bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of immunization of hamsters against recombinant P26h on fertility rates
Authors: Gaudreault C , Montfort L , Sullivan R
Published: March 2004

Despite the various contraceptive methods available, an effective and inexpensive method remains to be established. Immunocontraception may help to achieve this goal. P26h has been proposed as a candidate for the development of a male contraceptive vaccine. P26h, a hamster sperm protein, interacts with the zona pellucida. Furthermore, in vivo fertilization can be blocked completely by active immunization of male hamsters against P26h. Maltose binding protein (MBP)-P26 shares antigenic determinants with the native P26h present on cauda epididymal spermatozoa. The aim of the present study was to reproduce the immunocontraceptive properties of native P26h by immunizing male hamsters against a recombinant P26h fused with a maltose binding protein (MBP). Active immunization of male hamsters with the MBP-P26h showed that specific anti-P26h circulating IgGs could be generated. Mating of immunized male hamsters with superovulated females resulted in a significant decrease, 20-25%, in the fertilization rate. This result is in agreement with results from in vitro sperm-zona pellucida binding assays. Indeed, the anti-recombinant P26h IgGs showed lower inhibitory properties when compared with anti-native P26h IgG. Despite the high anti-P26h IgG titres generated in hamsters, histological studies showed that active immunization has no pathological sequelae to the reproductive tissues. The potential of P26h as a candidate for a contraceptive vaccine is discussed.
Quantitative assessment of testicular germ cell production and kinematic and morphometric parameters of ejaculated spermatozoa in the grey mouse lemur, Microcebus murinus
Authors: Aslam H , Schneiders A , Perret M , Weinbauer GF , Hodges JK
Published: March 2004

Germ cell production and organization of the testicular epithelium in a prosimian species, the grey mouse lemur, Microcebus murinus, was investigated to extend knowledge of comparative primate spermatogenesis. In addition, semen samples collected from adult male lemurs (body weight 53-92 g; n = 16) by rectal probe electroejaculation were evaluated using computer-assisted morphometric and kinematic analysis of spermatozoa. Epididymidal spermatozoa were collected from six animals after hemicastration; the testes were weighed and prepared for stereological analysis and flow cytometry. The relative testis mass (as a percentage of body weight) ranged between 1.17 and 5.6%. Twelve stages of testicular seminiferous epithelium as described for macaques were applied and only a single stage was observed in most of the seminiferous tubule cross-sections. On average (mean SD), a single testis contained 1870 +/- 829 x 10(6) germ cells and 35 +/- 12 x 10(6) Sertoli cells. Germ cell ratios (preleptotene:type B spermatogonia = 2, round spermatid:pachytene = 3; elongated spermatid:round spermatids = 1) indicated high spermatogenic efficacy. Sperm head dimensions and tail lengths of the ejaculated and epididymidal spermatozoa were similar. Percentages of defects (neck/mid-piece and tail) were low ( 10%) and similar for ejaculated and epididymidal spermatozoa. Spermatozoa were highly motile, characterized by extensive lateral head displacement, but relatively low progressive motility. In conclusion, the grey mouse lemur has unusually large testes with a highly efficient spermatogenic process and large sperm output. These features, together with the high proportion of morphologically normal and highly motile spermatozoa in the ejaculates, indicate that Microcebus murinus is a species in which sperm competition after ejaculation is likely to occur. The predominantly single spermatogenic stage system seems to be an ancestral feature among primates.
Tumour suppressor genes in sporadic epithelial ovarian cancer
Authors: Liu Y , Ganesan TS
Published: March 2004

Ovarian cancer is the most frequent cause of death from gynaecological malignancies in the western world, and sporadic epithelial ovarian cancer is its most predominant form. The aetiology of sporadic ovarian cancer remains unknown. Genetic studies have enabled a better understanding of the evolution of tumour progression. A major focus of research has been to identify tumour suppressor genes implicated in sporadic ovarian cancer over the past decade. Several tumour suppressor genes have been identified by strategies such as positional cloning and differential expression display. Further research is warranted to understand fully their contribution to the pathogenesis of sporadic ovarian cancer.
Characterization and biological roles of oviduct-specific, oestrogen-dependent glycoprotein
Authors: Buhi WC
Published: March 2004

During late follicular growth, oestrus, fertilization and early embryonic development, the oviduct, under specific hormonal control, produces fluid and contributes secretory macromolecules that optimize the microenvironment for gamete maturation and transport, fertilization and early cleavage-stage embryonic development. This review describes the state of knowledge concerning the physiological and biochemical characterization of the major oviduct secreted protein, the oviduct-specific, oestrogen-dependent glycoprotein. The identification, localization within the oviduct, binding and association with oocytes, embryos and spermatozoa, hormonal control of gene expression, cloning, gene organization, protein sequences and post-translational modifications of oviduct-specific, oestrogen-dependent glycoprotein are discussed. Identification of biological functions for this glycoprotein, its interactions with spermatozoa, oocytes and embryos and its potential as a paracrine regulator of fertilization and development are also discussed.
Steroidogenesis in bovine granulosa cells: the effect of short-term changes in dietary intake
Authors: Armstrong DG , Gong JG , Gardner JO , Baxter G , Hogg CO , Webb R
Published: March 2004

The nutritional status of a cow is a key factor in the regulation of both follicle growth and oocyte quality. In this study, the effect of diets designed to increase circulating insulin and insulin-like growth factor I (IGF-I) concentrations on steroid production by granulosa cells in vitro was examined to analyse the mechanisms through which these changes occur. Hereford x Friesian heifers (n = 24) were offered maintenance or twice maintenance diets during the experimental period (17 days). Circulating concentrations of FSH did not differ between the two dietary groups, whereas insulin and IGF-I concentrations showed significant diet x day of oestrous cycle interactions. Ovaries were collected on day 3 of the first follicle wave after synchronization of oestrus. Granulosa cells were isolated from small (1-4 mm) and medium-sized (4-8 mm) follicles and cultured in the presence of long R3-IGF-I or bFSH or both. After 4 days in culture, granulosa cells isolated from small follicles, but not medium-sized follicles, collected from cattle offered the twice maintenance diet secreted significantly higher (P < 0.05) amounts of oestradiol compared with granulosa cells collected from cattle offered the maintenance diet. The effect was apparent in either the presence or absence of FSH and long R3-IGF-I. This nutritional effect on aromatase activity in granulosa cells was not apparent after day 6 of culture. There was no effect of diet on progesterone production by granulosa cells after 4 or 6 days of culture. These results support the hypothesis that dietary-induced changes in circulating insulin and IGF-I concentrations have a direct effect on the steroidogenic potential of bovine granulosa cells from small follicles. The dietary-induced increases in aromatase activity in small follicles combined with the increased concentration of metabolic hormones are possible mechanisms through which short-term changes in nutrition may affect follicle dynamics.
Expression of a blood-brain barrier-specific antigen in the reproductive tract of the male rat
Authors: Ghabriel MN , Lu JJ , Hermanis G , Zhu C , Setchell BP
Published: March 2004

The endothelial barrier antigen (EBA) is a protein expressed specifically by the endothelial cells of the rat brain barrier vessels. This antigen has been described as a 'barrier protein' and is used as a marker for the competent blood-brain barrier. A blood-testis barrier has also been described. However, unlike the blood-brain barrier, which is formed by endothelial cells, the blood-testis barrier is formed mainly by the Sertoli cells, which provide an isolated environment for spermatogenic cells within the seminiferous tubules. Testicular blood vessels express the erythroid glucose transporter protein and other markers, which are strongly expressed in brain blood vessels, and may contribute to the blood-testis barrier. This study was carried out to determine whether Sertoli cells or testicular blood vessels express EBA. Tissues of other organs were used as controls for EBA expression. EBA was expressed by the endothelial cells in most microvessels of the testis, and in a few vessels of the epididymis, seminal vesicle, prostate gland, vas deferens and bladder-neck region. Furthermore, EBA was strongly and consistently detected in epithelial cells of the rete testis and dorsolateral prostate gland, and in a few epithelial cells of the ventral prostate gland, the seminal vesicle and the coagulating gland. However, Sertoli cells, which are the main site of the blood-testis barrier, were negative for EBA. In conclusion, EBA may have a wider role in rat tissues than has been previously appreciated.
Cytokine expression in the uterus of mice with pregnancy loss: effect of maternal immunopotentiation with GM-CSF
Authors: Savion S , Zeldich E , Orenstein H , Shepshelovich J , Torchinsky A , Carp H , Toder V , Fein A
Published: March 2004

It is believed that failure of the maternal immune system to actively support embryonic development, through production of the appropriate cytokine network, might be responsible for embryonic death. Thus, the aim of this study was to evaluate the possible involvement of cytokines such as tumour necrosis factor alpha (TNF-alpha) and transforming growth factor beta2 (TGF-beta2), which are crucial for normal embryonic development, in the early stages of mechanisms that mediate induced pregnancy loss. The early stages of the resorption process induced by lipopolysaccharide (LPS) were characterized by blood accumulation in the vicinity of the embryo, preceding any visible embryonic damage. At that time, immunohistochemical analysis revealed an increased expression of TNF-alpha in the primary and secondary decidua, which was reduced as the resorption process was completed. In contrast, TGF-beta2 expression was decreased in the primary and secondary decidua, as well as in the glandular epithelium, at all the times assessed. Maternal immunopotentiation with granulocyte macrophage-colony stimulating factor (GM-CSF), which controls maternal immune activities supporting normal embryonic development, decreased the resorption rate in LPS-treated mice while normalizing the expression of TNF-alpha and TGF-beta2 in the uterus of these animals throughout the ongoing resorption process. These results indicate a possible role for maternal immunopotentiation with GM-CSF in the mechanisms mediating the early stages of pregnancy loss, possibly via modulation of TNF-alpha and TGF-beta2 activity.
Abnormal expression of acid glycosidases in seminal plasma and spermatozoa from infertile men with varicocele
Authors: Corrales JJ , Burgo RM , Galindo P , Munoz-Barroso I , Miralles JM , Villar E
Published: March 2004

The activities of acid beta-glucuronidase, alpha-mannosidase, alpha-glucosidase, alpha-galactosidase, beta-galactosidase and beta-N-acetylglucosaminidase were analysed in seminal plasma and spermatozoa from 26 infertile men with varicocele and from 36 men of normal fertility. Semen samples from ten men with non-obstructive azoospermia were used as control specimens that contained the other components of semen. Spermatozoa were solubilized by both physical (homogenization) and chemical (Triton-X100) methods to obtain the soluble and non-soluble fractions. The activities of several glycosidases measured both in seminal plasma and spermatozoa were directly correlated with the numbers of spermatozoa and sperm motility, confirming previous studies. As some infertile patients with varicocele have normal semen parameters, whereas others have low numbers of spermatozoa and low sperm motility, the varicocele patients were prospectively divided into two groups: one (n = 15) with normal spermiograms and the other (n = 11) with abnormal spermiograms. The activities (expressed in mU ml(-1)) of alpha-mannosidase, beta-galactosidase and beta-N-acetylglucosaminidase in seminal plasma of normozoospermic infertile patients with varicocele were significantly higher than those of fertile controls, but not when expressed in U per 10(8) spermatozoa. The activities of beta-glucuronidase, alpha-mannosidase, beta-galactosidase and beta-N-acetylglucosaminidase in seminal plasma when expressed in U per 10(8) spermatozoa in varicocele patients with abnormal spermiograms were significantly higher than in those of men of normal fertility. The activity of alpha-mannosidase in the soluble fraction of sperm homogenates, expressed as U per 10(8) spermatozoa, was significantly higher in infertile patients with varicocele and abnormal spermiograms than in controls. In the non-soluble fraction of spermatozoa from infertile patients with varicocele, there was an increase in the expression of beta-galactosidase and beta-N-acetylglucosaminidase activities compared with the fraction of spermatozoa from fertile subjects. In summary, infertile patients with varicocele displayed an overexpression of acid alpha-mannosidase, beta-galactosidase and beta-N-acetylglucosaminidase activities in seminal plasma and spermatozoa that may be associated with functional defects in spermatozoa as these glycosidases play an important role in mammalian fertilization.
Role of PGF2alpha and oxytocin in parturition in the brushtail possum (Trichosurus vulpecula)
Authors: Veitch C , Brown L , Sernia C , Gemmell RT
Published: March 2004

Maturation of the fetal pituitary and adrenal glands allows the secretion of cortisol, which in turn leads to an increase in prostaglandin and mesotocin production. The production of prostaglandin and mesotocin results in an increase in uterine contractions and initiates birth in marsupials. The major metabolite of PGF(2alpha), 13,14-dihydro-15-keto-prostaglandin F(2alpha) (PGFM), has been found in the plasma of the possum at the time of birth and administration of PGF(2alpha) to female possums induced the adoption of the birth position. Evidence that mesotocin is an integral hormone of birth in the tammar wallaby indicates that both PGF(2alpha) and mesotocin or oxytocin are required for marsupial birth. The presence of PGF(2alpha) receptors in the uterus and corpus luteum of the possum, and the in vitro uterine responsiveness to PGF(2alpha) or oxytocin, were examined. PGF(2alpha) receptors were not observed in possum uteri and the inability of PGF(2alpha) to cause contractions indicates that PGF(2alpha) is not involved directly in contraction of the uterus at parturition. The presence of oxytocin and mesotocin receptors in the uterus of possoms and the ability of oxytocin to induce uterine contraction in vitro supports the view that mesotocin is required for expulsion of the young from the uterus. Low numbers of PGF(2alpha) receptors were found in the possum corpus luteum at birth, indicating an involvement of PGF(2alpha) in regression of the corpus luteum.
Developmental competence in vivo and in vitro of in vitro-matured equine oocytes fertilized by intracytoplasmic sperm injection with fresh or frozen-thawed spermatozoa
Authors: Choi YH , Love CC , Love LB , Varner DD , Brinsko S , Hinrichs K
Published: March 2004

This study was undertaken to evaluate the development of equine oocytes in vitro and in vivo after intracytoplasmic sperm injection (ICSI) with either fresh or frozen-thawed spermatozoa, without the use of additional activation treatments. Oocytes were collected from ovaries obtained from an abattoir and oocytes classified as having expanded cumulus cells were matured in M199 with 10% fetal bovine serum and 5 microU FSH ml(-1). After 24-26 h of in vitro maturation, oocytes with a first polar body were selected for manipulation. Fresh ejaculated stallion spermatozoa were used for the experiment after swim-up for 20 min in sperm-Tyrode's albumen lactate pyruvate. Frozen-thawed spermatozoa from the same stallion were treated in a similar way. Spermatozoa were immobilized and injected into the oocytes using a Piezo drill. Presumptive zygotes were cultured in G1.2 medium for 20 or 96 h after the injection was administered, or were transferred to the oviducts of recipient mares and recovered 96 h later. In addition, bovine oocytes with first polar bodies were injected with the two types of stallion spermatozoa and fixed 20 h after injection to examine pronuclear formation. Fertilization rate (pronucleus formation and cleavage) at 20 h after injection of spermatozoa was not significantly different between fresh and frozen-thawed sperm groups in either equine or bovine oocytes. Pronucleus formation after injection of spermatozoa into bovine oocytes was significantly higher than that for equine oocytes (P < 0.05). There were no significant differences in cleavage rate or average number of nuclei at 96 h between equine oocytes injected with fresh or frozen-thawed spermatozoa. However, embryos developed in vivo for 96 h had a significantly higher number of nuclei in both sperm treatments compared with those cultured in vitro. These results indicate that good activation rates may be obtained after injection of either fresh or frozen-thawed equine spermatozoa without additional activation treatment. Injection of frozen-thawed equine spermatozoa results in similar embryo development to that obtained with fresh equine spermatozoa. In vitro culture of equine zygotes in G1.2 medium results in a similar cleavage rate but reduced number of cells compared with in vivo culture within the oviduct. Bovine oocytes may be useful as models for assessing sperm function in horses.
Matrix metalloproteinases (MMPs) in the endometrium of bitches
Authors: Chu Py PY , Salamonsen LA , Lee CS , Wright PJ
Published: March 2004

The relationships between activities of matrix metalloproteinases (MMPs) in the canine uterus and the occurrence of degeneration of the luminal epithelium, cystic endometrial hyperplasia, pyometra and uterine remodelling post partum were determined. Mature bitches (n = 10) were ovariectomized, treated with hormones (oestradiol benzoate, progestagen) and investigated at stages simulating pro-oestrus (n = 2), oestrus (n = 2), dioestrus (n = 2), and mid- (n = 2) and late (n = 2) anoestrus (3 and 9 weeks, respectively, after cessation of treatment with progestagen). Untreated bitches (n = 1 per group) served as controls (Expt 1). An additional 10 ovariectomized bitches, at the end of treatment-induced simulated dioestrus, were treated each day for a further 3 weeks either with the same dose (standard dose, n = 3), a decreased dose (n = 3) or an increased dose (n = 3) of progestagen, or no treatment (withdrawal dose, n = 1). These bitches were then investigated (Expt 2). A suture was placed in the lumen of one uterine horn of another five bitches at ovariectomy. Three of these bitches were treated to induce simulated dioestrus and two bitches served as untreated controls. In the hormone-treated bitches, the suture resulted in cystic endometrial hyperplasia in one bitch and in cystic endometrial hyperplasia with pyometra in two bitches. The control bitches showed no cystic endometrial hyperplasia or pyometra (Expt 3). Four intact bitches were studied at 2 (n = 1), 3 (n = 2) and 11 (n = 1) weeks post partum. Uterine tissues were also collected from two bitches with naturally occurring cystic endometrial hyperplasia with pyometra (Expt 4). All uteri were examined histologically and the activities of MMP-2, -7 and -9 (latent and active forms) were assessed using zymography of extracts of endometrium. In Expts 1 and 2, marked degeneration of the luminal epithelium in six of 25 bitches (simulated mid-anoestrus, withdrawal dose and decreased dose groups) was not associated with changes in MMP activities. Markedly increased activities of MMP-2 (active form), -7 (latent form) and -9 (active and latent forms) were observed in the bitches with cystic endometrial hyperplasia with pyometra (but not with cystic endometrial hyperplasia alone) and in the bitches at 2 and 3 weeks post partum (but not at 11 weeks post partum). These results indicate that MMPs are not involved with degeneration of the luminal epithelium, but are involved with uterine remodelling in the postpartum bitch and with cystic endometrial hyperplasia with pyometra.
REDOX regulation of early embryo development
Authors: Harvey AJ , Kind KL , Thompson JG
Published: March 2004

Preimplantation embryonic development is associated with a change in preference in energy metabolism pathways. Although oxidative phosphorylation is obligatory in most species throughout preimplantation development, an increasing role for energy derived from glycolysis is associated with compaction and blastulation. Such a shift in metabolic pathway preference is desirable as the embryo faces an increasingly hypoxic environment in utero. We hypothesize that this shift in metabolic preference is associated with a change in the reduction-oxidation (REDOX) state within the embryo, affecting not only the energy production required for development, but also the activity of REDOX-sensitive transcription factors, which may alter gene expression patterns. Shifts in intracellular REDOX state may also contribute to spatial differences in cell activity, especially after compaction, and perhaps even major embryonic events such as fertilization, genome activation and cellular differentiation.
Behavioural significance of prolactin signalling in the central nervous system during pregnancy and lactation
Authors: Grattan DR
Published: March 2004

The role of prolactin in the regulation of mammary gland development and function during pregnancy and lactation is well established. However, in addition, prolactin appears to have a much wider role in the physiology of lactation. There is widespread expression of prolactin receptors in the hypothalamus during lactation, indicative of a multi-faceted role for prolactin in regulating hypothalamic function. During pregnancy and lactation, the maternal brain undergoes structural and functional modification, allowing the establishment of appropriate behaviour to feed and nurture the offspring, to adjust to the nutritional and metabolic demands of milk production, and to maintain appropriate hormone secretion to allow milk synthesis, secretion and ejection. The coordination of such a range of neurobiological and neuroendocrine adaptations requires an endocrine signalling mechanism, capable of communicating the reproductive state to the brain. Evidence indicates that prolactin is part of this mechanism.
Embryo development and establishment of pregnancy after embryo transfer in pigs: coping with limitations in the availability of viable embryos
Authors: King TJ , Dobrinsky  , Zhu J , Finlayson HA , Bosma W , Harkness L , Ritchie WA , Travers A , McCorquodale C , Day BN , Dinnyes A , De Sousa PA , Wilmut I
Published: March 2004

Embryo transfer and pregnancy maintenance strategies in pigs were evaluated with reference to situations in which limited numbers of viable embryos or micromanipulated embryos are available, such as pig cloning. Development of embryos with compromised zona pellucida was compared with development of embryos with intact zona pellucida. Micromanipulation had no effect on blastocyst production rates after development in vivo or in vitro, but development in vivo improved the number of embryos reaching the blastocyst stage. Transfer of embryos with compromised zona pellucida resulted in live piglets. Several hormone treatments to maintain pregnancy were tested in a model in which three embryos were transferred into unmated recipient gilts, compared with transfer of three embryos into mated recipients. None of the hormonal treatments resulted in pregnancy rates of more than 25% at term and no more than 9% of transferred embryos survived, in comparison with 50% of the mated recipients successfully carrying 25% of transferred embryos. Lastly, the developmental potential of parthenogenetic embryos was assessed and 62% of transferred embryos resulted in pregnancies, none of which continued beyond day 55 of gestation. After co-transfer of three fertilized embryos with 55-60 parthenogenetic embryos into each of six recipients, two live piglets were delivered. The results from the present study indicate that transfer of zona pellucida compromised embryos can yield litters of normal piglets. In addition, it was demonstrated in a model system involving the transfer of three fertilized embryos into mature gilts that hormonal pregnancy maintenance strategies support a low proportion of embryos to term. Lastly, the present study shows for the first time a comparably effective but novel alternative for pregnancy maintenance in the pig involving the co-transfer of parthenote embryos.
A possible extratubular origin of epididymal basal cells in mice
Authors: Holschbach C , Cooper TG
Published: March 2004

The origin of basal cells in mouse epididymis was examined by counting the numbers of basal cells, intratubular mitotic figures and peritubular cells during development of the epididymis. Putative precursors of basal cells were labelled with bromodeoxyuridine and the nuclei of daughter cells were examined. Histochemical localization of cytokines was performed to gauge their involvement in migration of basal cell precursors from extratubular sources. The results indicate that basal cells may arise from extratubular sources as: (i) there was a decrease in the number of mitotic figures as the number of basal cells increased; (ii) no mitotic figures were observed in the base of the epithelium; (iii) the increase in the number of peritubular cells did not parallel the number of basal cells in all epididymal regions; (iv) division of epithelial cells into daughter cells was circumferential and not radial; (v) bromodeoxyuridine-labelled basal cell nuclei were mostly not found in the vicinity of labelled principal cell nuclei and vice versa; and (vi) the percentage of labelled basal cell nuclei was higher than that of the other cells, which is indicative of their arrival from a more highly labelled pool. In addition, no age-dependent correlation was noted between any of the cytokines tested and appearance of basal cells in the epithelium, and basal cells expressed intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1), which provides further evidence of a relationship between basal cells and immunocytes. If basal cells have an immunological function, failure of their recruitment into the epididymal epithelium at about the time of puberty may have repercussions for immunological protection of spermatozoa and, ultimately, for fertility in the adult.
The importance of oxytocin mechanisms in the control of mouse parturition
Authors: Douglas AJ , Leng G , Russell JA
Published: March 2004

The role of oxytocin in parturition in mice was investigated. Pup birth profiles, blood samples and brains were collected from parturient mice observed under red light conditions in a reversed light:dark photoperiod. Peripheral administration of an oxytocin antagonist in a dose-dependent manner delayed the birth of subsequent pups, indicating that oxytocin is required for a normal pup birth profile. Oxytocin neurones were activated during birth as shown by both increased immediate early gene ( Fos) expression in oxytocin neurones in the supraoptic nucleus and increased plasma oxytocin concentrations during birth. In addition, the nucleus of the tractus solitarius and the olfactory bulbs, sites that process inputs to oxytocin neurones, become activated during parturition. Exposure to stress during parturition halted subsequent deliveries; at this stage plasma oxytocin concentrations were not higher than those of virgin mice, and birth was restored by administration of oxytocin. Administration of beta-adrenergic antagonist (propranolol) also restored stress-delayed birth, whereas administration of ritrodrine (beta-agonist) delayed birth in non-stressed mice, indicating that adrenergic mechanisms contribute to stress-delayed births in mice. Administration of morphine (mu-opioid agonist) delayed births transiently, but naloxone (opioid antagonist) did not prevent stress-delayed birth, indicating that endogenous opioids do not appear to contribute to neuroendocrine or uterine mechanisms that promote birth in mice. Therefore, despite evidence in oxytocin knockout mice that oxytocin is not essential for parturition in this species, the results of the present study indicate that oxytocin neurone activity and secretion contribute to the birth process in normal mice.
Kinetics of early in vitro development of bovine in vivo- and in vitro-derived zygotes produced and/or cultured in chemically defined or serum-containing media
Authors: Holm P , Booth PJ , Callesen H
Published: March 2004

The kinetics of the in vitro development of early embryos from bovine zygotes derived in vitro and in vitro were compared, investigating the effect of serum during in vitro maturation and fertilization (IVM-IVF) and in culture. Zygotes were collected from superovulated heifers or produced in vitro from immature oocytes with or without serum supplementation, and cultured subsequently in defined culture medium (SOFaaci) with or without serum supplementation. Time-lapse images were recorded every 0.5 h throughout the culture period. More in vivo- than in vitro-derived zygotes developed to the compact morula or blastocyst stages (87% versus 47-54%, respectively; P < 0.05). Embryo development was blocked predominantly at the second or fourth cell cycles (28 and 29%). However, blastomeres degenerated at all cleavage stages. Serum supplementation during IVM-IVF resulted in abnormally sized blastomeres at first cleavage (defined serum: 20-22% versus in vivo-derived: 8%, P < 0.05). The duration of the second, third and fifth cell cycles of in vivo-derived zygotes were 1-5 h shorter compared with those of in vitro-derived zygotes cultured under similar conditions (P < 0.05). However, the kinetics of embryo development was affected by serum during IVM-IVF and culture. The first and fourth cell cycles were prolonged by 4-5 h in the absence of serum during IVM-IVF, whereas the presence of serum during culture decreased the duration of the fourth cell cycle and triggered premature blastulation. The results of this study illustrate the differences and similarities between the morphology and developmental kinetics of in vivo- and in vitro-derived zygotes, and show how serum supplementation during IVM-IVF and culture can alter these parameters.
Birth of pups after transfer of mouse embryos derived from vitrified preantral follicles
Authors: dela Pena EC , Takahashi Y , Katagiri S , Atabay EC , Nagano M
Published: March 2004

Preantral follicles mechanically isolated from the ovaries of 12-day-old mice were exposed to 2 mol ethylene glycol l(-1) for 2 or 5 min and then to a vitrification solution containing 6 mol ethylene glycol l(-1) and 0.3 mol raffinose l(-1) for 0.5, 1.0 or 2.0 min before vitrification. The vitrified and fresh preantral follicles were treated with collagenase, and the oocyte-granulosa cell complexes (OGCs) obtained were cultured in vitro for 10 days in membrane inserts. Preantral follicles exposed to 2 mol ethylene glycol l(-1) for 5 min and then to the vitrification solution for 0.5 or 1.0 min showed the highest survival rates after warming. The follicular loss after warming was approximately 20%. After in vitro culture, the proportion of viable OGCs from the vitrified follicles was 10% lower than that of the fresh preantral follicles. There were no differences in the rates of maturation, fertilization and subsequent development to blastocysts between the oocytes derived from vitrified follicles and those derived from fresh preantral follicles; however, the developmental competence of the oocytes derived from both vitrified and fresh preantral follicles grown in vitro was lower than that of oocytes grown in vivo. One of the five recipient mice that received 20 blastocysts derived from vitrified preantral follicles gave birth to six live pups. The results of the present study demonstrate for the first time that mouse preantral follicles can be vitrified and that some of the embryos derived from vitrified preantral follicles can develop to live pups.
Effect of adrenocorticotrophic hormone on the development of oestrogen-induced changes and hyperplasia formation in the mouse uterus
Authors: Gunin AG , Emelianov V , Tolmachev AS
Published: March 2004

The aim of this study was to examine the role of adrenocorticotrophic hormone (ACTH) in proliferative and morphogenetic reactions in the uterus under continuous oestrogen treatment. Ovariectomized mice received injections of oestradiol dipropionate or vehicle once a week (2 microg per 100 g), and injections of ACTH (10 microg per 100 g) once a day or once a week for 30 days. Additional control mice received oestradiol and saline, vehicle of oestradiol, and ACTH once a day or once a week, or vehicle of ACTH, for 30 days. This study shows for the first time that ACTH affects oestrogen-dependent reactions in the uterus. Treatment with ACTH once a day resulted in a decrease in uterine mass, in cell proliferation (assessed by the number of mitotic and bromodeoxyuridine (BrdU)-labelled cells) and in the incidence of endometrial hyperplasia, in particular complex and atypical hyperplasia. Treatment with ACTH once a week led to a marked reduction in the incidence of endometrial hyperplasia, a slight increase in uterine mass and had almost no effect on cell proliferation. Daily treatment with ACTH reduced the concentration of oestrogen receptor alpha in all uterine compartments, but weekly ACTH administration had the opposite effect. Expression of glucocorticoid receptors, beta-catenin and glycogen synthase kinase-3beta in uterine tissues was lower in animals treated with oestradiol and ACTH once a day or once a week. When olive oil was used instead of oestradiol, treatments with ACTH did not produce detectable changes in all parameters examined. Thus, glucocorticoid receptor, oestrogen receptor alpha, beta-catenin and glycogen synthase kinase-3beta are involved in the effects of ACTH on oestrogen-induced changes in uterine mass, cell proliferation and the incidence of hyperplasia.
Matrix metalloproteinases and their specific tissue inhibitors in menstruation
Authors: Dong JC , Dong H , Campana A , Bischof P
Published: March 2004

The timely breakdown of extracellular matrix is essential for menstruation. Matrix metalloproteinases, which are able to degrade virtually all components of the extracellular matrix, are spatiotemporally expressed in the cyclic endometrium. The expression of most matrix metalloproteinases is regulated transcriptionally and their proteolytic activities are precisely controlled. The balance between matrix metalloproteinases and their specific tissue inhibitors is believed to be crucial for menstruation. This review focuses on the roles of matrix metalloproteinases and their tissue inhibitors in the initiation of menstruation and on the regulation of matrix metalloproteinase expression and activation. For example, the function of matrix metalloproteinases and their tissue inhibitors in endometrial re-epithelialization and angiogenesis during endometrial regeneration, when cell migration is facilitated to ensure endometrial repair, is discussed. This and other processes, although not fully resolved, serve to illustrate the involvement of matrix metalloproteinases and their tissue inhibitors in the process of menstruation.
Role of facilitative glucose uptake in the glucose-inorganic phosphate-mediated retardation and inhibition of development in different strains of mouse embryos
Authors: Scott L , Whittingham DG
Published: March 2004

Mouse embryos from different strains develop differently in vitro depending on the composition of the culture medium, and in particular on the presence or absence of glucose and inorganic phosphate. Glucose is both stimulatory and inhibitory in certain conditions. Glucose uptake by cells can be passive, down concentration gradients, or active, through sodium driven pumps, or can occur through facilitative transport. This study investigated the effects of inhibition of facilitative glucose transport on the glucose-inorganic phosphate-mediated blocks in development in three different strains of mouse embryo, CF-1, CD-1 and an F2 hybrid. Development of CF-1 and CD-1 embryos is blocked in medium containing glucose and inorganic phosphate but not in medium containing glucose alone, and F2 embryos are not affected. Inhibition of facilitated glucose transport to the eight-cell-morula stage in CF-1 and CD-1 embryos resulted in development in medium containing both glucose and inorganic phosphate, indicating that the prevention of facilitative glucose uptake can overcome the developmental block. Removal of inhibition before the eight-cell-morula stage resulted in total arrest of CF-1 embryos and minimum development of CD-1 embryos. F2 embryos are not affected by inorganic phosphate and glucose and showed no response to the transporter inhibitor at any stage. These data support the contention that facilitated glucose transport is active in embryos, is phosphate-dependent and that its inhibition can overcome the glucose-inorganic phosphate-mediated developmental blocks in mouse embryos.
Dynamics of intracellular calcium induced by lactate and glucose in rat pachytene spermatocytes and round spermatids
Authors: Reyes JG , Herrera E , Lobos L , Salas K , Lagos N , Jorquera RA , Labarca P , Benos DJ
Published: March 2004

Glycolytic metabolism in meiotic and post-meiotic spermatogenic cells shows differentiation-related changes. The developmental and physiological significance of these metabolic changes is not known. The aim of the present study was to test the hypothesis that glucose and lactate metabolism can modulate intracellular calcium [Ca2+](i) in spermatogenic cells in an opposing and dynamic manner. Fluorescent probes were used to measure [Ca2+](i) and pH(i), and HPLC was used to measure intracellular adenine nucleotides and mitochondrial sensing of ATP turnover. [Ca2+](i) in pachytene spermatocytes and round spermatids was modulated by changes in lactate and glucose concentrations in the media. The kinetics and magnitude of the [Ca2+](i) changes induced by lactate and glucose were different in meiotic and post-meiotic spermatogenic cells. The presence of glucose in the medium induced a decrease in pH(i) in spermatogenic cells. This glucose-induced pH(i) decrease occurred later than the changes in [Ca2+](i), which were also observed when the pH(i) decrease was inhibited, indicating that the glucose-induced [Ca2+](i) increase was not a consequence of pH(i) changes. Hexose phosphorylation in glycolysis was part of the mechanism by which glucose metabolism induced a [Ca2+](i) increase in spermatogenic cells. The sensitivity of [Ca2+](i) to carbohydrate metabolism was higher in round spermatids than in pachytene spermatocytes. Thus, differentiation-related changes in carbohydrate metabolism in spermatogenic cells determine a dynamic and differential modulation of their [Ca2+](i) by glucose and lactate, two substrates secreted by the Sertoli cells.
Seasonal changes in testicular size and serum LH, prolactin and testosterone concentrations in male polar bears (Ursus maritimus)
Authors: Howell-Skalla LA , Cattet MR , Ramsay MA , Bahr JM
Published: March 2004

Little is known about the reproductive endocrinology of the male polar bear, Ursus maritimus, except that serum testosterone concentrations are high in April and May during the mating season and are low from August to November during the non-mating season. The objective of this study was to describe the relationship between seasonal changes in testicular size and serum concentrations of testosterone, LH and prolactin. Blood samples and testicular measurements were obtained from free-ranging male polar bears in Canada in April (n = 5) and May (n = 15) near Resolute Bay, Northwest Territories and near Churchill, Manitoba in July (n = 15) and October (n = 22). Testis size was greater in May (39.4 +/- 3.5 cm(2)) than in October (27.3 +/- 2.0 cm(2)) (P = 0.002). Serum testosterone concentrations were approximately three-fold higher in April (5.8 +/- 0.8 ng ml(-1)) than in May (1.7 +/- 0.5 ng ml(-1)), July (0.6 +/- 0.2 ng ml(-1)) and October (1.1 +/- 0.2 ng ml(-1)). Similarly, serum LH concentrations were high in April (0.14 +/- 0.04 ng ml(-1)) and low in May (0.09 +/- 0.01 ng ml(-1)), July (0.10 +/- 0.02 ng ml(-1)) and October (0.08 +/- 0.00 ng ml(-1)). Serum prolactin concentrations were high in April (1.9 +/- 0.3 ng ml(-1)), highest in May (2.5 +/- 0.2 ng ml(-1)), lower in July (1.3 +/- 0.1 ng ml(-1)) and lowest in October (0.8 +/- 0.07 ng ml(-1)). The present study demonstrates a positive relationship between testicular size and serum concentrations of LH, prolactin and testosterone in the male polar bear and confirms the previously reported seasonal changes in serum testosterone concentrations. Data from the present study provide important baseline and comparative endocrine information that can be used to aid captive breeding programmes in zoos and to further ecological-behavioural studies of polar bears.
Roles of the ovarian surface epithelium in ovulation and carcinogenesis
Authors: Murdoch WJ , McDonnel AC
Published: March 2004

Although ovarian mechanisms of ovulation have been a subject of investigation for more than a century, essential regulatory pathways remain uncertain. A role for the ovarian surface epithelium in ovulation has recently been demonstrated. Ovarian surface epithelial cells in close contact with the apical wall of preovulatory ovine follicles secrete a urokinase-type plasminogen activator in response to surge concentrations of (locally delivered) gonadotrophins. Urokinase activates latent collagenases and stimulates release of tumour necrosis factor alpha from thecal endothelium. Tumour necrosis factor alpha progressively induces matrix metalloproteinase gene expression, apoptosis and inflammatory necrosis. Collagenolysis and cellular death are a prelude to stigma formation and ovarian rupture. Epithelium exfoliated from the dome of ovulatory follicles is replenished by generative stem cell replication and migration from the wound edges. Common epithelial ovarian cancer has been related to successive bouts of ovulation and mitosis. The integrity of the DNA of surface cells circumjacent to the ovarian rupture site is compromised during the ovulatory process. Clonal expansion of an epithelial cell with damaged (unrepaired) DNA is a putative factor in carcinogenesis. Ovarian cancer is a deadly insidious disease because typically it is asymptomatic until the malignancy has reached beyond the ovaries.
Expression and localization of inhibin alpha, inhibin/activin betaA and betaB and the activin type II and inhibin beta-glycan receptors in the developing human testis
Authors: Anderson RA , Cambray N , Hartley PS , McNeilly AS
Published: March 2004

Inhibins and activins have roles in the regulation of cell proliferation and differentiation in a variety of tissues. This study investigated the distribution of the three inhibin/activin subunits (alpha, betaA and betaB) and their receptors in the human testis between week 13 and week 19 of gestation using RT-PCR and immunohistochemistry. mRNA for all three subunits and for the activin type II receptors ActRIIA and ActRIIB was detected at all stages of gestation examined. Sertoli cells showed intense immunostaining for the alpha subunit and some staining for the betaB subunit, whereas only the betaB subunit was detected in gonocytes. No betaA subunit staining was detected within the tubules. All three subunits were localized to interstitial Leydig cells. Cells of the rete testis and the epididymal epithelium also showed immunostaining for betaB; however, staining for the other subunits was weak or absent. Peritubular cells showed intense immunostaining for the beta-glycan inhibin receptor, which was also localized to interstitial cells, but was not detected within the tubular compartment, rete testis or epididymal epithelium. ActRIIA was detected in gonocytes and in interstitial cells; ActRIIB was distributed widely. These data indicate that fetal Leydig and Sertoli cells have the potential to produce both activins and inhibins, whereas gonocytes may produce only activin B. The distribution of activin and inhibin receptors implies that the intratubular compartment and developing duct system are sites of action of activin B but not inhibin at this stage of development, whereas both activins and inhibins may be involved in the development and function of the peritubular and interstitial cells.
Insulin-like growth factor (IGF) system in the oocyte and somatic cells of bovine preantral follicles
Authors: Armstrong DG , Baxter G , Hogg CO , Woad KJ
Published: March 2004

Many studies have highlighted the role of the insulin-like growth factor (IGF) system in the control of antral follicular growth. However, much less is known about the involvement of the IGF system in the regulation of preantral follicular development. In an attempt to address this lack of knowledge, the present study describes the spatial and temporal patterns of expression of mRNA encoding components of the IGF system in bovine follicles during preantral stages of development. mRNA was detected by in situ hybridization using frozen sections (14 microm) of bovine ovarian tissue. Serial sections were probed with 35S-labelled bovine riboprobes. Type 1 IGF receptor mRNA was detected in granulosa cells and in the oocyte of preantral follicles; however, in this study, as in previous studies, it was not possible to detect mRNA encoding either IGF-I or -II. IGF binding protein (IGFBP)-2 mRNA was present in granulosa cells and oocytes of preantral follicles, and immunoreactive IGFBP-2 was detected around granulosa cells during this early stage of development. Occasionally, preantral follicles were identified in which there was no expression of IGFBP-2 in granulosa cells or the oocyte. IGFBP-3 mRNA was detected in the oocyte of preantral follicles and in the surrounding stromal tissue. mRNAs encoding IGFBP-2 and -3, and type 1 IGF receptor were first detected in type 2 follicles. In conclusion, although the IGF ligands are not expressed in preantral follicles, mRNAs encoding the type 1 IGF receptor, and IGFBP-2 and -3 were present and showed unique spatial patterns of expression within preantral follicles.