Differentiation of human endometrial stem cells encapsulated in alginate hydrogel into oocyte-like cells

Abstract

Introduction: Human endometrial mesenchymal stem cells (hEnMSCs) are a rich source of mesenchymal stem cells (MSCs) with multi-lineage differentiation potential, making them an intriguing tool in regenerative medicine, particularly for the treatment of reproductive and infertility issues. The specific process of germline cell-derived stem cell differentiation remains unknown, the aim is to study novel ways to achieve an effective differentiation method that produces adequate and functioning human gamete cells. Methods: We adjusted the optimum retinoic acid (RA) concentration for enhancement of germ cell-derived hEnSCs generation in 2D cell culture after 7 days in this study. Subsequently, we developed a suitable oocyte-like cell induction media including RA and bone morphogenetic protein 4 (BMP4), and studied their effects on oocyte-like cell differentiation in 2D and 3D cell culture media utilizing cells encapsulated in alginate hydrogel. Results: Our results from microscopy analysis, real-time PCR, and immunofluorescence tests revealed that 10 µM RA concentration was the optimal dose for inducing germ-like cells after 7 days. We examined the alginate hydrogel structural characteristics and integrity by rheology analysis and SEM microscope. We also demonstrated encapsulated cell viability and adhesion in the manufactured hydrogel. We propose that in 3D cell cultures in alginate hydrogel, an induction medium containing 10 µM RA and 50 ng/mL BMP4 can enhance hEnSC differentiation into oocyte-like cells. Conclusion: The production of oocyte-like cells using 3D alginate hydrogel may be viable in vitro approach for replacing gonad tissues and cells.

Silibinin ameliorates deoxycholic acid-induced pyroptosis in steatotic HepG2 cells by inhibiting NLRP3 inflammasome activation

“…Subsequently, the cells were incubated with primary antibodies overnight at 4 °C, followed by treatment with coraLite488-conjugated or coraLite594-conjugated secondary antibodies for 1 h at room temperature. The nuclei were stained with DAPI or PI, and the results were captured using a TCS SP8 confocal microscope (Leica, Mannheim, Germany) [ 38 ].…”