Recovery and maturation of immature oocytes in patients at risk for ovarian hyperstimulation syndrome.

PURPOSE: Our purpose was to examine the rate of immature oocyte recovery and their potential for in vitro maturation from canceled human menopausal gonadotropin cycles due to the risk of having ovarian hyperstimulation syndrome develop. METHODS: Patients underwent ultrasound-guided immature oocyte pickup. The number of oocytes recovered from these patients was recorded, and then cultured in vitro. Cumulus expansion and the stage of nuclear maturation were observed after 24 and 48 hr, respectively. RESULTS: Seventeen patients underwent 20 immature oocyte recoveries. A total of 162 oocytes (8.1 oocytes/patient) was obtained. All of the oocytes were enclosed in dense layers of cumulus cells. Among them, 78.4% showed cumulus expansion after 24 hr and 66% completed meiotic maturation to metaphase II after 48 hr in culture. There was only one immature oocyte pickup in which no oocytes were recovered (95% recovery rate). None of the patients had ovarian hyperstimulation syndrome develop. CONCLUSIONS: Immature oocytes can be recovered from canceled human menopausal gonadotropin cycles in patients who are at potential risk for severe hyperstimulation syndrome. These oocytes can be matured in vitro and can be used for clinical and research purposes as well.

Pregnancy after transfer of embryos which were generated from in-vitro matured oocytes.

In-vitro maturation of human oocytes is an important technique in assisted reproduction due to its potential for reducing the use of fertility drugs. We offered this technique as an alternative to cancelling the cycle to a patient who was at risk of ovarian hyperstimulation syndrome (OHSS) after treatment with gonadotrophin-releasing hormone analogue (GnRHa) and human menopausal gonadotrophin (HMG). The patient had 40 visible antral follicles with a maximum diameter of 13 mm and an oestradiol concentration of 14,000 pmol/l on cycle day 12. Immature oocytes were aspirated transvaginally under ultrasound guidance. Ten cumulus-enclosed oocytes were harvested and nine of them completed nuclear maturation to metaphase II after 48 h in culture. By 18 h after an intracytoplasmic sperm injection (ICSI) procedure, seven of these metaphase II stage oocytes displayed two distinct pronuclei and two polar bodies. All fertilized oocytes but one underwent cleaveage; four of these were transferred 2 days later. Endometrial priming was initiated with 8 mg oestradiol valerate daily from the day of oocyte retrieval and 50 mg progesterone was injected i.m. daily starting 2 days after that. A single intrauterine sac was seen containing one fetus with positive fetal heart beat on ultrasound at 7 weeks of gestation. Unfortunately, the pregnancy ended at 24 weeks shortly after premature rupture of membranes; a live healthy-looking girl was delivered who died 18 days later.

Progesterone does not potentiate the acrosome reaction in human spermatozoa: flow cytometric analysis using CD46 antibody.

The study was designed in order to investigate the action of progesterone on the spontaneous and ionophore-induced human spermatozoa acrosome reaction in vitro. The principle of the assay system is flow cytometric analysis of CD46 antibody binding to the inner acrosomal membrane. The technique is a simple and objective method of analysis, allowing fluorescent analysis of a large segment (5000 spermatozoa) of the spermatozoa population under investigation, with concomitant isolation of the live fraction of the spermatozoa population. Four concentrations of progesterone (1, 25, 50, and 100 microg/ml) were examined for their effects on spermatozoa capacitated for 4 and 24 h. In addition, motility parameters were examined by the CellSoft 2000 automated semen analyser system. Analysis of variance revealed that progesterone had no effect on either the spontaneous acrosome reaction or the ionophore-induced acrosome reaction at both 4 h and 24 h of spermatozoa capacitation times. Further, no effects on sperm motility parameters or on spermatozoa viability could be attributed to progesterone. We therefore conclude that progesterone has no objectively measurable effects on either the sperm acrosome reaction or sperm motility parameters, as measured in normal sperm populations.

Human leukocyte antigen typing and associated abnormalities in premature ovarian failure.

A case-controlled study was designed to assess the distribution of human leukocyte antigens (HLA) A, B, Cw, DR and DQ, and associated abnormalities in patients with premature ovarian failure (POF). A total of 37 patients in a tertiary care centre were diagnosed as having POF; all were < 37 years old. A subset was HLA-typed. The occurrence of associated diseases in patients with POF was recorded. A comparison of HLA typing was made between patients with POF and 100 organ donors from the same population. No statistically significant difference in the distribution of A, B, Cw, DR and DQ antigens was found between the study and control groups when corrected for the number of antigens tested. The commonest associated abnormality was positive autoimmune screen (43.3%). Abnormal thyroid function tests were detected in 23.3% of the patients. The observed high frequency of associated autoimmune phenomena in patients with POF points to an underlying autoimmune disorder in POF and warrants further studies to unravel the pathology of this condition.

Controlled comparison of interceed and amniotic membrane graft in the prevention of postoperative adhesions in the rabbit uterine horn model.

OBJECTIVE: To evaluate the effects of Interceed TC7 (Johnson & Johnson Medical, New Brunswick, NJ) and gamma-irradiated human amniotic membrane graft in prevention of postoperative adhesions in the rabbit uterine horn model. SETTING: A conventional animal laboratory. SUBJECTS: Virgin, female New Zealand white rabbits. INTERVENTIONS: Both uterine horns were subjected to clean transverse cuts 1 cm apart at the antimesenteric border. The left uterine horn was left uncovered in each case to act as a control. The right uterine horn was randomly covered with either Interceed TC7 or human amniotic membrane. OUTCOME MEASURES: Adhesion score was evaluated 4 weeks after the initial surgery. Samples of each uterine horn were collected for histopathological studies, and subserosal inflammatory score. RESULTS: Interceed resulted in higher adhesion scores and subserosal inflammation score than the control side (P = 0.03 and 0.05, respectively). A similar difference was found for amniotic membrane when compared with the uncovered control side (P = < 0.01 and 0.01, respectively). CONCLUSION: Interceed TC7 failed to have the preventive effect on adhesion formation it is claimed to possess. Amniotic membrane performed poorly too. Therefore, gentle tissue handling, irrigation and good surgical skills remain the most important prerequisite for adhesion prevention.

Is Percoll safe for in vivo use?

Fourteen virgin female rabbits were injected with 60% Percoll solution in the right ovary and uterine horn and sperm prepared with Percoll in the left ovary and uterine horn. Histologic examination after 4 weeks showed no inflammatory cell infiltration in either uterine horns or ovaries.

Pentoxifylline potentiates ionophore (A23187) mediated acrosome reaction in human sperm: flow cytometric analysis using CD46 antibody.

The study was designed to investigate the effects of pentoxifylline on the acrosome reaction of human spermatozoa in vitro, and to determine whether the reaction is differently modulated after sperm selection by multiple tube swim-up and Percoll buoyant density centrifugation. The acrosome reaction was induced in vitro by using calcium ionophore (A23187) and was detected by measuring the fluorescence of FITC-conjugated goat anti-mouse immunoglobulin bound to CD46 antibody (which binds to the CD46 receptor site on the inner acrosomal membrane) by flow cytometry. Spermatozoa separated on Percoll displayed significantly lower spontaneous acrosome reactions (P = 0.002) than did those separated by the swim-up technique. Pentoxifylline did not, by itself, induce acrosome reaction, but after induction with ionophore, it significantly increased the reaction (P = 0.003) and this increase was seen to be greater when Percoll separation was used as compared to the swim-up technique (P = 0.0002). We therefore conclude that Percoll selection of motile spermatozoa together with pentoxifylline treatment may be of value in assisted reproductive techniques, as an increased ARIC score arose after both treatments, and that flow cytometry allows a precise and rapid quantification of the acrosome reaction.

The value of luteal support with progesterone in gonadotropin-induced cycles.

OBJECTIVE: To evaluate the effect of luteal support with vaginal P suppositories in hMG- and hCG-induced cycles. DESIGN: Between March 1988 and February 1989 patients did not receive luteal support, while between March 1989 and January 1990 P was given routinely in the luteal phase. Induction protocol and patient selection remained otherwise unchanged. SETTING: Infertility clinic of a tertiary care hospital. PATIENTS: Twenty-seven patients with hypogonadotropic amenorrhea (World Health Organization [WHO] group I) (11 women with luteal support, 16 women without) and 102 patients with euprolactinemic clomiphene citrate (CC)-resistant anovulation (WHO group II) (52 women with luteal support, 50 women without). INTERVENTION: Vaginal P suppositories 200 mg/d in the luteal support group. MAIN OUTCOME MEASURES: Pregnancy rate (PR), pregnancy outcome. RESULTS: The overall PR in 118 cycles with luteal support was 26.3% whereas 10.4% pregnancies were achieved in 115 cycles of the control group. The influence of luteal support was more pronounced in patients with CC-resistant anovulation (25.2% versus 6.9%) than in patients with hypogonadotropic amenorrhea (33.3% versus 21.4%, not significant). The abortion rate was not significantly changed. CONCLUSION: Luteal support with P increases the PR after hMG and hCG induction. The need for supplementary P seems to be related to the underlying cause of ovarian disturbance.

Percoll semen preparation enhances human oocyte fertilization in male-factor infertility as shown by a randomized cross-over study.

The aim of this study was to compare two methods of semen preparation: multiple tube swim-up and Percoll separation, using a randomized cross-over clinical study, in which sperm parameters, oocyte fertilization rates, embryo quality and cell stage were analysed. Overall, there was no difference between the two preparation methods in the normozoospermic cycles. In the male-factor cycles, Percoll extracted a higher total number of spermatozoa (P = 0.02), increased the concentration of motile spermatozoa (P = 0.02), increased the total number of motile spermatozoa per sample (P = 0.02), and enhanced the recovery rate of motile spermatozoa (P = 0.04) compared to swim-up. There was a significant improvement in fertilization rates (P = 0.0006), in the percentage of embryos over 2-cell stage on day of transfer (P = 0.004), and in the number of replaced embryos per transfer (P = 0.01) in the Percoll as compared to swim-up cycles. There was no significant difference in embryo quality. We conclude, therefore, that in advanced reproductive procedures where sperm dysfunction exists, semen preparation with Percoll should replace the swim-up technique.

Artificial endometrial stimulation for frozen embryo replacement.

In this study, 18 cycles initiated for the transfer of cryopreserved embryos. Endometrium was prepared using a uniform protocol of exogenous E and P. This approach eliminated cycle cancellations caused by ovulatory dysfunction and gave complete cycle control. Fourteen cycles had embryo division and replacement resulting in three pregnancies. In these conception cycles, E was discontinued at 12 days and P at 26 days after ET. One aborted at 6 weeks gestation, whereas the other two pregnancies progressed uneventfully into the third trimester.