bluuebunny/crossref_metadata_2025_split huggingface dataset:
Production of chimeric rabbits from morulae by a simple procedure
Authors: Yang Xiangzhong , Foote R. H. 
Published: February 2005

AbstractExperiments were conducted to develop a simple and reliable technique to produce chimeric rabbits from morula stage embryos. In Experiments 1 and 2, an in‐vitro test of viability was initially performed by culturing embryos to the blastocyst stage. Ninety‐three percent of the “chimeric” embryos developed to the blastocyst stage compared to 94% for controls when embryos were manipulated soon after collection (Exp. 1). Eighty‐one percent chimeric embryos and 78% control embryos developed to blastocyst stage when embryos were held at room temperature for 4 hr (Exp. 2). In Experiment 3, enough morula‐stage embryos were available from true breeding Dutch‐belted and albino rabbits to form potentially 67 diverse “color” pairs. These micromanipulated pairs of morulae were successfully combined to produce 64 chimeric embryos (96%, 64/67). They were transferred to the uteri of seven recipient does and three became pregnant producing 13 young. Four of the young exhibited substantial overt chimerism (31%) and one more was a possible chimera.
Role for fucose‐sulfate‐rich carbohydrates in the penetration of zona‐pellucida‐free hamster eggs by hamster spermatozoa
Authors: Dravland J. E. , Mortimer D. 
Published: February 2005

AbstractThe interaction of acrosome‐reacted hamster spermatozoa and zona‐pellucida‐free hamster eggs was investigated by incubating gametes in the presence of a variety of simple and complex carbohydrates. Significant inhibition of gamete fusion was achieved only in the presence of fucoidan and ascophyllin, two algal polysaccharides containing fucose sulfate. These compounds did not interfere with sperm motility, capacitation, or acrosome reactions. It is concluded that these two compounds share common structural features with putative cell‐surface carbohydrates involved in sperm‐oolemmal interaction.
Hemizona assay using salt‐stored human oocytes: Evaluation of zona pellucida capacity for binding human spermatozoa
Authors: Franken Daniel R. , Burkman Lani J. , Oehninger Sergio C. , Coddington Charles C. , Veeck Lucinda L. , Kruger Thinus F. , Rosenwaks Zev , Hodgen Gary D. 
Published: February 2005

AbstractHuman oocytes were stored (25°C) in 1.5 M MgCl2 for 6–30 days, then utilized in the new hemizona assay (HZA) for tight binding of human spermatozoa [Burkman et al.: Fertil Steril 49:688–697, 1988]. We have compared 1) the ability of matching salt‐treated hemizonae or dimethylsulfoxide (DMSO)‐treated hemizonae to distinguish between sperm from semen having normal versus subnormal characteristics and, 2) the kinetics of fertile sperm binding to salt‐treated or DMSO‐treated hemizonae. After sperm preparation, one salt‐treated hemizona was incubated with normal spermatozoa and the matching hemizona was placed with sperm from the subnormal group. As a control, DMSO‐treated hemizonae were incubated in additional sperm droplets. After 4 hours, the number of sperm tightly bound to each hemizona was counted. Within the normal semen group, there was equivalent binding to salt‐ or DMSO‐treated hemizonae (54.0 ± 12 and 49 ± 14, respectively, mean ± SEM). Similarly, tight binding of sperm from the subnormal group was not affected by the zona storage method (21 ± 8 and 17 ± 5, respectively). For either storage approach, binding of subnormal sperm was significantly less (P < 0.01) compared with the number of normal sperm attached to the matching hemizona. For the kinetics study, the hemizona binding of proven fertile spermatozoa was followed throughout 8.5 hours. The shape of the binding curve was the same for zonae stored by either method and was consistent with our published kinetics data. Salt storage offers a simple and inexpensive means for accumulating and transporting human zonae pellucida; the resulting hemizonae function effectively in the HZA for estimating sperm binding potential.
Analysis of the chromosome complement in outbred mouse sperm fertilizing in vitro
Authors: Martin‐DeLeon Patricia A. 
Published: February 2005

AbstractThe chromosome complements in a population of mouse sperm from random‐bred ICR donors were analyzed at first‐cleavage metaphase after in vitro fertilization (IVF) of oocytes from females of the same strain. The sperm were aged as donations occurred within an average of 31 days, either since last mating or at arrival at the animal facility in the case of virgin males. Of a total of 598 sperm complements studied from 22 sexually mature males aged 10–26 weeks old, there was one diploid complement (0.17%). The frequencies of hyperhaploidy and structural aberrations that were studied in 338 complements were 4.4% and 3.6%, respectively, giving an overall frequency of 8.0%. The hyperhaploid complements consisted of n + 1, n + 2, n + 3, and n + 7 counts, while the structural abnormalities were of the chromosome type and included large and small fragments and a possible translocation. This is the highest frequency of sperm chromosome abnormalities reported for mouse sperm obtained from males under physiological conditions and fertilized in vitro or in vivo. Sperm aging, strain, and/or technique differences are among the factors that may be responsible for this high frequency. Since the 8.0% frequency of hyperhaploidy and structural abnormalities is similar to the frequency reported for human sperm after IVF, the outbred murine in vitro fertilization system may be a useful model to study the origin of human sperm chromosome abnormalities.
Meiotic resumption and gap junction modulation in the cultured rat cumulus‐oocyte complex
Authors: Wert Susan E. , Larsen William James 
Published: February 2005

AbstractIntercellular communication within the ovarian follicle has been implicated in the control of meiotic arrest and maturation in the mammalian oocyte. We have shown that a rapid down‐regulation of cumulus cell gap junctions is correlated temporally with meiotic resumption in the intact rat follicle [Larsen et al., Dev Biol, 113:517–521]. Here this relationship has been analyzed further by incubating isolated rat cumulus‐oocyte complexes (COCs) with agents known to influence germinal vesicle breakdown (GVBD) or that have been shown to modulate gap junction turnover in vitro. Quantitative freeze‐fracture analysis revealed that cumulus cell gap junction membrane decreased significantly prior to the initiation of GVBD in COCs incubated in medium lacking serum or other additives. The addition of serum and follicle‐stimulating hormone, an experimental condition that delayed GVBD, accelerated and augmented gap junction loss at both the cumulus cell and oocyte surface. The continuous elevation of cyclic adenosine monophosphate levels, which stimulates gap junction formation in other systems, maintained meiotic arrest but did not interfere with gap junction loss. Conversely, the complete inhibition of junctional loss by a microfilament destabilizing agent, dihydrocytochalasin B, did not alter the course of GVBD normally seen in its absence. Subsequent freeze‐fracture analysis and dye coupling experiments confirmed that cumulus and oocyte gap junctions in these preparations were intact and functional during the period of meiotic resumption. These findings suggest that factors other than cumulus and oocyte gap junction turnover are required for the control of meiotic arrest and maturation in the isolated COC. However, these results do support our earlier suggestion that gap junction loss within the cumulus oophorus is instrumental in isolating the oocyte from the regulatory influence of its underlying membrana granulosa cells during meiotic maturation in the intact preovulatory follicle [Larsen et al., Dev Biol, 122:61–71].
Human antral follicles: Oocyte nucleus and the karyosphere formation (electron microscopic and autoradiographic data)
Authors: Parfenov Vladimir , Potchukalina G. , Dudina L. , Kostyuchek D. , Gruzova M. 
Published: February 2005

AbstractThe functional organization of the nucleus in the oocytes from human antral follicles was examined by morphological and autoradiographic analysis methods at the light and electron microscopic level. According to the position of the nucleus, the level of its transcriptional activity, and the pattern of distribution of structures in it, oocytes fall into two groups. In the first one, the oocytes with the nucleus in the central position are characterized by the distribution of numerous structures all over the nucleus or by a different extent of aggregation of chromatin around the nucleolus. The nuclei of these oocytes are characterized by [3H]uridine incorporation, the label being localized over purely fibrillar, agranular nucleoli and over dispersed fibrillar chromatin adjacent to either the regions of densely packed chromatin or fibrillar‐granular material of the nucleolus‐like bodies. The latter, the same as condensed chromatin, do not incorporate [3H]uridine. In the second group, the nuclei are displaced towards the oocyte's periphery, and chromosomes surround the nucleolus as a continuous mass closely adjacent to its surface, thus forming a karyosphere. The karyosphere formation takes place on the background of cessation of nuclear transcriptional activity. A fully formed karyosphere represents a complex of closely associated inactivated structures: Nucleolus, chromosomes, and nucleolus‐like bodies. The karyosphere nucleolus bears no granules and consists of densely packed finely fibrillar material (fibrils 3 nm thick). Two zones (central and peripheral) can be distinguished in a nucleolus. Nucleolus‐like bodies, consisting of granules 20 nm in diameter embedded in finely fibrillar material, are often associated with chromosomes.In this study, data obtained by observations on the loss of association between the oocyte (with karyosphere) and corona radiata cells are evaluated. The relation of the karyosphere formation to the atresia process and the duration of karyosphere existence in human antral follicles are also discussed.
Continuous live‐dead discrimination of ram sperm during freezing
Authors: Fiser P. S. , Marcus G. J. 
Published: February 2005

AbstractUse of the dye amaranth (Color Index 16185) as a supravital stain for ram sperm is described. At a concentration of 0.4% in diluted semen, the dye was completely excluded by motile sperm and had no effect on sperm motility. The nuclei of immotile sperm were stained pink by amaranth. The decrease in sperm motility during 24‐h storage at 5°C was accompanied by a corresponding increase in stained sperm nuclei. The presence of the dye during freezing had no effect on sperm cryosurvival but tended to reduce sperm motility during post‐thaw incubation. Insemination of ewes with fresh semen containing amaranth or with semen frozen in the presence of amaranth resulted in pregnancies in 7/10 ewes in each group, compared to 6/9 in the case of ewes inseminated with fresh semen without dye.
Induction and inhibition of amphibian (Rana pipiens) oocyte maturation by protease inhibitor (TPCK)
Authors: Ishikawa Katsutoshi , Schuetz Allen W. , Francisco S. K. San 
Published: February 2005

AbstractWe report for the first time that oocyte nuclear and cytoplasmic maturation are triggered in vitro in non‐hormone‐treated amphibian (Rana pipiens) ovarian follicles following transient exposure to synthetic chymotrypsin inhibitor Nα‐tosyl‐L‐phenylalanine‐chloromethyl ketone (TPCK). The mechanism of action of TPCK in regulating oocyte maturation was investigated and compared to that induced by progesterone or pituitary hormone. Follicular oocytes failed to mature following continuous exposure to the same doses of TPCK in the presence or absence of progesterone. Continuous treatment of follicles with lower levels of TPCK occasionally induced GVBD in the absence of progesterone and augmented maturational effects of low levels of progesterone. TPCK induced maturation of intrafollicular oocytes without stimulating progesterone production and also induced maturation of naked oocytes. Stimulation of follicular progesterone synthesis following gonadotropin stimulation or addition of pregnenolone was inhibited by TPCK, indicating that TPCK affects metabolic processes in both the somatic and germinal components of the ovarian follicle. Oocyte maturation induced by either TPCK or progesterone was inhibited by cycloheximide, calcium‐deficient medium, and forskolin. Results suggest that TPCK induces oocyte maturation independent of steroidogenesis via mechanisms similar to those triggered by progesterone involving protein synthesis, formation of cytoplasmic maturation‐promoting factor (MPF), and changes in cAMP levels. Our data indicate that a chymotrypsin‐like protease plays a role(s) in regulating the oocyte meiotic maturation process.
Influence of serum and hormones on bovine oocyte maturation and fertilization in vitro
Authors: Younis A. I. , Brackett B. G. , Fayrer‐Hosken R. A. 
Published: February 2005

AbstractThree approaches were investigated for improvement of in vitro maturation (IVM), in vitro fertilization (IVF), and early embryonic development in cattle. These were: 1) Selection of oocytes, 2) medium supplementation with fetal calf serum (FCS) and cow sera (DO, Dl, D10, and D20 to correspond with estrus, metestrus, diestrus, and proestrus, respectively), and 3) addition of follicle‐stimulating hormone (FSH), luteinizing hormone (LH), and estradiol‐17β (E2)during maturation. Greater proportions (percentage) of oocytes initially selected for their compact cumulus cells completed IVM and IVF when compared to unselected oocytes (P < .05). Proportions (percentage) of selected oocytes that matured and cleaved after in vitro insemination according to serum type used for IVM were: FCS: 110/175 (62.9%) and 37/110 (33.6%) and DO: 130/145 (89.7%) and 52/130 (40.0%); D1 127/130 (97.7%) and 41/127 (32.3%); D10 95/98 (96.9%) and 35/95 (36.8%); D20:113/116 (97.4%) and 49/113 (43.4%). A higher proportion (P < .05) of embryos resulting from the D20 group reached four‐ and eight‐cell stages. In FCS‐supplemented maturation media with no hormones added during maturation (control), results of IVM and IVF were 157/265 (59.2%) and 39/157 (24.8%), respectively. With E2 (1 μg/ml) and FSH (5 μg/ml), comparable results were 189/215 (87.9%) and 71/189 (37.6%); with E2 (1 μg/ml) plus LH (10 μml), 280/327 (85.6%) and 111/280 (39.6%). Added hormones improved IVM results (P < .05) and, when FSH or LH was added with E2, in vitro development to four‐ and eight‐cell stages was markedly enhanced (P < .05). Selection of oocytes, D20 serum, and added E2 and FSH or LH for IVM improved in vitro development of bovine embryos after IVF.
Incorporation and fate of specific sperm plasma membrane components following insemination as revealed by ultrastructural immunocytochemistry
Authors: Longo Frank J. 
Published: February 2005

AbstractStudies have been carried out to 1) further characterize sperm specific plasma membrane polypeptides (33 and 35 kDa) that are recognized by a monoclonal antibody previously described (Longo, 1989) and 2) follow the incorporation and dispersal of these proteins within plasmalemmae of monospermic and polyspermic sea urchin (Arbacia punctuluta) eggs and oocytes utilizing immunocytochemical methods at the ultrastructural level of observation. Only sperm labeled when incubated with monoclonal antibody to the 33 and 35 kDa proteins followed by colloidal gold‐tagged second antibody. Colloidal gold label was observed on the egg plasma membrane immediately after gamete membrane fusion; the amount and extent of label, i.e., the distance from the site of sperm incorporation, increased with time postinsemination. By 20 min postinsemination approximately one hemisphere of the inseminated egg/oocyte was associated with label. The expanding distribution of colloidal gold label on inseminated eggs and oocytes vs. time reflects the free diffusion of 33 and 35 kDa sperm surface proteins among egg/oocyte plasma membrane components. Label was also found in forming endocytotic vesicles, suggesting that sperm plasma membrane proteins may be internalized.
Monospermy and polyspermy after partial zona dissection of reinseminated human oocytes
Authors: Matter Henry , Talansky Beth , Gordon Jon , Cohen Jacques 
Published: February 2005

AbstractPartial zona dissection (PZD), a zona drilling method that uses mechanical force to open the zona pcllucida while the oocyte is shrunken in a sucrose solution, was applied to 121 unfertilized 1‐day‐old mature human oocytes prior to reinsemination. The 115 surviving oocytes were divided into three groups in which the duration between sucrose addition and reinsemination was varied: I) Less than 20 minutes, II) 21 to 45 minutes, and III) longer than 45 minutes. There was a trend toward a reduced fertilization and polyspermy rate as the time between sucrose exposure and insemination in sucrose‐free medium increased. Moreover, there was a statistically significant reduction in the number of oocytes penetrated by more than four sperm in group III (0/41) versus group I (7/34), and in group III, parthcnogcnctic development was observed. The incidence of polyspermy was also increased in oocytes manipulated more than 25 hours after retrieval compared with those manipulated 21‐24 hours after recovery, supporting the idea that aged oocytes have a reduced ability to block polyspermy. Oocyte contraction in sucrose occurred in three different patterns: spherical, pear‐shaped, and crenated. Both the fertilization and polyspermy rates were significantly higher in the crenated group.These results indicate that changes resembling activation occur following sucrose exposure and that sucrose activation can be used to reduce the risk of polyspermic fertilization in zona drilling procedures. In addition, the pattern of shrinkage in sucrose can be used as an indicator of oocyte receptivity to sperm penetration.
Glycerylphosphorylcholine diesterase activity of uterine fluid in conditions inducing secondary sex ratio change in the rat
Authors: Mitra Jayashree , Chowdhury Mridula 
Published: February 2005

AbstractFood restriction or decreasing the ratio of [Na+, K+] to Ca++, Mg++J in the diet of female rats before conception favoured the production of female offspring. Seven days of food deprivation decreased uterine fluid GPC diesterase activity in female rats, whereas long‐term food restriction (21 days), rather than decreasing the enzyme activity, apparently stimulated it. Dietary Ca and Mg supplementation, likewise, produced significant decrease in GPC di esterase activity. A significantly positive correlation was observed between the levels of uterine GPC diesterase and secondary sex ratio change, which indicates that these dietary techniques of sex‐ratio manipulation may modulate the uterine fluid GPC diesterase activity and influence sex determination in utero.
Cumulus cell dispersion induced by estradiol in mouse oviduct in vitro
Authors: Roblero Luis S. , Riffo Marta D. , Croxatto Horacio B. 
Published: February 2005

AbstractDispersion of cumulus cells in nonmated mice is completed in the oviduct 15–20 h after ovulation. Oviducts, isolated 1 h after ovulation (13 h post‐human chorionic gonaditropin), were cultured in vitro for 40 h. In these oviducts, denuded oocytes were first seen at 30 h of culture, indicating that cumulus dispersion proceeded at a slower rate in vitro.Oocyte denudation was accelerated in a dose‐dependent manner by the addition of estiadiol to the culture medium in which oviducts were incubated. The addition of progesterone or cycloheximide to the culture medium strongly inhibited oocyte denudation even in the presence of estradiol. When isolated cumuli were incubated in the absence of oviductal tissue, the rate of cell dispersion was slower than that of cumuli incubated inside the oviduct and the addition of estradiol to the culture failed to accelerate this process.On the basis of these data, we propose that cumulus cell dispersion is accelerated by an estrogen‐dependent protein produced by the oviduct and that this effect of estrogen is antagonized by progesterone.
Acrosomal function of human spermatozoa with normal and abnormal head morphology
Authors: Fukuda Masaru , Morales Patricio , Overstreet James W. 
Published: February 2005

AbstractIn previous studies we have shown differences in the function of morphologically normal and abnormal sperm by evaluating their flagellar movements and swimming trajectories. In this study we have compared the capability of morphologically normal and abnormal human sperm to undergo an acrosome reaction after incubation with human follicular fluid. Semen samples were studied from 6 research donors and 21 semen evaluation patients. All men had normal semen by clinical criteria. Semen was prepared either by a two‐step Percoll gradient centrifugation or the sperm werediluted, washed, and centrifuged three times. Sperm suspensions were incubated for 24 hours in a modified Tyrode's medium, containing 2.6% bovine serum albumin, prior to dilution with human follicuiar fluid. The percentage of acrosome reactions among viable sperm was assessed after 15 minutes using the supra vital Hoescht stain and fluoresceinated pea lectin. Sperm head size was measured with an ocular micrometer and normal values were defined as length 3–5 μm and width 2–3 μm. At least 25 viable normal sperm, and 25 viable abnormal sperm were analyzed for acrosome reactions on each slide. With Percoll separation the percentage of acrosome reactions (mean ± sem) for normal sperm was 38i ± 3% vs. 22 ± 2% for abnormal sperm (P < 0.005). After washing, the comparable values were 12 ± 1 % vs. 5 ± 1 % (P < 0.005). The incidence of spontaneous acrosome reactions (24 hours of incubation, no follicular fluid) was also higher for normal sperm than abnormal sperm (9 ± 1 % vs. 4 ± 1 %, P < 0.01). These data demonstrate an association between normal sperm morphology and acrosomal function.
Heterogeneity of epididymal spermatozoa of the hamster
Authors: Sullivan Robert , Robitaille Gilles 
Published: February 2005

AbstractEpididymal transit is involved in the acquisition of fertilizing ability by mammalian spermato‐zoa. The epididymis is implicated in the addition and/or modification of sperm surface proteins functionally involved in the processes leading to fertilization. In order to characterize these sperm components, we have studied the modifications of sperm membrane proteins during epididymal maturation. Hamster spermatozoa were collected from the caput, corpus, and proximal and distal cauda of the epididymis and submitted to a continuous Percoll gradient centrifugation. Independently of their epididymal origin, the spermatozoa were distributed into two bands of buoyant densities of 1,045 and 1,088 on the gradient. However, the proportion of more dense spermatozoa increased progressively along the epididymis. This proportion is 87–fold higher in the distal cauda compared to the caput. SDS‐PAGE electrophoresis demonstrated that although they originate from different regions of the epididymis, spermatozoa of the same density exhibited similar membrane protein patterns. In contrast, the electrophoretic patterns of spermatozoa with densities of 1,045 and 1,088 were very different One of these differences involves a 26 kD protein that is implicated in zona pellucida recognition (Sullivan and Bleau, Gamete Res 12.101–116,1985). This protein is present in dense sperm obtained from all regions of the epididymis but is absent in spermatozoa of low density. If we consider that this 26 kD protein is a ‘marker’ of surface changes occurring during sperm maturation, we can hypothesize that in the proximal segments of the epididymis a certain proportion of spermatozoa are indeed mature but that this proportion increases considerably along the epididymis.