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Cite this page: Hill, M.A. (2024, June 21) Embryology Fertilization. Retrieved from


Acrosin is essential for sperm penetration through the zona pellucida in hamsters PNAS February 4, 2020 117 (5) 2513-2518

Mammalian oocytes are surrounded by the zona pellucida, a glycoprotein coat that protects the oocyte and embryo from mechanical damage during their preimplantation development within the oviduct. Fertilizing spermatozoa must penetrate the zona, but we do not know the exact mechanisms underlying this process. Sperm proteases were thought to work as zona lysins, but gene-knockout studies in mice did not support this assumption. In this study, we generated hamsters without acrosin, the major acrosomal protease, to examine its role in both in vivo and in vitro fertilization. Surprisingly, mutant male hamsters were completely infertile because their spermatozoa were unable to penetrate the zona. We thus demonstrated that, at least in hamsters, acrosin is essential for sperm penetration through the zona.


Male Contraceptive Development: Update on Novel Hormonal and Nonhormonal Methods

Long JE1, Lee MS2, Blithe DL2. Author information Abstract BACKGROUND: Development of new methods of male contraception would address an unmet need for men to control their fertility and could increase contraceptive options for women. Pharmaceutical research and development for male contraception was active in the 1990s but has been virtually abandoned. The Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD) has supported a contraceptive development program since 1969 and supports the majority of hormonal male contraceptive development. Nonhormonal methods are also in development but are at earlier stages. CONTENT: Several hormonal male contraceptive agents have entered clinical trials. Single-agent products being evaluated include dimethandrolone undecanoate, 11β-methyl-nortestosterone dodecyl carbonate, and 7α-methyl-19-nortestosterone. A contraceptive efficacy trial of Nestorone® gel and testosterone gel in a single application will begin in 2018. Potential nonhormonal methods are at preclinical stages of development. Many nonhormonal male contraceptive targets that affect either sperm production or sperm function have been identified. Targeted pathways include the retinoic acid pathway, bromodomain and extraterminal proteins, and pathways for Sertoli cell-germ cell adhesion or sperm motility. Druggable targets include CatSper, the sperm Na+/K+-exchanger, TSSK, HIPK4, EPPIN, and ADAMs family proteins. Development of a procedure to reversibly block the vas deferens (initially developed in India in the 1980s) is undergoing early stage research in the US under the trade name Vasalgel™. SUMMARY: NICHD has supported the development of reversible male contraceptive agents. Other organizations such as the World Health Organization and the Population Council are pursuing male contraceptive development, but industry involvement remains dormant. © 2018 American Association for Clinical Chemistry.

Long JE, Lee MS & Blithe DL. (2019). Male Contraceptive Development: Update on Novel Hormonal and Nonhormonal Methods. Clin. Chem. , 65, 153-160. PMID: 30602479 DOI.


Harper JC, Aittomäki K, Borry P, Cornel MC, de Wert G, Dondorp W, Geraedts J, Gianaroli L, Ketterson K, Liebaers I, Lundin K, Mertes H, Morris M, Pennings G, Sermon K, Spits C, Soini S, van Montfoort APA, Veiga A, Vermeesch JR, Viville S & Macek M. (2018). Recent developments in genetics and medically assisted reproduction: from research to clinical applications. Eur. J. Hum. Genet. , 26, 12-33. PMID: 29199274 DOI.

Selected genes in Male Infertility
Gene abbreviation Name Gene Location Online Mendelian
Inheritance in Man (OMIM)
HUGO Gene Nomenclature
Committee (HGNC)
GeneCards (GCID) Diagnosis
AURKC Aurora kinase C 19q13.43 603495 11391 GC19P057230 Macrozoospermia
CATSPER1 Cation channel sperm-associated 1 11q13.1 606389 17116 GC11M066034 Asthenozoospermia
CFTR Cystic fibrosis transmembrane conductance regulator 7q31.2 602421 1884 GC07P117465 Obstructive azoospermia
DNAH1 Dynein axonemal heavy chain 1 3p21.1 603332 2940 GC03P052350 Asthenozoospermia
DPY19L2 Dpy-19-like 2 gene 12q14.2 613893 19414 GC12M063558 Globozoospermia
GALNTL5 Polypeptide N-acetylgalactosaminyltransferase-like 5 7q36.1 615133 21725 GC07P151956 Asthenozoospermia
MAGEB4 MAGE family member B4 Xp21.2 300153 6811 GC0XP030260 Azoospermia
NANOS1 Nanos C2HC-type zinc finger 1 10q26.11 608226 23044 GC10P119029 Azoospermia
NR0B1 Nuclear receptor subfamily 0 group B member 1 Xp21.2 300473 7960 GC0XM030322 Azoospermia
NR5A1 Nuclear receptor subfamily 5 group A member 1 9q33.3 184757 7983 GC09M124481 Azoospermia
SOHLH1 Spermatogenesis and oogenesis-specific basic helix–loop–helix 1 9q34.3 610224 27845 C09M135693 Azoospermia
vSPATA16 Spermatogenesis-associated 16 3q26.31 609856 29935 GC03M172889 Globozoospermia
SYCE1 Synaptonemal complex central element protein 1 10q26.3 611486 28852 GC10M133553 Azoospermia
TAF4B TATA-box binding protein-associated factor 4b 18q11.2 601689 11538 GC18P026225 Azoospermia
TEX11 Testis expressed 11 Xq13.1 300311 11733 GC0XM070528 Azoospermia
TEX15 Testis expressed 15, meiosis and synapsis associated 8p12 605795 11738 GC08M030808 Azoospermia
WT1 Wilms tumour 1 8p12 607102 12796 GC11M032365 Azoospermia
ZMYND15 Zinc-finger MYND-type containing 15 17p13.2 614312 20997 GC17P004740 Azoospermia
  Table data source[1] (table 1)    Links: fertilization | spermatozoa | testis | Male Infertility Genes | Female Infertility Genes | oocyte | ovary | Genetic Abnormalities | ART

  Asthenozoospermia - (asthenospermia) term for reduced spermatozoa motility. Azoospermia - term for no spermatozoa located in the ejaculate. Globozoospermia - term for spermatozoa with a round head and no acrosome.

Selected Female Infertility Genes
Gene abbreviation Name Gene Location Online Mendelian
Inheritance in Man (OMIM)
HUGO Gene Nomenclature
Committee (HGNC)
GeneCards (GCID) Diagnosis
BMP15 Bone morphogenetic protein 15 Xp11.22 300247 1068 GC0XP050910 Primary ovarian insufficiency
CLPP Caseinolytic mitochondrial matrix peptidase proteolytic subunit 19p13.3 601119 2084 GC19P006369 Primary ovarian insufficiency
EIF2B2 Eukaryotic translation initiation factor 2B subunit beta 14q24.3 606454 3258 GC14P075002 Primary ovarian insufficiency
FIGLA Folliculogenesis-specific BHLH transcription factor 2p13.3 608697 24669 GC02M070741 Primary ovarian insufficiency
FMR1 Fragile X mental retardation 1 Xq27.3 309550 3775 GC0XP147912 Primary ovarian insufficiency
FOXL2 Forkhead box L2 3q22.3 605597 1092 GC03M138944 Primary ovarian insufficiency
FSHR Follicle stimulating hormone receptor 2p16.3 136435 3969 GC02M048866 Primary ovarian insufficiency
GALT Galactose-1-phosphate uridylyltransferase 9p13.3 606999 4135 GC09P034636 Primary ovarian insufficiency
GFD9 Growth differentiation factor 9 5q31.1 601918 4224 GC05M132861 Primary ovarian insufficiency
HARS2 Histidyl-TRNA synthetase 2, mitochondrial 5q31.3 600783 4817 GC05P141975 Primary ovarian insufficiency
HFM1 HFM1, ATP-dependent DNA helicase homolog 1p22.2 615684 20193 GC01M091260 Primary ovarian insufficiency
HSD17B4 Hydroxysteroid 17-beta dehydrogenase 4 5q23.1 601860 5213 GC05P119452 Primary ovarian insufficiency
LARS2 Leucyl-TRNA synthetase 2, mitochondrial 3p21.31 604544 17095 GC03P045405 Primary ovarian insufficiency
LHCGR Luteinizing hormone/choriogonadotropin receptor 2p16.3 152790 6585 GC02M048647 Primary ovarian insufficiency
LHX8 LIM homeobox 8 1p31.1 604425 28838 GC01P075128 Primary ovarian insufficiency
MCM8 Minichromosome maintenance 8 homologous recombination repair factor 20p12.3 608187 16147 GC20P005926 Primary ovarian insufficiency
MCM9 Minichromosome maintenance 9 homologous recombination repair factor 6q22.31 610098 21484 GC06M118813 Primary ovarian insufficiency
NOBOX NOBOX oogenesis homeobox 7q35 610934 22448 GC07M144397 Primary ovarian insufficiency
NOG Noggin 17q22 602991 7866 GC17P056593 Primary ovarian insufficiency
PMM2 Phosphomannomutase 2 16p13.2 601785 9115 GC16P008788 Primary ovarian insufficiency
POLG DNA polymerase gamma, catalytic subunit 15q26.1 174763 9179 GC15M089316 Primary ovarian insufficiency
REC8 REC8 meiotic recombination protein 14q12 608193 16879 GC14P024171 Primary ovarian insufficiency
SMC1B Structural maintenance of chromosomes 1B 22q13.31 608685 11112 GC22M045344 Primary ovarian insufficiency
SOHLH1 Spermatogenesis and oogenesis-specific basic helix–loop–helix 1 9q34.3 610224 27845 GC09M135693 Primary ovarian insufficiency
STAG3 Stromal antigen 3 7q22.1 {{Chr 608489 11356 GC07P100177 Primary ovarian insufficiency
SYCE1 Synaptonemal Complex Central Element Protein 1 10q26.3 611486 28852 GC10M133553 Primary ovarian insufficiency
TLE6 Transducin-like enhancer of split 6 19p13.3 612399 30788 GC19P002976 Embryonic lethalithy
TUBB8 Tubulin beta 8 Class VIII 10p15.3 616768 20773 GC10M000048 Oocyte maturation arrest
TWNK Twinkle MtDNA helicase 10q24.31 606075 1160 GC10P100991 Primary ovarian insufficiency
  Table data source[1] (table 1)    Links: fertilization | oocyte | ovary | | Female Infertility Genes | spermatozoa | testis | Male Infertility Genes | Genetic Abnormalities | ART

 Primary ovarian insufficiency - depletion or dysfunction of ovarian follicles with cessation of menses before age 40 years.
 Oocyte maturation arrest - arrest of human oocytes may occur at different stages of meiosis.

Oocyte-specific deletion of Gsα induces oxidative stress and deteriorates oocyte quality in mice

Exp Cell Res. 2018 Jul 16. pii: S0014-4827(18)30472-5. doi: 10.1016/j.yexcr.2018.07.023. [Epub ahead of print]

Xie Y1, Wu B2, Jin Y1, Zhang A1, Sun X1, Zhang X1, Gao X1, Dong R1, Li H3, Gao J4.


The stimulatory heterotrimeric Gs protein alpha subunit (Gsα) is a ubiquitous guanine nucleotide-binding protein that regulates the intracellular cAMP signaling pathway and consequently participates in a wide range of biological events. In the reproductive system, despite Gsα being associated with oocyte meiotic arrest in vitro, the exact role of Gsα in female fertility in vivo remains largely unknown. Here, we generated oocyte-specific Gsα knockout mice by using the Cre/LoxP system. We observed that the deletion of Gsα caused complete female infertility. Exclusion of post-implantation abnormalities, oogenesis, fertilization, and early embryo development was subsequently monitored; meiosis in Gsα-deficient oocytes precociously resumed in only 43% of antral follicles from mutant mice, indicating that alteration of meiotic pause was not the key factor in infertility. Ovulation process and number were normal, but the rate of morphological abnormal oocytes was apparently increased; spindle organization, fertilization, and early embryo development were impaired. Furthermore, the level of ROS (reactive oxygen species) and the mitochondrial aggregation increased, and antioxidant glutathione (GSH) content, ATP level, mtDNA copy number, and mitochondrial membrane potential decreased in Gsα-deficient oocytes. GV oocytes from mutant mice showed early-stage apoptosis. Meanwhile, the Gsα knockout-induced decline in oocyte quality and low developmental potential was partially rescued by antioxidant supplementation. To sum up, our results are the first to reveal that the profile of Gsα oocyte-specific deletion caused female infertility in vivo, and oxidative stress plays an important role in this event. KEYWORDS: G(s)α; conditional knockout; infertility; mitochondria; mouse oocyte; oxidative stress PMID: 30026030 DOI: 10.1016/j.yexcr.2018.07.023

Zinc sparks induce physiochemical changes in the egg zona pellucida that prevent polyspermy

Integr Biol (Camb). 2017 Feb 20;9(2):135-144. doi: 10.1039/c6ib00212a.

Que EL1, Duncan FE2, Bayer AR3, Philips SJ4, Roth EW5, Bleher R5, Gleber SC6, Vogt S6, Woodruff TK7, O'Halloran TV8. Author information

Abstract During fertilization or chemically-induced egg activation, the mouse egg releases billions of zinc atoms in brief bursts known as 'zinc sparks.' The zona pellucida (ZP), a glycoprotein matrix surrounding the egg, is the first structure zinc ions encounter as they diffuse away from the plasma membrane. Following fertilization, the ZP undergoes changes described as 'hardening', which prevent multiple sperm from fertilizing the egg and thereby establish a block to polyspermy. A major event in zona hardening is cleavage of ZP2 proteins by ovastacin; however, the overall physiochemical changes contributing to zona hardening are not well understood. Using X-ray fluorescence microscopy, transmission and scanning electron microscopy, and biological function assays, we tested the hypothesis that zinc release contributes to ZP hardening. We found that the zinc content in the ZP increases by 300% following activation and that zinc exposure modulates the architecture of the ZP matrix. Importantly, zinc-induced structural changes of the ZP have a direct biological consequence; namely, they reduce the ability of sperm to bind to the ZP. These results provide a paradigm-shifting model in which fertilization-induced zinc sparks contribute to the polyspermy block by altering conformations of the ZP matrix. This adds a previously unrecognized factor, namely zinc, to the process of ZP hardening. PMID: 28102396 PMCID: PMC5439353 DOI: 10.1039/c6ib00212a


A Specific Transitory Increase in Intracellular Calcium Induced by Progesterone Promotes Acrosomal Exocytosis in Mouse Sperm

Biol Reprod. 2016 Jan 27. pii: biolreprod.115.136085. [Epub ahead of print]

Romarowski A, Sánchez-Cárdenas C, Ramírez-Gómez HV, Puga Molina LD, Treviño CL, Hernández Cruz A, Darszon AI, Buffone MG.


During capacitation, sperm acquire the ability to undergo the acrosome reaction (AR), an essential step in fertilization. Progesterone produced by cumulus cells has been associated with various physiological processes in sperm, including stimulation of AR. An increase in intracellular Ca2+ ([Ca2+]i) is necessary for AR to occur. In this study, we investigated the spatio-temporal correlation between the changes in ([Ca2+]i and AR in single mouse spermatozoa in response to Progesterone. We found that Progesterone stimulates an ([Ca2+]i increase in five different patterns: gradual increase, oscillatory, late transitory, immediate transitory and sustained. We also observed that the ([Ca2+]i increase promoted by Progesterone starts at either the flagellum or the head. We validated the use of FM4-64 as an indicator for the occurrence of the AR by simultaneously detecting its fluorescence increase and the loss of EGFP in transgenic EGFPAcr sperm. For the first time, we have simultaneously visualized the rise in ([Ca2+]i and the process of exocytosis in response to Progesterone and found that only a specific transitory increase in ([Ca2+]i originated in the sperm head promotes the initiation of AR. Copyright 2016 by The Society for the Study of Reproduction. KEYWORDS: Acrosome; Acrosome reaction; Gamete biology; Progesterone/Progesterone receptor; Sperm

PMID 26819478


Binding of sperm protein Izumo1 and its egg receptor Juno drives Cd9 accumulation in the intercellular contact area prior to fusion during mammalian fertilization

Development. 2014 Oct;141(19):3732-9. doi: 10.1242/dev.111534. Epub 2014 Sep 10.

Chalbi M1, Barraud-Lange V2, Ravaux B1, Howan K1, Rodriguez N3, Soule P3, Ndzoudi A2, Boucheix C4, Rubinstein E4, Wolf JP2, Ziyyat A2, Perez E1, Pincet F1, Gourier C5.


Little is known about the molecular mechanisms that induce gamete fusion during mammalian fertilization. After initial contact, adhesion between gametes only leads to fusion in the presence of three membrane proteins that are necessary, but insufficient, for fusion: Izumo1 on sperm, its receptor Juno on egg and Cd9 on egg. What happens during this adhesion phase is a crucial issue. Here, we demonstrate that the intercellular adhesion that Izumo1 creates with Juno is conserved in mouse and human eggs. We show that, along with Izumo1, egg Cd9 concomitantly accumulates in the adhesion area. Without egg Cd9, the recruitment kinetics of Izumo1 are accelerated. Our results suggest that this process is conserved across species, as the adhesion partners, Izumo1 and its receptor, are interchangeable between mouse and human. Our findings suggest that Cd9 is a partner of Juno, and these discoveries allow us to propose a new model of the molecular mechanisms leading to gamete fusion, in which the adhesion-induced membrane organization assembles all key players of the fusion machinery. © 2014. Published by The Company of Biologists Ltd. KEYWORDS: CD9; Fertilization; Izumo1; Juno; Membrane dynamics; Membrane organization

PMID 25209248

Versatile action of picomolar gradients of progesterone on different sperm subpopulations

PLoS One. 2014 Mar 10;9(3):e91181. doi: 10.1371/journal.pone.0091181. eCollection 2014.

Uñates DR, Guidobaldi HA, Gatica LV, Cubilla MA, Teves ME, Moreno A, Giojalas LC.


High step concentrations of progesterone may stimulate various sperm physiological processes, such as priming and the acrosome reaction. However, approaching the egg, spermatozoa face increasing concentrations of the hormone, as it is secreted by the cumulus cells and then passively diffuses along the cumulus matrix and beyond. In this context, several questions arise: are spermatozoa sensitive to the steroid gradients as they undergo priming and the acrosome reaction? If so, what are the functional gradual concentrations of progesterone? Do spermatozoa in different physiological states respond differentially to steroid gradients? To answer these questions, spermatozoa were confronted with progesterone gradients generated by different hormone concentrations (1 pM to 100 µM). Brief exposure to a 10 pM progesterone gradient stimulated priming for the acrosome reaction in one sperm subpopulation, and simultaneously induced the acrosome reaction in a different sperm subpopulation. This effect was not observed in non-capacitated cells or when progesterone was homogeneously distributed. The results suggest a versatile role of the gradual distribution of very low doses of progesterone, which selectively stimulate the priming and the acrosome reaction in different sperm subpopulations.

PMID 24614230

Juno is the egg Izumo receptor and is essential for mammalian fertilization

Fertilization occurs when sperm and egg recognize each other and fuse to form a new, genetically distinct organism. The molecular basis of sperm–egg recognition is unknown, but is likely to require interactions between receptor proteins displayed on their surface. Izumo1 is an essential sperm cell-surface protein, but its receptor on the egg has not been described. Here we identify folate receptor 4 (Folr4) as the receptor for Izumo1 on the mouse egg, and propose to rename it Juno. We show that the Izumo1–Juno interaction is conserved within several mammalian species, including humans. Female mice lacking Juno are infertile and Juno-deficient eggs do not fuse with normal sperm. Rapid shedding of Juno from the oolemma after fertilization suggests a mechanism for the membrane block to polyspermy, ensuring eggs normally fuse with just a single sperm. Our discovery of an essential receptor pair at the nexus of conception provides opportunities for the rational development of new fertility treatments and contraceptives.


Roles of the oviduct in mammalian fertilization

Reproduction. 2012 Dec;144(6):649-60. doi: 10.1530/REP-12-0279. Epub 2012 Oct 1.

Coy P, García-Vázquez FA, Visconti PE, Avilés M. Source Department of Physiology, Faculty of Veterinary, University of Murcia, Campus Mare Nostrum, Campus de Espinardo, Murcia 30071, Spain.


The oviduct or Fallopian tube is the anatomical region where every new life begins in mammalian species. After a long journey, the spermatozoa meet the oocyte in the specific site of the oviduct named ampulla and fertilization takes place. The successful fertilization depends on several biological processes that occur in the oviduct some hours before this rendezvous and affect both gametes. Estrogen and progesterone, released from the ovary, orchestrate a series of changes by genomic and nongenomic pathways in the oviductal epithelium affecting gene expression, proteome, and secretion of its cells into the fluid bathing the oviductal lumen. In addition, new regulatory molecules are being discovered playing important roles in oviductal physiology and fertilization. The present review tries to describe these processes, building a comprehensive map of the physiology of the oviduct, to better understand the importance of this organ in reproduction. With this purpose, gamete transport, sperm and oocyte changes in the oviductal environment, and other interactions between gametes and oviduct are discussed in light of recent publications in the field.

PMID 23028122

Live cell imaging of the oviduct

Methods Enzymol. 2012;506:415-23. doi: 10.1016/B978-0-12-391856-7.00045-7.

Kölle S. Source Department of Urology, Ludwig Maximilians University of Munich, Munich, Germany.


In the oviduct, the integrity of oocyte and sperm transport, fertilization, and early embryonic ontogenesis is essential for successful reproduction. Up to now, most of the knowledge on oocyte and sperm transport, gamete interaction and embryonic development has in most cases been gained exclusively by in vitro studies. In addition, especially the mechanisms of gameto-maternal interaction and embryo-maternal communication in the oviduct are still unknown. Recent techniques of live cell imaging and digital videomicroscopy allow for the first time to provide actual new insights in the mechanisms of sperm transport, sperm storage, oocyte transport, fertilization, gameto-maternal interaction and embryo-maternal crosstalk under near in vivo conditions. Detailed knowledge of these important events in the oviduct is the prerequisite to develop new therapeutic concepts for subfertility and infertility and to increase the success rates of the actual techniques of assisted reproduction (ART). Additionally the effects of drugs and hormones used in ART can be effectively studied using a functional oviductal epithelium. The guidelines for live cell imaging in the oviduct presented here should enable researches to establish a functional digital analysis system which allows to study physiological and pathological events in the oviduct under near in vivo conditions. Copyright © 2012 Elsevier Inc. All rights reserved.

PMID 22341236

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

Ciliary activity in the oviduct of cycling, pregnant, and muscarinic receptor knockout mice

Biol Reprod. 2012 Apr 19;86(4):120. doi: 10.1095/biolreprod.111.096339. Print 2012 Apr.

Noreikat K, Wolff M, Kummer W, Kölle S. Source Institute of Veterinary Anatomy, Histology, and Embryology, Justus-Liebig University, Giessen, Germany.


The transport of the oocyte and the embryo in the oviduct is managed by ciliary beating and muscular contractions. Because nonneuronally produced acetylcholine influences ciliary beating in the trachea via the muscarinic receptors M2 and M3, we supposed that components of the cholinergic system may also modulate ciliary activity in the oviduct. To address this issue, we analyzed the expression profile of muscarinic receptors (CHRMs) in the murine oviduct by RT-PCR and assessed ciliary beat frequency (CBF) and cilia-driven particle transport speed (PTS) on the mucosal surface of opened oviductal segments in correlation with histomorphological investigations. RT-PCR of laser-assisted microdissected epithelium revealed expression of Chrm subtypes Chrm1 and Chrm3. In opened isthmic segments, particle transport was barely seen, correlating with a significantly lower number of ciliated cells compared to the ampulla. In the ampulla, basal PTS and CBF were high (71 μm/sec and 21 Hz, respectively) both in cycling and pregnant wild-type mice and in mice with targeted deletion of the Chrm genes Chrm1, Chrm3, Chrm4, and Chrm5. In contrast to the trachea, where basal ciliary activity was low and largely enhanced by muscarinic stimulation, muscarinic agonists and antagonists did not affect the high ampullar PTS. Our results imply that this high oviductal autonomous ciliary activity is independent from the intrinsic cholinergic system and serves to maintain optimal clearance of the tube throughout all stages of the estrous cycle and early pregnancy.

PMID 22302687

The relative contributions of propulsive forces and receptor-ligand binding forces during early contact between spermatozoa and zona pellucida of oocytes

J Theor Biol. 2012 Feb 7;294:139-43. Epub 2011 Nov 15.

Kozlovsky P, Gefen A. Source Department of Biomedical Engineering, Faculty of Engineering, Tel Aviv University, Tel Aviv 69978, Israel.


When reaching the zona pellucida (ZP) of the oocyte, spermatozoa apply propulsive forces produced by the motion of their flagella, which push against the ZP and theoretically should contribute to their penetration into the ZP. Additionally, specific receptors on the spermatozoon head bind to ZP3 ligands located on the surface of the ZP, which locks the sperm's head onto the oocyte. Both mechanisms are important cofactors in the initial sperm penetration into the ZP, which is required for successful fertilization of the oocyte, but it is unclear which forces-mechanical thrust or biochemical binding-are more influential at this stage. To address this question, we developed a biomechanical sperm-oocyte contact model, which is based on the Johnson-Kendall-Roberts model adopted from the contact mechanics theory. The modeling predicted that during the early stage of penetration into the ZP, biochemical binding forces acting on spermatozoa, which are swimming at a (normal) velocity of 100μm/s are ∼4.2-times to ∼16.7-times less than the mechanically-generated propulsive forces. In a simulated pathology of a low number of properly functioning receptors (50 out of 300receptors/μm(2)), the biochemical binding forces are ∼63-times less than the propulsive forces for the normally swimming sperm. It is suggested that such dominance of the propulsive forces over the biochemical binding forces can prevent efficient binding of spermatozoa to the ZP of the oocyte due to continuous movement of the sperm (which is not necessarily perpendicular to the ZP surface, and can cause sliding of sperm over the ZP). Thus, our theoretical analysis indicates that a sufficiently large density of receptors to ZP3 ligands on the sperm head is critical at the stage of early sperm-oocyte contact, in order to allow an efficient acrosome reaction to follow, so that the spermatozoon can start penetrating into the ZP. Copyright © 2011 Elsevier Ltd. All rights reserved.

PMID 22100500

The nerve of ovulation-inducing factor in semen

Proc Natl Acad Sci U S A. 2012 Sep 11;109(37):15042-7. Epub 2012 Aug 20.

Ratto MH, Leduc YA, Valderrama XP, van Straaten KE, Delbaere LT, Pierson RA, Adams GP. Source Faculty of Veterinary Sciences and Faculty of Agricultural Sciences, Universidad Austral de Chile, Valdivia, Chile.


A component in seminal fluid elicits an ovulatory response and has been discovered in every species examined thus far. The existence of an ovulation-inducing factor (OIF) in seminal plasma has broad implications and evokes questions about identity, tissue sources, mechanism of action, role among species, and clinical relevance in infertility. Most of these questions remain unanswered. The goal of this study was to determine the identity of OIF in support of the hypothesis that it is a single distinct and widely conserved entity. Seminal plasma from llamas and bulls was used as representative of induced and spontaneous ovulators, respectively. A fraction isolated from llama seminal plasma by column chromatography was identified as OIF by eliciting luteinizing hormone (LH) release and ovulation in llamas. MALDI-TOF revealed a molecular mass of 13,221 Da, and 12-23 aa sequences of OIF had homology with human, porcine, bovine, and murine sequences of β nerve growth factor (β-NGF). X-ray diffraction data were used to solve the full sequence and structure of OIF as β-NGF. Neurite development and up-regulation of trkA in phaeochromocytoma (PC(12)) cells in vitro confirmed NGF-like properties of OIF. Western blot analysis of llama and bull seminal plasma confirmed immunorecognition of OIF using polyclonal mouse anti-NGF, and administration of β-NGF from mouse submandibular glands induced ovulation in llamas. We conclude that OIF in seminal plasma is β-NGF and that it is highly conserved. An endocrine route of action of NGF elucidates a previously unknown pathway for the direct influence of the male on the hypothalamo-pituitary-gonadal axis of the inseminated female.

PMID 22908303


Gas6 downregulation impaired cytoplasmic maturation and pronuclear formation independent to the MPF activity

PLoS One. 2011;6(8):e23304. Epub 2011 Aug 5.

Kim KH, Kim EY, Kim Y, Kim E, Lee HS, Yoon SY, Lee KA. Source Department of Biomedical Science, College of Life Science, Fertility Center, CHA Research Institute, CHA University, CHA General Hospital, Seoul, Korea.


Previously, we found that the growth arrest-specific gene 6 (Gas6) is more highly expressed in germinal vesicle (GV) oocytes than in metaphase II (MII) oocytes using annealing control primer (ACP)-PCR technology. The current study was undertaken to investigate the role of Gas6 in oocyte maturation and fertilization using RNA interference (RNAi). Interestingly, despite the specific and marked decrease in Gas6 mRNA and protein expression in GVs after Gas6 RNAi, nuclear maturation including spindle structures and chromosome segregation was not affected. The only discernible effect induced by Gas6 RNAi was a change in maturation promoting factor (MPF) activity. After parthenogenetic activation, Gas6 RNAi-treated oocytes at the MII stage had not developed further and arrested at MII (90.0%). After stimulation with Sr(2+), Gas6-silenced MII oocytes had markedly reduced Ca(2+) oscillation and exhibited no exocytosis of cortical granules. In these oocytes, sperm penetration occurred during fertilization but not pronucleus (PN) formation. By roscovitine and colcemid treatment, we found that the Gas6 knockdown affected cytoplasmic maturation directly, independent to the changed MPF activity. These results strongly suggest that 1) the Gas6 signaling itself is important to the cytoplasmic maturation, but not nuclear maturation, and 2) the decreased Gas6 expression and decreased MPF activity separately or mutually influence sperm head decondensation and PN formation.

PMID 21850267

Non-genetic contributions of the sperm nucleus to embryonic development

Asian J Androl. 2011 Jan;13(1):31-5. Epub 2010 Oct 18. Yamauchi Y, Shaman JA, Ward WS. Source Department Anatomy and Reproductive Biology, Institute for Biogenesis Research, John A. Burns School of Medicine, University of Hawaii at Manoa, Honolulu, HI, USA.


Recent data from several laboratories have provided evidence that the newly fertilized oocyte inherits epigenetic signals from the sperm chromatin that are required for proper embryonic development. For the purposes of this review, the term epigenetic is used to describe all types of molecular information that are transmitted from the sperm cell to the embryo. There are at least six different forms of epigenetic information that have already been established as being required for proper embryogenesis in mammals or for which there is evidence that it may do so. These are (i) DNA methylation; (ii) sperm-specific histones, (iii) other chromatin-associated proteins; (iv) the perinuclear theca proteins; (v) sperm-born RNAs and, the focus of this review; and (vi) the DNA loop domain organization by the sperm nuclear matrix. These epigenetic signals should be considered when designing protocols for the manipulation and cryopreservation of spermatozoa for assisted reproductive technology as necessary components for effective fertilization and subsequent embryo development.

PMID 20953203

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

Most fertilizing mouse spermatozoa begin their acrosome reaction before contact with the zona pellucida during in vitro fertilization

Proc Natl Acad Sci U S A. 2011 Mar 22;108(12):4892-6. Epub 2011 Mar 7.

Jin M, Fujiwara E, Kakiuchi Y, Okabe M, Satouh Y, Baba SA, Chiba K, Hirohashi N.

Department of Biological Sciences, Department of Genetic Counseling, and Glycoscience Institute, Ochanomizu University, Tokyo 112-8610, Japan. Abstract To fuse with oocytes, spermatozoa of eutherian mammals must pass through extracellular coats, the cumulus cell layer, and the zona pellucida (ZP). It is generally believed that the acrosome reaction (AR) of spermatozoa, essential for zona penetration and fusion with oocytes, is triggered by sperm contact with the zona pellucida. Therefore, in most previous studies of sperm-oocyte interactions in the mouse, the cumulus has been removed before insemination to facilitate the examination of sperm-zona interactions. We used transgenic mouse spermatozoa, which enabled us to detect the onset of the acrosome reaction using fluorescence microscopy. We found that the spermatozoa that began the acrosome reaction before reaching the zona were able to penetrate the zona and fused with the oocyte's plasma membrane. In fact, most fertilizing spermatozoa underwent the acrosome reaction before reaching the zona pellucida of cumulus-enclosed oocytes, at least under the experimental conditions we used. The incidence of in vitro fertilization of cumulus-free oocytes was increased by coincubating oocytes with cumulus cells, suggesting an important role for cumulus cells and their matrix in natural fertilization.

PMID 21383182

Sperm Proteasomes Degrade Sperm Receptor on the Egg Zona Pellucida during Mammalian Fertilization

PLoS One. 2011 Feb 23;6(2):e17256.

Zimmerman SW, Manandhar G, Yi YJ, Gupta SK, Sutovsky M, Odhiambo JF, Powell MD, Miller DJ, Sutovsky P.

Division of Animal Science, and Departments of Obstetrics, Gynecology, and Women's Health, University of Missouri-Columbia, Columbia, Missouri, United States of America. Abstract Despite decades of research, the mechanism by which the fertilizing spermatozoon penetrates the mammalian vitelline membrane, the zona pellucida (ZP) remains one of the unexplained fundamental events of human/mammalian development. Evidence has been accumulating in support of the 26S proteasome as a candidate for echinoderm, ascidian and mammalian egg coat lysin. Monitoring ZP protein degradation by sperm during fertilization is nearly impossible because those few spermatozoa that penetrate the ZP leave behind a virtually untraceable residue of degraded proteins. We have overcome this hurdle by designing an experimentally consistent in vitro system in which live boar spermatozoa are co-incubated with ZP-proteins (ZPP) solubilized from porcine oocytes. Using this assay, mimicking sperm-egg interactions, we demonstrate that the sperm-borne proteasomes can degrade the sperm receptor protein ZPC. Upon coincubation with motile spermatozoa, the solubilized ZPP, which appear to be ubiquitinated, adhered to sperm acrosomal caps and induced acrosomal exocytosis/formation of the acrosomal shroud. The degradation of the sperm receptor protein ZPC was assessed by Western blotting band-densitometry and proteomics. A nearly identical pattern of sperm receptor degradation, evident already within the first 5 min of coincubation, was observed when the spermatozoa were replaced with the isolated, enzymatically active, sperm-derived proteasomes. ZPC degradation was blocked by proteasomal inhibitors and accelerated by ubiquitin-aldehyde(UBAL), a modified ubiquitin protein that stimulates proteasomal proteolysis. Such a degradation pattern of ZPC is consistent with in vitro fertilization studies, in which proteasomal inhibitors completely blocked fertilization, and UBAL increased fertilization and polyspermy rates. Preincubation of intact zona-enclosed ova with isolated active sperm proteasomes caused digestion, abrasions and loosening of the exposed zonae, and significantly reduced the fertilization/polyspermy rates after IVF, accompanied by en-mass detachment of zona bound sperm. Thus, the sperm borne 26S proteasome is a candidate zona lysin in mammals. This new paradigm has implications for contraception and assisted reproductive technologies in humans, as well as animals.

PMID 21383844


Mechanisms of sperm-egg interactions: between sugars and broken bonds

Sci Signal. 2010 Oct 5;3(142):pe35.

Visconti PE, Florman HM.

Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst, MA 01003, USA. Abstract A model of the early events of mammalian fertilization has emerged during the past 30 years. However, studies during the past decade have used newly available mouse models to readdress these processes. Here, we will consider these new data in light of the existing model and point to areas of reconciliation and of controversy.

PMID 20923932;3/142/pe35

Paper suggests the following 3 models for Mouse Models of sperm binding to the ZP.

1. The glycan model - Sperm bind by interacting selectively with O-glycans conjugated to ZP3 at Ser332, Ser334, or both residues. Sperm fail to bind after fertilization because of the enzymatic modification of the adhesion glycan by an egg glycosidase.

2. The supermolecular structure model - The sperm binding site is not specified (shown as including elements of ZP2 and ZP3). Access to this binding site is regulated by proteolytic cleavage of ZP2, such that after fertilization sperm are restricted from this site and cannot bind to the ZP.

3. A hybrid model - Sperm bind to an O-glycan that is conjugated to ZP3 at a site other than Ser332 or Ser334. Sperm access to this glycan is regulated by the proteolytic cleavage state of ZP2.

The role and regulation of sperm gelsolin prior to fertilization

J Biol Chem. 2010 Oct 11.

Finkelstein M, Etkovitz N, Breitbart H.

Bar-Ilan University, Israel. Abstract In order to acquire fertilization competence, spermatozoa should undergo several biochemical changes in the female reproductive tract, known as capacitation. The capacitated spermatozoon can interact with the egg zona-pellucida resulting in the occurrence of the acrosome reaction, a process which allowed its penetration into the egg and fertilize it. Sperm capacitation requires actin polymerization, while F-actin must disperse prior to the acrosome reaction. Here we suggest that the actin severing protein, gelsolin, is inactive during capacitation and is activated prior to the acrosome reaction. The release of bound gelsolin from phosphatidylinositol 4,5-bisphosphate (PIP(2)) by PBP10, a peptide containing the PIP2-binding domain of gelsolin, or by activation of PLC which hydrolyses PIP(2), caused rapid Ca(2+)-dependent F-actin depolymerization as well as enhanced acrosome reaction. Using immunoprecipitation assays, we showed that the tyrosine kinase SRC and gelsolin coimmunoprecipitate, and activating SRC by adding 8Br-cAMP enhanced the amount of gelsolin in this precipitate. Moreover, 8Br-cAMP enhanced tyrosine phosphorylation of gelsolin and its binding to PIP(2(4,5)), both of which inactivated gelsolin, allowing actin polymerization during capacitation. This actin polymerization was blocked by inhibiting the Src-family-kinases, suggesting that gelsolin is activated under these conditions. These results are further supported by our finding that PBP10 was unable to cause complete F-actin breakdown in the presence of 8Br-cAMP or vanadate. In conclusion, inactivation of gelsolin during capacitation occurs by its binding to PIP(2) and tyrosine phosphorylation by SRC. The release of gelsolin from PIP(2) together with its dephosphorylation enables gelsolin activation, resulting in the acrosome reaction.

PMID 20937821

Sperm head binding to epithelium of the oviduct isthmus is not an essential preliminary to mammalian fertilization - review

Zygote. 2010 Jul 21:1-5.

Hunter RH.

Institute for Reproductive Medicine, Hannover Veterinary University, Bünteweg 15, D-30559 Hannover, Germany. Abstract SummaryIn endeavouring to understand the nature of sperm-oviduct interactions in mammals, attention was focused on experimental models in which fertilization can occur without a preliminary phase of sperm head binding to the isthmus epithelium. The ovarian endocrine milieu imposed on the oviduct tissues plays an important role in the binding phenomenon, although less so after the time of ovulation. Nonetheless, a sperm suspension introduced into the peritoneal cavity or surgical insemination directly into the oviduct ampulla before ovulation can result in fertilization, as can a surgical model in which the isthmus has been resected and the remaining portions of the duct reanastomosed. Mating or artificial insemination after ovulation in pigs permits rapid sperm transport to the site of fertilization, and the frequency of polyspermic penetration increases with the post-ovulatory age of eggs.Strategies underlying sperm binding were considered, especially in terms of preovulatory sperm storage and suppression of full membranous maturation. These, in turn, raised the problem of how sperm binding in vitro to oviduct cells from prepuberal animals or to cells harvested during the luteal phase of the estrous cycle, or to cells from the ampulla or even the tracheal epithelium, can act to regulate sperm storage and maturation with precision. In an evolutionary perspective, preovulatory binding of diverse populations of cells to the endosalpinx may have developed as a form of fine tuning to assist in sperm selection, to synchronize completion of capacitation with the events of ovulation, and to promote monospermic fertilization by a controlled release of competent gametes.

PMID 20663263


Reduction of mouse egg surface integrin alpha9 subunit (ITGA9) reduces the egg's ability to support sperm-egg binding and fusion

Biol Reprod. 2009 Apr;80(4):833-41. Epub 2009 Jan 7.

Vjugina U, Zhu X, Oh E, Bracero NJ, Evans JP.

Department of Biochemistry and Molecular Biology, Division of Reproductive Biology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA. Abstract The involvement of egg integrins in mammalian sperm-egg interactions has been controversial, with data from integrin inhibitor studies contrasting with evidence from knockouts showing that specific integrin subunits are not essential for fertility. An alpha(4)/alpha(9) (ITGA4/ITGA9) integrin subfamily member has been implicated in fertilization but not extensively examined, so we tested the following three hypotheses: 1) an ITGA4/ITGA9 integrin participates in sperm-egg interactions, 2) short-term acute knockdown by RNA interference of integrin subunits would result in a fertilization phenotype differing from that of chronic depletion via knockout, and 3) detection of a fertilization phenotype is sensitive to in vitro fertilization (IVF) assay conditions. We show that mouse and human eggs express the alpha(9) integrin subunit (ITGA9). RNA interference-mediated knockdown resulted in reduced levels of Itga9 mRNA and surface protein in mouse eggs. RNA interference attempts to knockdown ITGA9's likely beta partner, beta(1) (ITGB1), resulted in reduced Itgb1 mRNA but no reduction in ITGB1 surface protein. Therefore, studies using a function-blocking anti-ITGB1 antibody tested the hypothesis that ITGB1 participates in gamete interactions. Analyses of sperm-egg interactions with Itga9-knockdown eggs and anti-ITGB1 antibody-treated eggs in IVF assays using specific sperm:egg ratios revealed the following: 1) a reduction, but not complete loss, of sperm-egg binding and fusion was observed and 2) the reduction of sperm-egg binding and fusion was not detected in inseminations with high sperm:egg ratios. These data demonstrate that ITGA9 and ITGB1 participate in sperm-egg interactions but clearly are not the only molecules involved. This also shows that careful design of IVF parameters allows detection of deficiencies in gamete interactions.

PMID 19129508

Etiology of sperm immunity in women

Fertil Steril. 2009 Feb;91(2):639-43. Epub 2008 Feb 20.

Clarke GN.

Andrology Unit, Royal Women's Hospital, Melbourne, Victoria, Australia.

Abstract Sperm immunity in females can reduce the likelihood of natural conception, and sperm antibodies from female sera have been shown to inhibit IVF in humans and in several animal models. The etiology of sperm immunity in human females is unknown, but several possible mechanisms have been proposed, including cross-reactivity with microbial antigens and interferon gamma-mediated potentiation of the antisperm immune response in women whose male partners have sperm autoantibodies in their semen. This article reviews these ideas and postulates a novel hypothesis based on the potential for the generation of anti-idiotype antibodies in women whose partners have sperm antibodies in their semen.

PMID 18281044

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



Vol. 52 Nos. 5/6 (2008) Fertilization

Participation of cysteine-rich secretory proteins (CRISP) in mammalian sperm-egg interaction

Int J Dev Biol. 2008;52(5-6):737-42.

Cohen DJ, Busso D, Da Ros V, Ellerman DA, Maldera JA, Goldweic N, Cuasnicu PS.

Instituto de Biología y Medicina Experimental (IBYME-CONICET), Buenos Aires, Argentina. Abstract Mammalian fertilization is a complex multi-step process mediated by different molecules present on both gametes. CRISP1 (cysteine-rich secretory protein 1) is an epididymal protein thought to participate in gamete fusion through its binding to egg-complementary sites. Structure-function studies using recombinant fragments of CRISP1 as well as synthetic peptides reveal that its egg-binding ability resides in a 12 amino acid region corresponding to an evolutionary conserved motif of the CRISP family, named Signature 2 (S2). Further experiments analyzing both the ability of other CRISP proteins to bind to the rat egg and the amino acid sequence of their S2 regions show that the amino acid sequence of the S2 is needed for CRISP1 to interact with the egg. CRISP1 appears to be involved in the first step of sperm binding to the zona pellucida, identifying a novel role for this protein in fertilization. The observation that sperm testicular CRISP2 is also able to bind to the egg surface suggests a role for this protein in gamete fusion. Subsequent experiments confirmed the participation of CRISP2 in this step of fertilization and revealed that CRISP1 and CRISP2 interact with common egg surface binding sites. Together, these results suggest a functional cooperation between CRISP1 and CRISP2 to ensure the success of fertilization. These observations contribute to a better understanding of the molecular mechanisms underlying mammalian fertilization.

PMID 18649285

Laparoscopic observation of spontaneous human ovulation

Lousse JC, Donnez J.

Fertil Steril. 2008 Sep;90(3):833-4. Epub 2008 Apr 28.

Department of Gynecology, Université Catholique de Louvain, 1200 Brussels, Belgium. Abstract We report laparoscopic observation of spontaneous human ovulation, illustrated by protrusion of the mature follicle and oocyte release into the peritoneal cavity.

PMID: 18440526

Sperm binding to the zona pellucida is not sufficient to induce acrosome exocytosis.

Baibakov B, Gauthier L, Talbot P, Rankin TL, Dean J. Development. 2007 Mar;134(5):933-43. PMID: 17293534

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

inactive X chromosome

Cell cycle-dependent localization of macroH2A in chromatin of the inactive X chromosome. Chadwick BP, Willard HF. J Cell Biol. 2002 Jun 24;157(7):1113-23. Epub 2002 Jun 24. PMID: 12082075 \ JCB

Histone modifications and nuclear architecture: a review.

Bártová E, Krejcí J, Harnicarová A, Galiová G, Kozubek S.

J Histochem Cytochem. 2008 Aug;56(8):711-21. Epub 2008 May 12. Review.PMID: 18474937


Establishment of the mammalian membrane block to polyspermy: evidence for calcium-dependent and -independent regulation

Reproduction. 2007 Feb;133(2):383-93.

Gardner AJ, Williams CJ, Evans JP.

Division of Reproductive Biology, Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, Room W3606, 615 N. Wolfe St., Maryland, USA. Abstract One crucial result of egg activation is the establishment of blocks on the zona pellucida and the egg plasma membrane to prevent fertilization by additional sperm. The mechanism(s) by which a mammalian egg regulates the establishment of the membrane block to polyspermy is largely unknown. Since Ca(2+) signaling regulates several egg activation events, this study investigates how sperm-induced Ca(2+) transients affect the membrane block to polyspermy, building on our previous work (Biology of Reproduction 67:1342). We demonstrate that mouse eggs that experience only one sperm-induced Ca(2+) transient establish a membrane block that is less effective, than in eggs that experience normal sperm-induced Ca(2+) transients but that is more effective than in eggs with completely suppressed [Ca(2+)](cyt) increases. Sperm-induced increases in [Ca(2+)](cyt) regulate the timing of membrane block establishment, as this block is established more slowly in eggs that experience one or no sperm-induced Ca(2+) transients. Finally, our studies produce the intriguing discovery that there is also a Ca(2+)-independent event that is associated with fertilization in the pathway leading to membrane block establishment. Taken together, these data indicate that Ca(2+) plays a role in facilitating membrane block establishment by regulating the timing with which this change in egg membrane function occurs, and also that the membrane block differs from other post-fertilization egg activation responses as Ca(2+) is not the only stimulus. The membrane block to polyspermy in mammalian eggs is likely to be the culmination of multiple post-fertilization events that together modify the egg membrane's receptivity to sperm.

PMID 17307906


Time from insemination to first cleavage predicts developmental competence of human preimplantation embryos in vitro

Hum Reprod. 2002 Feb;17(2):407-12.

Fenwick J, Platteau P, Murdoch AP, Herbert M. Source Reproductive Medicine, BioScience Centre, ICFL, Times Square, Newcastle upon Tyne NE1 4EP, UK.


BACKGROUND: The absence of reliable markers for the identification of viable embryos for transfer at the early cleavage stage is likely to contribute to the generally low implantation rates and high incidence of multiple gestation in IVF treatment. In this study, we investigate the relationship between timing of first cleavage and the incidence of blastocyst formation in vitro. METHODS: Couples (n = 70) with at least one embryo remaining after transfer were included in the analyses. All embryos (n = 579) were examined for early cleavage at 25 h after insemination. Following embryo transfer, the remaining embryos (n = 426) were cultured until day 7 of development, and assessed for blastocyst formation. RESULTS: Eighty-five embryos (14.7%) cleaved to the 2-cell stage within 25 h of insemination; 26 of these were selected for transfer on day 2. Of the 59 embryos remaining in culture, 19 (32.2%) developed to the blastocyst stage; this was a significantly higher number than was observed in embryos (61/367; 16.6%) that failed to cleave within 25 h of insemination (P < 0.01). Within these two groups of embryos the proportion of hatched blastocysts was 11/59 (18.6%) and 26/367 (7.1%) respectively (P < 0.005). CONCLUSIONS: These findings indicate that early cleavage is indicative of increased developmental potential in human embryos and may be useful as an additional criterion in the selection of embryos for transfer.

PMID 11821286


Magnetic resonance imaging of male and female genitals during coitus and female sexual arousal

BMJ. 1999 Dec 18-25;319(7225):1596-600.

Schultz WW, van Andel P, Sabelis I, Mooyaart E.

Department of Gynaecology, University Hospital Groningen, PO Box 30 001, 9700 RB Groningen, Netherlands. Abstract OBJECTIVE: To find out whether taking images of the male and female genitals during coitus is feasible and to find out whether former and current ideas about the anatomy during sexual intercourse and during female sexual arousal are based on assumptions or on facts.

DESIGN: Observational study.

SETTING: University hospital in the Netherlands.

METHODS: Magnetic resonance imaging was used to study the female sexual response and the male and female genitals during coitus. Thirteen experiments were performed with eight couples and three single women.

RESULTS: The images obtained showed that during intercourse in the "missionary position" the penis has the shape of a boomerang and 1/3 of its length consists of the root of the penis. During female sexual arousal without intercourse the uterus was raised and the anterior vaginal wall lengthened. The size of the uterus did not increase during sexual arousal.

CONCLUSION: Taking magnetic resonance images of the male and female genitals during coitus is feasible and contributes to understanding of anatomy.

PMID: 10600954


Post-ovulatory ageing of the human oocyte and embryo failure

Hum Reprod. 1998 Feb;13(2):394-7.

Wilcox AJ, Weinberg CR, Baird DD. Source Epidemiology Branch, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709, USA. Abstract We carried out a prospective study of 221 healthy women who were attempting pregnancy. During the study, women collected daily urine samples and kept daily records of intercourse. Ovulation and early pregnancy losses were later identified by immunoassays of urinary human chorionic gonadotrophin and steroid metabolites. We have used these data to examine whether the risk of early pregnancy loss was higher with post-ovulatory ageing of the oocyte. 192 pregnancies were ranked by the probability that the oocyte might have aged before fertilization. There was a statistically significant increase in the risk of early loss as the likelihood of oocyte ageing increased (P < 0.05). No similar risk was observed for clinical miscarriages. Post-ovulatory ageing of the oocyte prior to fertilization may cause early pregnancy failure in humans as it does in several other mammalian species.

PMID 9557845


Preliminary observations on polar body extrusion and pronuclear formation in human oocytes using time-lapse video cinematography

Hum Reprod. 1997 Mar;12(3):532-41.

Payne D, Flaherty SP, Barry MF, Matthews CD. Source Reproductive Medicine Unit, The University of Adelaide, The Queen Elizabeth Hospital, South Australia.


In this study, we have used time-lapse video cinematography to study fertilization in 50 human oocytes that had undergone intracytoplasmic sperm injection (ICSI). Time-lapse recording commenced shortly after ICSI and proceeded for 17-20 h. Oocytes were cultured in an environmental chamber which was maintained under standard culture conditions. Overall, 38 oocytes (76%) were fertilized normally, and the fertilization rate and embryo quality were not significantly different from 487 sibling oocytes cultured in a conventional incubator. Normal fertilization followed a defined course of events, although the timing of these events varied markedly between oocytes. In 35 of the 38 fertilized oocytes (92%), there were circular waves of granulation within the ooplasm which had a periodicity of 20-53 min. The sperm head decondensed during this granulation phase. The second polar body was then extruded, and this was followed by the central formation of the male pronucleus. The female pronucleus formed in the cytoplasm adjacent to the second polar body at the same time as, or slightly after, the male pronucleus, and was subsequently drawn towards the male pronucleus until the two abutted. Both pronuclei then increased in size, the nucleoli moved around within the pronuclei and some nucleoli coalesced. During pronuclear growth, the organelles contracted from the cortex towards the centre of the oocyte, leaving a clear cortical zone. The oocyte decreased in diameter from 112 to 106 microm (P < 0.0001) during the course of the observation period. The female pronucleus was significantly smaller in diameter than the male pronucleus (24.1 and 22.4 microm respectively, P = 0.008) and contained fewer nucleoli (4.2 and 7.0 respectively, P < 0.0001). After time-lapse recording, oocytes were cultured for 48 h prior to embryo transfer or cryopreservation. Embryo quality was related to fertilization events and periodicity of the cytoplasmic wave, and it was found that good quality embryos arose from oocytes that had more uniform timing from injection to pronuclear abuttal and tended to have a longer cytoplasmic wave. In conclusion, we have shown that time-lapse video cinematography is an excellent tool for studying fertilization and early embryo development, and have demonstrated that human fertilization comprises numerous complex dynamic events.

PMID 9130755

A role for the disintegrin domain of cyritestin, a sperm surface protein belonging to the ADAM family, in mouse sperm-egg plasma membrane adhesion and fusion

J Cell Biol. 1997 Apr 7;137(1):105-12.

Yuan R, Primakoff P, Myles DG.

Molecular and Cellular Biology, University of California, Davis 95616, USA. Abstract Sperm-egg plasma membrane fusion is preceded by sperm adhesion to the egg plasma membrane. Cell-cell adhesion frequently involves multiple adhesion molecules on the adhering cells. One sperm surface protein with a role in sperm-egg plasma membrane adhesion is fertilin, a transmembrane heterodimer (alpha and beta subunits). Fertilin alpha and beta are the first identified members of a new family of membrane proteins that each has the following domains: pro-, metalloprotease, disintegrin, cysteine-rich, EGF-like, transmembrane, and cytoplasmic domain. This protein family has been named ADAM because all members contain a disintegrin and metalloprotease domain. Previous studies indicate that the disintegrin domain of fertilin beta functions in sperm-egg adhesion leading to fusion. Full length cDNA clones have been isolated for five ADAMs expressed in mouse testis: fertilin alpha, fertilin beta, cyritestin, ADAM 4, and ADAM 5. The presence of the disintegrin domain, a known integrin ligand, suggests that like fertilin beta, other testis ADAMs could be involved in sperm adhesion to the egg membrane. We tested peptide mimetics from the predicted binding sites in the disintegrin domains of the five testis-expressed ADAMs in a sperm-egg plasma membrane adhesion and fusion assay. The active site peptide from cyritestin strongly inhibited (80-90%) sperm adhesion and fusion and was a more potent inhibitor than the fertilin beta active site peptide. Antibodies generated against the active site region of either cyritestin or fertilin beta also strongly inhibited (80-90%) both sperm-egg adhesion and fusion. Characterization of these two ADAM family members showed that they are both processed during sperm maturation and present on mature sperm. Indirect immunofluorescence on live, acrosome-reacted sperm using antibodies against either cyritestin or fertilin beta showed staining of the equatorial region, a region of the sperm membrane that participates in the early steps of membrane fusion. Collectively, these data indicate that a second ADAM family member, cyritestin, functions with fertilin beta in sperm-egg plasma membrane adhesion leading to fusion.

PMID 9105040

  1. 1.0 1.1 Harper JC, Aittomäki K, Borry P, Cornel MC, de Wert G, Dondorp W, Geraedts J, Gianaroli L, Ketterson K, Liebaers I, Lundin K, Mertes H, Morris M, Pennings G, Sermon K, Spits C, Soini S, van Montfoort APA, Veiga A, Vermeesch JR, Viville S & Macek M. (2018). Recent developments in genetics and medically assisted reproduction: from research to clinical applications. Eur. J. Hum. Genet. , 26, 12-33. PMID: 29199274 DOI.