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. 2021 Dec 2;601(7894):600–605. doi: 10.1038/s41586-021-04267-8

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Extended Data Fig. 8 — a. Immunofluorescence stainings for CDH1 (Magenta) and a ciliated cell marker acetylated α-tubulin (Yellow) in OFELs (left). Y-Z plane shows the apical location of the cilia (right). Scale bar: 50 μm. b. Immunofluorescence staining for FOXA2 (Yellow) marking the endometrial glandular cells in OFELs. Scale bar: 50 μm. c. Immunofluorescence staining for PAEP (Yellow) in non-stimulated (left) and stimulated (right) OFELs. d. qRT-PCR measurement of the expression levels of window-of-implantation markers in OFELs cultured with different media. Ctrl: Control medium, E: Estradiol, P: Progesterone, C: cAMP, X: XAV-939. Expression levels were normalized relative to the housekeeping gene GAPDH and the control condition. n = 2 independent experiments. The colors depict the data from 3 different donors. e. Heatmap of key cell cycle and secretory epithelial genes differentially expressed between stimulated and non-stimulated OFELs in bulk transcriptome. f. Staining for incorporated EdU (Red) reflective of cell proliferation in a stimulated OFEL (left). Scale bar: 50 μm. Quantification of the number of EdU⁺ cells in non-stimulated and stimulated OFELs (right). Counterstain with Hoechst marking DNA. n=4 independent experiments. mean± S.D.; Unpaired two-tailed t-test, *** is P = 0.0009. g. Quantification of blastoid attachment onto OFELs prepared using endometrial organoids from 3 different donors. n=3 independent experiments for donor 1 and n=2 independent experiments for donor 2 and 3; mean± S.D.; Unpaired two-tailed t-test, ** is P =0.0011. h. Immunofluorescence stainings for MUC1 (Magenta), a glycoprotein that highly expresses at the luminal epithelial surface of endometrium in the receptive phase⁴⁶, with an attached GFP+ blastoid (48 h after deposition onto an OFEL). Dashed lines indicate the area that trophoblast cells repelled endometrial cells. Scale bar: 200 μm i. Quantification of blastoid attachment onto non-stimulated, stimulated OFELs, and OFELs additionally exposed to the contraceptive Levonorgestrel (LNG, 10 μM). n=3 independent experiments. mean± S.D.; one-way Anova and Tukey’s multiple comparisons test, * is P = 0.0211, *** is P = 0.0006.

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