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Improved Murine Blastocyst Quality and Development in a Single Culture Medium Compared to Sequential Culture Media

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Abstract

Objective

Validate single versus sequential culture media for murine embryo development.

Design

Prospective laboratory experiment.

Setting

Assisted Reproduction Laboratory.

Animals

Murine embryos.

Interventions

Thawed murine zygotes cultured for 3 or 5 days (d3 or d5) in single or sequential embryo culture media developed for human in vitro fertilization.

Main Outcome Measures

On d3, zygotes developing to the 8 cell (8C) stage or greater were quantified using 4’,6-diamidino-2-phenylindole (DAPI), and quality was assessed by morphological analysis. On d5, the number of embryos reaching the blastocyst stage was counted. DAPI was used to quantify total nuclei and inner cell mass nuclei. Localization of ubiquitin C-terminal hydrolase L1 (UCHL1) and ubiquitin C-terminal hydrolase L3 (UCHL3) was reference points for evaluating cell quality.

Results

Comparing outcomes in single versus to sequential media, the odds of embryos developing to the 8C stage on d3 were 2.34 time greater (P = .06). On d5, more embryos reached the blastocyst stage (P = <.0001), hatched, and had significantly more trophoblast cells (P = .005) contributing to the increased total cell number. Also at d5, localization of distinct cytoplasmic UCHL1 and nuclear UCHL3 was found in high-quality hatching blastocysts. Localization of UCHL1 and UCHL3 was diffuse and inappropriately dispersed throughout the cytoplasm in low-quality nonhatching blastocysts.

Conclusions

Single medium yields greater cell numbers, an increased growth rate, and more hatching of murine embryos. Cytoplasmic UCHL1 and nuclear UHCL3 localization patterns were indicative of embryo quality. Our conclusions are limited to murine embryos but one might speculate that single medium may also be more beneficial for human embryo culture. Human embryo studies are needed.

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Author information

Authors and Affiliations

  1. Division of Reproductive and Perinatal Research, Department of Obstetrics, Gynecology and Women’s Health, School of Medicine, 1 Hospital Dr, Dc051.00, Columbia, MO, USA

    Justin M. Hennings MS, Randall L. Zimmer MS, Henda Nabli MS, Peter Sutovsky PhD & Kathy L. Sharpe-Timms PhD

  2. Division of Animal Sciences, College of Agriculture, Food and Natural Resources, Columbia, MO, USA

    Justin M. Hennings MS, Peter Sutovsky PhD, Miriam Sutovsky MS & Kathy L. Sharpe-Timms PhD

  3. Department of Health Management and Informatics, School of Medicine, The University of Missouri, Columbia, MO, USA

    J. Wade Davis PhD

  4. Department of Statistics, College of Arts and Sciences, The University of Missouri, Columbia, MO, USA

    J. Wade Davis PhD

  5. Biostatistics and Research Design, Galena Hall, Columbia, MO, USA

    J. Wade Davis PhD

Authors
  1. Justin M. Hennings MS
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  2. Randall L. Zimmer MS
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  3. Henda Nabli MS
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  4. J. Wade Davis PhD
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  5. Peter Sutovsky PhD
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  6. Miriam Sutovsky MS
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  7. Kathy L. Sharpe-Timms PhD
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Corresponding author

Correspondence to Kathy L. Sharpe-Timms PhD.

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Hennings, J.M., Zimmer, R.L., Nabli, H. et al. Improved Murine Blastocyst Quality and Development in a Single Culture Medium Compared to Sequential Culture Media. Reprod. Sci. 23, 310–317 (2016). https://doi.org/10.1177/1933719115618281

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