Mouse follicular oocytes, denuded and intact, were cultured in pyruvate salt sol and glutamine salt sol supplemented bovine serum albumin to compare the maturation rate. Glutamine has no effect on maturation of the denuded mouse oocyte but has an effect on maturation of the intact oocyte by increasing the maturation rate, depending on the increased concentration of glutamine (0.4 mM to 2 mM). Changes in osmolarity of the operation medium from 280 mOsm to 310 mOsm has no discernible effect on the oocyte maturation. A high frequency of abnormal 1st polar bodies was observed in pyruvate salt sol. and this may be due to the increased energy source in the cytoplasm of the 1st polar body when the po1ar body was extruded into the perivitelline space after the 1st meiosis.
Animal ; Cell Division ; Female ; Glutamine/metabolism ; In Vitro ; Mice ; Oocytes/cytology ; Oocytes/metabolism* ; Ovum/metabolism* ; Pyruvates/metabolism

Mouse follicular oocytes, denuded and intact, were cultured in pyruvate salt sol and glutamine salt sol supplemented bovine serum albumin to compare the maturation rate. Glutamine has no effect on maturation of the denuded mouse oocyte but has an effect on maturation of the intact oocyte by increasing the maturation rate, depending on the increased concentration of glutamine (0.4 mM to 2 mM). Changes in osmolarity of the operation medium from 280 mOsm to 310 mOsm has no discernible effect on the oocyte maturation. A high frequency of abnormal 1st polar bodies was observed in pyruvate salt sol. and this may be due to the increased energy source in the cytoplasm of the 1st polar body when the po1ar body was extruded into the perivitelline space after the 1st meiosis.
Animal ; Cell Division ; Female ; Glutamine/metabolism ; In Vitro ; Mice ; Oocytes/cytology ; Oocytes/metabolism* ; Ovum/metabolism* ; Pyruvates/metabolism

OBJECTIVETo analyze sex chromosome mosaicisms in early cleavage-stage human embryos and blastocysts with poor embryo quality score based on the numbers of pronucleus(PN) zygotes using X,Y dual color fluorescence in situ hybridization (FISH), and to discuss the possible mechanisms.

METHODSFresh or frozen-thawed early cleavage-stage human embryos and blastocysts with poor embryo quality score not suitable for embryo transfer were studied with dual color FISH.

RESULTSDouble signal rate of 2PN among early cleavage-stage embryos was 66.67%, which was significantly higher than 1PN and 3PN embryos. Single signal rate of 1PN early cleavage-stage embryos was 90.41%, which was significantly higher than 2PN and 3PN ones. Three signal rate of 3PN early cleavage-stage embryos was 28.00%, which was significantly higher than 1PN and 2PN ones. Double signal rate of 3PN ones was 46.00%, which was significantly higher than 1PN ones. The polyploid rate of frozen-thawed early cleavage-stage embryos was 23.53%, which was slightly higher than that of fresh embryos, but with no statistical significance. The mosaicism rate of 24 blastocysts was 100.00% and the double signal dominant (≥ 50%) rate was 62.50%, which was significantly higher than the rate of early cleavage-stage embryos.

CONCLUSIONUsing 2PN as the criterion for embryo quality score cannot guarantee the selection of normal fertilized embryo for transplantation. Frozen-thawed embryos may harbor more polyploid cells. To avoid the selection of embryos with abnormal chromosomes, combinations of pre-implantation genetic diagnosis (PGD) and prenatal diagnosis are necessary. Meanwhile, blastocysts with poor quality scores may provide an important source for embryo stem cells.

Blastocyst ; metabolism ; Cleavage Stage, Ovum ; metabolism ; Fertilization in Vitro ; Humans ; In Situ Hybridization, Fluorescence ; Mosaicism ; embryology ; Sex Chromosomes

The luminal fluid environment of the female reproductive tract is considered critical for the sperm to undergo a series of molecular events leading to the final acquisition of their fertilizing capacity. It has been shown that the fluid in the female reproductive tract contains high content of HCO3- and it plays an important role in sperm functions including sperm motility, capacitation, hyperactivation and acrosome reaction. This review summarizes the effects of HCO3- on sperm functions occurring in the female reproductive tract and discusses the transport mechanisms involved in mediating uterine HCO3- secretion. New evidence is also presented to show possible cause of female infertility due to defective HCO3- transporting mechanism.
Animals ; Bicarbonates ; metabolism ; Female ; Fertilization ; physiology ; Humans ; Male ; Sperm Capacitation ; physiology ; Sperm-Ovum Interactions ; physiology ; Uterus ; metabolism ; secretion

Dicer is an RNAse III endonuclease that is essential for the biogenesis of microRNAs and small interfering RNAs. These small RNAs transcriptionally and post-transcriptionally regulate mRNA expression through RNA interference mechanisms. Recently, the function of Dicer in female reproduction has begun to be elucidated through the use of knockout mouse models. Several latest studies have indicated that Dicer gene plays a key role in female reproductive processes such as oocyte maturation, early embryonic development and implantation and steroidgenesis. When Dicer expression is decreased in female reproductive tissues or cells, it will cause infertility. In this article, author discuss the role of Dicer gene in female reproductive tract, and advance of Dicer gene study in female reproductive events.
Animals ; Embryonic Development ; genetics ; Female ; Humans ; MicroRNAs ; Ovary ; metabolism ; Ovum ; metabolism ; RNA Interference ; Reproduction ; genetics ; Ribonuclease III ; genetics ; metabolism ; Uterus ; metabolism

Fertilization is a complex process involving multiple steps, of which sperm-egg fusion is most important. This article presents a detailed review of some of the key sperm membrane proteins closely related with fertilization, such as the Izumo, the ADAMs gene family and the Crisp gene family proteins, which is of practical significance for deeper insights into the mechanisms of sperm-egg fusion, as well as for the improvement of clinical diagnosis of male infertility and development of novel contraceptive drugs.
Animals ; Cell Fusion ; Gene Expression ; Humans ; Male ; Membrane Proteins ; genetics ; metabolism ; Oocytes ; cytology ; metabolism ; Seminal Plasma Proteins ; genetics ; metabolism ; Sperm-Ovum Interactions ; Spermatozoa ; cytology ; metabolism

Sulfogalactosylglycerolipid (SGG) is the main glycolipid in male mammalian germ cells, which is selectively and highly expressed in mammalian testes and helps form the lipid bilayer of cell membrane. In the process of spermatogenesis, SGG is involved in the meiosis of spermiocytes. Either deficiency or accumulation of SGG will lead to male infertility. SGG homeostasis in the testis is the premise of normal spermatogenesis. In the process of sperm-zona binding, SGG becomes a component of lipid raft and provides a platform for signal transduction. The SGG binding protein plays a role in sperm-egg recognition and membrane fusion. SGG has a great research value and application prospect in male reproduction.
Animals ; Cell Membrane ; Galactolipids ; physiology ; Humans ; Infertility, Male ; etiology ; Lipid Bilayers ; metabolism ; Male ; Signal Transduction ; Sperm-Ovum Interactions ; physiology ; Spermatogenesis ; physiology ; Spermatozoa ; metabolism ; Testis ; physiology

For studying the effect of integrin on the [Ca(2+)](i) of mouse eggs and its transmembrane signaling mechanism, zona-free mouse eggs were loaded with calcium probe Fluo-3/AM and the intensity of fluorescence of the eggs treated with different factors was measured through laser confocal microscopy. The results showed that the [Ca(2+)](i) of zona-free mouse eggs was increased when the eggs were treated with RGD peptide, fibronectin (Fn) and anti-mouse integrin subunit alpha(6) and beta(1) monoclonal antibodies, respectively. The [Ca(2+)](i) of the mouse eggs was also increased when the eggs were placed in calcium-free medium and treated with RGD peptide or Fn. The changes in the mouse egg [Ca(2+)](i) caused by RGD and Fn were similar to those caused by sperm. However, the concentration of Ca(2+) of the zona-free mouse eggs pretreated with tyrosine kinase inhibitor was not increased when the eggs were treated in the same way, and, neither was the intracellular calcium increased in those eggs pretreated with PKC inhibitor when the eggs were treated with RGD peptide. It is therefore suggested that the occupancy of integrins on the membrane of mouse eggs by their ligands mediates the release of Ca(2+) and then the increase in the [Ca(2+)](i) of eggs, which is one of the early events of egg activation. The tyrosine kinase signaling pathway and PKC are involved in this process as well.
Animals ; Calcium ; metabolism ; Calcium Channels ; metabolism ; Female ; Integrins ; physiology ; Ion Transport ; drug effects ; Mice ; Oligopeptides ; pharmacology ; Oocytes ; metabolism ; Ovum ; metabolism ; Protein Kinase C ; metabolism ; Protein-Tyrosine Kinases ; metabolism ; Signal Transduction

Germ cells make two major decisions when they move from an indeterminate state to their final stage of gamete production. One decision is sexual commitment for sperm or egg production, and the other is to maintain mitotic division or entry into meiosis. It is unclear whether the two decisions are made as a single event or separate events, because there has been no evidence for the presence of germ cell sex prior to meiosis. Here we report direct evidence in the fish rainbow trout that gonia have distinct sexuality. We show that dazl expression occurs in both male and female gonia but exhibits differential intracellular distribution. More strikingly, we show that boule is highly expressed in male gonia but absent in female gonia. Therefore, mitotic gonia possess sex, sperm/egg decision and mitosis/meiosis decision are two independent events, and sperm/egg decision precedes mitosis/meiosis decision in rainbow trout, making this organism a unique vertebrate model for mechanistic understanding of germ cell sex differentiation and relationship between the two decisions.
Animals ; Female ; Fish Proteins ; genetics ; Gene Expression Regulation ; Male ; Meiosis ; Oncorhynchus mykiss ; genetics ; physiology ; Ovary ; cytology ; metabolism ; Ovum ; cytology ; metabolism ; RNA, Messenger ; genetics ; metabolism ; RNA-Binding Proteins ; genetics ; Sex Determination Processes ; Spermatozoa ; cytology ; metabolism ; Testis ; cytology ; metabolism

Objective: To elucidate the possible role of human lysozyme-like protein 4 (LYZL4) in fertilization and characterize its enzymatic properties. METHODS: The localization of LYZL4 in human spermatozoa was investigated by immunofluorescence staining, the sources of LYZL4 on the sperm surface examined by RT-PCR, and the role of LYZL4 in fertilization assessed by the zona-free hamster egg penetration test. The recombinant plasmid pPIC9K-LYZL4 was constructed and its expression induced with methanol after transformed into competent Pichia pastoris GS115. The recombinant LYZL4 protein (rLYZL4) was purified from the fermentation supernatant and subsequently identified by Western blot. The hyaluronan binding ability of rLYZL4 was determined by ELISA and the muramidase activity, hyaluronidase activity, and free radical scavenging ability examined by spectrophotometric methods. RESULTS: Immunodetection with a specific antiserum localized LYZL4 on the acrosomal membrane of mature spermatozoa, which was exclusively secreted from the testis and epididymis as shown by RT-PCR. Immunoneutralization of LYZL4 significantly decreased the number of human spermatozoa bound to zona-free hamster eggs in a dose-dependent manner in vitro. The recombinant protein was expressed successfully by the P. pastoris strain GS115. Purified rLYZL4 exhibited a potent hyaluronan binding ability and a strong free radical scavenging ability but no muramidase or hyaluronidase activity. CONCLUSIONS LYZL4 secreted from the testis and epididymis is localized on the acrosomal membrane of mature spermatozoa and plays a role in sperm-egg binding as well as in binding hyaluronan and scavenging free radicals, which suggests that it might be a multi-functional molecule contributive to sperm protection and sperm-egg binding.
Acrosome ; enzymology ; Animals ; Blotting, Western ; Cricetinae ; Enzyme-Linked Immunosorbent Assay ; Epididymis ; Female ; Fertilization ; physiology ; Free Radical Scavengers ; metabolism ; Humans ; Hyaluronic Acid ; metabolism ; Male ; Muramidase ; analysis ; physiology ; Pichia ; Plasmids ; metabolism ; Recombinant Proteins ; analysis ; metabolism ; Sperm-Ovum Interactions ; physiology ; Spermatozoa ; enzymology ; Testis