Paula C. Morton

Birth after cryopreservation of immature oocytes with subsequent in vitro maturation

OBJECTIVE To establish the clinical feasibility of using cryostored germinal vesicle oocytes for IVF and ET. DESIGN Case report. SETTING Private infertility clinic. PATIENT(S) A 28-year-old woman with tubal infertility undergoing IVF therapy. INTERVENTION(S) Oocytes collected after ovarian stimulation were frozen without insemination or were inseminated, fertilized, and frozen as cleavage stage embryos. No fresh oocyte or embryo transfer was undertaken. All oocytes were thawed, and those that survived were used for IVF-ET. MAIN OUTCOME MEASURE(S) Oocyte cryosurvival, in vitro maturation, fertilization, embryo development, and pregnancy outcome. RESULT(S) None of 16 mature oocytes survived thawing; however, three of 13 germinal vesicle oocytes survived. After 30 hours in vitro maturation two oocytes had matured and underwent intracytoplasmic sperm injection with the partners sperm. Both fertilized normally and were transferred to the patient. The woman delivered an apparently healthy female infant at 40 weeks. CONCLUSION(S) This case report proves the feasibility if not the efficiency of using immature oocytes for cryostorage, coupling both cryopreservation and in vitro maturation.

Leuprolide acetate: serum and follicular fluid concentrations and effects on human fertilization, embryo growth, and granulosa-lutein cell progesterone accumulation in vitro

Data from various animal models have demonstrated significant extrapituitary effects of gonadotropin-releasing hormone agonists. The purpose of this study was to determine the effects of therapeutic concentrations of leuprolide acetate on human granulosa-lutein cell steroidogenesis, fertilization, and embryo growth rates in vitro. During leuprolide administration, mean serum concentrations of leuprolide were less than 50 ng/ml and were undetectable 48 hours after cessation of administration. There was no effect of leuprolide on progesterone (P) secretion by granulosa-lutein cells cultured in the presence or absence of human chorionic gonadotropin. The effect of leuprolide on embryo growth rates was evaluated with the mouse two-cell embryo culture model and a retrospective review of early embryo growth rates in humans receiving adjunctive leuprolide therapy. There was no measurable effect of leuprolide on early embryo growth in either species. These data indicate that (1) serum and follicular and peritoneal fluid concentrations are undetectable 2 days after discontinuation of leuprolide; (2) there is no measurable effect of leuprolide on human or murine embryo growth rates up to the 8 cell stage in vitro; and (3) there is no measurable effect of leuprolide on granulosa-lutein cell P accumulation.

Practical evolution and application of direct intracytoplasmic sperm injection for male factor and idiopathic fertilization failure infertilities

OBJECTIVE To analyze the introduction of a new assisted fertilization technique for the treatment of severe male factor and idiopathic fertilization failure infertilities. DESIGN Retrospective analysis of 16-month clinical application of IVF-ET where insemination was performed solely by direct intracytoplasmic sperm injection. SETTING Clinical IVF-ET program. PATIENTS Ninety-two couples undergoing 105 cycles of sperm injection. RESULTS One hundred embryo transfers yielded 28 viable pregnancies (28%) from which eight normal deliveries have occurred to date. Complete cleavage arrest or fertilization failure occurred in four cycles, and one couple had all embryos cryopreserved. One thousand one hundred forty-three eggs were injected of which 173 (15%) degenerated. Four hundred seventy-nine of the surviving 970 eggs became normally fertilized (49%), and 381 of these zygotes (79.5%) developed suitably for cryopreservation or for transfer. Thirty-four of 310 embryos transferred implanted, yielding an implantation rate of 11%. Both testicular and epididymal sperm were used successfully to achieve fertilization and pregnancies, as was sperm retrieved by electroejaculation. Older women and couples suffering from prior idiopathic fertilization failure had a markedly poorer outcome. CONCLUSIONS These results confirm that the intracytoplasmic sperm injection technique is a successful form of assisted fertilization that can be applied to a wide range of couples at significant risk from fertilization failure.

CRYOPRESERVATION OF HUMAN EMBRYOS AND OOCYTES

Human embryo cryopreservation is now a fully established adjunct to assisted reproduction, with thawed embryos implanting at a rate approaching that of fresh embryos. As with fresh embryos, the quality of frozen-thawed embryos is affected by patient age at the time of oocyte retrieval, and by culture conditions. Ovarian stimulation incorporating gonadotropin-releasing hormone analogs does not appear to be detrimental to cryopreservation success, and natural or artificial thaw cycles seem equally effective. Successful cryopreservation seems feasible for one- to eight-cell embryos and blastocysts using a variety of protocols. Selection criteria for embryos to be frozen can vary, and embryo quality has the most significant impact on post-cryopreservation viability. Unfertilized oocyte cryopreservation is coming closer to routine clinical application.

Dizygotic twin delivery following in vitro fertilization and transfer of thawed blastocysts cryopreserved at day 6 and 7

OBJECTIVE To report the first conception and delivery following transfer of thawed human blastocysts maintained in extended in vitro culture with cryopreservation at day 6 and 7. DESIGN Case report. SETTING Major urban infertility referral center. PATIENT(S) A 26-year-old woman with pelvic endometriosis and two prior unsuccessful in vitro fertilization/embryo transfer (IVF-ET) attempts. INTERVENTION(S) The patient underwent controlled ovarian hyperstimulation using a combined FSH + hMG protocol, and 24 oocytes were retrieved. MAIN OUTCOME MEASURE(S) Dizygotic twin delivery after IVF and intracytoplasmic sperm injection (ICSI), assisted embryo hatching, and ultrasound-guided transfer of cryopreserved blastocysts. RESULT(S) After three embryos were subjected to assisted hatching, they were transferred fresh on day 3, but no implantation occurred. All nontransferred embryos (n = 11) were observed during extended in vitro culture and three blastocysts were selected for cryopreservation on day 6 and 7; thaw and transfer occurred the following month and a pregnancy was achieved. Dizygotic twins (female/female) were delivered by cesarean in the early third trimester. CONCLUSION(S) Substantial advancements have been made in the field of embryo cryogenics and in vitro fertilization, but controversy remains regarding the value of freezing late-developing human blastocysts. Here we describe the first reported live births with IVF after extended in vitro culture and cryopreservation at day 6 and 7 after fertilization.

Contraceptive Effect of Two Anti-Sperm Monoclonal Antibodies, Administered Singly and in Combination, in the Mouse

Although many reports in the literature suggest an association between infertility and anti-sperm antibodies, the particular sperm antigens) that may be responsible is unknown. In an effort to sort out this dilemma, as well as to identify sperm components involved specifically at discrete steps of the gamete interaction process, we have generated antimouse sperm monoclonal antibodies (mAbs).

Micromanipulation in assisted reproduction: a review of current technology.

Since the first successful application of micromanipulationin human in-vitro fertilization 10 years ago, the breadth of its application has expanded to encompass single sperm injection, zona ablation, polar body and blastomere biopsy, and very recently the first reports of both cytoplasmic and nuclear transfer.