Asherman’s syndrome (AS) is one of the gynecological disorders caused by the destruction of the endometrium. For some cases of AS available surgical methods and hormonal therapy are ineffective. Stem cell transplantation may offer a potential alternative for AS cure. Human endometrial mesenchymal stem cells (eMSC) organized in spheroids were transplanted in rats with damaged endometrium modeled on AS. Treatment response was defined as pregnancy outcome and litter size. Application of eMSC in spheroids significantly improved the rat fertility with the AS model. eMSC organized in spheroids retain all properties of eMSC in monolayer: growth characteristics, expression of CD markers, and differentiation potential. Synthesis of angiogenic and anti-inflammatory factors drastically increased in eMSC assembled into spheroids. Human endometrial mesenchymal stem cells (eMSC) can be successfully applied for Asherman’s syndrome (AS) treatment in the rat model. eMSC organized in spheroids were more therapeutically effective than the cells in monolayer. After transplantation of eMSC in spheroids the pregnancy outcome and litter size in rats with AS was higher than in rats that received autologous rat bone marrow cells. It suggests the therapeutic plausibility of heterologous eMSC in case of failure to use autologous cells.

Endometrial mesenchymal stem/stromal cells (eMSCs) hold great promise in bioengineering and regenerative medicine due to their high expansion potential, unique immunosuppressive properties and multilineage differentiation capacity. Usually, eMSCs are maintained and applied as a monolayer culture. Recently, using animal models with endometrial and skin defects, we showed that formation of multicellular aggregates known as spheroids from eMSCs enhances their tissue repair capabilities. In this work, we refined a method of spheroid formation, which makes it possible to obtain well-formed aggregates with a narrow size distribution both at early eMSC passages and after prolonged cultivation. The use of serum-free media allows this method to be used for the production of spheroids for clinical purposes. Wound healing experiments on animals confirmed the high therapeutic potency of the produced eMSC spheroids in comparison to the monolayer eMSC culture.

Human-endometrium mesenchymal stem cells (eMSCs) are a promising source of stem cells for regenerative medicine. A large amount of these cells accumulated by in vitro cultivation are usually required for transplantation into patients. We established several cell eMSC lines and cultivated them over a long period to examine the possibility of spontaneous transformation. All cell lines exhibit limited lifespan, undergo replicative senescence, and die. Karyotypic analysis upon different passages reveals that most cells display karyotypic stability. Thus, extended in vitro cultivation of eMSCs does not lead to spontaneous transformation, which makes therapeutic application of these cells safe for patients. During long-term cultivation, eMSCs maintain the expression of surface markers.

The mechanisms underlying the therapeutic potential of MSCs are the focus of intense research. We studied human MSCs isolated from desquamated endometrium (eMSCs), which, as previously shown, have high regenerative potential in various disease models. The aim was to evaluate the role of secreted VEGF in stimulating angiogenesis and maintaining eMSC viability and migration, which is important for improving the therapeutic properties of MSCs. We compared three eMSC cultures differing in the level of VEGF secretion: 3D spheroids, monolayer eMSCs, and monolayer eMSCs with VEGF knockdown. Spheroid eMSCs produced higher amounts of VEGF and had the strongest paracrine effect on HUVEC. eMSCs with VEGF knockdown did not stimulate angiogenesis. Monolayered eMSCs expressed VEGFR1, while spheroid eMSCs expressed both VEGFR1 and VEGFR2 receptors. The knockdown of VEGF caused a significant decrease in the viability and migration of eMSCs. eMSCs from 3D spheroids enhanced proliferation and migration in response to exogenous VEGF, in contrast to monolayered eMSCs. Our results suggest that the VEGF–VEGFR1 loop appears to be autocrine-involved in maintaining the viability of eMSCs, and VEGFR2 expression enhances their response to exogenous VEGF, so the angiogenic potential of eMSC can be up- or downregulated by intrinsic VEGF signals.

Stem cell transplantation, which is based on the application of mesenchymal stem/stromal cells (MSCs), is a rapidly developing approach to the regenerative therapy of various degenerative disorders characterized by brain and heart failure, as well as skin lesions. In comparison, the use of stem cell transplantations to treat infertility has received less attention. One of the causes of miscarriages and fetal growth delay is the loss of the decidual reaction of endometrial cells. The present study modeled decidualization processes in pseudopregnant rats. For cell transplantation experiments, the rats were transplanted with MSCs established from endometrial fragments in menstrual blood (eMSCs). These cells express common MSC markers, are multipotent and are able to differentiate into various tissue lineages. Cell therapy frequently requires substantial cell biomass, and cultivation of MSCs may be accompanied by significant changes to their properties, including malignant transformation. In order to minimize the potential for malignant transformation, the proliferation of eMSCs was irreversibly suppressed by irradiation and mitomycin C treatment. Transplantation of the rats with viable, non-proliferating eMSCs stimulated the development of all elements of decidual tissue. Conversely, transplantation of the rats with cells killed using 95% ethanol did not result in the development of decidual tissue. The present study demonstrated the potential for applying eMSCs to the cellular therapy of infertility associated with endometrial disorders characterized by decidualization insufficiency and implantation failure. In addition, the transplantation of viable but non-proliferating cells ensured that their oncogenic potential was limited.