Derivation of hormone-responsive human endometrial organoids and stromal cells from cryopreserved biopsies

The human endometrium is a dynamic tissue that undergoes cyclic changes in response to sex steroid hormones to provide a receptive status for embryo implantation. Disruptions in this behavior may lead to implantation failure and infertility; therefore, it is essential to develop an appropriate in vitro model to study endometrial changes in response to sex hormones. In this regard, the first choice would be human endometrial cells isolated from biopsies that could be used as monolayer cell sheets or to generate endometrial organoids. However, the need for fresh samples and short-time viability of harvested endometrial biopsy limits these approaches. In order to overcome these limitations, we sought to develop an efficient, simple, robust and reproducible method to cryopreserve human endometrial biopsies that could be stored and/or shipped frozen and later thawed to generate endometrial organoids and endometrial stromal cells (EnSCs). These cryopreserved biopsies could be thawed and used to generate simple endometrial organoids or organoids for co-culture with matched stromal cells that are functionally responsive to sex hormones as similar as the organoids generated from fresh biopsy. An optimal endometrial tissue cryopreservation method would allow the possibility for endometrial tissue biobanking to enable future organoid generation from both healthy tissues and pathological conditions, and open new venues for generate endometrial assembloids, consisting of epithelial organoids and primary stromal cells. •Human endometrial organoids can be generated and long-term cultured from cryopreserved biopsies.•Cryopreserved human endometrial biopsies-derived organoids and stromal cells are functionally responsive to sex hormones.•Co-cultured human endometrial organoids with matched stromal cells, the assembloids, respond to sex hormone treatment.