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Biomedical subjects

J C Osborn

Publications and source records attributed to J C Osborn.

At least 19 recordsLinked to original sources

Follicular steroidogenesis and oocyte maturation after superovulation of goats (Capra hircus) with gonadotrophins.

Follicles were sampled at three different times after treatment with 1200 iu pregnant mares' serum gonadotrophin (PMSG) or 12 mg ovine follicle-stimulating hormone (FSH), and from untreated control animals. The meiotic status and protein synthesis of the oocyte from each follicle was determined and correlated with the intrafollicular concentration of oestradiol and progesterone. Significantly higher amounts of oestradiol were present in PMSG-treated animals at sponge withdrawal than in FSH-treated and control goats. Twenty hours later, both oestradiol and progesterone concentrations in the PMSG group were higher than those in the FSH group, and were equivalent to control animals at the onset of oestrus. At 18 h after the administration of human chorionic gonadotrophin (hCG), oestradiol decreased markedly in all three treatment groups, whereas progesterone remained significantly higher in PMSG-treated follicles. Although these high concentrations of intrafollicular steroids were associated with a higher incidence of premature condensation of chromatin in oocytes, the two events were not causally related. Moreover, cytoplasmic maturation was not prematurely activated in these oocytes and a changed pattern of protein synthesis was observed in oocytes from all three treatment groups after the hCG injection. Whereas disturbances in follicular steroidogenesis of oestradiol and progesterone occur in vivo in goats superovulated with PMSG, they do not underlie the premature activation of the initial stages of nuclear maturation in oocytes but are associated with normal cytoplasmic maturation.

Administration, Intravaginal

Ultrasound-guided embryo transfer: a controlled trial.

Embryo transfer (ET) was performed in 94 patients using transvaginal ultrasound guidance. Pregnancy rates were increased over a control group of 246 patients, although statistical significance was reached only in the subgroup of single ETs. Advantages over the traditional blind technique of ET were observed. Acceptance of the procedure by both clinicians and patients was high.

Embryo Transfer

Luteinizing hormone and follicle stimulating hormone induce premature condensation of chromatin in goat (Capra hircus) oocytes.

This study tested the hypothesis that premature condensation of chromatin in goat oocytes following superovulation with 1200 i.u. pregnant mare serum gonadotrophin (PMSG) is mediated by the high luteinizing hormone (LH) activity inherent in this gonadotrophin. Goats were treated with either a standard (3.95 mL) or high (7.90 mL) dose of a highly purified follicle-stimulating hormone (FSH) preparation (Ovagen), and different doses of human chorionic gonadotrophin (hCG) were added to increase the level of LH bioactivity during superovulation. The meiotic status of oocytes obtained at sponge withdrawal was compared between different treatments and correlated with profiles of LH bioactivity in peripheral plasma. Injection of 100 i.u. hCG (which gave a plasma LH profile comparable to 1200 i.u. PMSG) or 200 i.u. hCG resulted in significantly more oocytes showing premature condensation of chromatin without germinal vesicle breakdown than with 25 i.u. hCG or treatment with FSH alone. Nevertheless, nuclear maturation was also prematurely activated in a significant number of oocytes with a high dose of FSH alone, even though LH bioactivity was not detected in plasma. It is concluded that high LH bioactivity during superovulation of goats with gonadotrophins activates the initial stages of nuclear maturation in oocytes. However, highly purified FSH preparations in high doses can also induce this apparent abnormality in the timing of oocyte maturation through mechanisms unrelated to any LH contamination.

Animals

Premature condensation of chromatin induced in goat (Capra hircus) oocytes after gonadotrophin treatment.

In comparison with ovine follicle stimulating hormone (FSH), superovulation of goats with pregnant mare serum gonadotrophin (PMSG) produced premature ovulations within 48 h of drug administration. To test the hypothesis that this may be associated with a differential effect of the two drugs on oocyte maturation, we have compared the meiotic status of oocytes obtained at three different time intervals from animals treated with 1200 i.u. PMSG or 12 mg ovine FSH and from untreated control animals. Significantly more oocytes from PMSG-treated, compared with control and FSH-treated, animals showed premature condensation of chromatin at both the time of sponge withdrawal and 20 h later. The chromatin condensation was, however, not associated with germinal vesicle breakdown. In contrast, when oocytes were examined 6 h before the expected time of ovulation following human chorionic gonadotrophin (hCG) injection, no significant difference was found in the proportion of oocytes undergoing germinal vesicle breakdown between the three treatment groups, with most oocytes being at the metaphase I or II stage of meiosis. We conclude that superovulation of goats with PMSG at a dose resulting in a high incidence of premature ovulations is associated with premature activation of the initial stages of nuclear maturation in oocytes. In contrast, although treatment with 12 mg ovine FSH did not cause premature ovulations, it was not totally devoid of premature chromatin-condensing activity in oocytes.

Animals

The hypo-osmotic swelling test and the sperm mucus penetration test in determining fertilization of the human oocyte.

The usefulness of the hypo-osmotic swelling (HOS) test and the sperm mucus penetration (SMP) test as sperm function tests for in-vitro fertilization was analysed in 56 couples. Using logistic regression analysis only the SMP test was independently related to fertilization (P = 0.004), no false negative results were obtained, i.e. no fertilization if sperm from the ejaculate failed to penetrate mucus. The HOS test was of no predictive value. The results justify a further examination of the SMP test in other IVF centres.

Cervix Mucus

Factors associated with accidental fractures of the zona pellucida and multipronuclear human oocytes following in-vitro fertilization.

A total of 101 multipronuclear oocytes (7.4% of fertilizations) were retrospectively identified in this in-vitro fertilization programme. The use of a manual syringe suction system, instead of an electric pump, to aspirate follicles, was associated with a significant increase in the proportion of oocytes with fractured zonae pellucidae (P less than 0.001), a lower normal fertilization rate (P less than 0.05) and a higher proportion of multipronuclear fertilizations (P less than 0.001). Irrespective of the mode of follicular aspiration, significantly more multipronuclear fertilizations occurred following stimulation with a combination of clomiphene and human menopausal gonadotrophin, than after clomiphene alone (P less than 0.05). It was concluded that the aspiration pressures, created by syringe suction, were more likely to rupture the zona pellucida of some oocytes, while in others it predisposed to an increased multipronuclear fertilization rate.

Female

Modification of food cariogenicity in rats by mineral-rich concentrates from milk.

Dairy products, including milk, cheese, and casein, can reduce the caries-causing potential of cariogenic substrates as measured in various animal, plaque acidity, and in vitro systems. Although the mechanisms responsible for protection are not completely identified, substances containing Ca and P may contribute to the protective potential by reducing demineralization and/or promoting remineralization of enamel. Casein may reduce demineralization by forming a protective coat on the enamel surface. By means of a rat model, this study evaluated the ability of three casein-free milk mineral concentrates with various levels of whey protein, calcium, and phosphate to modify the cariogenicity of a powdered diet containing 20% sucrose. Analysis of these data indicates that there were no significant differences among groups for weight gain, total food consumption, or feeding frequency, as monitored by a computer-based infrared activity monitor. All three mineral concentrates significantly reduced buccal caries, and two of the three reduced sulcal caries by from 10 to 30%. The analysis further shows that casein-free milk mineral fractions can modify the cariogenicity of sucrose-containing foods in a rat model.

Animals

Effect of alterations in follicular steroidogenesis on the nuclear and cytoplasmic maturation of ovine oocytes.

The effects of inhibitors of follicular steroidogenesis on biochemical changes occurring in oocytes maturing in vitro were studied using radiolabelling and polyacrylamide gel electrophoresis. These effects were correlated with previously investigated developmental abnormalities induced by the same inhibitors. The most severe effects were generated by inhibition of 17 alpha-hydroxylase with the drug SU10603 which resulted in a greatly increased ratio of progesterone to testosterone and oestrogen. Such treatment halved the rate of meiotic maturation. Treated oocytes were analysed individually on SDS-PAGE gels and quantitative analysis showed that the drug had induced synthetic abnormalities even in those oocytes that resumed meiosis. This conclusion was confirmed by separation of oocyte proteins on two-dimensional gels. The effects of SU were reduced by delaying addition of the drug until 6h after the beginning of maturation but were not alleviated by the addition of exogenous oestrogen to the culture medium. When oocytes from SU-treated follicles were transferred to inseminated, recipient ewes and recovered 24h later, two-dimensional electrophoresis again revealed abnormalities in their protein synthetic patterns. Almost total abolition of steroid secretion by aminoglutethimide (AG) had much less effect on oocyte protein synthesis, although the proportion of oocytes maturing was reduced from 65% to 46%. The aromatase inhibitor, androstatriendione (AST) although eliminating follicular oestrogen secretion, had no effect on the rate of maturation and very little effect on protein synthesis. These results correlate well with the effects of steroid inhibitors on fertilization and early cleavage.

Aminoglutethimide

cAMP in ovine oocytes: localization of synthesis and its action on protein synthesis, phosphorylation, and meiosis.

In the first series of experiments, the source of cAMP in the sheep oocyte was studied. Cholera toxin was shown to be a potent stimulator of cAMP in isolated sheep oocytes, demonstrating the presence of adenyl cyclase. There was no evidence for transmission of cAMP from stimulated myocardial cell monolayers to cumulus-enclosed oocytes even though the existence of a concentration gradient of cAMP and of intercellular communication were demonstrated. However, gonadotrophin-stimulated follicle shells were able to induce a rise in the cAMP content of denuded or cumulus-enclosed oocytes in the same dish, independently of cell contact. Further experiments were designed to study the effects of a cholera toxin-stimulated rise in cAMP on the maturation of oocytes. When applied to cumulus-oocyte complexes, cholera toxin did not block germinal vesicle breakdown (GVBD), nor the accompanying changes in protein synthesis and phosphorylation, although there was evidence for a delaying effect. There were, however, indications that the toxin was inducing abnormalities that became gross when the concentration was raised to 1 microgram/ml. This high concentration of cholera toxin was able to block the maturation of oocytes in intact, gonadotrophin-treated follicles, although once again abnormalities were evident. We conclude that the role of cAMP in the maturation of the sheep oocyte is different from that proposed in the mouse.

Animals

Dental cariogenic evaluation of foods using human plaque pH and an experimental rat-caries model.

Five well-characterized foods were used to compare two systems for estimating the cariogenic potential of food: human plaque-acidity using interdental metallic touch electrodes, and cariogenicity testing in rats. Both systems identified the same food as the least cariogenic or acidogenic food and identified the same three foods as being both cariogenic in rats and acidogenic in man, although the ranking of these foods differed. The systems differed in their assessment of potato chips, a food high in cooked starch and low in free sugars. The results confirmed that non-acidogenic foods are non-cariogenic, but that foods acidogenic in man may exhibit a greater range of cariogenicity in rats. Data from both human plaque pH studies and rat caries models may permit better assessment of cariogenic potential of foods containing fermentable carbohydrates.

Adult

Gonadotrophin-induced abnormalities in sheep oocytes after superovulation.

The development of preovulatory follicles involves an initial phase of somatic cell differentiation and a final phase, initiated by the LH surge, when both the somatic and germinal compartments alter. Abnormalities in this pattern of compartmental development after superovulation have been identified by examining follicles from control, PMSG- and FSH-treated sheep. The pattern of proteins synthesized by oocytes from untreated sheep did not differ after culture of follicles in hormone-free medium from that of germinal vesicle oocytes in vivo. Similarly, 93.5% of oocytes from sheep injected with a pituitary gonadotrophin (FSH-P) synthesized the unchanged germinal vesicle pattern of proteins during culture in an hormonally neutral culture environment. By contrast, the administration of the placental gonadotrophin, PMSG, induced in 28% of oocytes changes in the pattern of synthesis which are normally associated with maturation. An examination of follicular steroidogenesis showed that both total output and particularly oestrogen secretion was over twice as high in follicles from PMSG-treated as compared with FSH-treated animals (P less than 0.01). We conclude that the compartmental pattern of development and steroidogenesis is grossly perturbed in many follicles from PMSG-treated animals. Premature activation of the germinal compartment results in aged or abnormal oocytes and a hostile reproductive tract.

Animals

Role of calcium ions in the control of embryogenesis of Xenopus. Changes in the subcellular distribution of calcium in early cleavage embryos after treatment with the ionophore A23187.

Treatment of stage 5 Xenopus embryos with the ionophore A23187 for only 10 min, in the absence of extracellular Mg2+ and Ca2+, causes cortical contractions and a high incidence of abnormal embryos during subsequent development. Cation analysis shows that divalent ions are not lost from the embryos, but that Ca2+ is redistributed within the subcellular fractions. Ca2+ is probably released from yolk platelets and/or pigment granules by the action of A23187, [Ca2+] rises in the cytosol, and the mitochondria attempt to take up this free Ca2+. The mitochondria concomitantly undergo characteristic ultrastructural transformations, changing towards energized-twisted and energized-zigzag conformations. A23187 allows these changes to be demonstrated in situ. Extracellular divalent cations (10(-4) M) interfere with this intracellular action of A23187. Intracellular accumulation of Na+ (by treatment with ouabain) or Li+ also causes abnormal development, probably by promoting a release of Ca2+ from the mitochondria. It is suggested (a) that all these treatments cause a rise in [Ca2+]i which interferes with normal, integrated cell division, so causing, in turn, abnormal embryogenesis, (b) that levels of [Ca2+]i are of importance in regulating cleavage, (c) that the mitochondria could well have a function in regulating [Ca2+]i during embryogenesis in Xenopus, and (d) that vegetalizing agents may well act by promoting a rise in [Ca2+]i in specific cells in the amphibian embryo.

Animals