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Biomedical subjects

V N Bolton

Publications and source records attributed to V N Bolton.

At least 19 recordsLinked to original sources
PubMed2003

Live births after intracytoplasmic sperm injection in the management of oligospermia and azoospermia in Nigeria.

Intracytoplasmic sperm injection has revolutionised the management of male infertility. We report two cases that demonstrate the successful application of this technology in Nigeria in the management of both oligospermia and azoospermia. The first case relates to the treatment of a 31-year-old woman who required intracytoplasmic sperm injection of her husband's sperm for the treatment of both tubal fertility and male infertility. She had three embryos transferred on 9th June 1999 and was delivered of healthy male and female infants by caesarean section in January 2000 at 33 weeks gestation. The second case describes a 38-year-old woman who required intracytoplasmic sperm injection of the husband's surgically collected sperm for the management of azoospermia. She had two embryos transferred on 16th December 1999 and was delivered of a healthy male infant by caesarean section on 19th July 2001.

Adult↗
PubMed1999

A prospective randomized study comparing the outcome of in-vitro fertilization and embryo transfer following culture of human embryos individually or in groups before embryo transfer on day 2.

A prospective randomized trial of in-vitro fertilization and embryo transfer was undertaken to investigate the reported beneficial effects of culturing preimplantation human embryos in groups, rather than individually. A total of 159 treatment cycles, in which the women were matched for age, basal gonadotrophin concentrations and number of previous attempts, were included in the study. Of these, 78 cycles were randomized to the 'individual culture' group, and 81 cycles were randomized to the 'group culture' group. The groups did not differ in terms of the median number of oocytes or embryos obtained per cycle. There was no statistically significant difference between the two groups in terms of treatment outcome, as assessed by pregnancies or clinical pregnancies.

Adult↗
PubMed1999

A prospective comparison of 'in house' and commercially prepared Earle's balanced salt solution in human in-vitro fertilization.

A prospective, randomized study was undertaken to compare the use of Earle's balanced salt solution (EBSS) prepared 'in house' with that produced commercially, in 448 cycles of therapeutic in-vitro fertilization. Outcome was assessed in terms of fertilization and cleavage rates, embryo morphology, and implantation rates following embryo transfer. The only differences that were found between the two media in any of the outcome parameters were in the number of cycles with failed fertilization (1/218 in 'in house' medium compared with 10/230 in commercially prepared medium; P = 0.0186), and in the rate at which embryos cleaved. Thus, while the median number of blastomeres per embryo was no different in the two groups at 46-49 h post insemination (three in embryos cultured in 'in-house' medium, compared with four in those cultured in commercially prepared medium; P > 0.1), the number of embryos per cycle that had cleaved to the 4-cell stage by 46-49 h post insemination was significantly greater in the Medi-Cult than in the EBSS medium (P < 0.001).

Adult↗
PubMed1999

Synthetic profiles of polypeptides of human oocytes and normal and abnormal preimplantation embryos.

There is considerable variation in the rate of development in vitro of individual preimplantation human embryos. The relationship between the rate of development and patterns of polypeptide synthesis in individual embryos was examined using SDS-PAGE and autoradiography. After incubation in [35S]methionine, 19 polypeptide bands were identified that change between fertilization and the morula stage. Although changes in two of the bands occurred in embryos that were developing normally and in ageing oocytes, and are thus independent of fertilization, the changes identified in the remaining 17 bands occurred only after fertilization. In embryos that were developing abnormally, as assessed by delayed cleavage, cleavage arrest or extensive fragmentation, the alteration in polypeptide synthetic profiles increased with increasing abnormality.

Autoradiography↗
PubMed1998

Cytogenetic analysis of human preimplantation embryos following developmental arrest in vitro.

The relationship between chromosomal abnormalities in the human preimplantation embryo and developmental arrest in vitro was investigated. Cytogenetic analysis of 171 embryos that had arrested between the pronucleate and the 8-cell stages demonstrated that the overall incidence of chromosomal abnormality among these embryos was 63.4%. Of the embryos that arrested at the pronucleate stage (n = 48), 47.9% were chromosomally abnormal, compared with 59.5% of those that arrested between the 2- and 4-cell stages (n = 50), and 82.8% of those arrested between the 5- and 8-cell stage (n = 73). The rate of abnormality in embryos with poor morphology (irregular shaped blastomeres and considerable extracellular fragmentation) was significantly higher (86.8%; n = 33) than those with good morphology (60%; n = 51; P<0.005). These results suggest that there is an association between chromosomal abnormality, developmental arrest in vitro, and poor morphology.

Blastomeres↗
PubMed1998

Distribution of alpha3, alpha5 and alpha(v) integrin subunits in mature and immature human oocytes.

The distribution of three integrin subunits, alpha3, alpha5 and alpha(v), in immature and mature human oocytes has been examined using immunofluorescence and confocal microscopy. The results demonstrate that both alpha5 and alpha(v) are present at the germinal vesicle stage, while alpha3 was only detected in oocytes after germinal vesicle breakdown, in metaphase I and II stage oocytes. The cortical concentration of integrin subunits alpha3 and alpha5 is consistent with their localization in the oolemma. In contrast, the homogeneous distribution of alpha(v) throughout the oocyte suggests the existence of cytoplasmic reservoirs of this protein in the oocyte.

Antigens, CD↗
PubMed1996

The relationship between chromosomal abnormality in the human preimplantation embryo and development in vitro.

The relationship between the survival of the human preimplantation embryo in vitro and chromosomal abnormality was investigated by cytogenetic analysis of a total of 250 embryos of varying morphology between the pronucleate stage and the 8-cell stage. The overall incidence of chromosomal abnormality among these embryos was 49%. At the pronucleate stage (n = 46) the incidence was 65.2%, at the 2-4-cell stage (n = 126) it was 54.6%, and at the 5-8-cell stage (n = 78) it was 27.4%. Cleavage-stage embryos with poor morphology (irregular shaped blastomeres with severe extracellular fragmentation) showed a higher incidence of chromosomal abnormality (62%; 54 of 87 analysed) than those with good morphology (22.2%; 26 of 117 analysed). This study demonstrates: (i) that there is progressive loss of chromosomally-abnormal embryos during preimplantation development; and (ii) that there is an association between chromosomal abnormality and embryo morphology.

Adult↗
PubMed1996

The timing of pronuclear formation, DNA synthesis and cleavage in the human 1-cell embryo.

The timing of pronuclear formation and breakdown, DNA synthesis and cleavage during the first cell cycle of human embryogenesis are described. Pronuclei formed between 3 and 10 h post-insemination (hpi; median 8 hpi). S-phase commenced between 8 and 14 hpi, and was completed between 10 and 18 hpi. M-phase was observed between 22 and 31 hpi (median duration 3 h), and cleavage to the 2-cell stage took place between 25 and 33 hpi. The timing of the same events was determined in 1-cell embryos derived from re-inseminated human oocytes that had failed to fertilize during therapeutic in-vitro fertilization (IVF). In these embryos, pronuclei formed between 3 and 8 h post-re-insemination (hpr-i), coinciding with the beginning of S-phase. While S-phase was completed as early as 10 hpr-i in some embryos, it extended until at least 16 hpr-i in others. Pronuclear breakdown and cleavage occurred from 23 and 26 hpr-i respectively; however, they did not occur in some embryos until after 46 hpr-i. The results demonstrate a markedly greater degree of variation in the timing of these events in embryos derived from re-inseminated oocytes compared with embryos derived from conventional IVF, and thus throw into question the validity of using the former as models for studies of the first cell cycle of human embryogenesis.

Cell Cycle↗
PubMed1995

Superovulation, IGFBP-1 and birth weight.

In this study, the effect of superovulation on the circulating levels of insulin-like growth factor binding protein-1 (IGFBP-1) has been investigated. IGFBP-1 levels were measured in singleton pregnancies achieved either naturally (n = 203) or following superovulation, in-vitro fertilisation and embryo transfer (IVF-ET) with either pituitary desensitisation with buserelin and superovulation with human menopausal gonadotrophin (b/hMG) followed by IVF-ET (n = 15) or with clomiphene citrate and hMG (CC/hMG) followed by IVF-ET (n = 15, 1st trimester only). The circulating levels of IGFBP-1 were similar in all three groups during the first trimester, and in both normal and b/hMG pregnancies in the second, but were significantly higher during the third trimester in b/hMG pregnancies than in normal pregnancies (P = 0.0002). The birth weights were significantly lower in the b/hMG group (P = 0.04), but not in the CC/hMG group compared with natural conceptions. Gestational age at delivery was similar in control and b/hMG pregnancies, but significantly reduced in CC/hMG pregnancies (P = 0.04). These data suggest that pregnancies achieved following superovulation with b/hMG are associated with elevated levels of IGFBP-1 during the third trimester of pregnancy and reduced birth weight.

Adult↗
PubMed1995

The effect of temperature fluctuations on the cytoskeletal organisation and chromosomal constitution of the human oocyte.

The effect of temperature fluctuation on spindle integrity and chromosomal organisation in the human oocyte, and the consequences of such effects on the chromosomal constitution of resulting parthenotes, were investigated. A total of 340 oocytes were stained immunocytochemically with an antibody to alpha-tubulin, and 502 were activated parthenogenetically. Exposure of oocytes to room temperature for 2, 10 or 30 min caused disruption of the spindle in 77% (n = 26), 72% (n = 18) and 89% (n = 19) of cases respectively, with evidence of chromosomal dispersal in 50%, 56% and 52.6% respectively. These effects were reversed when oocytes were returned to 37 degrees C after exposure to room temperature for 2 min, but not after 10 min or 30 min. Temperature reduction affected rates of parthenogenetic activation of oocytes (2 min: 67%, n = 27; 10 min: 68%, n = 28; 30 min: 54%, n = 35) and cleavage of resulting parthenotes, but only if oocytes were exposed to room temperature for 30 min (30 min: 53%, n = 19). There is a direct association between temperature-induced spindle damage in the oocyte (70%, 50 of 63) and chromosomal abnormalities in parthenotes developed from oocytes exposed to room temperature (56%, 23 of 41; p < 0.01).

Chromosome Aberrations↗
PubMed1994

The relationship between chromosomal abnormalities in the human oocyte and fertilization in vitro.

The relationship between chromosomal abnormalities in the human oocyte and fertilization in vitro was investigated by cytogenetic analysis of an unselected population of oocytes, where failure to achieve fertilization was attributed to dysfunctional spermatozoa. The results demonstrated that 47% of such oocytes were chromosomally abnormal. These data were used to calculate that the incidence of chromosomal abnormalities in oocytes that do, and those that do not develop pronuclei following insemination in vitro is 26.6% and 20.4% respectively. Statistical analysis demonstrated no relationship between chromosomal abnormality in the oocyte and its capacity to achieve fertilization in vitro.

Adult↗
PubMed1993

Polypeptide profiles of human oocytes and preimplantation embryos.

The polypeptides that direct fertilization and early development until activation of the embryonic genome occurs, at the 4-8 cell stage in the human, are exclusively maternal in origin, and are either synthesized during oogenesis or translated later from maternal mRNA. Using sodium dodecyl sulphate-polyacrylamide gel electrophoresis and silver stain, we have visualized and compared the polypeptides present in different populations of human oocytes and cleavage stage embryos obtained after superovulation and insemination in vitro. Two polypeptide patterns were resolved, differing in the region of mol. wt 69 kDa. The distribution of these patterns showed no correlation with the ability of individual oocytes to achieve fertilization and develop normally to the 8-cell stage.

Blastocyst↗
PubMed1993

Immaturity and chromosomal abnormalities in oocytes that fail to develop pronuclei following insemination in vitro.

A total of 293 oocytes that failed to develop pronuclei after insemination in vitro with apparently normal, fertile spermatozoa were obtained from 87 women undergoing therapeutic in-vitro fertilization. The oocytes were investigated cytogenetically to determine the incidence of immaturity and chromosomal abnormalities. Cytogenetic examination was possible in 81% of the preparations, in which immaturity and chromosomal abnormalities were present in 29.5 and 58.7% respectively.

Cell Nucleus↗
PubMed1993

Interactions between the embryo and corpus luteum.

A total of 102 patients who had become pregnant following in-vitro fertilization (IVF) and embryo transfer were studied at weekly intervals between 4 and 14 weeks gestation. The pregnancies were classified as follows: (i) normal singleton, n = 52; (ii) normal twin, n = 24; (iii) heterotopic, n = 4 (weeks 4-8 only); and (iv) anembryonic with a viable intra-uterine singleton, n = 22. The serum concentrations of human chorionic gonadotrophin (hCG), Schwangerschaft protein-1 (SP-1) and pregnancy-associated plasma protein-A (PAPP-A), oestradiol and progesterone were measured. The mean serum concentrations of HCG, SP-1 and PAPP-A were significantly less in heterotopic than in singleton, singleton/anembryonic or twin pregnancies (P < 0.01-0.05), while those of progesterone and oestradiol were not different at any time. There were no significant differences between the serum concentrations of any of the substances analysed in singleton/anembryonic and singleton pregnancies, but the concentrations of all the substances analysed were significantly greater in twin pregnancies from as early as 7 weeks (P < 0.01-0.05). These data show that in heterotopic pregnancies trophoblast function is reduced, as suggested by the lower concentrations of the placental proteins. Despite this the concentrations of oestradiol and progesterone, derived predominantly from the corpus luteum between 4 and 8 weeks, are equivalent to those found in twin pregnancies, and greater than those found in singleton and singleton/anembryonic pregnancies. These findings support the notion that although HCG may rescue the corpus luteum it does not subsequently have a direct effect on its function, and suggest that the embryo may influence corpus luteum function.

Chorionic Gonadotropin↗
PubMed1992

Preimplantation diagnosis of a human beta-globin transgene in biopsied trophectoderm cells and blastomeres of the mouse embryo.

The preimplantation diagnosis of a HbSA-globin transgene in biopsied trophectoderm cells and blastomeres in embryos using a transgenic mouse model for the trait of human sickle-cell anaemia has been undertaken. A sensitive procedure was developed for the amplification of the human beta-globin gene sequence flanking the sickle mutation. Polymerase chain reaction (PCR) assays were undertaken on one to five biopsied trophectoderm cells and isolated blastomeres of the preimplantation mouse embryo. After biopsy the blastocysts were cultured whilst the cells were analysed for the presence of the transgene, and a high proportion (82-91%) were viable as assessed by the presence of a blastocoele cavity within a 5-h period. The majority of the biopsied cultured blastocysts were frozen and used to confirm the diagnosis; 90 biopsied cultured blastocysts were transferred to pseudopregnant recipients and 34% established pregnancy. Material from day 13.5 post-coitum fetuses was also used to confirm the original diagnosis. The time (4-5 h) required to carry out the analysis obviates a need for extended culture or cryopreservation of the biopsied embryo. In individual experiments under optimal conditions, the presence of the transgene in biopsied cells was detected with 100% accuracy, and the PCR analysis was sensitive at the 1-cell level. The overall success rate of diagnosis and confirmation of the presence or absence of the human beta-globin sequence in the biopsied embryo was 70%. Over the entire experimental period (14 months) DNA contamination from a variety of sources did occasionally occur; the methods used to overcome this problem are discussed.

Anemia, Sickle Cell↗
PubMed1992

Laparoscopic zygote intrafallopian transfer using augmented local anesthesia.

In this study, 29 laparoscopic ZIFTs were performed in 21 patients using local anesthesia augmented with intravenous analgesia. The technique was well tolerated; significant discomfort arose only when the fallopian tubes were manipulated and was minimized by transferring zygotes to one tube only. Seven pregnancies resulted, of which three have delivered and one is ongoing.

Adult↗
PubMed1991

Pregnancies after in vitro fertilization and transfer of human blastocysts.

In a study of 29 cycles of IVF, ET was performed on day 5 after oocyte recovery when embryos had developed to the morula/blastocyst stage. Three preclinical pregnancies and three live births resulted (2 singleton and 1 twin), giving a viable PR per ET of 10%. It is concluded that while day 5 ET may well be important in terms of embryo biopsy for the preimplantation diagnosis of genetic disease, day 2 ET remains preferable for therapeutic IVF. Although these data would not support the introduction of day 5 ET into routine therapeutic IVF, delayed ET should be considered as an alternative approach to preimplantation diagnosis. Indeed, because the latter will generally involve the treatment of normal, fertile couples, it might be predicted that embryo survival rates, and thus the rate of pregnancy after day 5 ET, would be better than those presented here.

Embryo Transfer↗