two primary steroidogenic cell types. The theca cells (and to a lesser extent, the stroma) are responsible for androgen synthesis, and the granulosa cells are responsible for conversion of androgens to estrogens, as well as progesterone synthesis. These cells undergo a transformation in the luteal phase of the menstrual cycle, converting them from estrogen producing, to predominantly progesterone producing cells. Understanding the molecular mechanisms regulating these cells is essential in understanding the regulation of steroidogenesis and reproduction. Creation of appropriate in vitro cell model systems can provide important tools for the study of ovarian function. This has led to the development of ovarian steroidogenic cell lines in several laboratories. Developing theca cell lines has met with limited success. Conversely, numerous human and animal granulosa cell lines have been developed. This review will discuss the existing granulosa cell lines and their characteristics.