Differentiation of mouse bone marrow mesenchymal stem cells into germ-like cells by retinoic acid, titanium nanotubes-coated fibrin formation, and ultraviolet radiation as inductions in vitro

Abstract

Abstract Background Although advances in reproductive medicine-relative stem cell biology are focused on multipotent stem cells differentiation into artificial gametes, which is low-efficiency-functional of the germ-like cells. Our purpose is to survey and promote the potential of mouse bone marrow mesenchymal stem cells (mBMMSCs) differentiation into late-stage germ-like cells (GLCs) by increasing the expression of the germline stem cells markers. Materials and Methods In this experimental study, the multipotent differentiation potential of mBMMSCs assesmented by Alizarin red-S and Oil red-O staining. Furthermore, we investigated the suitable conditions which promoted mBMMSCs differentiation into GLCs by applying the effective concentration of retinoic acid (RA), culturing them in titanium nanotubes (TNTs)-coated fibrin (F + TNT) formation as a 2D scaffold, and ultraviolet (UV) radiation inductions afterward. The biocompatibility and morphology of the TNTs and the F + TNT properties were examined by MTT and Scanning Electron Microscope (SEM) analysis, respectively. Results The optimum concentration biocompatibility of TNTs was 50 µg/ml for differentiation after 14 days. The morphology of the TNTs and F + TNT were confirmed using SEM and Raman spectrum, respectively. After culturing the multipotent of mBMMSCs in RA and F + TNT formation with RA and UV radiation, male and female germline stem cell markers, Mvh / Ddx4, Dazl, and Plzf expression, and the cells' integrity in fibrin were revealed using Immunofluorescence analysis, Western Blots, and Real-Time quantitative PCR (RT-qPCR). Conclusion We demonstrate that mBMMSCs can be differentiated into GLCs in a medium containing 10− 5 M RA in which the specific marker was expressed properly in 2D F + TNT formation. In addition, F + TNT and UV radiation could offer a proper 2D scaffold for mBMMSCs-derived germ-like cells regarding in vitro maturation (IVM) of assisted reproductive technology (ART).