bluuebunny/crossref_metadata_2025_split huggingface dataset:
Selective binding of mouse and human spermatozoa to beads coated with extracellular matrix components
Authors: De Felici Massimo , Camaioni Antonella , Russo Mario A. , Siracusa Gregorio 
Published: February 2005

AbstractThe ability of mouse epididymal and human ejaculated spermatozoa to bind to beads coated with various extracellular matrix components was examined. Mouse spermatozoa preferentially bound to beads coated with heparin (average values ranging between 6.2 and 8.8 sperm per bead were obtained in different experiments) and with chondroitinsulfate (6.2–7.0), and also, although with significant differences across replicate experiments, to beads coated with laminin (7.9–15.6 sperm per bead) and with collagen type I (6.1–18.5). Human spermatozoa bound to collagen‐coated beads (15.4–22.6 sperm per bead) and, to a much lower extent, to chondroitinsulfate‐coated beads (3.2–4.7); they were also able to bind heparin‐coated beads, although with ample differences between individual sperm donors (ranging between 0.8 and 18.7 sperm per bead). Very few human and mouse sperm bound fibronectin‐coated beads; beads coated with albumin, hyaluronic acid, and chondronectin were always totally free of adhering sperm. The possible physiological role of the interactions between spermatozoa and extracellular matrix components are discussed.
Asynchrony between pronuclear development and protein synthetic changes in zygotes of the mouse derived from oocytes aged postovulation in vivo and fertilized in vivo
Authors: Boerjan M. L. , Op Het Veld C. W. , Koopman M. B. H. 
Published: February 2005

AbstractThe pattern of proteins synthesized by one‐cell embryos derived from unaged oocytes and oocytes aged postovulation in vivo was analyzed by means of 35S‐methionine labeling and gel electrophoresis. The oocytes were obtained after ovulation induction by an injection of luteinizing hormone‐releasing hormone (LHRH) at proestrus or after a superovulation procedure. The analysis was performed in unfertilized aged and unaged secondary oocytes and in zygotes derived from them. The patterns of proteins synthesized by secondary oocytes from all experimental groups were very similar: The oocytes showed a predominant synthesis of 35 kDa proteins. Zygotes from aged as well as unaged LHRH‐induced oocytes also showed a predominant synthesis of one group of polypeptides with a relative molecular weight of about 35 kDa. The proteins of the 35 kDa protein complex migrated in an upper (u), middle (m), or lower (l) band in 10% polyacrylamide sodium dodecyl sulfate (SDS) gels. The u‐ and m‐band 35 kDa proteins were shown to be synthesized by secondary oocytes. Early pronuclear zygotes from unaged LHRH‐induced oocytes synthesized u‐ and m‐ but no l‐band 35 kDa proteins. In contrast, part (38%, n = 47) of the early pronuclear zygotes from aged LHRH‐induced oocytes did synthesize the l‐band 35 kDa proteins. Late pronuclear zygotes (LPZ) from aged as well as unaged oocytes synthesized predominantly the l‐band 35 kDa proteins. However, although only 5.8% (n = 51) of LPZ from unaged oocytes synthesize m‐ and l‐band 35 kDa proteins, these bands of proteins are present in 25% (n = 24) of the LPZ from aged oocytes. Thus, in zygotes from aged oocytes, at least part of the fertilization‐dependent changes in the 35 kDa protein synthetic pattern are advanced with reference to the morphological stage of pronuclear progression. Most (80.6%, n = 36) of the early pronuclear zygotes from unaged superovulated oocytes synthesized u‐ and m‐band 35 kDa proteins, like early pronuclear zygotes from unaged LHRH‐induced oocytes. However, 16.7% did not synthesize any of the 35 kDa proteins at all. Thus with respect to the protein synthetic pattern of 35 kDa proteins, zygotes from unaged superovulated oocytes are different from zygotes derived from unaged LHRH‐induced oocytes.
Subzonal microinjection of mouse spermatozoa: Insufficient sperm motility might induce phagocytosis
Authors: Klemm Martina , Engel Wolfgang 
Published: February 2005

AbstractAcrosome‐reacted CB6F1 mouse spermatozoa with slight flagellar motility were microinjected under the zona pellucida of CB6F1 mouse oocytes. Electron microscopy revealed the presence of swollen and decondensed sperm heads in the oocyte cytoplasm. Sixty‐one percent of the microinjected oocytes reached a morphologically apparent two‐cell stage, but chromosomal analysis demonstrated only haploid chromosomal complements in all cases. The exposure of microinjected oocytes to suspensions of spermatozoa of mice homozygous for a 2,4 reciprocal translocation resulted in normal fertilization and embryonic development with a maternally as well as a paternally derived haploid genome. Identical results were obtained with oocytes microinjected with medium and subjected to in vitro fertilization thereafter. Thus it can be suggested that the microinjected spermatozoa with insufficient flagellar motility are incorporated into the oocyte cytoplasm by phagocytosis. These spermatozoa do not induce a polyspermy block but induce the oocyte to parthenogenetic development.
Further observations on the phagocytosis of Candida albicans by Hamster and Human Oocytes
Authors: Koeheler James K. , Clark Judy M. , Dianne Smith W. 
Published: February 2005

AbstractPathogenic yeast, Candida albicans, were incubated with hamster and human oocytes for up to 21 hours in order to determine the nature and time course of phagocytosis of these organisms. Aliquotes of the interacting cells were taken at various time intervals for electron microscopic examination. Some specimens had their zona pellucidae enzymatically removed prior to incubation with yeast, and these specimens showed the most extensive interaction and phagocytosis of Candida. The zona pellucida appears to be an effective barrier to yeast, at least over the time span studied. The observations are consistent with the hypothesis of an initial attachment of yeast via a surface component to oocyte microvilli followed by phagocytic uptake into an endosome. There is no compelling evidence of lysosomal degradation of the yeast over the time course of this study; however, the oocytes appear to undergo some degenerative changes at long incubation times.
Isolation, characterization, and localization of the zona pellucida binding proteins of boar sperm
Authors: Hanqing Mo , Tai‐Ying Yan , Zhao‐Wen Shen 
Published: February 2005

AbstractIn order to isolate the putative zona pellucida‐binding proteins (ZPBPs) from boar spermatozoa, a new, simple method has been developed. The new isolation strategy made the most of the highly specific interactions between the components of the gametes. Detergent‐extracted boar sperm proteins were submitted to affinity chromatography on a ZP‐Sepharose column. SDS‐PAGE analyses of the retained fraction under reducing conditions revealed that in addition to a component of Mr 38,000, the predominant ZPBPs contained at least three low molecular weight proteins (Mr <20 kDa). The isolated ZPBPs were effective in blocking sperm‐oocyte binding in vitro. Using immunofluorescence microscopy, the ZPBPs were shown to be localized primarily in the sperm head, especially in the acrosomal cap region.
Distribution of filipin‐sterol complexes in the plasma membrane of stallion spermatozoa during the epididymal maturation process
Authors: López A. M. L. , De Souza W. 
Published: February 2005

AbstractThe presence and distribution of cholesterol in mature and immature epididymal spermatozoa was analyzed using filipin as a cytochemical tool in freeze‐fracture replicas and thin section preparations. The polyenic‐antibiotic filipin formed complexes with 3,β‐OH sterols, producing characteristic protrusions, or pits, that were heterogeneously distributed in the plasma membrane of stallion spermatozoa, revealing a specific organization in a functionally specialized area of the gamete. The acrosomal region of the sperm head presented a significantly higher density of filipin sterol complexes than the postacrosomal region, which was usually free of these complexes. The plasma membrane of the flagellum also showed filipin sterol complexes randomly distributed in freeze‐fracture replicas. The strong filipin labeling observed in the membrane of spermatozoa obtained from the caput region of the epididymis decreased significantly during epididymal passage. The significance of these changes is not completely understood, but they might contribute to establishing the molecular organization necessary for sperm transit and storage in the epididymis as well as to development of motile spermatozoa that are able to fertilize the oocyte and induce normal embryonic development.
A preferential site for sperm‐egg fusion in mammals
Authors: Talansky Beth E. , Malter Henry E. , Cohen Jacques 
Published: February 2005

AbstractThe tendency of mammalian sperm‐egg fusion to occur at a site away from the first polar body was investigated in a homologous (mouse oocytes and mouse spermatozoa) and in a heterologous model (hamster oocytes and mouse spermatozoa). Following micromanipulation of the zona pellucida either in proximity to or opposite the first polar body, in vitro fertilization was performed and subsequent differences in sperm‐egg interaction were evaluated. Since spermatozoa from random‐bred mice do not readily penetrate intact zonae pellucidae in vitro, it is likely that zona penetration occurred through the artificial holes in both models. The creation of a gap in the zona pellucida opposite the first polar body was associated with levels of sperm fusion that were significantly higher than those resulting from manipulation near the first polar body. Spermatozoa were rarely found to penetrate the hole completely, and in general few spermatozoa were observed in the perivitelline space. The proximity between pronuclei following sperm penetration was correlated with the position of the incision with respect to the polar body. The findings suggest that breaching the zona pellucida for microsurgical fertilization should be performed away from the microvillus‐free area of the oocyte.
Studies on zona hardening in rat oocytes that are matured in vitro in a serum‐free medium
Authors: Zhang X. , Rutledge J. , Armstrong D. T. 
Published: February 2005

AbstractThe present study confirms that zonae pellucidae of rat oocytes became resistant to chymotrypsin digestion (zona hardening) after undergoing in vitro maturation (IVM) in a serum‐free medium. However, zona hardening and did not occur when empty zonae without oocytes were cultured in the same IVM conditions, suggesting that oocyte‐derived factor(s) is responsible for zona hardening in oocytes matured in the serum‐free medium. Zona hardening occurred primarily after dictyate oocytes were cultured for 6–8 hours in the IVM medium without serum. Zona hardening could be prevented or alleviated if oocytes were cultured in the IVM medium containing bovine foetal calf serum, a soybean trypsin inhibitor, or β‐mercaptoethanol, and in vitro fertilization rates for such oocytes were normal. Possible mechanisms of the phenomenon of zona hardening in oocytes matured in serum‐free conditions are discussed in relation to its possible relevance to the cortical reaction and the physiological block to polyspermy.
Maturation of the rat cumulus‐oocyte complex: Structure and function
Authors: Phillips David M. , Dekel Nava 
Published: February 2005

AbstractThe cumulus cells that surround the mammalian oocyte become dispersed following the preovulatory surge of the pituitary gonadotropin, luteinizing hormone (LH). We have examined cumulus‐oocyte complexes of PMSG‐primed immature rats before and at 1, 2, 3, 4, 6, and 8 hr after injection of human chorionic gonadotropin (hCG), which acts on the rat ovary like the pituitary gonadotropin. Associations between projections of the cumulus cells and the oocyte were analyzed in thin sections. We observed that some cumulus projections were greatly enlarged where they associate with the oocyte. These enlarged regions were filled with numerous small vesicles. Gap junctions between cumulus cell projections and the oocytes were small. We quantitated the number and size of gap junctions between cumulus cells. The number of small gap junctions (<1 μM) between cumulus cells did not change significantly over the 8‐hr period after hCG administration. Large gap junctions, however, showed a general downward trend beginning after the third hour post hCG. Light microscopic observations of plastic sections revealed that dispersion of the cumulus oophorus is not observed until after 4 hr post‐hCG, but between 4 and 8 hr after gonadotropin administration the cumulus becomes markedly dispersed. In the majority of the oocytes in these complexes the germinal vesicle (GV) displayed some irregularity in shape at 2 hr post‐hCG, although absence of the GV was not observed until later.Our observations suggest a new means of communication in the cumulus‐oocyte complex by the vesicle‐filled enlargements of the cumulus cell projections at the oocyte surface. They further indicate that the decrease in metabolic coupling observed in rat cumulus‐oocyte complexes soon after exposure to LH is not associated with a change in number and size of the gap junctions between the cumulus cells. We suggest that it is either the disruption of the gap junctions at the region of contact of the cumulus cell projections with the oocyte surface or the operation of a gating mechanism that blocks the junctional channels without affecting their morphological appearance that is responsible for uncoupling of the oocyte from the cumulus cells.
Expression of the rig gene in mouse oocytes and early embryos
Authors: Taylor Kent D. , Pikó Lajos 
Published: February 2005

AbstractA clone selected from a two‐cell mouse embryo cDNA library has been sequenced and identified as rig cDNA. The rig gene codes for a highly conserved nuclear protein, which may have a general role in cell growth or replication (Shiga et al.: Proc Natl Acad Sci USA 87:3594, 1990). The quantitative changes in rig mRNA were studied in blot hybridization experiments with total RNA from oocytes and early embryos. The amount and relative abundance of rig mRNA change considerably during early development. There are about 1.6 × 104 rig mRNA molecules in a late growth‐stage oocyte; this number is reduced to about one‐tenth in the ovulated egg but increases about twenty‐fold during cleavage through the blastocyst stage. In F9 embryonal carcinoma cells, the relative abundance of rig mRNA is similar to that in blastocysts (about 0.1% of the mRNA population), but it is about eight‐fold higher in the mouse myeloma cell line MOPC‐104E. The high level of rig mRNA in late growth‐stage oocytes suggests that the rig gene product may be important for cverall transcriptional activity rather than DNA replication and mitosis. Alternatively, the rig protein may be a storage product of oogenesis and have a role in the initiation of development.
Genomic imprinting: Male mice with uniparentally derived sex chromosomes
Authors: Handel Mary Ann , Hunt Patricia A. 
Published: February 2005

AbstractAlthough it has been known that there is an X‐chromosome imprinting effect during early embryogenesis in female mammals, it remains unknown if parental origin of the X chromosome has an effect in males. Furthermore, it has not been possible to produce animals with normal sex chromosomes of uniparental origin to further evaluate such imprinting effects. We have devised a breeding scheme to produce male mice, designated XPYP males, in which both the X and Y chromosomes are paternally inherited. To our knowledge, these are the first mammals produced that have a normal sex chromosome constitution but with both sex chromosomes derived from one parent. Development and reproduction in these XPYP males and the sex ratio and chromosome constitution of their offspring appeared normal; thus there is no apparent effect in males of having both sex chromosomes derive from one parent of of having the X chromosome derived from an inappropriate parent. Although we have detected no X‐chromosome imprinting effect in these males, evidence from other sources suggests that the X chromosome is parentally imprinted. Thus detection and definition of an imprint can depend on the assay used.
Oxygen transport to embryos in microdrop cultures
Authors: Baltz Jay M. , Biggers John D. 
Published: February 2005

AbstractThe standard method for culturing small preimplantation mammalian embryos is a system in which they are placed into microdrops of culture medium under oil. Thus the source of oxygen for the embryos lies beyond two liquid phases—medium and oil. If transport of oxygen is not sufficiently rapid to replace that consumed by the embryos, the medium could become depleted of oxygen. Over small distances, the dominant means by which oxygen is transported through liquid is by diffusion. Our calculations show that diffusion alone is sufficient to supply oxygen to mouse embryos; there is virtually no perturbation of the oxygen concentration when there are up to 10 embryos in the drop, and 50 embryos produce a drop in oxygen tension that is not large enough to have a deleterious effect. Furthermore, diffusion is probably not the dominant mechanism by which oxygen is transported to the embryos; on the scale of these microdrops, convection is faster and would serve to mix the drop so that anoxic regions cannot develop. Therefore, we conclude that even a relatively large number of embryos in a culture drop do not significantly deplete oxygen.
Formation of the perinuclear theca in spermatozoa of diverse mammalian species: Relationship of the manchette and multiple band polypeptides
Authors: Longo Frank J. , Cook Susan 
Published: February 2005

AbstractThe perinuclear theca is a novel cytoskeletal consisting of a densely layered lamina that surrounds the nucleus of mammalian sperm. Using antibodies specific for the multiple band polypeptides present in the perinuclear theca of bull sperm, we show that a heterogeneous group of immunological related proteins are present in the sperm heads of other mammals with greatly different morphologies, including guinea pig, hamster, rat, and mouse. In none of the species were identical groups of immunoreactive polypeptides found, although immunoreactive proteins of molecular weights 65,000 to 80,000 were present in the sperm heads of all species examined. Immunoreactive proteins less than Mr 55,000 were prominent in rat sperm heads and mouse sperm; guinea pig, hamster, and rat sperm heads and mouse sperm had one band in common at approximately Mr 50,000. Different immunoreactive proteins were present in isolated sperm tails. The perinuclear theca first appeared in the subacrosomal space of round to elongating spermatids. Later, with the caudal movement of the manchette, the postacrosomal segment of the perinuclear theca was deposited in a cephalad to caudal direction along the sperm nucleus. Concomitantly, the cytoplasmic space between the nuclear envelope and the plasma membrane narrowed such that only the theca occupied this portion of the sperm head. Immunoreactivity accompanied the ultrastructural appearance of the subacrosomal layer and the postacrosomal segment. The periods of spermiogenesis, in which sub‐ and post‐acrosomal components of the perinuclear theca are formed and the morphogenesis of sperm organelles with which these elements are associated, suggest that components of this cytoskeletal structure function to join the acrosome and the postacrosomal plasma membrane to the nucleus.
Electroporation of bovine spermatozoa to carry foreign DNA in oocytes
Authors: Gagné Marc B. , Pothier François , Sirard Marc‐André 
Published: February 2005

AbstractIn the present study, electroporation was used to test the ability of spermatozoa to carry foreign DNA into the bovine oocytes. Frozen‐thawed bovine spermatozoa (107/ml) were electroporated using six different combinations of voltge (500, 1,000, or 1,500 V) and capacitance (1 or 25 μ Farads) in the presence of 1 mg/ml of plasmid pRGH527. The portions of plasmids retained by sperm cells after three washings (stable for ten washings) were 4.3, 5.5, 5.1, 6.0, 6.8, and 5.8% for 1 μFarad, 500, 1,000, and 1,500 V and 25 μFarads, 500, 1,000, and 1,500 V, respectively. Nonelectroporated cells have retained only 1% of plasmids. In the same experiment, electroporated spermatozoa were acrosome reacted by treatment with ionophore A23187 to evaluate the fraction of marked plasmids joined at the acrosomal membrane. The results show that 3.5, 5.0, 4.4, 5.0, 6.3, and 4.4% remain tied to the ionophore‐treated sperm. Only 0.7% of plasmid was retained after removal of the acrosome of nonelectroporated cells. Acrosome reaction was not significantly induced by the electrical field (EF) (P < 0.005). EF decrease motility significantly for > 100 V in 0.3 M mannitol (M) and mannitol‐TALP (MT) (1/1) media and ≥500 V (P<0.05) in TALP medium. The retained plasmid rate was compared between TALP medium M and MT media and resulted in a percentage of 1.0, 2.5, 6.5 at 1 μFarads, 100 V, and 0.9, 3.8, and 3.8 at 25 μFarads, 100 V in TALP, MT, and M medium, respectively. Sperm cells electroporated at 1 μFarad, 500 or 1,000 V, 25 μFarad, 500 V or 1,000 in TALP medium hold plasmids in proportion of 5.2, 5.4, 7.4, and 6.0%. Electroporation above 100 V in M and MT killed the cells. In a part of this experiment, spermatozoa electroporated in the presence of radiolabeled plasmids have been treated with DNase I and results revealed that 35, 28, 54, 58, and 3% of marked DNA remains in sperm cells following digestion after electroporation in TALP (1,000 V, 1 and 25 μFarads), M medium (100 V, 1 and 25 μFarads), and control, respectively. Using in vitro matured bovine oocytes, the electroporation conditions were correlated with the fertilization rate (85% for control and 55% for electroporated spermatozoa). Autoradiography of embryos following fertilization indicated the presence of plasmids in the cytoplasm and in the zona pellucida. Finally, the use of Polymerase Chain Reaction (PCR) revealed the presence of plasmids in blastocyst cells in 12% for control (n=67) and 22, 17, and 19% for electroporated spermatozoa in TALP (n=188,25μFarads, 1,000 V), MT (n=29, 25 μFarads, 100 V) and M (n=21, 25 μFarads, 100 V), respectively. In conclusion, our results indicate A) that electroporation results in an increase absorption of DNA by the spermatozoa, B) that the male gametes can carry foreing DNA in oocytes at fertilisation, and C) that an increased fraction of day 5 embryos is showing a positive response for the inserted gene with the polymerase chain reaction.