An assessment of the differentiation potential of epithelial stem cells in adult endometrium

Background: We are at the beginning of a new era where regenerative medicine is really starting to take hold. An appreciation of adult stem cells and their role in tissue maintenance have heralded novel insight into endometrial remodelling. The idea that human endometrium could be a plentiful source of adult stem/progenitor cells (ASPCs) has captured the imagination of many stem cell biologists. Not only that, they are also hypothesised to be involved in endometrial proliferative diseases, such as endometriosis. However, work remains preliminary and is stunted by a lack of specific endometrial stem cell markers. Recently, stage specific embryonic antigen -1 (SSEA-1) was proposed as a candidate epithelial ASPC marker in adult endometrium. In order to validate this, stem cell assays are required. These assays include measurements of clonogenicity, prolonged self-renewal and differentiation potential. Aim: To assess the differentiation potential of SSEA-1+ cells in vitro. Methods: All endometrial culture samples were collected from pre-menopausal women undergoing surgery for benign disease at Liverpool Women’s Hospital (LWH). Epithelial cells were cultured in 3D MatrigelTM to produce gland-like structures. Gland-like structures were characterised via immuno-histochemistry, to validate their resemblance to glands in endometrial tissue in vivo. To assess multi-potency, magnetic cell sorted (MACS) SSEA-1 enriched and depleted cell fractions were cultured in adipogenic and osteogenic inducing media. In order to assess pluripotency, SSEA-1 enriched and depleted fractions were cultured in neurogenic media. Results: SSEA-1 enriched cell fractions had a greater propensity to produce gland-like structures in 3D culture than SSEA-1 depleted cell fractions. Gland-like structures best resembled glands in post-menopausal (PM) endometrium. Non-gland like structures also formed in 3D culture from endometriosis samples, which stained exclusively and intensely for CK5/6. SSEA-1 enriched and depleted fractions were unable to differentiate into other cell types of mesodermal lineage. However, both enriched and depleted fractions were able to produce ectodermal derived neural-like PGP9.5 positive cells. Discussion: This work confirms SSEA-1 is a reliable epithelial ASPC marker in adult endometrium. SSEA-1+ cells possess an increased ability to produce gland-like structures, reminiscent of in vivo endometrial glands. This therefore confirms unipotency. The presence of CK5/6 in non-gland-like structures from endometriosis samples suggests a potential role in endometriosis aetiology, and warrants further work. SSEA-1 enriched and depleted cells were unable to undergo multi-lineage differentiation, but were capable of producing neural-like cells. As both cell populations were unable to demonstrate multipotency, it unlikely SSEA-1 enriched/depleted cells are pluripotent. More likely, a neural progenitor may exist in adult endometrium, one that has been included in both cell populations. Given the close proximity of endometrial glands to the placenta, the commitment of epithelial stem cells could be an ancillary mechanism to prevent maternal microchimersim in offspring, which is associated with a number of pathologies.