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Biomedical subjects

W R Dukelow

Publications and source records attributed to W R Dukelow.

At least 37 records · Page 2Linked to original sources
PubMed1988

The chromosomal normality of in vitro-fertilized rabbit oocytes.

The chromosomal normality of rabbit oocytes fertilized in vitro was examined. Ovum donors were superovulated with pregnant mare's serum gonadotropin and human chorionic gonadotropin (hCG). Follicular oocytes were collected laparoscopically 9-10 h after hCG treatment and incubated in vitro with spermatozoa capacitated in vivo. Of 267 aspirated oocytes, 191 (71.4%) were fertilized in vitro and developed to the 2- to 8-cell stage 24-48 h after insemination. In the chromosomal studies, 121 (63.4%) were examined. Of these, 94 (77.7%) had a normal diploid complement of chromosomes (2n = 44) and 6 (5%) showed aneuploidy. Of the remaining 21, 20 were triploid and one was tetraploid. The incidence of triploid oocytes after in vitro fertilization was higher than the rate in vivo (16.5% vs. 9.0%, respectively). These triploid oocytes were suspected to be the result of polyspermic fertilization in vitro. In addition, at the Metaphase II stage, 62 (89.9%) of 69 induced, preovulatory oocytes had a normal number of chromosomes.

Animals↗
PubMed1987

Ovaries remain functional in squirrel monkeys (Saimiri sciureus) immunized with porcine zona pellucida 55,000 macromolecule.

Fifty female squirrel monkeys were each immunized with 200 micrograms of a purified preparation of the 55,000 macromolecule (ZP3) from porcine zona pellucida. The fertility status of these immunized monkeys, as well as the effect of ZP3 antibodies on ovarian function, was monitored. High anti-ZP3 titers were achieved (greater than 75% binding levels as determined by radioimmunoassay) and remained high (approximately 67% binding level) for the duration of this study. Hormonal evaluations indicated initial disturbances in normal ovarian steroid secretion and function that were confirmed by laparoscopic observation and oocyte production data. Histological examination of ovaries at 6-7 mo post-injection suggested an interference in folliculogenesis. No pregnancies were observed in the immunized monkeys during this period. By 10-15 mo post-immunization, hormonal and laparoscopic data indicated that ovarian function was recovering in injected monkeys despite the continued presence of high titers to ZP3. Collectively, these results demonstrate that although immunization with ZP3 initially produces disturbances in normal ovarian function that inhibit fertility, these effects are reversible. Such findings encourage the continued intensive investigation of purified porcine zona macromolecules for immunocontraceptive purposes.

Animals↗
PubMed1985

Chemically defined and serum supplemented media effects on mouse embryo development.

Mouse embryos at various stages of development were used to test three types of media: TC199, CMRL-1066 and TALP. The effect of 20% human cord serum (HCS) and fetal calf serum (FCS) were also compared in TC199 and CMRL-1066 media. Embryos were recovered at the two, four and eight cell stages and assessed for at least one cleavage progression in 24 hours. There was no difference in embryo growth rates between the media for four and eight cell embryos, however TALP significantly increased the proportion of two cell embryos which underwent at least one cleavage division. HCS significantly promoted a greater number of cleavage divisions compared with FCS. This study indicates that the defined medium (TALP) can be employed for equal or increased growth rates of early mouse embryos cultured in vitro compared to two serum supplemented media (TC199 and CMRL-1066).

Journal Article↗
PubMed1985

The use of vital dyes to assess embryonic viability in the hamster, Mesocricetus auratus.

Experiments were designed to assess the use of the vital dyes trypan blue and fluorescein diacetate as indicators of the viability of hamster ova and embryos. Exclusion of trypan blue and fluorescence with fluorescein diacetate showed high correlations with uptake of [3H]uridine by ova and further development of embryos in vitro. Ova killed by freezing and thawing incorporated [3H]uridine at background levels. Trypan blue exclusion and fluorescein diacetate uptake were highly correlated with each other (r = 0.99). Trypan blue and fluorescein diacetate serve as excellent indices of viability in ova and early embryos of hamsters.

Animals↗
PubMed1985

Effect of progesterone and medroxyprogesterone acetate on pregnancy length and litter size in mink.

The objective of these studies was to determine the effect of progesterone and medroxyprogesterone injected at varying times post coitum on gestation length and litter size. During a 3-year period mink females of "Standard" strain were given progesterone at a dose 5 mg at 15 and 20 days in the first year and at 17-20 days in the second year, after the last mating. The respective control groups were given the vehicle. Medroxyprogesterone acetate was given in the second year at a dose of 4 mg at 14-19 days following the last mating and in the third year at a dose of 2 mg at 8 days after the last mating. The results on pregnancy length and litter size after progesterone injections in the experimental and respective control groups were as follows: 52.7 days and 4.3 kits, 51.3 days and 4.3 kits, 52.2 days and 5.8 kits, 50.2 days and 4.8 kits, 44.7 days and 5.6 kits, 46.0 days and 6.1 kits. A dose of 4 mg MPA resulted in the blockage of parturition in pregnant females. After administration of MPA at doses of 2 mg at 8 days following the last mating, the pregnancy length and litter size in the experimental group were: 48.0 days and 6.1 kits and in the control: 52.2 days and 4.8 kits.

Animals↗
PubMed1985

Xenogenous and in vitro fertilization of frozen/thawed primate oocytes and blastomere separation of embryos.

Little research has been done on the in vitro and xenogenous fertilization of cryopreserved primate oocytes. This study reports the development of freezing and thawing methods for squirrel monkey oocytes with subsequent successful fertilization by these two methods. Preliminary results on techniques for blastomere separation using the hamster and squirrel monkey as models are also given. These studies have important implications relative to the long-term frozen storage of human oocytes, their subsequent thawing, in vitro fertilization and embryo transfer, and the use of the blastomere separation procedure, in conjunction with in vitro fertilization, in the diagnosis of embryonic normality and possible congenital defects prior to implantation.

Animals↗
PubMed1983

Preimplantation development of the primate embryo after in vitro fertilization.

The developing use of in vitro fertilization (IVF) as a human clinical procedure has prompted the exploitation of nonhuman primates to assess the chromosomal and biochemical normality of embryos produced by IVF. Of 1995 oocytes recovered from squirrel monkeys, 628 (31.5%) matured and 339 (54.0%) fertilized. Fertility can be significantly enhanced by the addition of 1 or 10 microM dbcAMP to the culture medium. Chromosome analysis of oocytes and embryos used in these studies revealed an incidence of abnormality between 7 and 25%, comparable with that found for both in vivo and in vitro fertilized embryos from other laboratory species. There is no evidence that the IVF technique increases chromosomal abnormality. There was a decrease in protein synthesis of oocytes at maturation and during the early embryonic development stages, but an increase in the rate of RNA synthesis as development progressed. There was steroid uptake in early preimplantation embryos. The temporal relationships of early embryonic developmental events in the squirrel monkey have been determined.

Animals↗
PubMed1983

Heteroimmunization of squirrel monkeys (Saimiri sciureus) with a purified porcine zona antigen (PPZA): immune response and biologic activity of antiserum.

The potential for utilization as a contraceptive vaccine of a 60,000 Mr glycoprotein component, purified porcine zona antigen (PPZA), isolated from porcine zonae, was investigated in the squirrel monkey. Immunization resulted in production and maintenance of high antibody titers for at least 1 year. Comparable immune profiles were obtained using either monkey or pig zonae in assay systems, but dose-dependent variations in immune response were not observed. In situ antibody binding to monkey zonae was detected, but significantly fewer ovulated eggs were obtained from immunized monkeys than from controls. Exposure of antibody-pretreated pig, monkey, and human zonae to homologous sperm resulted in total inhibition of sperm attachment for the respective species. Thus, the contraceptive potential of PPZA antibodies in these species is demonstrated.

Animals↗
PubMed1983

Nonhuman primate in vitro fertilization: biochemical changes associated with embryonic development.

Little research has focused on the biochemistry of primate embryos fertilized in vitro. These studies were designed to examine alterations in cellular incorporation of leucine, uptake of steroid hormones, and overall viability of such embryos. The results may be applied to certain investigations of embryonic development in humans. Squirrel monkey ova and embryos were incubated in medium supplemented with either 3H-leucine, 3H-17 beta-estradiol (E2), or 3H-progesterone (P) and processed for autoradiographic analysis or assayed for radioactivity. Cellular incorporation of 3H-leucine (as an indication of protein synthesis), declined with oocyte maturation in vitro and remained constant at in vitro fertilization (IVF). The use of the vital dyes trypan blue and fluorescein diacetate showed high correlations with the cellular incorporation of 3H-leucine by ova, therefore demonstrating viability. Uptake of E2 and P increased at IVF. The results indicate normal metabolic development of primate embryos fertilized in vitro that is comparable to preimplantation development of other mammalian species.

Animals↗
PubMed1982

Chromosomal analyses after in vitro fertilization of squirrel monkey (Saimiri sciureus) oocytes.

The effect of 1 microM dibutyril cAMP (cAMP media) on the chromosomal normality of in vitro fertilized squirrel monkey oocytes was examined. A total of 877 oocytes were collected laparoscopically 15-16 h after hCG injection from hormonally-induced follicles of 65 squirrel monkeys. Of these, 330 (37.6%) were prepared between 6 and 30 h after insemination and analyzed. Sperm penetration and the presence of a swollen sperm head in the ooplasm were observed by 6 h, and the nucleate stage by 10 h. Addition of 1 microM dbcAMP did not affect the oocyte maturation rate, however the in vitro fertilization rate was increased 26.5% over controls (46.3 vs. 36.6%). The first cleavage metaphase was found at 16 h after insemination. Between 24 and 30 h post-insemination, 76.6% of fertilized oocytes had reached the first cleavage metaphase. An incidence of triploidy (including dispermy) of 11.7% was observed. After chromosomal analysis of metaphase I, univalents were observed in 16.7% of the cases. Aneuploidy (8.9%) was also found in metaphase II. Of the first cleavage metaphases, 88.0% had the diploid number of chromosomes. Aneuploidy occurred 12.0% of the time and 5 cases of triploidies were observed.

Animals↗
PubMed1982

Time relations of squirrel monkey (Saimiri sciureus) sperm capacitation and ovum maturation in an in-vitro fertilization system.

First polar body expulsion in the squirrel monkey ovum occurred at an average of 13.1 h after hCG injection when observed in culture but could occur as early as 5 h after hCG. the mean time for extrusion of the second polar body was 4.7 h after insemination of the culture with freshly ejaculated spermatozoa (16.3 h after hCG). The earliest time, and therefore also that of sperm capacitation, was 2 h after insemination. First cleavage occurred 16.2 h after insemination.

Animals↗
PubMed1982

Serum estradiol 17 beta, progesterone, and relative luteinizing hormone levels in Saimiri sciureus: cyclic variations and the effect of laparoscopy and follicular aspiration.

With increasing use of the squirrel monkey (Saimiri sciureus) in reproductive studies, a precise knowledge of the hormonal variations within the cycle becomes essential for timing of ovulation. The frequent utilization of laparoscopy for follicle aspiration and oocyte recovery warrants analysis of hormonal alterations. In the first study, estradiol 17 beta, progesterone, and luteinizing hormone (LH) levels were quantitated daily in the same group of 14 animals for 16 days. Serum estradiol and LH levels showed significant increases on day 4 of the cycle. Progesterone levels peaked on day 6. Based upon the hormonal peak intervals, the reproductive cycle of the squirrel monkey was estimated to be 8-9 days in length. Laparoscopy did not alter normal serum levels of estradiol, progesterone, and LH. Aspiration of the follicular contents, however, resulted in a significant decrease in estradiol and LH, and a rise in progesterone within 2 h.

Animals↗
PubMed1982

Nonhuman primate in vitro fertilization: seasonality, cumulus cells, cyclic nucleotides, ribonucleic acid, and viability assays.

Since establishing our squirrel monkey (Saimiri sciureus) in vitro fertilization (IVF) system in 1972, 1788 oocytes have been laparoscopically recovered from follicles with 669 matured and 332 fertilized. Oocyte maturation was higher (P less than 0.05) from March to June. Among the oocytes, 50% showed first polar body extrusion by 21 hours after recovery. Maturation was significantly higher (P less than 0.05) in oocytes with corona radiata cells, compared with denuded oocytes. Sperm motility studies indicated that 2.6 and 5.2 mM caffeine enhanced motility by 10% to 30% over controls for the first 5 hours. Addition of 1 and 10 microM dibutyryl cyclic adenosine 3':5'-monophosphate (dbcAMP) enhanced fertilization by 50.3% and 50.0%, respectively. Preliminary studies of 3H-urindine incorporation and uptake by unfertilized oocytes showed levels of 3.43 +/- 0.21 and 55.01 +/- 7.16 fmole/embryo/3 hours, respectively. Exclusion of the vital dye trypan blue by oocytes and embryos gave a high correlation with morphology and survival. Autoradiographic analysis indicated nuclear grain densities in oocytes cultured with 3H-uridine. The results of the studies indicated that oocyte maturation was dependent on season and corona radiata cells, and that addition of dbcAMP enhanced IVF. Trypan blue was found be a useful embryo viability test.

Animals↗