Publikationsdatenbank

Establishment of a murine 3D cell culture model of the endometrium (2019)

Art

Poster

Autoren

Peter, Dominique (WE 11)

Erickson, N. A. (WE 11)

Mundhenk, L (WE 12)

Michel, Geert

Na, Ellen

Bienert, Michaela

Buck, Volker

Wittler, Lars

Thoene-Reineke, C (WE 11)

Forschungsprojekt

Save – IVF; Entwicklung eines 3D in-vitro Fertilisationszellkulturmodell in der Maus

Kongress

7th Annual GSCN Conference of the German Stem Cell Network (GSCN)

Berlin, Germany, 23.09. – 25.12.2019

Quelle

Kontakt

Institut für Tierschutz, Tierverhalten und Versuchstierkunde

Königsweg 67
14163 Berlin
+49 30 838 61146
tierschutz@vetmed.fu-berlin.de

Abstract / Zusammenfassung

Establishment of a murine 3D-cell-culture-model of the endometrium for early embryo implantation

Due to the complexity of embryo implantation, animal studies are not fully replaceable. In vitro models may at least partially mirror early embryo implantation. This study aims to establish a murine 3D-cell-culture-model - combining endometrial cells with an embryoid body generated in vitro from murine trophoblast stem cells (TS cells) or an ex vivo blastocyst - resembling first embryo-maternal interaction to analyze the mechanisms of early embryo implantation.
Endometrial stromal and epithelial cells were isolated simultaneously in separate suspensions (1) from uteri of mice 3.5 days after plug detection. For 3D-formation, stromal cells were suspended in GrowDex® or Matrigel® with apical epithelial cell seeding after gel solidification and cultivation for up to six days. Matrix and cellular conservation were compared macroscopically and microscopically by hematoxylin-eosin staining after cryo-, paraformaldehyde- or methanol-carnoy-fixation. Cells were immunohistophenotyped via cytokeratin AE1/AE3-, Ki67-, and active caspase 3- antibodies. For embryoid body generation, TS cells were cultured with proliferation-inhibited mouse embryonal fibroblasts (MEF), passaged, and banked (2).
Compared to GrowDex®, Matrigel® offered adequate handling and stability. In contrast to other fixation methods, only PFA revealed best macroscopic and histologic preservation of gel structure. Some stromal cells formed extensions in Matrigel®. Cytokeratin-positive epithelial cells lacked high prismatic morphology, tended to aggregate apically, whilst forming gland-like structures in Matrigel®. Some epithelial cells showed positive Ki67 signals, whilst most of the cells were caspase negative. TS cells grew in typical roundish colonies with defined demarcation toward MEF cells and increasing cell density over time.