Talk:Oocyte Development

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Cite this page: Hill, M.A. (2024, June 24) Embryology Oocyte Development. Retrieved from

10 Most Recent Papers

Note - This sub-heading shows an automated computer PubMed search using the listed sub-heading term. References appear in this list based upon the date of the actual page viewing. Therefore the list of references do not reflect any editorial selection of material based on content or relevance. In comparison, references listed on the content page and discussion page (under the publication year sub-headings) do include editorial selection based upon relevance and availability. (More? Pubmed Most Recent)

Oocyte Development

<pubmed limit=5>Oocyte Development</pubmed>

Oocyte Meiosis

<pubmed limit=5>Oocyte+Meiosis</pubmed>

Polar Body

<pubmed limit=5>Polar+Body</pubmed>

Oocyte Quality

<pubmed limit=5>Oocyte Quality</pubmed>


Xie Y, Wu B, Jin Y, Zhang A, Sun X, Zhang X, Gao X, Dong R, Li H & Gao J. (2018). Oocyte-specific deletion of Gsα induces oxidative stress and deteriorates oocyte quality in mice. Exp. Cell Res. , , . PMID: 30026030 DOI.

Oocyte stage-specific effects of MTOR determine granulosa cell fate and oocyte quality in mice

Proc Natl Acad Sci U S A. 2018 May 21. pii: 201800352. doi: 10.1073/pnas.1800352115. [Epub ahead of print]

Guo J1, Zhang T1, Guo Y1, Sun T1, Li H1, Zhang X1, Yin H1, Cao G1, Yin Y1, Wang H1, Shi L1, Guo X1, Sha J1, Eppig JJ2, Su YQ3,4,5.

Abstract MTOR (mechanistic target of rapamycin) is a widely recognized integrator of signals and pathways key for cellular metabolism, proliferation, and differentiation. Here we show that conditional knockout (cKO) of Mtor in either primordial or growing oocytes caused infertility but differentially affected oocyte quality, granulosa cell fate, and follicular development. cKO of Mtor in nongrowing primordial oocytes caused defective follicular development leading to progressive degeneration of oocytes and loss of granulosa cell identity coincident with the acquisition of immature Sertoli cell-like characteristics. Although Mtor was deleted at the primordial oocyte stage, DNA damage accumulated in oocytes during their later growth, and there was a marked alteration of the transcriptome in the few oocytes that achieved the fully grown stage. Although oocyte quality and fertility were also compromised when Mtor was deleted after oocytes had begun to grow, these occurred without overtly affecting folliculogenesis or the oocyte transcriptome. Nevertheless, there was a significant change in a cohort of proteins in mature oocytes. In particular, down-regulation of PRC1 (protein regulator of cytokinesis 1) impaired completion of the first meiotic division. Therefore, MTOR-dependent pathways in primordial or growing oocytes differentially affected downstream processes including follicular development, sex-specific identity of early granulosa cells, maintenance of oocyte genome integrity, oocyte gene expression, meiosis, and preimplantation developmental competence. KEYWORDS: Sertoli-like cells; female infertility; granulosa cell; meiosis; oocyte-expressed MTOR PMID: 29784807 DOI: 10.1073/pnas.1800352115

Regulation of germ cell development by intercellular signaling in the mammalian ovarian follicle

Wiley Interdiscip Rev Dev Biol. 2018 Jan;7(1). doi: 10.1002/wdev.294. Epub 2017 Sep 11.

Clarke HJ1.


Prior to ovulation, the mammalian oocyte undergoes a process of differentiation within the ovarian follicle that confers on it the ability to give rise to an embryo. Differentiation comprises two phases-growth, during which the oocyte increases more than 100-fold in volume as it accumulates macromolecules and organelles that will sustain early embryogenesis; and meiotic maturation, during which the oocyte executes the first meiotic division and prepares for the second division. Entry of an oocyte into the growth phase appears to be triggered when the adjacent granulosa cells produce specific growth factors. As the oocyte grows, it elaborates a thick extracellular coat termed the zona pellucida. Nonetheless, cytoplasmic extensions of the adjacent granulosa cells, termed transzonal projections (TZPs), enable them to maintain contact-dependent communication with the oocyte. Through gap junctions located where the TZP tips meet the oocyte membrane, they provide the oocyte with products that sustain its metabolic activity and signals that regulate its differentiation. Conversely, the oocyte secretes diffusible growth factors that regulate proliferation and differentiation of the granulosa cells. Gap junction-permeable products of the granulosa cells prevent precocious initiation of meiotic maturation, and the gap junctions also enable oocyte maturation to begin in response to hormonal signals received by the granulosa cells. Development of the oocyte or the somatic compartment may also be regulated by extracellular vesicles newly identified in follicular fluid and at TZP tips, which could mediate intercellular transfer of macromolecules. Oocyte differentiation thus depends on continuous signaling interactions with the somatic cells of the follicle. WIREs Dev Biol 2018, 7:e294. doi: 10.1002/wdev.294 This article is categorized under: Gene Expression and Transcriptional Hierarchies > Cellular Differentiation Signaling Pathways > Cell Fate Signaling Early Embryonic Development > Gametogenesis.

PMID: 28892263 PMCID: PMC5746469

   [Available on 2019-01-01]




Complete in vitro oogenesis: retrospects and prospects

Cell Death Differ. 2017 Nov;24(11):1845-1852. doi: 10.1038/cdd.2017.134. Epub 2017 Aug 25.

Wang JJ1, Ge W1, Liu JC1, Klinger FG2, Dyce PW3, De Felici M2, Shen W1.


Precise control of mammalian oogenesis has been a traditional focus of reproductive and developmental biology research. Recently, new reports have introduced the possibility of obtaining functional gametes derived in vitro from stem cells. The potential to produce functional gametes from stem cells has exciting applications for regenerative medicine though still remains challenging. In mammalian females ovulation and fertilization is a privilege reserved for a small number of oocytes. In reality the vast majority of oocytes formed from primordial germ cells (PGCs) will undergo apoptosis, or other forms of cell death. Removal occurs during germ cell cyst breakdown and the establishment of the primordial follicle (PF) pool, during the long dormancy at the PF stage, or through follicular atresia prior to reaching the ovulatory stage. A way to solve this limitation could be to produce large numbers of oocytes, in vitro, from stem cells. However, to recapitulate mammalian oogenesis and produce fertilizable oocytes in vitro is a complex process involving several different cell types, precise follicular cell-oocyte reciprocal interactions, a variety of nutrients and combinations of cytokines, and precise growth factors and hormones depending on the developmental stage. In 2016, two papers published by Morohaku et al. and Hikabe et al. reported in vitro procedures that appear to reproduce efficiently these conditions allowing for the production, completely in a dish, of a relatively large number of oocytes that are fertilizable and capable of giving rise to viable offspring in the mouse. The present article offers a critical overview of these results as well as other previous work performed mainly in mouse attempting to reproduce oogenesis completely in vitro and considers some perspectives for the potential to adapt the methods to produce functional human oocytes. PMID: 28841213 PMCID: PMC5635224 DOI: 10.1038/cdd.2017.134

Dynamic changes in mitochondrial DNA, distribution and activity within cat oocytes during folliculogenesis

Reprod Domest Anim. 2017 Apr;52 Suppl 2:71-76. doi: 10.1111/rda.12851. Epub 2017 Jan 22.

Songsasen N1, Henson LH1, Tipkantha W2,3, Thongkittidilok C1, Henson JH4, Chatdarong K3, Comizzoli P1.


Mitochondria play fundamental roles during oocyte development. The accumulation and spatial redistribution of these energy-producing organelles have been linked to the developmental competence of mammalian oocytes. Here, we assessed the copy number, distribution and activity of mitochondria within cat oocytes during folliculogenesis. In Experiment 1, oocytes were recovered from primordial (n = 152), primary (112), secondary (95), early (131), small (118), antral (86) and advanced antral (5) stages follicles, and mitochondria DNA extracted and quantified using qPCR. In Experiment 2, oocytes from pre-antral (n = 44), early antral (n = 66), small antral (n = 59), antral (n = 41) and advanced antral (n = 21) follicles were isolated and stained with CMXRos MitoTracker dye to assess mitochondrial distribution pattern and activity levels. Oocyte's mitochondria DNA (mtDNA) copy numbers gradually increased as folliculogenesis progressed, with a significant shift at the small antral stage (0.5 to <1 mm in diameter). The location of mitochondria gradually shifted from a homogeneous distribution throughout the cytoplasm in pre-antral oocytes to a pericortical concentration in the advanced antral stage. Quantification of CMXRos fluorescent intensity revealed a progressive increase in mitochondrial activity in oocytes from the pre-antral to the large antral follicles. Taken together, these findings demonstrated that cat oocytes undergo dynamic changes in mitochondrial copy number, distribution and activity during folliculogenesis. These significant modifications to this crucial cytoplasmic organelle are likely associated with the acquisition of developmental competency by cat oocytes. © 2017 Blackwell Verlag GmbH.

KEYWORDS: cat oocyte; follicle development; mitochondria copy number; mitochondrial distribution

PMID: 28111812 DOI: 10.1111/rda.12851


Spatial Characterization of Bioenergetics and Metabolism of Primordial to Preovulatory Follicles in Whole Ex Vivo Murine Ovary

Biol Reprod. 2016 Dec;95(6):129. doi: 10.1095/biolreprod.116.142141. Epub 2016 Sep 28.

Cinco R1, Digman MA2,3, Gratton E2,3, Luderer U4,5,6.


Previous work characterizing ovarian bioenergetics has defined follicular metabolism by measuring metabolic by-products in culture media. However, culture conditions perturb the native state of the follicle, and these methods do not distinguish between metabolism occurring within oocytes or granulosa cells. We applied the phasor approach to fluorescence lifetime imaging microscopy (phasor FLIM) at 740-nm two-photon excitation to examine the spatial distribution of free and protein-bound nicotinamide adenine dinucleotide hydride (NADH) during primordial through preovulatory stages of follicular development in fresh ex vivo murine neonatal and gonadotropin stimulated prepubertal ovaries. We obtained subcellular resolution phasor FLIM images of primordial through primary follicles and quantified the free/bound NADH ratio (relative NADH/NAD+) separately for oocyte nucleus and oocyte cytoplasm. We found that dynamic changes in oocyte nucleus free/bound NADH paralleled the developmental maturation of primordial to primary follicles. Immunohistochemistry of NAD+-dependent deacetylase SIRTUIN 1 (SIRT1) in neonatal ovary revealed that increasing SIRT1 expression in oocyte nuclei was inversely related to decreasing free/bound NADH during the primordial to primary follicle transition. We characterized oocyte metabolism at these early stages to be NADH producing (glycolysis/Krebs). We extended the results of prior studies to show that cumulus and mural granulosa cell metabolism in secondary through preovulatory follicles is mainly NADH producing (glycolysis/Krebs cycle), while oocyte metabolism is mainly NADH consuming (oxidative phosphorylation). Taken together, our data characterize dynamic changes in free/bound NADH and SIRT1 expression during early follicular development and confirm results from previous studies defining antral and preovulatory follicle metabolism in culture. © 2016 by the Society for the Study of Reproduction, Inc.

KEYWORDS: FLIM; Krebs cycle; NADH; cumulus cells; folliculogenesis; glycolysis; granulosa cells; oocyte; oxidative phosphorylation; phasor approach to fluorescence lifetime imaging microscopy; sirtuin 1

PMID 27683265 PMCID: PMC5315427 [Available on 2017-12-01] DOI: 10.1095/biolreprod.116.142141


A stereological study on organelle distribution in human oocytes at prophase I

Zygote. 2015 Jul 14:1-9. [Epub ahead of print]

Pires-Luís AS1, Rocha E2, Bartosch C3, Oliveira E1, Silva J4, Barros A4, Sá R1, Sousa M5.


The ultrastructural analysis of human oocytes at different maturation stages has only been descriptive. The aim of this study was to use a stereological approach to quantify the distribution of organelles in oocytes at prophase I (GV). Seven immature GV oocytes were processed for transmission electron microscopy and a classical manual stereological technique based on point-counting with an adequate stereological grid was used. The Kruskal-Wallis test and Mann-Whitney U-test with Bonferroni correction were used to compare the means of the relative volumes occupied by organelles in oocyte regions: cortex (C), subcortex (SC) and inner cytoplasm (IC). Here we first describe in GV oocytes very large vesicles of the smooth endoplasmic reticulum (SER), vesicles containing zona pellucida-like materials and coated vesicles. The most abundant organelles were the very large vesicles of the SER (6.9%), mitochondria (6.3%) and other SER vesicles (6.1%). Significant differences in organelle distribution were observed between ooplasm regions: cortical vesicles (C: 1.3% versus SC: 0.1%, IC: 0.1%, P = 0.001) and medium-sized vesicles containing zona pellucida-like materials (C: 0.2% versus SC: 0.02%, IC: 0%, P = 0.004) were mostly observed at the oocyte cortex, whereas mitochondria (C: 3.6% versus SC: 6.0%, IC: 7.2%, P = 0.005) were preferentially located in the subcortex and inner cytoplasm, and SER very large vesicles (IC: 10.1% versus C: 0.9%, SC: 1.67%, P = 0.001) in the oocyte inner cytoplasm. Further quantitative studies are needed in immature metaphase-I and mature metaphase-II oocytes, as well as analysis of correlations between ultrastructural and molecular data, to better understand human oocyte in vitro maturation. KEYWORDS: Cell polarity; Human germinal vesicle oocytes; Organelle distribution; Stereology; Ultrastructure

  1. coated vesicles were observed at the oocyte surface, which indicates that human oocytes at prophase- I exchange materials with the exterior by receptor- mediated endocytosis.
  2. exocytosis vesicles containing ZP- like materials were observed in the oocyte cortex. distinct from the dense cortical vesicles
  3. SER-VLV were found mainly in the inner cytoplasm. represent the organelle with the highest relative volume occupied in the oocyte

Cortical vesicles and medium-sized vesicles containing ZP-like materials were mostly observed at the oocyte cortex, whereas mitochondria were preferentially located in the subcortex and inner cytoplasm, and SER-VLV in the oocyte inner cytoplasm.

PMID 26170179

Piwi Proteins and piRNAs in Mammalian Oocytes and Early Embryos

Cell Rep. 2015 Mar 31;10(12):2069-82. doi: 10.1016/j.celrep.2015.02.062. Epub 2015 Mar 26.

Roovers EF1, Rosenkranz D2, Mahdipour M3, Han CT4, He N5, Chuva de Sousa Lopes SM5, van der Westerlaken LA6, Zischler H2, Butter F7, Roelen BA3, Ketting RF8.


Germ cells of most animals critically depend on piRNAs and Piwi proteins. Surprisingly, piRNAs in mouse oocytes are relatively rare and dispensable. We present compelling evidence for strong Piwi and piRNA expression in oocytes of other mammals. Human fetal oocytes express PIWIL2 and transposon-enriched piRNAs. Oocytes in adult human ovary express PIWIL1 and PIWIL2, whereas those in bovine ovary only express PIWIL1. In human, macaque, and bovine ovaries, we find piRNAs that resemble testis-borne pachytene piRNAs. Isolated bovine follicular oocytes were shown to contain abundant, relatively short piRNAs that preferentially target transposable elements. Using label-free quantitative proteome analysis, we show that these maturing oocytes strongly and specifically express the PIWIL3 protein, alongside other, known piRNA-pathway components. A piRNA pool is still present in early bovine embryos, revealing a potential impact of piRNAs on mammalian embryogenesis. Our results reveal that there are highly dynamic piRNA pathways in mammalian oocytes and early embryos. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

PMID 25818294


The road to maturation: somatic cell interaction and self-organization of the mammalian oocyte

Nat Rev Mol Cell Biol. 2013 Mar;14(3):141-52. doi: 10.1038/nrm3531.

Li R, Albertini DF. Source 1] Stowers Institute for Medical Research, 1000 East 50th Street, Kansas City, Missouri 64110, USA. [2] Department of Molecular and Integrative Physiology, University of Kansas Medical Center, 3901 Rainbow Boulevard, Kansas City, Kansas 66160, USA.


Mammalian oocytes go through a long and complex developmental process while acquiring the competencies that are required for fertilization and embryogenesis. Recent advances in molecular genetics and quantitative live imaging reveal new insights into the molecular basis of the communication between the oocyte and ovarian somatic cells as well as the dynamic cytoskeleton-based events that drive each step along the pathway to maturity. Whereas self-organization of microtubules and motor proteins direct meiotic spindle assembly for achieving genome reduction, actin filaments are instrumental for spindle positioning and the establishment of oocyte polarity needed for extrusion of polar bodies. Meiotic chromatin provides key instructive signals while being 'chauffeured' by both cytoskeletal systems.

PMID 23429793

Cumulus and granulosa cell markers of oocyte and embryo quality

Fertil Steril. 2013 Mar 15;99(4):979-97. doi: 10.1016/j.fertnstert.2013.01.129.

Uyar A, Torrealday S, Seli E. Source Department of Obstetrics, Gynecology, and Reproductive Sciences, Yale University School of Medicine, New Haven, Connecticut. Abstract Lack of an objective, accurate, and noninvasive embryo assessment strategy remains one of the major challenges encountered in in vitro fertilization. Cumulus and mural granulosa cells reflect the characteristics of the oocyte, providing a noninvasive means to assess oocyte quality. Specifically, transcriptomic profiling of follicular cells may help identify biomarkers of oocyte and embryo competence. Current transcriptomics technologies include quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) for analysis of individual genes and microarrays and high-throughput deep sequencing for whole genome expression profiling. Recently, using qRT-PCR and microarray technologies, a multitude of studies correlated changes in cumulus or granulosa cell gene expression with clinically relevant outcome parameters, including in vitro embryo development and pregnancy. While the initial findings are promising, a clinical benefit from the use of identified biomarker genes remains to be demonstrated in randomized controlled trials. Copyright © 2013 American Society for Reproductive Medicine. All rights reserved. PMID 23498999

T-Type Ca2+ Current Activity during Oocyte Growth and Maturation in the Ascidian Styela plicata

PLoS One. 2013;8(1):e54604. doi: 10.1371/journal.pone.0054604. Epub 2013 Jan 22.

Gallo A, Russo GL, Tosti E. Source Animal Physiology and Evolution Laboratory, Stazione Zoologica Anton Dohrn, Napoli, Italy.


Voltage-dependent calcium currents play a fundamental role during oocyte maturation, mostly L-type calcium currents, whereas T-type calcium currents are involved in sperm physiology and cell growth. In this paper, using an electrophysiological and pharmacological approach, we demonstrated, for the first time in oocytes, that T-type calcium currents are present with functional consequences on the plasma membrane of growing immature oocytes of the ascidian Styela plicata. We classified three subtypes of immature oocytes at the germinal vesicle stage on the basis of their size, morphology and accessory cellular structures. These stages were clearly associated with an increased activity of T-type calcium currents and hyperpolarization of the plasma membrane. We also observed that T-type calcium currents oscillate in the post-fertilization embryonic stages, with minimal amplitude of the currents in the zygote and maximal at 8-cell stage. In addition, chemical inhibition of T-type calcium currents, obtained by applying specific antagonists, induced a significant reduction in the rate of cleavage and absence of larval formation. We suggest that calcium entry via T-type calcium channels may act as a potential pacemaker in regulating cytosolic calcium involved in fertilization and early developmental events.

PMID 23349937


Ovastacin, a cortical granule protease, cleaves ZP2 in the zona pellucida to prevent polyspermy

J Cell Biol. 2012 Apr 2;197(1):37-44. doi: 10.1083/jcb.201112094.

Burkart AD1, Xiong B, Baibakov B, Jiménez-Movilla M, Dean J.


The mouse zona pellucida is composed of three glycoproteins (ZP1, ZP2, and ZP3), of which ZP2 is proteolytically cleaved after gamete fusion to prevent polyspermy. This cleavage is associated with exocytosis of cortical granules that are peripherally located subcellular organelles unique to ovulated eggs. Based on the cleavage site of ZP2, ovastacin was selected as a candidate protease. Encoded by the single-copy Astl gene, ovastacin is an oocyte-specific member of the astacin family of metalloendoproteases. Using specific antiserum, ovastacin was detected in cortical granules before, but not after, fertilization. Recombinant ovastacin cleaved ZP2 in native zonae pellucidae, documenting that ZP2 was a direct substrate of this metalloendoprotease. Female mice lacking ovastacin did not cleave ZP2 after fertilization, and mouse sperm bound as well to Astl-null two-cell embryos as they did to normal eggs. Ovastacin is a pioneer component of mouse cortical granules and plays a definitive role in the postfertilization block to sperm binding that ensures monospermic fertilization and successful development.

PMID 22472438

Reproductive biology. Potential egg stem cells reignite debate

Science. 2012 Mar 2;335(6072):1029-30.

Vogel G.

Reproductive biologist Jonathan Tilly of Massachusetts General Hospital has challenged the conventional wisdom that female mammals are born with all the egg cells, or oocytes, they will ever have, arguing that in mice—and perhaps also in humans—there must be an ongoing source of new eggs. In 2005, he proposed that bone marrow was a source of eggs in mice. (That idea was discredited a year later.) But this week online in Nature Medicine, Tilly and colleagues report isolating rare cells in ovarian tissue from adult women that can grow in lab dishes and form immature oocytes. This latest claim is earning some cautious acceptance.

PMID 22383817

Oocyte formation by mitotically active germ cells purified from ovaries of reproductive-age women

Nat Med. 2012 Feb 26;18(3):413-21. doi: 10.1038/nm.2669.

White YA, Woods DC, Takai Y, Ishihara O, Seki H, Tilly JL. Source Vincent Center for Reproductive Biology, Massachusetts General Hospital Vincent Department of Obstetrics and Gynecology, Massachusetts General Hospital, Boston, Massachusetts, USA. Abstract Germline stem cells that produce oocytes in vitro and fertilization-competent eggs in vivo have been identified in and isolated from adult mouse ovaries. Here we describe and validate a fluorescence-activated cell sorting-based protocol that can be used with adult mouse ovaries and human ovarian cortical tissue to purify rare mitotically active cells that have a gene expression profile that is consistent with primitive germ cells. Once established in vitro, these cells can be expanded for months and can spontaneously generate 35- to 50-μm oocytes, as determined by morphology, gene expression and haploid (1n) status. Injection of the human germline cells, engineered to stably express GFP, into human ovarian cortical biopsies leads to formation of follicles containing GFP-positive oocytes 1-2 weeks after xenotransplantation into immunodeficient female mice. Thus, ovaries of reproductive-age women, similar to adult mice, possess rare mitotically active germ cells that can be propagated in vitro as well as generate oocytes in vitro and in vivo. Comment in

Nat Med. 2012 Mar;18(3):353-4.

PMID 22366948

Retransplantation of cryopreserved ovarian tissue: the first live birth in Germany

Dtsch Arztebl Int. 2012 Jan;109(1-2):8-13. Epub 2012 Jan 9.

Müller A, Keller K, Wacker J, Dittrich R, Keck G, Montag M, Van der Ven H, Wachter D, Beckmann MW, Distler W.

Abstract BACKGROUND: Cryopreserved ovarian tissue can be retransplanted to restore fertility after radiation or chemotherapy. To date, 15 live births after retransplantation have been reported worldwide. We report the first pregnancy and the first live birth after retransplantation in Germany. CASE REPORT: A 25-year-old female patient received initial chemotherapy and radiation of the mediastinum for Hodgkin's lymphoma in 2003 and suffered a relapse two years later. Ovarian tissue was laparoscopically removed and cryopreserved, and she was then treated with high-dose chemotherapy and stem cell transplantation. She remained in remission for 5 years and she could not conceive during this time. The cryopreserved ovarian tissue was thawed and laparoscopically retransplanted into a peritoneal pouch in the ovarian fossa of the right pelvic wall. Three months later, her menopausal symptoms resolved, and she had her first spontaneous menstruation. Six months after retransplantation, after two normal menstrual cycles, low-dose follicle stimulating hormone (FSH) treatment induced the appearance of a dominant follicle in the tissue graft. Ovulation was then induced with human chorionic gonadotropin (HCG), whereupon the patient conceived naturally. After an uncomplicated pregnancy, she bore a healthy child by Caesarean section on 10 October 2011. Histological examination of biopsy specimens revealed that the ovarian tissue of the graft contained follicles in various stages of development, while the original ovaries contained only structures without any reproductive potential. CONCLUSION: This was the first live birth after retransplantation of cryopreserved ovarian tissue in Germany and also the first case with histological confirmation that the oocyte from which the patient conceived could only have come from the retransplanted tissue. In general, young women who will be undergoing chemotherapy and/or radiotherapy for cancer must be informed and counseled about the available options for fertility preservation.

PMID 22282711

Recent advances in oocyte and ovarian tissue cryopreservation and transplantation

Best Pract Res Clin Obstet Gynaecol. 2012 Jan 31. [Epub ahead of print] Rodriguez-Wallberg KA, Oktay K. Source Karolinska Institute, Department of Clinical Science, Intervention and Technology, Division of Obstetrics and Gynecology K57, Karolinska University Hospital Huddinge SE-141 86 Stockholm, Sweden; Karolinska University Hospital Huddinge, Fertility Unit, Stockholm, Sweden.


Options for preserving fertility in women include well-established methods such as fertility-sparing surgery, shielding to reduce radiation damage to reproductive organs, and emergency in-vitro fertilisation after controlled ovarian stimulation, with the aim of freezing embryos. The practice of transfering frozen or thawed embryos has been in place for over 25 years, and today is a routine clinical treatment in fertility clinics. Oocytes may also be frozen unfertilised for later thawing and fertilisation by intracytoplasmic sperm injection in vitro. In recent years, oocyte cryopreservation methods have further developed, reaching promising standards. More than 1000 children are born worldwide after fertilisation of frozen and thawed oocytes. Nevertheless, this technique is still considered experimental. In this chapter, we focus on options for fertility preservation still in development that can be offered to women. These include freezing of oocytes and ovarian cortex and the transplantation of ovarian tissue. Copyright © 2012 Elsevier Ltd. All rights reserved.

PMID 22301053


Mechanism of the chromosome-induced polar body extrusion in mouse eggs

Cell Div. 2011 Aug 25;6:17. doi: 10.1186/1747-1028-6-17.

Wang Q, Racowsky C, Deng M. Source Department of Obstetrics and Gynecology and Reproductive Biology, 75 Francis Street, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA. Abstract BACKGROUND: An oocyte undergoes two rounds of asymmetric division to generate a haploid gamete and two small polar bodies designed for apoptosis. Chromosomes play important roles in specifying the asymmetric meiotic divisions in the oocytes but the underlying mechanism is poorly understood. RESULTS: Chromosomes independently induce spindle formation and cortical actomyosin assembly into special cap and ring structures in the cortex of the oocyte. The spindle and the cortical cap/ring interact to generate mechanical forces, leading to polar body extrusion. Two distinct force-driven membrane changes were observed during 2nd polar body extrusion: a protrusion of the cortical cap and a membrane invagination induced by an anaphase spindle midzone. The cortical cap protrusion and invagination help rotate the spindle perpendicularly so that the spindle midzone can induce bilateral furrows at the shoulder of the protruding cap, leading to an abscission of the polar body. It is interesting to note that while the mitotic spindle midzone induces bilateral furrowing, leading to efficient symmetric division in the zygote, the meiotic spindle midzone induced cytokinetic furrowing only locally. CONCLUSIONS: Distinct forces driving cortical cap protrusion and membrane invagination are involved in spindle rotation and polar body extrusion during meiosis II in mouse oocytes.

PMID 21867530

A 4D movie showing the DNA bead-induced ectopic polar body extrusion during meiosis II. The DNA beads (shown in blue) were injected at 10 O'clock position. Microtubules were labeled by microinjection of rhodamine conjugated tubulin. Note that the DNA bead spindle underwent rotation from horizontal to vertical position and the DNA beads were kept in the eggs after ectopic polar body extrusion. Maternal chromosomes are out of the z-sections.

The biology and dynamics of mammalian cortical granules

Reprod Biol Endocrinol. 2011 Nov 17;9:149.

Liu M. Source Department of Life Science and Graduate Institute of Biotechnology, Private Chinese Culture University, Taipei, Republic of China.


Cortical granules are membrane bound organelles located in the cortex of unfertilized oocytes. Following fertilization, cortical granules undergo exocytosis to release their contents into the perivitelline space. This secretory process, which is calcium dependent and SNARE protein-mediated pathway, is known as the cortical reaction. After exocytosis, the released cortical granule proteins are responsible for blocking polyspermy by modifying the oocytes' extracellular matrices, such as the zona pellucida in mammals. Mammalian cortical granules range in size from 0.2 um to 0.6 um in diameter and different from most other regulatory secretory organelles in that they are not renewed once released. These granules are only synthesized in female germ cells and transform an egg upon sperm entry; therefore, this unique cellular structure has inherent interest for our understanding of the biology of fertilization. Cortical granules are long thought to be static and awaiting in the cortex of unfertilized oocytes to be stimulated undergoing exocytosis upon gamete fusion. Not till recently, the dynamic nature of cortical granules is appreciated and understood. The latest studies of mammalian cortical granules document that this organelle is not only biochemically heterogeneous, but also displays complex distribution during oocyte development. Interestingly, some cortical granules undergo exocytosis prior to fertilization; and a number of granule components function beyond the time of fertilization in regulating embryonic cleavage and preimplantation development, demonstrating their functional significance in fertilization as well as early embryonic development. The following review will present studies that investigate the biology of cortical granules and will also discuss new findings that uncover the dynamic aspect of this organelle in mammals.

PMID 22088197

Predictive value of oocyte morphology in human IVF: a systematic review of the literature

Hum Reprod Update. 2011 Jan-Feb;17(1):34-45. Epub 2010 Jul 16.

Rienzi L, Vajta G, Ubaldi F. Source G.EN.E.R.A Centre for Reproductive Medicine, Clinica Valle Giulia, Via G. De Notaris 2, 00197 Rome, Italy. Abstract BACKGROUND: Non-invasive selection of developmentally competent human oocytes may increase the overall efficiency of human assisted reproduction and is regarded as crucial in countries where legal, social or religious factors restrict the production of supernumerary embryos. The purpose of this study was to summarize the predictive value for IVF success of morphological features of the oocyte that can be obtained by light or polarized microscopic investigations. METHODS: Studies about oocyte morphology and IVF/ICSI outcomes were identified by using a systematic literature search. RESULTS: Fifty relevant articles were identified: 33 analysed a single feature, 9 observed multiple features and investigated the effect of these features individually, 8 summarized the effect of individual features. Investigated structures were the following: meiotic spindle (15 papers), zona pellucida (15 papers), vacuoles or refractile bodies (14 papers), polar body shape (12 papers), oocyte shape (10 papers), dark cytoplasm or diffuse granulation (12 papers), perivitelline space (11 papers), central cytoplasmic granulation (8 papers), cumulus-oocyte complex (6 papers) and cytoplasm viscosity and membrane resistance characteristics (2 papers). None of these features were unanimously evaluated to have prognostic value for further developmental competence of oocytes. CONCLUSIONS: No clear tendency in recent publications to a general increase in predictive value of morphological features was found. These contradicting data underline the importance of more intensive and coordinated research to reach a consensus and fully exploit the predictive potential of morphological examination of human oocytes.

PMID 20639518 Figure 3 Metaphase II human oocyte observed using polarized light microscopy

CD9 tetraspanin generates fusion competent sites on the egg membrane for mammalian fertilization

Proc Natl Acad Sci U S A. 2011 Jun 20. [Epub ahead of print]

Jégou A, Ziyyat A, Barraud-Lange V, Perez E, Wolf JP, Pincet F, Gourier C. Source Laboratoire de Physique Statistique, Ecole Normale Supérieure de Paris, Université Pierre et Marie Curie, Université Paris Diderot, Centre National de la Recherche Scientifique, 24 rue Lhomond, 75005 Paris, France.


CD9 tetraspanin is the only egg membrane protein known to be essential for fertilization. To investigate its role, we have measured, on a unique acrosome reacted sperm brought in contact with an egg, the adhesion probability and strength with a sensitivity of a single molecule attachment. Probing the binding events at different locations of wild-type egg we described different modes of interaction. Here, we show that more gamete adhesion events occur on Cd9 null eggs but that the strongest interaction mode disappears. We propose that sperm-egg fusion is a direct consequence of CD9 controlled sperm-egg adhesion properties. CD9 generates adhesion sites responsible for the strongest of the observed gamete interaction. These strong adhesion sites impose, during the whole interaction lifetime, a tight proximity of the gamete membranes, which is a requirement for fusion to take place. The CD9-induced adhesion sites would be the actual location where fusion occurs.

PMID: 21690351

Defining ovarian reserve to better understand ovarian aging

Reprod Biol Endocrinol. 2011 Feb 7;9:23.

Gleicher N, Weghofer A, Barad DH. Source Center for Human Reproduction-New York, NY, USA.

Abstract Though a widely utilized term and clinical concept, ovarian reserve (OR) has been only inadequately defined. Based on Medline and PubMed searches we here define OR in its various components, review genetic control of OR, with special emphasis on the FMR1 gene, and discuss whether diminished OR (DOR) is treatable. What is generally referred to as OR reflects only a small portion of total OR (TOR), a pool of growing (recruited) follicles (GFs) at different stages of maturation. Functional OR (FOR) depends on size of the follicle pool at menarche and the follicle recruitment rate. Both vary between individuals and, at least partially, are under genetic control. The FMR1 gene plays a role in defining FOR at all ages. Infertility treatments have in the past almost exclusively only centered on the last two weeks of folliculogenesis, the gonadotropin-sensitive phase. Expansions of treatments into earlier stages of maturation will offer opportunity to significantly improve ovarian stimulation protocols, especially in women with DOR. Dehydroepiandrosterone (DHEA) may represent a first such intervention. Data generated in DHEA-supplemented women, indeed, suggest a new ovarian aging concept, based on aging of ovarian environments and not, as currently is believed, aging oocytes.

PMID: 21299886


Downregulation of both gene expression and activity of Hsp27 improved maturation of mouse oocyte in vitro

Liu JJ, Ma X, Cai LB, Cui YG, Liu JY. Reprod Biol Endocrinol. 2010 May 14;8:47.

BACKGROUND: Heat shock protein 27 (Hsp27), a member of the small heat shock protein family, is an apoptosis regulator. Our previous proteomic study showed that Hsp27 mainly expressed in human oocyte, and that Hsp27 expression was downregulated in the ovaries derived from women with the polycystic ovary syndrome (PCOS), a well known endocrinal disorder with abnormal apoptotic activity and folliculogenesis. However, the exact effects of Hsp27 downregulation on oocyte development have not yet been clarified.

CONCLUSIONS: Downregulation of Hsp27 improved the maturation of mouse oocytes, while increased early stage of apoptosis in oocytes by inducing the activation of extrinsic, caspase 8-mediated pathway.

PMID: 20465849

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

(good annexin and cleaved caspase immunofluorecence images)

Three-dimensional in vitro follicle growth: overview of culture models, biomaterials, design parameters and future directions

Reprod Biol Endocrinol. 2010 Oct 14;8(1):119.

Desai N, Alex A, Abdelhafez F, Calabro A, Goldfarb J, Fleischman A, Falcone T.

Abstract ABSTRACT: In vitro ovarian follicle culture is a new frontier in assisted reproductive technology with tremendous potential, especially for fertility preservation. Folliculogenesis within the ovary is a complex process requiring interaction between somatic cell components and the oocyte. Conventional two-dimensional culture on tissue culture substrata impedes spherical growth and preservation of the spatial arrangements between oocyte and surrounding granulosa cells. Granulosa cell attachment and migration can leave the oocyte naked and unable to complete the maturation process. Recognition of the importance of spatial arrangements between cells has spurred research in to three-dimensional culture system. Such systems may be vital when dealing with human primordial follicles that may require as long as three months in culture. In the present work we review pertinent aspects of in vitro follicle maturation, with an emphasis on tissue-engineering solutions for maintaining the follicular unit during the culture interval. We focus primarily on presenting the various 3-dimensional culture systems that have been applied for in vitro maturation of follicle:oocyte complexes. We also try to present an overview of outcomes with various biomaterials and animal models and also the limitations of the existing systems.

PMID 20946661

Loss of maternal ATRX results in centromere instability and aneuploidy in the mammalian oocyte and pre-implantation embryo

PLoS Genet. 2010 Sep 23;6(9). pii: e1001137.

Baumann C, Viveiros MM, De La Fuente R.

Female Germ Cell Biology Group, Department of Clinical Studies, University of Pennsylvania, Kennett Square, Pennsylvania, United States of America. Abstract

The α-thalassemia/mental retardation X-linked protein (ATRX) is a chromatin-remodeling factor known to regulate DNA methylation at repetitive sequences of the human genome. We have previously demonstrated that ATRX binds to pericentric heterochromatin domains in mouse oocytes at the metaphase II stage where it is involved in mediating chromosome alignment at the meiotic spindle. However, the role of ATRX in the functional differentiation of chromatin structure during meiosis is not known. To test ATRX function in the germ line, we developed an oocyte-specific transgenic RNAi knockdown mouse model. Our results demonstrate that ATRX is required for heterochromatin formation and maintenance of chromosome stability during meiosis. During prophase I arrest, ATRX is necessary to recruit the transcriptional regulator DAXX (death domain associated protein) to pericentric heterochromatin. At the metaphase II stage, transgenic ATRX-RNAi oocytes exhibit abnormal chromosome morphology associated with reduced phosphorylation of histone 3 at serine 10 as well as chromosome segregation defects leading to aneuploidy and severely reduced fertility. Notably, a large proportion of ATRX-depleted oocytes and 1-cell stage embryos exhibit chromosome fragments and centromeric DNA-containing micronuclei. Our results provide novel evidence indicating that ATRX is required for centromere stability and the epigenetic control of heterochromatin function during meiosis and the transition to the first mitosis.

PMID 20885787


Sperm chromatin-induced ectopic polar body extrusion in mouse eggs after ICSI and delayed egg activation

PLoS One. 2009 Sep 29;4(9):e7171. doi: 10.1371/journal.pone.0007171.

Deng M, Li R. Source Stowers Institute for Medical Research, Kansas City, Missouri, United States of America.


Meiotic chromosomes in an oocyte are not only a maternal genome carrier but also provide a positional signal to induce cortical polarization and define asymmetric meiotic division of the oocyte, resulting in polar body extrusion and haploidization of the maternal genome. The meiotic chromosomes play dual function in determination of meiosis: 1) organizing a bipolar spindle formation and 2) inducing cortical polarization and assembly of a distinct cortical cytoskeleton structure in the overlying cortex for polar body extrusion. At fertilization, a sperm brings exogenous paternal chromatin into the egg, which induces ectopic cortical polarization at the sperm entry site and leads to a cone formation, known as fertilization cone. Here we show that the sperm chromatin-induced fertilization cone formation is an abortive polar body extrusion due to lack of spindle induction by the sperm chromatin during fertilization. If experimentally manipulating the fertilization process to allow sperm chromatin to induce both cortical polarization and spindle formation, the fertilization cone can be converted into polar body extrusion. This suggests that sperm chromatin is also able to induce polar body extrusion, like its maternal counterpart. The usually observed cone formation instead of ectopic polar body extrusion induced by sperm chromatin during fertilization is due to special sperm chromatin compaction which restrains it from rapid spindle induction and therefore provides a protective mechanism to prevent a possible paternal genome loss during ectopic polar body extrusion.

PMID 19787051

Chromatin configurations in the germinal vesicle of mammalian oocytes

Mol Hum Reprod. 2009 Jan;15(1):1-9. Epub 2008 Nov 18.

Tan JH, Wang HL, Sun XS, Liu Y, Sui HS, Zhang J.

Laboratory for Animal Reproduction and Embryology, College of Animal Science and Veterinary Medicine, Shandong Agricultural University, Tai-an City, Shandong Province 271018, Peolple's Republic of China.

Abstract In all the studied mammalian species, chromatin in the germinal vesicle (GV) is initially decondensed with the nucleolus not surrounded by heterochromatin (the NSN configuration). During oocyte growth, the GV chromatin condenses into perinucleolar rings (the SN configuration) or other corresponding configurations with or without the perinucleolar rings, depending on species. During oocyte maturation, the GV chromatin is synchronized in a less condensed state before germinal vesicle breakdown (GVBD) in species that has been minutely studied. Oocytes may also take on a SN/corresponding configuration during early atresia, but they undergo GVBD at the advanced stage of atresia. As not all the species show the SN configuration while in all the species, gene transcription always stops at the late stage of oocyte growth, it is suggested that not the formation of perinucleolar rings but a thorough condensation of GV chromatin is essential for transcriptional repression. The GV chromatin configuration is highly correlated with oocyte competence; oocytes must end the NSN configuration before they gain the full meiotic competence, and they must take on the SN/corresponding configurations and stop gene transcription before they acquire the competence for early embryonic development. While factors inhibiting follicle atresia tend to synchronize oocytes in a chromatin configuration toward maturation, factors inducing follicle atresia tend to synchronize oocytes in a chromatin configuration reminiscent of early atresia. Furthermore, although condensation of GV chromatin is associated with transcriptional repression, both processes may not be associated with histone deacetylation during oocyte growth.

PMID 19019837


Meiosis in oocytes: predisposition to aneuploidy and its increased incidence with age

Hum Reprod Update. 2008 Mar-Apr;14(2):143-58. Epub 2007 Dec 14.

Jones KT. Source Institute for Cell and Molecular Biosciences, The Medical School, University of Newcastle, Framlington Place, Newcastle, NE2 4HH, UK. Abstract Mammalian oocytes begin meiosis in the fetal ovary, but only complete it when fertilized in the adult reproductive tract. This review examines the cell biology of this protracted process: from entry of primordial germ cells into meiosis to conception. The defining feature of meiosis is two consecutive cell divisions (meiosis I and II) and two cell cycle arrests: at the germinal vesicle (GV), dictyate stage of prophase I and at metaphase II. These arrests are spanned by three key events, the focus of this review: (i) passage from mitosis to GV arrest during fetal life, regulated by retinoic acid; (ii) passage through meiosis I and (iii) completion of meiosis II following fertilization, both meiotic divisions being regulated by cyclin-dependent kinase (CDK1) activity. Meiosis I in human oocytes is associated with an age-related high rate of chromosomal mis-segregation, such as trisomy 21 (Down's syndrome), resulting in aneuploid conceptuses. Although aneuploidy is likely to be multifactorial, oocytes from older women may be predisposed to be becoming aneuploid as a consequence of an age-long decline in the cohesive ties holding chromosomes together. Such loss goes undetected by the oocyte during meiosis I either because its ability to respond and block division also deteriorates with age, or as a consequence of being inherently unable to respond to the types of segregation defects induced by cohesion loss.

PMID 18084010


Germline stem cells and neo-oogenesis in the adult human ovary

Dev Biol. 2007 Jun 1;306(1):112-20. Epub 2007 Mar 12. Liu Y, Wu C, Lyu Q, Yang D, Albertini DF, Keefe DL, Liu L.

College of Life Sciences, Sun Yat-Sen University, Guangzhou 510275, China. Abstract It remains unclear whether neo-oogenesis occurs in postnatal ovaries of mammals, based on studies in mice. We thought to test whether adult human ovaries contain germline stem cells (GSCs) and undergo neo-oogenesis. Rather than using genetic manipulation which is unethical in humans, we took the approach of analyzing the expression of meiotic marker genes and genes for germ cell proliferation, which are required for neo-oogenesis, in adult human ovaries covering an age range from 28 to 53 years old, compared to testis and fetal ovaries served as positive controls. We show that active meiosis, neo-oogenesis and GSCs are unlikely to exist in normal, adult, human ovaries. No early meiotic-specific or oogenesis-associated mRNAs for SPO11, PRDM9, SCP1, TERT and NOBOX were detectable in adult human ovaries using RT-PCR, compared to fetal ovary and adult testis controls. These findings are further corroborated by the absence of early meiocytes and proliferating germ cells in adult human ovarian cortex probed with markers for meiosis (SCP3), oogonium (OCT3/4, c-KIT), and cell cycle progression (Ki-67, PCNA), in contrast to fetal ovary controls. If postnatal oogenesis is confirmed in mice, then this species would represent an exception to the rule that neo-oogenesis does not occur in adults.

PMID 17428461

How eggs arrest at metaphase II: MPF stabilisation plus APC/C inhibition equals Cytostatic Factor

Balbiani body

  • (Henneguy, 1887) is a cytoplasmic structure present in young immature oocytes in mammals and many other species
  • transient structure composed of a large number of mitochondria, Golgi complexes, endoplasmic reticulum.
  • ill-defined electron-dense granulofibrillar material that congregates at a perinuclear location
  • as the oocyte matures it moves to a peripheral location in the oocyte cortex (in some species corresponding to the future vegetal pole) and disperses (reviewed by Kloc et al., 2004).


  • in some species contribute to the assembly and transportation of the germ plasm determinants to the cortex of the mature oocyte
  • In mammals ( “inductive”, “regulative” or “epigenetic” mode of germ cell formation) do not have germ plasm determinants, the function remains largely obscure may include
    • the controlled amplification of mitochondria or the creation of an asymmetry or polarity in the oocyte.


Henneguy F. (1887). Note sur la vesicle de Balbiani. C. R. Hebd. Seances Soc. Biol. Ses. Fil. 39, 69.

Kloc M., Bilinski S. and Etkin L.D. (2004). The Balbiani body and germ cell determinants: 150 years later. Curr. Top. Dev. Biol. 59, 1-36.


Egg coat proteins activate calcium entry into mouse sperm via CATSPER channels

Xia J, Ren D. Biol Reprod. 2009 Jun;80(6):1092-8. Epub 2009 Feb 11. PMID 19211808

ZP2 and ZP3 traffic independently within oocytes prior to assembly into the extracellular zona pellucida

Hoodbhoy T, Avilés M, Baibakov B, Epifano O, Jiménez-Movilla M, Gauthier L, Dean J. Mol Cell Biol. 2006 Nov;26(21):7991-8. PMID 17047254