bluuebunny/crossref_metadata_2025_split huggingface dataset:
Ejaculate‐hormonal traits in the leopard cat (Felis bengalensis) and sperm function as measured by in vitro penetration of zona‐free hamster ova and zona‐intact domestic cat oocytes
Authors: Howard Jogayle , Wildt David E. 
Published: February 2005

AbstractElectroejaculate traits and circulating follicle‐stimulating hormone (FSH), luteinizing hormone (LH), and testosterone concentrations were analyzed in adult leopard cats (Felis bengalensis), a rare felid species indigenous to east Asia. The ability of leopard cat sperm to bind and penetrate zona‐free hamster ova and zonaintact domestic cat oocytes in vitro was examined as a means of testing sperm function. The influence of culture media [Biggers, Whitten, Whittingham (BWW) vs. modified Krebs Ringer bicarbonate (mKRB)], seminal plasma removal, and swim‐up separation on sperm motility, sperm morphology, and oocyte penetration also were assessed. Sperm treatments included dilution of raw semen (DR), ejaculate centrifugation, and either resuspension (NS) or swim‐up processing (SU). The percentage of oocytes penetrated (penetration rate) and the number of penetrated sperm/oocyte (penetration index) were determined. Ejaculates from each male consisted of at least a 50% sperm motility rating, and hormone concentrations in individual males were unrelated to any ejaculate trait measured concurrently on the same day. The SU technique improved (P< 0.05) percent sperm motility and the proportion of structurally normal sperm compared to DR and NS treatments. Leopard cat spermatozoa were capable of binding to and penetrating hamster ova and domestic cat oocytes; however, penetration was influenced by culture medium and seminal processing. In the hamster assay, a higher (P< 0.05) penetration rate and penetration index were achieved when mKRB was used for gamete incubation instead of BWW. NS processing also increased (P< 0.05) overall penetration compared to DR and SU. In the cat oocyte assay, zona penetration rate was similar (P< 0.05) in the DR, NS, and SU aliquots; however, the zona penetration index was increased (P< 0.05) by the NS compared to the DR and SU treatments. This study (1) provides baseline ejaculate and endocrine norms for the leopard cat, (2) demonstrates that leopard cat sperm undergo nuclear decondensation in hamster ova and penetrate zona‐intact domestic cat oocytes, (3) indicates that seminal plasma removal enhances leopard cat sperm fertilizing ability and ovam penetration, and (4) suggests that heterologous oocyte penetration is effective for assessing factors influencing fertilization and sperm function in this nondomestic felid.
Regulation of mouse preimplantation development: Inhibitory effect of the calmodulin antagonist W‐7 on the first cleavage
Authors: Poueymirou William T. , Schultz Richard M. 
Published: February 2005

AbstractThe calmodulin antagonist W‐7 inhibits cleavage of 1‐cell mouse embryos in a concentration‐dependent manner. This inhibition is likely to be specific for a calmodulin‐mediated process, since the less active congener W‐5 does not inhibit cleavage when used at concentrations of W‐7 that do. Concentrations of W‐7 that inhibit cleavage and do not inhibit either the uptake or incorporation of [35S]methionine do inhibit [3H]thymidine incorporation; similar concentrations of W‐5 do not inhibit [3H]thymidine incorporation. Consistent with W‐7′s ability to inhibit cleavage by inhibiting DNA synthesis is that addition of W‐7 at later times that correspond with exit from S phase results in cleavage to the 2‐cell stage. Although W‐7 does inhibit cleavage of 1‐cell embryos, it does not inhibit transcriptional activation, which occurs in the 2‐cell embryo and is characterized by the synthesis of a group of proteins of Mr = 70,000. Results of these experiments suggest a role for calmodulin in the first cell cycle of the mouse embryo and provide another example in which zygotic gene activation is not dependent on progression through the first cell cycle.
Early nuclear events of in vitro fertilization in the domestic fowl (Gallus domesticus)
Authors: Nakanishi Akira , Utsumi Kyozo , Iritani Akira 
Published: February 2005

AbstractIn vitro fertilization in the domestic fowl (Gallus domesticus) was investigated by observation of the early nuclear events. Ova retrieved from the fimbria following ovulation were inseminated in vitro with 106 − 107 spermatozoa in Dulbecco's modified Eagle's medium (DMEM) for 10 min and then further incubated in DMEM + albumen for 1, 2, 3, or 4 hr. These eggs were histologically examined by epifluorescent microscopy after staining with 4′,6′‐diamidino‐2‐phenylindole (DAPI). Nuclei of spermatozoa at various stages of transformation were observed in the ova incubated for 1–3 hr. Close pairing of two pronuclei, presumed to be male and female juxtaposition, was detected in ova incubated for 4 hr. These data provide direct evidence for the in vitro fertilization of fowl eggs and suggested that the early process of in vitro fertilization is comparable to that of in vivo fertilization.
Identification of heparin‐binding proteins in bovine seminal plasma
Authors: Chandonnet L. , Roberts K. D. , Chapdelaine A. , Manjunath P. 
Published: February 2005

AbstractA group of four similar proteins, BSP‐A1, BSP‐A2, BSP‐A3, and BSP‐30‐kDa, represent the major acidic proteins found in bovine seminal plasma (BSP). These proteins are secretory products of the seminal vesicles; they bind to spermatozoa upon ejaculation and could represent decapacitation factors. It has been shown that the glycosaminoglycans present in the female reproductive tract are involved in the capacitation of spermatozoa. Therefore, it was of interest to investigate whether BSP‐A1, ‐A2, ‐A3, and ‐30‐kDa proteins of bovine seminal fluid interact with heparin. Chromatography of alcohol precipitates of bovine seminal fluid on a heparin–Sepharose column resolved these proteins into three peaks. Peaks 1 and 2 (retarded proteins) were eluted upon extensive washing of the column with 0.05 M phosphate buffer, pH 7.4 (equilibrating buffer), and accounted for approximately 25% of the applied proteins. Proteins in peak 3 represented adsorbed proteins and were eluted with phosphate buffer containing 1 M NaCl. Proteins in each peak were characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE) under reducing conditions. Peak 1 contained proteins with molecular weights ranging from 8 to 350 kDa, peak 2 contained a single protein with a molecular weight of 14 kDa, and peak 3 contained proteins with molecular weights of 15.5, 16, 25, and 30 kDa. The proteins in peak 3 were further resolved into unadsorbed (peak 4) and adsorbed (peak 5) proteins on a gelatin–Agarose column. Separation of the proteins of peak 3 and peak 5 by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and reducing agents followed by transfer to nitrocellulose and probing with antibodies against the previously well‐characterized BSP proteins indicated the presence of BSP‐A1, BSP‐A2, BSP‐A3, and BSP‐30‐kDa proteins. These results demonstrate that BSP‐A1, ‐A2, ‐A3, and ‐30‐kDa proteins bind to heparin or glycosaminoglycans and we postulate interaction of these spermatozoa‐bound proteins with glycosaminoglycans in the female reproductive tract.
Immunocytochemical studies of hamster oocyte activation
Authors: Sinosich M. J. , Lanzendorf S. E. , Hodgen G. D. 
Published: February 2005

AbstractBy indirect immunofluorescence, using rabbit anti‐heparin‐binding placental protein (HBPP) antiserum, we studied HBPP expression by physiologically and non‐physiologically (microsurgically) activated hamster gametes. Whereas mature gametes (sperm, metaphase II oocytes) were negative, in vivo conceived preimplantation embryos, from pronuclear to two‐ and four‐cell stages, were HBPP positive. No HBPP was demonstrated in the zona pellucida, but HBPP‐dependent immunofluorescence was localized in the perivitelline space. Oocytes incubated with hyaluronidase demonstrated variable responses from negative to positive. (Diluent or sperm) microinjected oocytes were all activated and HBPP positive within 4 h after stimulation. Thus neither activation by microinjection nor HBPP expression required paternal gametes. These kinetics suggest that HBPP may be a cortical granule secretogogue which can be applied to monitor oocyte responses during in vitro manipulations.
Injection of isolated centriolar complex to induce mitosis in starfish eggs
Authors: Yamada Hisafumi , Kuraishi Ritsu , Amikura Reiko M. , Nagano Hiroshi 
Published: February 2005

AbstractWhen centriolar complex isolated from starfish spermatozoa was injected into starfish immature oocytes (fully grown, germinal vesicle stage) followed by treating the latter with 1‐methyladenine, mitotic events such as condensation and division of chromosomes of the female pronucleus and cytokinesis following completion of meiosis were observed. No cortical reaction detected in the oocytes. Essentially the same was noted for mature oocytes (pronuclear stage) into which the centriolar complex had been injected. The oocytes that had received sperm tail fraction or buffer alone did not initiate cleavage. It would thus appear that sperm centriolar complex is significantly essential to the initiation of cleavage.
Expression of genes for insulin and insulin‐like growth factors and receptors in early postimplantation mouse embryos and embryonal carcinoma cells
Authors: Telford Nancy A. , Hogan Aileen , Franz Carolyne R. , Schultz Gilbert A. 
Published: February 2005

AbstractThe expression of genes for insulin and insulin‐like growth factors (IGFs) and their receptors was examined in early postimplantation mouse embryos and differentiating F9 embryonal carcinoma cells using mRNA phenotyping. Messenger RNA phenotyping involves the reverse transcription of RNA followed by amplification of specific target cDNA sequences using the polymerase chain reaction (PCR). The identities of the resulting PCR fragments were confirmed using at least two of the following methods: (1) size determination by agarose gel electrophoresis, (2) the presence of diagnostic restriction sites, (3) hybridization with radio‐labeled cDNA probes, (4) sequencing of the PCR fragment. Transcripts for insulin receptors, IGF‐I receptors, and IGF‐II receptors were detected in RNA samples from day 7.5 to day 9.5 mouse embryos and in F9 cells, although the level of insulin receptor mRNA in F9 cells was very low. Transcripts for both IGF‐I and IGF‐II ligands were also detectable in the embryo and F9 RNA samples, but transcripts for insulin ligand were undetectable in either set of material. The results suggest that insulin does not act as a paracrine or autocrine growth factor in early postimplantation embryos or F9 cells but that both embryos and F9 cells have the potential to respond to exogenous (e.g., maternal) sources of insulin. Both IGF‐I and IGF‐II could act as paracrine or autocrine growth factors, and IGF‐II is the more abundant growth factor in differentiating F9 cells.
Expression of the insulin‐like and platelet‐derived growth factor genes in human uterine tissues
Authors: Boehm Keith D. , Daimon Makoto , Gorodeski Israel G. , Sheean Leon A. , Utian Wulf H. , Ilan Judith 
Published: February 2005

AbstractThe human uterus repeatedly exhibits cyclic biochemical and cytological changes during the reproductive period of life. These changes are the result of a well‐characterized endocrine network involving the hypothalamus, pituitary, and ovary. The exact nature of the mechanism(s) by which the sex steroids act on the uterus remains to be elucidated. Possible local mediators of hormonal action on the uterus include polypeptide growth factors. Using the method of RNA transfer blot hybridization, we have analyzed tissue samples from the cycling human endometrium and tissue samples of human myometrium and myometrial benign tumor (leiomyoma) for the presence of platelet‐derived growth factor (PDGF) and insulin‐like growth factor (IGF) RNA. All the uterine tissues examined possessed RNA for PDGF‐B chain and IGF‐I and ‐II. Two transcripts were observed for PDGF‐B chain, four were observed for IGF‐I, and eight were observed for IGF‐II. Overall, the relative abundance of PDGF‐B chain RNA was consistent in all of the uterine tissues examined. In contrast, IGF RNA relative abundance varied. IGF‐I RNA was highest in late proliferative stage endometrium, and IGF‐II RNA was highest in early proliferative stage endometrium. Both IGF‐I and IGF‐II RNAs were greater in amount in leiomyoma than in myometrium. The increased IGF‐I RNA in late proliferative–stage human endometrium correlates with the known elevation of estradiol secretion by the ovary and the increased concentration of uterine estradiol receptors during this stage of the menstrual cycle. Furthermore, these data are consistent with reports of increased IGF‐I RNA in the rat uterus in response to administration of estradiol. Elevated levels of IGF RNA in leiomyomas may be related to the genesis and/or progression of these myometrial tumors.
Assessment of the viability and acrosome status of fresh and frozen‐thawed human spermatozoa using single‐wavelength fluorescence microscopy
Authors: Centola G. M. , Mattox J. H. , Burde S. , Leary J. F. 
Published: February 2005

AbstractThe purpose of this study was to use fluorescence microscopy to determine the viability and acrosome status of fresh and frozen‐thawed human spermatozoa. Sperm cells were stained with the viability stains Hoechst 33258 (H33258) alone, or propidium iodide (PI) alone, and PI in combination with FITC‐conjugated Pisum sativum agglutinin (PSA). The PSA stains the acrosome contents of permeabilized acrosome‐intact sperm. Viability by fluorescence microscopy was compared to conventional eosin nigrosin staining. The overall viability using H33258 was not significantly different from that using PI. Therefore, PI was used in combination with PSA for simultaneous measurement of viability and acrosome status at the same excitation wavelength (488 nm). By combining PI and PSA, four subgroups of cells could be detected: group I, PI‐neg/PSA‐neg—viable, physiologic acrosome reacted (AR); group II, PI‐neg/PSA–pos—viable, non‐AR; group III, PI‐pos/PSA‐neg–‐nonviable, non‐AR; group IV, PI‐pos/PSA–neg—nonviable, degenerative AR. The postthaw sperm exhibited a significantly greater percent of sperm that were acrosome reacted (both viable and degenerative) (groups I and IV) than the fresh semen. We conclude that frozen‐thawed sperm may undergo premature break‐down of the acrosome prior to interaction with the oocyte, thus explaining the reduced fertility potential of cryopreserved semen.
Relationships between follicular fluid steroid hormone concentrations, oocyte maturity, in vitro fertilization and embryonic development in the rhesus monkey
Authors: Morgan Patricia M. , Boatman Dorothy E. , Bavister Barry D. 
Published: February 2005

AbstractOocytes and matched samples of follicular fluid (FF) were obtained from 70 follicles of five rhesus monkeys stimulated with either pregnant mare serum gonadotropin or human menopausal gonadotropin. Follicular aspiration was performed 30–32 h after human chorionic gonadotropin administration. The concentrations of estradiol (E2), progesterone (P), testosterone (T), and dihydrotestosterone (DHT) in FF were measured. Twenty‐six percent of oocytes were classified as mature (M), 41% matured in vitro (Miv), 13% were dysmature, and 20% atretic. M oocytes were associated with significantly higher levels of P and a higher P:E2 ratio. There were no differences in hormone levels associated with fertilized and nonfertilized oocytes. Thirty‐five embryos developed to the six‐ to eight‐cell stage in vitro, of which 13 exhibited optimal cleavage rates. Significantly lower levels of E2 and higher P:E2 ratios were associated with the more rapidly cleaving embryos. Proportionally more embryos showing optimal cleavage rates developed from M compared to Miv oocytes, and only embryos derived from M oocytes developed to blastocysts in culture. Optimal cleavage rates to the six‐ to eight‐cell stage in vitro, rather than fertilization rates, are a better indicator of (subsequent) developmental capacity, and, in this study, embryonic development was closely associated with the maturity of the oocyte at recovery.
Localization of bindin expression during sea urchin spermatogenesis
Authors: Nishioka David , Ward Rita D. , Poccia Dominic , Kostacos Connie , Minor Joseph E. 
Published: February 2005

AbstractExpression of the bindin gene was examined in testicular cells of the sea urchin Strongylocentrotus purpuratus. In situ hybridization studies, using an 35S‐labeled antisense RNA probe transcribed from a bindin cDNA, reveal that bindin mRNAs are localized in spermatogenic cells displaced towards the lumens of maturing testicular acini. Little or no hybridization is observed in spermatogenic cells displaced towards the perivisceral epithelium or in somatic cells of the testis. A similar localization of the bindin protein itself is observed using a rhodamine‐conjugated polyclonal antibody against bindin, which shows a punctate immunofluorescence pattern in late spermatogenic cells. Immunogold labeling of ultrathin sections and electron microscopy reveal that this punctate immunofluorescence is an apparent result of localized deposits of bindin in intracellular vesicles. Through the terminal stages of spermatogenesis, these bindin‐containing vesicles apparently fuse to form the single acrosomal vesicle of the mature spermatozoon. These results indicate (1) that bindin mRNAs are transcribed relatively late in spermatogenesis, (2) that bindin is translated soon after production of its mRNA, (3) that bindin quickly associates with intracellular vesicles during or soon after its synthesis, and (4) that these vesicles fuse to form the single acrosomal vesicle during the terminal stage of spermatogenesis.
Regulation of laminin gene expression in the expansion of mouse blastocysts
Authors: Kang Hae Mook , Kim Kyungjin , Kwon Hyuk Bang , Cho Wan Kyoo 
Published: February 2005

AbstractThis study aims to elucidate gene expression of laminin and its role in expansion of the blastocyst during mouse early embryogenesis. The gene expression of laminin, in particular the B1 subunit and the synthesis of laminin polypeptides, was examined during the expansion of blastocyst by a RNA‐blot hybridization with 32P‐labeled laminin B1 cDNA and immunoprecipitation followed by a SDS‐PAGE, respectively. Laminin B1 transcript was actively expressed in the blastocyst stage of embryos. The gene expression of laminin B1 and the synthesis of laminin protein were also increased when blastocyst was expanded. Treatments of cAMP analogue, isobutylmethylxanthine, forskolin, and cholera toxin, which are known to stimulate the blastocyst expansion, increased laminin B1 transcript levels and synthesis of laminin polypeptides. Treatment with retinoic acid, a known regulator of laminin gene expression, not only increased the gene expression of laminin but stimulated the blastocoel expansion without a significant increase in intracellular cAMP levels.These results indicate that laminin gene expression may play an important role in the process of blastocyst expansion in the mouse preimplantation embryos.
Development of preimplantation rabbit embryos in uterine flushing–supplemented culture media
Authors: Fischer Bernd , Jung Thomas , Hegele‐Hartung Christa , Beier Henning M. 
Published: February 2005

AbstractThe development of cultured rabbit preimplantation embryos grown in standard media (Ham's F‐10 or BSM II supplemented with bovine serum albumin (BSA) or homologous serum) or in Ham's medium supplemented with uterine flushings was compared. The uterine flushings derived from donors of 0.5–6 years of age. Uterine flushing supplemented media were used natively or after treatments like sterilization by filtration, lyophilization, three times freezing/thawing, heat denaturation, dialysis, or ultrafiltration. Compared with in vivo controls, embryonic growth was substantially reduced during in vitro culture, demonstrably by smaller diameters and impaired cell proliferation (measured by thymidine incorporation). The growth retardation was more pronounced in blastocyts (recovered at day 4 post coitum [p.c.]) than in morulae (recovered at day 3 p.c.). Development in uterine flushing media was notably better than in standard media but did not comply with in vivo development. Highest thymidine incorporation was observed in media with increased concentrations of uterine secretions and after sequential supplementation of flushings from subsequent progestational stages. Advanced donor ages, heating up to 80°C, freezing, and lyophilizing did not affect incorporation data statistically significantly, whereas sterilization by filtration, ultrafiltration, and dialysis led to a significantly reduced thymidine incorporation in the cultured embryos. The positive effects of uterine flushing supplementation are attributed to the supply of components more adjusted to the needs of the cultured embryos and/or to a reduction of pathological effects in vitro like washing out of nutritive and regulatory components from the embryo into the surrounding culture medium.
Combined effects of protein synthesis and phosphorylation inhibitors on maturation of mouse oocytes in vitro
Authors: Motlík Jan , Rimkevičová Zora 
Published: February 2005

AbstractIn denuded mouse oocytes, neither 3 nor 5 hours of preincubation in dbcAMP (1 mM) and cycloheximide (10 μg/ml), followed by further 3 hours in cycloheximide only, lowered the rate of GVBD (93% and 92%, respectively). It means that 3 and 5 hours preincubation in cycloheximide did not impair the ability of mouse oocytes to resume meiosis in medium with the protein synthesis inhibitor. To test the combined effects of inhibition of protein phosphorylation and protein synthesis, oocytes were cultured for 3, 4, or 5 hours in 2 mM of 6‐DMAP and subsequently for 3 hours in 10 μg/ml cycloheximide. The incubation in 6‐DMAP for 4 or 5 hours diminished (63% or 35% of GVBD, respectively) the ability of mouse oocytes to resume meiosis when subsequent protein synthesis was blocked by cycloheximide. However, the highly condensed bivalents were always visible in GVs. Thus the above treatment did not prevent chromatin condensation although GVBD was blocked.