bluuebunny/crossref_metadata_2025_split huggingface dataset:
Further characterization of the mouse sperm surface zona‐binding site with galactosyltransferase activity
Authors: Benau Danny A. , McGuire Edward J. , Storey Bayard T. 
Published: February 2005

AbstractOne of the mouse sperm surface binding sites for zona pellucida ligands exhibits galactosyltransferase (GT) enzyme activity. The present study was undertaken to ascertain whether the GT site behaves as a noncatalytic binding site in its physiological capacity, with no glycosylation of zona ligands, or whether glycosylation of zona ligands is an integral part of sperm‐zona binding. The effects of Mn2+, the obligatory cation for GT catalysis, on enzyme activity and sperm‐zona binding were examined. With uridine‐5′‐diphosphogalactose (UDPgal) as galactose donor, and N‐acetylglucosamine (GlcNAc) as galactose acceptor, increasing concentrations of Mn2+ in the range of 0.1–10 mM increased GT enzyme activity, with half‐maximal activation at 0.65 mM Mn2+ (Vmax = 20 pmol/hr/106 cells). In the presence of 0–2 mM Mn2+, sperm‐zona binding was inhibited in a concentration‐dependent manner; 50% inhibition occurred at 1.25 mM Mn2+. At this concentration, GT enzyme activity was at 65% Vmax. To determine the specificity of the GT site for glycoprotein terminal carbohydrate residues, spermatozoa were incubated with, asialo‐ovine submaxillary mucin (N‐acetylgalactosamine residues), asialo‐, ‐α1‐acid glycoprotein (β1–4 galactose residues) ovalbumin (Ov; GlcNAc residues), and asialo‐agalacto‐/α1‐acid glycoprotein (AsAgAGP; GlcNAc residues). Only Ov and AsAgAGP acted as acceptors for galactose in the enzyme assay and inhibitors in the sperm‐zona binding assay. The kinetics of the interaction of AsAgAGP with the GT site were determined: the Km was 3.6 mg/ml, with Vmax of 33 pmol/hr/106 cells. In the absence of Mn2+ and UDPgal, sperm‐zona binding was inhibited in a hyperbolic manner by 0–8 mg/ml AsAgAGP; 50% inhibition was observed with 0.25 mg/ml AsAgAGP. Our model for this site predicts that, in the absence of Mn2+, UDPgal and glycoproteins with GlcNAc terminal residues compete for zona ligands to inhibit sperm‐zona binding; the presence of Mn2+ causes a change in the site from substrate binding conformation to catalytic conformation, which is unfavorable for sperm‐zona interaction.
Evaluation of rabbit sperm acrosomal integrity and fertilizing ability by use of vital stains
Authors: McBride C. E. , Fayrer‐Hosken R. A , Srivastava P. N. , Brackett B. G. 
Published: February 2005

AbstractThree staining procedures to detect sperm acrosome integrity were compared via electron microscopy. Stains were applied to epididymal, freshly ejaculated, in vivo capacitated, and sonicated sperm cells in addition to spermatozoa displaying sequentially removed plasma and outer and inner acrosomal membranes. Sequential membrane removal procedures resulted in removal of plasma membranes from 73% of all sperm cells, removal of plasma and outer acrosomal membranes from 74% of all sperm cells, and removal of plasma and outer and inner acrosomal membranes from 87% of all sperm cells as determined by electron microscopy. Live/dead staining results were not statistically different from subjective microscopic motility evaluations (P < 0.005) for epiddymal, sonicated, freshly ejaculated, and in vivo capacitated sperm samples. All three stains assessed were similarly capable of detecting the acrosome status of freshly ejaculated and of sonicated spermatozoa compared to data obtained by electron microscopy (P = 0.010). However, only the Bryan‐Akruk stain afforded data that were closely correlated with data obtained via electron microscopy for all sperm types assessed; the latter included in vivo capacitated spermatozoa and sperm cells rendered free of plasma membranes. Results confirmed an earlier report by successfully effecting sequential removal of rabbit acrosomal membrances and documented use of the Bryan‐Akruk acrosomal stain for evaluation of sperm cell popualtions for fertilizing ability. These findings should prove useful in further investigations of mechanisms involved in achievement of fertilizing ability by rabbit spermatozoa.
Origin and early development of female germ cells in Eoleptestheria ticinensis Balsamo‐Crivelli, 1859 (Crustacea, Branchiopoda, Conchostraca)
Authors: Sabelli Franca Scanabissi , Tommasini Stefano 
Published: February 2005

AbstractWe have studied the early stages of the development of the female germ cells in the Conchostraca Eoleptestheria ticinensis Balsamo‐Crivelli, 1859. The gametes originate in a scattered way throughout the tubular units of the gonad, with no development gradient recognizable. The female germ cells arise from successive karyokineses not followed by cytokinesis, within an unorganised area in which a sort of plasmodium is formed. Each primordial nucleus of this germarium develops and then forms an individual plasmic membrane. Subsequently, the usual organules differentiate in the cytoplasm. The presence of synaptinemal complexes and the beginning of the endogenous vitellogenesis by the rough endoplasmic reticulum and the Golgi apparatus qualify the previtellogenesis. The general characteristics of this early development phase of the gametes, as well as several substantial differences in the gametogenesis with respect to the other Phyllopoda studied, lead us to suggest the systematic positioning of the Leptestheriidae.
Effect of culture conditions and media on the frequency of chromosomal abnormalities in human sperm chromosome complements
Authors: Martin Renee H. , Templado Cristina , Ko Evelyn , Rademaker Alfred 
Published: February 2005

AbstractHuman sperm chromosomes can be visualized after fusion with hamster eggs. Most laboratories use one of two methods of sperm treatment for capacitation: incubation in a modified Krebs‐Ringer medium (BWW) for 5–7 h at 37°C or storage in a TES‐Tris yolk buffer (TYB) for 24–72 h at 4°C. To determine whether data from the two methods were comparable, we performed a series of controlled experiments on one normal donor in which ejaculates were split and one aliquot of sperm was capacitated in BWW for 5–7 h at 37°C (fresh) and the second aliquot was capacitated in TYB for 48 h at 4°C (TYB). After capacitation, the technique used to obtain human sperm chromosome complements was identical for both aliquots. Both fresh and TYB sperm were further subdivided into two groups, which were subjected to either a short (1 h) or a long (3 h) gamete coincubation in BWW. This experiment was performed to determine if the longer incubation in BWW might induce chromosomal fragile sites and breaks because of nutritional depletion of the medium. A total of 458 human sperm chromosome complements was analysed. There was no significant difference in the frequency of sperm chromosomal abnormalities or in the sex ratio in the sperm coincubated with eggs for a short (1 h) or long (3 h) time in BWW. When sperm pretreatments were compared, there was a significant increase in the frequency of total sperm chromosomal abnormalities after TYB storage compared to fresh treatment. This difference was due to an increased frequency of structural chromosomal abnormalities in sperm stored in TYB, not in hyperhaploid or hypohaploid sperm. There was no difference in the sex ratio of fresh (47.2% X) or TYB sperm (47.2% X). Thus our results suggest that storing sperm at 4°C for 48 h in TYB does not alter the frequency of aneuploidy but significantly increases the frequency of structural chromosomal abnormalities.
Quantitative changes in cytoskeletal β‐ and γ‐actin mRNAS and apparent absence of sarcomeric actin gene transcripts in early mouse embryos
Authors: Taylor Kent D. , Pikó Lajos 
Published: February 2005

AbstractActin is known to be synthesized both during oogenesis and in cleavage‐stage embryos in mice. Cytoskeletal β‐actin appears to be the major component, followed by γ‐actin, but the synthesis of α‐actin has also been inferred from protein electrophoretic patterns. We have studied the expression of cytoskeletal (β‐ and γ‐) and sarcomeric (α‐cardiac and α‐skeletal) actin genes at the level of the individual mRNAs in blot hybridization experiments using isoform‐specific RNA probes. The results show that there are about 2 × 104 β‐actin mRNA molecules in the fully grown oocyte; this number drops to about one‐half in the egg and less than one‐tenth in the late two‐cell embryo but increases rapidly during cleavage to about 3 × 105 molecules in the late blastocyst. The amount of γ‐actin mRNA is similar to that of β‐actin in oocytes and eggs but only about 40% as much in late blastocysts, indicating a differential accumulation of these mRNAs during cleavage. The developmental pattern of β‐ and γ‐actin mRNA provides a striking example of the transition from maternal to embryonic control that occurs at the two‐cell stage and involves the elimination of most or all of the maternal actin mRNA. There was no detectable α‐cardiac or α‐skeletal mRNA (i.e., < 1,000 molecules per embryo) at any stage from oocyte to late blastocyst, suggesting that the sarcomeric actin genes are silent during preimplantation development.
Immunoelectron microscopical visualization of ribonucleoproteins in the chromatoid body of mouse spermatids
Authors: Biggiogera M. , Fakan S. , Leser G. , Martin T. E. , Gordon J. 
Published: February 2005

AbstractThe chromatoid body (CB), a cytoplasmic organelle present only in germ cell line, was studied at the electron microscopic level in mouse spermatids using cytochemical techniques and specific antibodies directed against sn‐RNPs, hnRNPs, and ribosomal proteins. We found that specific staining for DNA as well as the use of monoclonal anti‐DNA antibodies show a complete absence of DNA in the CB. The CB remains stained, however, after the application of the ethidium bromide‐PTA technique, suggesting the presence of RNA within this organelle. snRNP as well as hnRNP proteins are demonstrated within the CB by means of specific monoclonal or polyclonal antibodies, especially during earlier spermiogenic stages. Monoclonal antibodies directed against the large ribosomal subunit proteins P1/P2 detect these antigens on the CB essentially along the internal threads of dense fibrillar material. Our findings suggest that the CB may function as a source of mRNA and/or of its partially processed precursors during the late stages of spermiogenesis, when the spermatid nucleus becomes gradually inactive.
Dimethylsulphoxide affects the organisation of microfilaments in the mouse oocyte
Authors: Vincent C. , Pickering S. J. , Johnson M. H. , Quick S. J. 
Published: February 2005

AbstractThe effect of dimethylsulphoxide (DMSO) on microfilament organisation has been studied in the mouse oocyte after staining with (NBD)‐phallacidin. The cortical actin meshwork was disrupted by exposure of oocytes to 1.5 M DMSO at 37°C, and this disruption was associated with changes in the cell surface, especially microvilli length and distribution, as observed by scanning electron microscopy. The irregular distribution of actin filaments observed also appears to lead to an irregular expansion of the cell after DMSO removal. However, when exposure to DMSO was combined with cooling, the effects on the microfilament system were much reduced. The reversibility of DMSO action is considered and the potential implications of microfilament disruption on the viability and functions of the oocyte discussed.
Chymotrypsin‐like enzymes are involved in sperm penetration through the vitelline coat of Ciona intestinalis egg
Authors: Pinto Maria Rosaria , Hoshi Motonori , Marino Rita , Amoroso Alessandro , De Santis Rosaria 
Published: February 2005

AbstractIn Ciona intestinalis, sperm penetration through the egg vitelline coat is an essential event of fertilization. We investigated whether trypsin‐ and chymotrypsin‐like enzymes are involved in this event. Inhibitors and peptide substrates for chymotrypsin‐like enzymes blocked the overall process of fertilization in a concentration‐dependent manner. The inhibitory activity was specifically exerted on the step of sperm penetration. Chymotrypsin‐like protease activity was identified in spermatozoa with the fluorogenic synthetic substrate Suc‐Ala‐Ala‐Phe‐AMC, which was the most effective substrate in blocking sperm penetration. These data indicate that a chymotrypsin‐like protease activity is a sperm lysin of Ciona intestinalis.
Stage‐specific expression of phosphoprotein p19 during spermatogenesis in the rat
Authors: Amat Jose A. , Fields Kay L. , Schubart Ulrich K. 
Published: February 2005

AbstractThe expression of phosphoprotein p19, a 19‐kDa cytosolic substrate for cyclic adenosine monophosphate (cAMP)‐dependent protein kinase, occurs abundantly in brain and testis and is developmentally regulated. In the present study we have identified the cell types of adult rat testis that contain p19. Using cryostat sections, which were first incubated with rabbit anti‐p19 for immunohistochemistry followed by counterstaining with periodic acid‐Schiff (PAS)‐hematoxylin to reveal nuclear morphology, we demonstrate that immunoreactive p19 is detectable only in germ cells and is restricted to a limited stage of spermatogenesis. Expression first appears after the differentiating gametes have entered the prophase of meiosis, is abundant in spermatocytes until meiosis is completed, and declines to undetectable levels in maturing spermatids. We have ruled out immunocross‐reactivity with SCG10, a 22‐kDa protein that is closely related in structure to p19, by demonstrating, using Northern blot analysis, that RNA transcripts encoding SCG10 are not detectable in adult rat testis, whereas p19 is abundantly expressed. The transient expression of p19 during spermatogenesis suggests that the protein plays a role during male gamete differentiation.
Ectopic expression of chloramphenicol acetyltransferase (CAT) in the cerebellum in mice transgenic for a carbonic anhydrase II promoter‐CAT construct that is without apparent phenotypic effect
Authors: Erickson Robert P. , Bevilacqua Arturo , Venta Patrick J. , Karolyi Jill , Tashian Richard E. 
Published: February 2005

AbstractWe have developed six transgenic lines of mice with constructs containing presumptive 5′ regulatory regions of carbonic anhydrase II (CA II). Four of the lines contained 1,100 bases of the 5′ flanking region of the human CA II gene, and two transgenic lines resulted from a construct containing 500 bases of the 5′ flanking region of the mouse CA II gene. Tissue‐specific expression of the chloramphenicol acetyltransferase (CAT) gene was not obtained in any of the transgenic lines. One of the transgenic lines was found to have high levels of expression of CAT in cerebellum. This expression persisted through multiple generations and was independent of the parental origin of the transgene. On the assumption that the expression was due to the insertion of the transgene in or near a gene expressed normally in cerebellum, homozygous mice were bred for the transgenic insert to see if a mutation might have been induced. Homozygous mice were found and seemed to be normal in all aspects of their phenotype studied. Thus, in this case, neither the insertion of the gene nor the ectopic expression of CAT seemed to be harmful to the animals.
Potential of hypertonic medium treatment for embryo micromanipulation: I. Survival of rabbit embryos in vitro and in vivo following sucrose treatment
Authors: Yang X. , Chen Y. , Chen J. , Foote R. H. 
Published: February 2005

AbstractRabbit zygotes and embryos were exposed to hypertonic sucrose in phosphate‐buffered saline (SPBS). In experiment one, 144 zygotes shrank to 32–36% of their initial volume in 1.0 M SPBS within 30 min. Neither hypertonic treatment with 0.5 M or 1.0 M SPBS nor micropuncture of the zona pellucida after shrinkage affected embryo development into blastocysts in vitro (88%, 83%, and 82%, respectively), compared to that of the controls (93%, P > .05). In experiment two, 252 two‐ to four‐cell‐ and 177 morula‐stage embryos were exposed to isotonic PBS control or 0.5 M, 1.0 M, or 1.5 M SPBS for 30, 60, 90, 120, and 150 min before transfer to PBS (290 mOsm). Embryo development was significantly reduced (P < .05) when embryos were exposed in 0.5 M and 1.0 M SPBS for more than 60 min or in 1.5 M SPBS for more than 30 min. In experiment 3, morulae exposed for 60 min to 0.5 M or 1.0 M SPBS shrank to 37–39% or 32–35% of their initial volume and then expanded to 87–94% or 81–90% of their initial volume, respectively, after being returned to isotonic PBS for 60 min, but embryos in 1.5 M SPBS had erratic osmotic behavior. In experiment four, 192 two‐ to four‐cell embryos exposed to 0.5 M SPBS for 0, 30, and 60 min before transfer to oviducts of recipients resulted in the production of 39%, 42% and 31% young, respectively (P > .05). Exposure of embryos to 0.5 M sucrose for 60 min clearly does not compromise developmental potential and can simplify and speed up micromanipulation procedures.
Ultrastructure of porcine embryos following development in vitro versus in vivo
Authors: Hyttel P. , Niemann H. 
Published: February 2005

AbstractThe present study examined the ultrastructural appearance of porcine embryos from the four‐cell stage to the blastocyst grown either in vivo or in vitro. Embryos were collected at slaughter from superovulated gilts and were fixed for transmission electron microscopy either immediately or after various periods of in vitro culture. In general, the morphology of in vivo and in vitro grown embryos was similar. In vivo grown four‐cell stages contained dense fibrillar nucleoli. At the eight‐cell stage the nucleoli possessed increasing amounts of chromatin and granules. In both stages the mitochondria were spherical or ovoid in shape and had only few cristae. In morulae and blastocysts the nucleoli were mainly of the fibrillogranular type, and the mitochondria were filamentous and possessed more cristae, of which many were tubular. Two major ultrastructural deviations were observed in about half of the in vitro cultured embryos. First, nucleolus‐like structures were found outside the nuclei in the cytoplasm of blastomeres. These structures were spherical and composed of chromatin‐like material containing characteristically a single large and several small vacuoles. The structures were frequently associated with profiles of smooth endoplasmic reticulum (SER). A second type of deviation was aggregates of SER appearing as spiral coils or multiangular complexes. Some embryos displayed both types of deviations. The physiological significance of these deviations remains speculative. They may be involved in the considerably reduced capability of porcine embryos to develop to piglets following in vitro culture.
Sperm DNA and sex chromosome differences between two geographical populations of the creeping vole, Microtus oregoni
Authors: Johnson Lawrence A. , Clarke Robert N. 
Published: February 2005

AbstractThe relative DNA content of the “O” and Y chromosome‐bearing sperm is presented for the creeping vole, Microtus oregoni. The animals had been trapped in Oregon and in Washington State. The two populations had very similar autosomal chromosome relationships but differed greatly in the size of their X chromosome (which is not carried by vole sperm) and in their Y chromosome. The greater size and banding differences of the Y chromosome of the Washington State vole compared to the Oregon vole paralleled the greater differences in sperm DNA between the Y‐bearing sperm and the sperm carrying no sex chromosome (O). The actual DNA differences between O and Y sperm was 12.5% for the sperm from the Washington State voles and 9.1% for sperm from the Oregon voles. The difference in sperm DNA content (12.5%) for Washington State voles was far greater than the difference shown for other voles or other mammals.
Chromosome constitution of highly motile mouse sperm
Authors: Estop A. , Catala V. , Santalo J. 
Published: February 2005

AbstractIn this study, we address the relationship between motility and genetic content of mouse sperm. The chromosome complements of highly motile mouse sperm, selected using the swim‐up technique, were analyzed after in vitro fertilization, at the first cleavage state. They were compared to those of unselected sperm. Identification of male and female chromosome sets was possible because of their differential condensation at the first mitotic division. In vitro fertilization, swim‐up separation, chromosome preparation, and staining were carried out using standard techniques. The results indicate that highly motile mouse sperm did not differ in types and frequencies of chromosomal abnormalities from those not selected for motility. Moreover, separation of motile sperm does not deviate the sex ratio from the theoretical 1:1.
Changes in exposed membrane proteins during in vitro capacitation of boar sperm
Authors: Berger Trish 
Published: February 2005

AbstractExposed plasma membrane proteins were labeled with 125I before and after incubation of boar sperm under capacitating conditions. Labeled protein profiles were compared to the ability of the sperm to penetrate zona‐free hamster ova. Quantitatively, the labeled sperm membrane proteins were primarily low Mr prior to capacitation. The majority of the labeled seminal plasma protein was also low Mr. After capacitation, two new proteins (64,000 Mr and 78,000 Mr) were labeled. Sperm did not exhibit these exposed membrane proteins when incubated under noncapacitating conditions. Appearance of these proteins was not correlated to the percentage of acrosome‐reacted sperm. Although the 64,000 Mr protein was not consistently observed, the relative labeling of the 78,000 Mr protein was highly correlated with the ability of sperm to fuse with zonafree hamster ova. The 78,000 Mr protein may be a sperm protein involved in fusion with the egg plasma membrane.
Effects of follicle stimulating hormone and purines on rat oocyte maturation
Authors: Törnell Jan , Brännström Mats , Magnusson Claes , Billig Håkan 
Published: February 2005

AbstractThe presented data demonstrate a dose‐dependent inhibition of spontaneous meiosis of cumulus‐enclosed rat oocytes by guanosine, hypoxanthine, and adenosine. The inhibition by adenosine was transient whereas guanosine and hypoxanthine exerted a persistent effect over 24 h of incubation. The order of potency of the substances was guanosine>hypoxanthine>adenosine and the inhibition was reversible. The inhibitory effect was reduced when the cumulus cells around the oocyte were removed. The inhibition during the first 12 h of incubation was potentiated by FSH. However, at 24 h of incubation FSH partially overcame the inhibitory effect by hypoxanthine but did not influence the inhibitory effect by guanosine. Also 8BrcAMP potentiated the inhibitory effect observed by guanosine, hypoxanthine, and adenosine, suggesting that the potentiating effect of FSH was mediated via cAMP. Our data demonstrate that adenosine, hypoxanthine, and guanosine synergized with FSH in inhibiting spontaneous rat meiosis, as previously shown in mouse. FSH could partially overcome the inhibitory effect exerted by hypoxanthine but did not counteract the inhibitory effect of guanosine.