A method for isolating and culturing ectopic epithelial and stromal cells to study human adenomyosis

Purpose Although adenomyosis is a common and benign gynecological disease, the specific pathogenesis of this condition is yet to be fully elucidated. It is difficult to culture primary cells of the ectopic endometrial epithelia and stroma from human adenomyosis lesions. Most of the previous of studies on adenomyosis were based on primary eutopic endometrium cells. However, as yet, no efficient protocols have been developed for the isolation, culture or purification of primary ectopic epithelial and stromal cells from human adenomyosis lesions. Therefore, the present study aimed to develop an efficient protocol for the isolation and culture of primary ectopic epithelial and stromal cells from human adenomyosis lesions.

In the present study, we aimed to obtain ectopic endometrium tissue from human adenomyosis foci and use a simple and operable type I collagenase digestion method for primary culture. Cells were isolated by sterile cell strainer filtration and flow cytometry was performed to identify, purify, and evaluate the viability of isolated ectopic endometrial cells.

Using our method, we successfully isolated and cultured highly purified and active ectopic endometrial epithelial and stromal cells from human adenomyosis foci. Ep-CAM was expressed in ectopic epithelial cells of human adenomyosis with a purity of 93.74% and a viability of 80.58%. In addition, CD10 were robustly expressed by ectopic stromal cells in human adenomyosis. Cellular purity and viability were determined to be 96.37 and 93.49%, respectively.

Our method provides a new experimental model for studying the molecular pathogenesis of human adenomyosis. Similar content being viewed by others Data availability All the data we did were uploaded and saved in our hospital’s library. As it involves patient privacy and other relevant national policies, it is not provided in this manuscript. However, if anyone is interested in it, please contact the corresponding author directly.

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This work was funded by National Key R&D Program of China (Grant number: 2022YFC2704003), National Natural Science Foundation of China (Grant numbers: 81974225, 82001518 and 82171636). Author information Authors and Affiliations Contributions ZF: conceptualization, writing—original draft, data curation, methodology, writing—review and editing, validation, resources. JW and TL: conceptualization, methodology, writing—review and editing, validation, supervision. MY and YP: data curation, methodology, resources, validation. XZ: supervision, conceptualization, writing—review and editing, project administration, validation. All the authors read the submitted version and approved it. Corresponding author Ethics declarations Conflict of interest The authors report no conflict of interest. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. Additional information Publisher's Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Rights and permissions Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law. About this article Cite this article Fang, Z., Wang, J., Li, T. et al. A method for isolating and culturing ectopic epithelial and stromal cells to study human adenomyosis. Arch Gynecol Obstet 309, 551–563 (2024). https://doi.org/10.1007/s00404-023-07254-8 Received: Accepted: Published: Version of record: Issue date: DOI: https://doi.org/10.1007/s00404-023-07254-8