The experience of extended in vitro human embryo cultivation in a culture medium containing endometrium cells. A pilot study

Abstract

BACKGROUND: At present, the knowledge on initial human embryogenesis stages is limited to the period of development from zygote to blastocyst. The creation of models based on the interaction between the embryo and endometrium in vitro, which accurately imitate the in vivo processes, represents the major way for implantation and post-implantation evaluation. Currently, no reports have been made on models reflecting the both processes simultaneously: the interaction of the embryo with the substrate, which represents many aspects of normal implantation, and the early post-implantation embryo development. The creation of a relevant model would allow investigation of implantation and early post-implantation processes as a whole.

AIM: The aim of this study was to evaluate the vitality and developmental potential of human embryos from the day 6 blastocyst stage during their extended co-incubation with the endometrium in a culture medium specifically designed for cultivation to the blastocyst stage.

MATERIALS AND METHODS: Embryos obtained in assisted reproductive technology programs were cultivated from the day 6 blastocyst stage up to 14 days of development in vitro in a culture medium designed for cultivation to the blastocyst stage, in the presence of endometrium cells. On day 14 of development, embryos and endometrial samples were first evaluated under an inverted microscope using Hoffman modulation contrast, then transferred to a special mold and impregnated with paraffin for cytoblock preparation. Obtained blocks were sliced, stained with hematoxylin and eosin and morphologically assessed.

RESULTS: The first sample visual assessment on day 14 of cultivation in a culture medium with endometrium cells revealed a viable developing embryo with no signs of degradation. During the histological examination, the endometrial sample corresponded to the secretory phase of the cycle. The morphological assessment of the conceptus detected trophoblast cells. The second sample visual assessment on the day 14 of cultivation in a culture medium with endometrium cells revealed a viable embryo with no signs of degradation, which was in direct contact with the endometrial component. A histological examination detected a secretory endometrial fragment of the surface (luminal) epithelium. During the morphological assessment of the embryo, trophoblast cells were detected.

CONCLUSIONS: The data obtained indicate the ability of the embryo to further develop from the day 6 blastocyst stage up to 14 days in a culture medium specifically designed for cultivation to the blastocyst stage, in the presence of endometrium cells. The latter can serve as an experimental model for both in vitro endometrial receptivity evaluation and intercellular interactions during implantation investigation.

Keywords

embryo / cultivation / endometrium / receptivity / extended cultivation / implantation

References

[1] Evans J, Walker KJ, Bilandzic M, et al. A novel “embryo-endometrial” adhesion model can potentially predict “receptive” or “non-receptive” endometrium. J Assist Reprod Genet. 2020;37(1):5–16. DOI: 10.1007/s10815-019-01629-0

[2] Evans J., Walker K.J., Bilandzic M., et al. A novel “embryo-endometrial” adhesion model can potentially predict “receptive” or “non-receptive” endometrium // J. Assist. Reprod. Genet. 2020. Vol. 37. No. 1. P. 5–16. DOI: 10.1007/s10815-019-01629-0

[3] You Y, Stelzl P, Zhang Y, et al. Novel 3D in vitro models to evaluate trophoblast migration and invasion. Am J Reprod Immunol. 2019;81(3):e13076. DOI: 10.1111/aji.13076

[4] You Y., Stelzl P., Zhang Y., et al. Novel 3D in vitro models to evaluate trophoblast migration and invasion // Am. J. Reprod. Immunol. 2019. Vol. 81. No. 3. P. e13076. DOI: 10.1111/aji.13076

[5] Berneau SC, Ruane PT, Brison DR, et al. Characterisation of osteopontin in an in vitro model of embryo implantation. Cells. 2019;8(5):432. DOI: 10.3390/cells8050432

[6] Berneau S.C., Ruane P.T., Brison D.R., Kimber S.J., et al. Characterisation of osteopontin in an in vitro model of embryo implantation // Cells. 2019. Vol. 8. No. 5. P. 432. DOI: 10.3390/cells8050432

[7] Zambuto SG, Clancy KBH, Harley BAC. A gelatin hydrogel to study endometrial angiogenesis and trophoblast invasion. Interface Focus. 2019;9(5):20190016. DOI: 10.1098/rsfs.2019.0016

[8] Zambuto S.G., Clancy K.B.H., Harley B.A.C. A gelatin hydrogel to study endometrial angiogenesis and trophoblast invasion // Interface Focus. 2019. Vol. 9. No. 5. P. 20190016. DOI: 10.1098/rsfs.2019.0016

[9] Stern-Tal D, Achache H, Jacobs Catane L, et al. Novel 3D embryo implantation model within macroporous alginate scaffolds. J Biol Eng. 2020;14:18. DOI: 10.1186/s13036-020-00240-7

[10] Stern-Tal D., Achache H., Jacobs Catane L., Reich R., Tavor Re’em T. Novel 3D embryo implantation model within macroporous alginate scaffolds // J. Biol. Eng. 2020. Vol. 14. P. 18. DOI: 10.1186/s13036-020-00240-7

[11] ISSCR. Guidelines for the Conduct of Human Embryonic Stem Cell Research. 2006. Version 1: December 21, 2006. [cited 2022 Aug 15]. Available from: https://www.isscr.org/docs/default-source/all-isscr-guidelines/hesc-guidelines/isscrhescguidelines2006.pdf?sfvrsn=0

[12] ISSCR. Guidelines for the Conduct of Human Embryonic Stem Cell Research. 2006. Version 1: December 21, 2006. [дата обращения: 15.08.2022]. Доступ по ссылке: https://www.isscr.org/docs/default-source/all-isscr-guidelines/hesc-guidelines/isscrhescguidelines2006.pdf?sfvrsn=0

[13] ISSCR. Guidelines for Stem Cell Research and Clinical Translation update. 2021. Version 1.0, May, 2021. [cited 2022 Aug 15]. Available from: https://www.isscr.org/docs/default-source/all-isscr-guidelines/2021-guidelines/isscr-guidelines-for-stem-cell-research-and-clinical-translation-2021.pdf?sfvrsn=979d58b1_4

[14] ISSCR. Guidelines for Stem Cell Research and Clinical Translation update. 2021. Version 1.0, May, 2021. [дата обращения: 15.08.2022]. Доступ по ссылке: https://www.isscr.org/docs/default-source/all-isscr-guidelines/2021-guidelines/isscr-guidelines-for-stem-cell-research-and-clinical-translation-2021.pdf?sfvrsn= 979d58b1_4

[15] Pera MF, de Wert G, Dondorp W, et al. What if stem cells turn into embryos in a dish? Nat Methods. 2015;12(10):917–919. DOI: 10.1038/nmeth.3586

[16] Pera M.F., de Wert G., Dondorp W., et al. What if stem cells turn into embryos in a dish? // Nat. Methods. 2015. Vol. 12. No. 10. P. 917–919. DOI: 10.1038/nmeth.3586

[17] Shahbazi MN, Jedrusik A, Vuoristo S, et al. Self-organisation of the human embryo in the absence of maternal tissues. Nat Cell Biol. 2016;18(6):700–708. DOI: 10.1038/ncb3347

[18] Shahbazi M.N., Jedrusik A., Vuoristo S., et al. Self-organisation of the human embryo in the absence of maternal tissues // Nat. Cell Biol. 2016. Vol. 18. No. 6. P. 700–708. DOI: 10.1038/ncb3347

[19] Deglincerti A, Croft GF, Pietila LN, et al. Self-organization of the in vitro attached human embryo. Nature. 2016;533:251–254. DOI: 10.1038/nature17948

[20] Deglincerti A., Croft G.F., Pietila L.N., et al. Self-organization of the in vitro attached human embryo // Nature. 2016. Vol. 533. P. 251–254. DOI: 10.1038/nature17948

[21] Teklenburg G, Salker M, Molokhia M, et al. Natural selection of human embryos: Decidualizing endometrial stromal cells serve as sensors of embryo quality upon implantation. PLoS One. 2010;5(4):2–7. DOI: 10.1371/journal.pone.0010258

[22] Teklenburg G., Salker M., Molokhia M., et al. Natural selection of human embryos: Decidualizing endometrial stromal cells serve as sensors of embryo quality upon implantation // PLoS One. 2010. Vol. 5. No. 4. P. 2–7. DOI: 10.1371/journal.pone.0010258

[23] Brosens JJ, Salker MS, Teklenburg G, et al. Uterine selection of human embryos at implantation. Sci Rep. 2014;4:4–11. DOI: 10.1038/srep03894

[24] Brosens J.J., Salker M.S., Teklenburg G., et al. Uterine selection of human embryos at implantation // Sci. Rep. 2014. Vol. 4. P. 4–11. DOI: 10.1038/srep03894

[25] Izmailova LSh, Vorotelyak EA, Vasiliev AV. Modeling of early development of mouse and human embryos in vitro. Ontogenez. 2020;51(5):323–337. (In Russ.). DOI: 10.31857/S0475145020050043

[26] Измайлова Л.Ш., Воротеляк Е.А., Васильев А.В. Моделирование раннего развития эмбрионов мыши и человека in vitro // Онтогенез. 2020. Т. 51. № 5. С. 323–337. DOI: 10.31857/S0475145020050043

[27] Kolahi KS, Donjacour A, Xiaowei L, et al. Effect of substrate stiffness on early mouse embryo development. PLoS One. 2012;7(7):e41717. DOI: 10.1371/journal.pone.0041717

[28] Kolahi K.S., Donjacour A., Xiaowei L., et al. Effect of substrate stiffness on early mouse embryo development // PLoS One. 2012. Vol. 7. No. 7. P. e41717. DOI: 10.1371/journal.pone.0041717

[29] Hiramatsu R, Matsuoka T, Kimura-Yoshida C, et al. External mechanical cues trigger the establishment of the anterior-posterior axis in early mouse embryos. Dev Cell. 2013;27(2):131–144. DOI: 10.1016/j.devcel.2013.09.026

[30] Hiramatsu R., Matsuoka T., Kimura-Yoshida C., et al. External mechanical cues trigger the establishment of the anterior-posterior axis in early mouse embryos // Dev. Cell. 2013. Vol. 27. No. 2. P. 131–144. DOI: 10.1016/j.devcel.2013.09.026

[31] Koot YEМ, Teklenburg G, Salker MS, et al. Molecular aspects of implantation failure. Biochim Biophys Acta. 2012;1822(12):1943–1950. DOI: 10.1016/j.bbadis.2012.05.017

[32] Koot Y.E.М, Teklenburg G., Salker M.S., et al. Molecular aspects of implantation failure // Biochim. Biophys. Acta. 2012. Vol. 1822. No. 12. P. 1943–1950. DOI: 10.1016/j.bbadis.2012.05.017

[33] Weimar CH, Post Uiterweer ED, Teklenburg G, et al. In-vitro model systems for the study of human embryo-endometrium interactions. Reprod BioMed Online. 2013;27(5):461–476. DOI: 10.1016/j.rbmo.2013.08.002

[34] Weimar C.H., Post Uiterweer E.D., Teklenburg G., et al. In-vitro model systems for the study of human embryo-endometrium interactions // Reprod. BioMed. Online. 2013. Vol. 27. No. 5. P. 461–476. DOI: 10.1016/j.rbmo.2013.08.002

RIGHTS & PERMISSIONS

Eсо-Vector