In mammals, the sperm deliver mRNA of unknown function into the oocytes during fertilization. The role of sperm microRNAs (miRNAs) in preimplantation development is unknown. miRNA profiling identified six miRNAs expressed in the sperm and the zygotes but not in the oocytes or preimplantation embryos. Sperm contained both the precursor and the mature form of one of these miRNAs, miR-34c. The absence of an increased level of miR-34c in zygotes derived from α-amanitin-treated oocytes and in parthenogenetic oocytes supported a sperm origin of zygotic miR-34c. Injection of miR-34c inhibitor into zygotes inhibited DNA synthesis and significantly suppressed first cleavage division. A 3′ UTR luciferase assay and Western blotting demonstrated that miR-34c regulates B-cell leukemia/lymphoma 2 (Bcl-2) expression in the zygotes. Coinjection of anti-Bcl-2 antibody in zygotes partially reversed but injection of Bcl-2 protein mimicked the effect of miR-34c inhibition. Oocyte activation is essential for the miR-34c action in zygotes, as demonstrated by a decrease in 3′ UTR luciferase reporter activity and Bcl-2 expression after injection of precursor miR-34c into parthenogenetic oocytes. Our findings provide evidence that sperm-borne miR-34c is important for the first cell division via modulation of Bcl-2 expression.

Fertilization in humans is initiated by binding of spermatozoa to a selectin ligand on the egg’s extracellular matrix.

MicroRNAs (miRNAs) are small non-coding RNAs that regulate the expression of other genes by transcriptional inhibition or translational repression. miR-34a is a known tumor suppressor gene and inhibits abnormal cell growth. However, its role in other tumorigenic processes is not fully known. This study aimed to investigate the action of miR-34a on cell invasion. We found that miR-34a is expressed at various levels in cervical cancer (HeLa, SiHa, C4I, C33a and CaSki) and trophoblast (BeWo and JAR) cell lines. Transient forced expression of miR-34a did not affect the proliferation of these cell lines. Computational miRNA target prediction suggested that Notch1 and Jagged1 were targets of miR-34a. By using functional assays, miR-34a was demonstrated to bind to the 3' untranslated regions of Notch1 and Jagged1. Forced expression of miR-34a altered the expression of Notch1 and Jagged1 protein as well as Notch signaling as shown by the response of Hairy Enhancer of Split-1 protein to these treatments using western blot analysis. Forced expression of miR-34a suppressed the invasiveness of HeLa and JAR cells. By using gamma-secretase inhibitor (N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester) that interfered Notch signaling and RNA interference that knockdown Notch1 expression, we confirmed that downregulation of Notch1 reduced the invasiveness of the cells. Transfection of intracellular domain of Notch nullifies the effect of miR-34a on the invasiveness of the cells. Besides, we identified that miR-34a affected cell invasion by regulating expression of urokinase plasminogen activator through Notch. Our results provide evidence that miR-34a inhibits invasiveness through regulation of the Notch pathway and its downstream matrix degrading enzyme.

Problem In mammals, implantation involves interactions between an activated blastocyst and a receptive endometrium. There are controversies on the role of microRNAs in preimplantation embryo development. The actions of endometrial microRNAs on implantation are beginning to be understood. Method of Study Review of literature on microRNAs in preimplantation embryos and endometrium. Results Emerging evidence suggests a role of microRNAs in blastocyst activation and implantation. Differential expression of microRNAs is found between receptive and non‐receptive endometria. Members of the let‐7, miR‐200, miR‐30 families, and the miR‐17‐92 clusters are more commonly found to be associated with endometrial receptivity. Experimental studies show that the targets of the differentially expressed microRNAs affect endometrial receptivity, decidualization, and embryo implantation. Free and exosome/microvesicle containing microRNAs have been detected in human and ovine uterine luminal fluid (ULF). They may serve as mediators of embryo–endometrium dialog. Some observations suggest that the microRNAs in ULF may be used as biomarkers in infertility treatment. Conclusion MicroRNAs in endometrium and blastocysts are involved in the implantation process.

Acrosome reaction is crucial to the penetration of spermatozoa through the zona pellucida (ZP). Glycosylation of ZP glycoproteins is important in spermatozoa-ZP interaction. Human ZP glycoprotein-3 (ZP3) is believed to initiate acrosome reaction. Recently, human ZP4 was also implicated in inducing acrosome reaction. These studies were based on recombinant human ZP proteins with glycosylation different from their native counterparts. In the present study, the effects of native human ZP3 and ZP4 on acrosome reaction and spermatozoa-ZP binding were investigated. Native human ZP3 and ZP4 were immunoaffinity-purified. They induced acrosome reaction and inhibited spermatozoa-ZP binding time- and dose-dependently to different extents. These biological activities of human ZP3 and ZP4 depended partly on their glycosylation, with N-linked glycosylation contributing much more significantly than O-linked glycosylation. Studies with inhibitors showed that both human ZP3- and ZP4-induced acrosome reactions were protein kinase-C, protein tyrosine kinase, T-type Ca2+ channels, and extracellular Ca2+ dependent. G-protein also participated in human ZP3- but not in ZP4-induced acrosome reaction. On the other hand, protein kinase-A and L-type Ca2+ channels took part only in human ZP4-induced acrosome reaction. This manuscript describes for the first time the actions of purified native human ZP3 and ZP4 on acrosome reaction and spermatozoa-ZP binding.

Glycodelin is a human glycoprotein with four reported glycoforms, namely glycodelin-A (GdA), glycodelin-F (GdF), glycodelin-C (GdC), and glycodelin-S (GdS). These glycoforms have the same protein core and appear to differ in their N-glycosylation. The glycosylation of GdA is completely different from that of GdS. GdA inhibits proliferation and induces cell death of T cells. However, the glycosylation and immunomodulating activities of GdF and GdC are not known. This study aimed to use ultra-high sensitivity mass spectrometry to compare the glycomes of GdA, GdC, and GdF and to study the relationship between the immunological activity and glycosylation pattern among glycodelin glycoforms. Using MALDI-TOF strategies, the glycoforms were shown to contain an enormous diversity of bi-, tri-, and tetraantennary complex-type glycans carrying Gal␤1-4GlcNAc (lacNAc) and/or GalNAc␤1-4GlcNAc (lacdiNAc) antennae backbones with varying levels of fucose and sialic acid substitution. Interestingly, they all carried a family of Sda (NeuAc␣2-3(GalNAc␤1-4)Gal)-containing glycans, which were not identified in the earlier study because of less sensitive methodologies used. Among the three glycodelins, GdA is the most heavily sialylated. Virtually all the sialic acid on GdC is located on the Sda antennae. With the exception of the Sda epitope, the GdC N-glycome appears to be the asialylated counterpart of the GdA/GdF glycomes. Sialidase activity, which may be responsible for transforming GdA/GdF to GdC, was detected in cumulus cells. Both GdA and GdF inhibited the proliferation, induced cell death, and suppressed interleukin-2 secretion of Jurkat cells and peripheral blood mononuclear cells. In contrast, no immunosuppressive effect was observed for GdS and GdC.

No abstract

Glycodelin is an example of a glycoprotein whose complex-type glycans mediate biological actions in human reproduction and immune reactions. Being attached to an identical protein backbone, glycodelin oligosaccharides vary significantly from one reproductive tissue to another and have an effect on its own secretion and role in cell communication. For instance, uterine glycodelin-A inhibits sperm-oocyte interaction by binding on the sperm head. This is a glycosylation-dependent phenomenon, in which fucosyltransferase-5 plays a key role. Glycodelin-S from seminal plasma binds evenly around the sperm head and maintains an uncapacitated state in the spermatozoa, until the isoform is detached during sperm passage through the cervix. Glycodelin-F from follicular fluid and Fallopian tube binds to the acrosomal region of the sperm head, thereby inhibiting both the sperm-oocyte binding and premature progesterone-induced acrosome reaction. The cumulus cells surrounding the oocyte can capture glycodelin-A and -F from the surrounding environment and convert these isoforms to a cumulus cell isoform, glycodelin-C. It differs by glycosylation from the other isoforms, and it too attaches on the sperm head, with the highest density in the equatorial region. Glycodelin-C is capable of detaching the sperm-bound inhibitory isoforms so that the sperm-oocyte binding is enhanced. Glycodelin-A also has immunosuppressive actions directed to cellular, humoral and innate immunity. Although these actions depend mainly on the protein backbone, glycosylation also plays a part. Glycosylated glycodelin may be involved in the protection of spermatozoa against maternal immune reactions, and glycodelin also has apoptogenic activity. Some glycosylation patterns of glycodelin may mask its apoptogenic domain. This review updates the recent research and clinical associations of glycodelin, highlighting the role of glycosylation.