Justin M Hennings1, Randall L Zimmer2, Henda Nabli2, J Wade Davis3, Peter Sutovsky1, Miriam Sutovsky4, Kathy L Sharpe-Timms5. 1. Division of Reproductive and Perinatal Research, Department of Obstetrics, Gynecology and Women's Health, School of Medicine, Columbia, MO, USA Division of Animal Sciences, College of Agriculture, Food and Natural Resources, Columbia, MO, USA. 2. Division of Reproductive and Perinatal Research, Department of Obstetrics, Gynecology and Women's Health, School of Medicine, Columbia, MO, USA. 3. Department of Health Management and Informatics, School of Medicine, The University of Missouri, Columbia, MO, USA Department of Statistics, College of Arts and Sciences, The University of Missouri, Columbia, MO, USA Biostatistics and Research Design, Galena Hall, Columbia, MO, USA. 4. Division of Animal Sciences, College of Agriculture, Food and Natural Resources, Columbia, MO, USA. 5. Division of Reproductive and Perinatal Research, Department of Obstetrics, Gynecology and Women's Health, School of Medicine, Columbia, MO, USA Division of Animal Sciences, College of Agriculture, Food and Natural Resources, Columbia, MO, USA timmsk@health.missouri.edu.
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.
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.
Authors: Su Jin Kim; Ok Jae Koo; Dae Kee Kwon; Jung Taek Kang; Sol Ji Park; Ma Ninia Gomez; Mohammad Atikuzzaman; Goo Jang; Byeong-Chun Lee Journal: Zygote Date: 2013-02-27 Impact factor: 1.442
Authors: Martha Luna; Alan B Copperman; Marlena Duke; Diego Ezcurra; Benjamin Sandler; Jason Barritt Journal: Fertil Steril Date: 2007-05-25 Impact factor: 7.329
Authors: Iratxe López-Pelayo; Javier María Gutiérrez-Romero; Ana Isabel Mangano Armada; María Mercedes Calero-Ruiz; Pablo Javier Moreno de Acevedo-Yagüe Journal: JBRA Assist Reprod Date: 2018-06-01