Nonsurgical Embryonic Selective Thinning And Transfer (NEST): A New Method To Enhance Implantation Rates Following Assisted Reproduction
S.A. Brody, M.L. Lydic, F. Hendee, A. Lachica, P. Quinn. In Vitro Fertilization program, Alvarado Hospital Medical Center, San Diego, CA and Division of Endocrinology & Metabolism, UCSD School of Medicine, La Jolla, CA.
Objectives: Failure of implantation remains an important problem limiting higher fecundity rates following assisted reproduction. Reproductive failure on this basis may be due to in vitro culture conditions which are known to be associated with delayed development and impaired hatching rates in human and other mammalian embryos. furthermore, genetic incompetence may represent the final common denominator for failed implantation in up to 45% of embryos. This study was conducted to determine the role of active management of embryonic growth and development utilizing newly defined growth media for embryo culture and selective thinning.
Design: Preclinical studies were performed on the rate of hatching in mouse embryos using pronase or acid Thyrode's (AT) for selective thinning. We used newly developed formulations of Quinn's HTF culture media characterized by the absence of glucose and phosphate and the addition of antioxidants and alternative energy sources, including glutamine. Clinical studies involved the active management of embryonic growth and development utilizing selective thinning. The most morphologically normal embryos were transferred on day 3 following selective thinning using acid Thyrode's. If there was an insufficient number of good quality embryos, then selective thinning was performed on day 2 post OPU.
Materials And Methods: Preclinical studies: The survival of preimplantation mouse embryos was determined following selective thinning. Selective thinning was performed on d2 utilizing pronase (N=25) (1 mcg/ml) or AT (N=25). Culture media contained 0.5% human serum albumin (HSA). In vivo fertilized zygotes were cultured for 24-48 hours in HTF and then exposed to pronase for 8 hours, or AT for approximately 10 minutes. The embryos were then cultured for an additional 48-72 hours. For the clinical studies IVF patients underwent luteal phase down regulation with leuprolide acetate A combination of pure FSH and human menopausal gonadotropins was used for controlled ovarian hyper-stimulation. Transvaginal ultrasound-guided aspirations of multiple follicles was performed, followed by washing, insemination, and in vitro fertilization. selective thinning was performed on the day of embryo transfer. Thinning was achieved on one pole of the embryo with the highest amount of debris or irregular cell size. The amount of thinning was inversely proportional to the thickness of the zona.
Results: Preclinical results: In mouse studies the application of pronase or AT resulted in no deterioration in embryo growth and development. Results of clinical studies revealed an unusually high fecundity rate following the active management of embryonic growth and development. Our first 15 patients were compared with historic controls from the previous 12-month period. The average number of oocytes obtained was: 15 ± 2 vs 6.1 ± 1.4 (P>.05). Fertilization in the two groups were 72% vs 67% (P>.05). The average number of grade 1 or 2 embryos obtained were: d2 post OPU, 4.7 ± 0.8 vs 6.1 ± 1.4 (P>.05); d3 post OPU, 6.1 ± 0.6 vs 4.5 ± 0.9 (P>.05). The average number of embryos transferred between the two groups was 4.5 ± 0.5 vs 3.9 ± 0.5 (P>.05). Clinical pregnancy rates between the two groups were 80% vs 27% (P>.05).
Conclusion: Nonsurgical embryonic selective thinning and transfer is a technique for the active management of early embryonic growth and development. It utilizes newly defined growth media which facilitates embryonic cleavage so that selective thinning and transfer can be performed on the third day post ovum pickup when genetic competence may be more easily assessed on morphologic grounds. Unlike assisted hatching, selective thinning can also be performed on d2 post OPU, since the zona is not fully breeched. The combination of embryonic thinning and selective transfer results in a dramatic increase in implantation rates, cycle fecundity rates, and overall pregnancy rates.
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