Bone tissue regeneration by 58S bioactive glass scaffolds containing exosome: an in vivo study.

Exosomes, the naturally secreted nanocarriers of cells, have recently been demonstrated to have therapeutic benefits in a variety of disease models where parent cells are not present. However, the use of exosomes in bone defect regeneration has been unusual, and little is documented about the underlying processes. In recent study we produced and characterized exosomes derived human endometrial mesenchymal stem stromal cells and 58S bioactive glass scaffolds; in following, in this research exosome loaded scaffolds synthetized and release of exosome, porosity and bioactivity of them were assessed. More over the effect of scaffolds on repair of critical-size bone defects in rat's calvaria was evaluated by histological examination and micro computed tomography (µ CT). The findings confirmed that constructed porous scaffolds consistently release exosomes; additionally, in vivo findings including Hematoxilin & Eosin staining, Immunohistochemistry, Masson's trichrome, histomorphometric analysis, and µ CT clarified that our implant has osteogenic properties. We discovered that Exo-treated scaffolds might promote osteogenesis especially compared to pure scaffolds, indicating that produced scaffolds containing exosomes could be a potential replacement in bone tissue engineering.

Optimizing human endometrial mesenchymal stem cells for maximal induction of angiogenesis.

Human endometrial mesenchymal stem cells (hEMSCs) have been shown to promote neo-vascularization; however, its angiogenic function lessens with age. To determine the optimal conditions for maximizing hEMSC angiogenic capacity, we examined the effects of serial passaging on hEMSC activity. hEMSCs were cultured from passages (P) 3, 6, 9, and 12, and analyzed for proliferation, migration, differentiation and senescence, as well as their capacity to induce angiogenesis. The results showed that hEMSC proliferation and migration significantly decreased after P12. Furthermore, hEMSC differentiation into adipogenic and osteogenic lineages, as well as their proangiogenic capacity, gradually decreased from P9-12, while senescence only occurred after P12. Evaluation of angiogenic-related protein levels showed that both transforming growth factor ß2 and Tie-2 was significantly reduced in hEMSCs at P12, compared to P3, possibly serving as the basis behind their lowered angiogenic capacity. Furthermore, in vivo angiogenesis evaluation with Matrigel plug assay showed that the optimal hEMSC to HUVEC ratio, for maximizing vessel formation, was 1:4. This study showed that hEMSC passaging was associated with lowered cellular functioning, bringing them closer to a senescent phenotype, especially after P12, thereby defining the optimal time period for cultivating fully functional hEMSCs for therapeutic applications.

Differentiation of human menstrual blood-derived endometrial mesenchymal stem cells into oocyte-like cells.

Human endometrial mesenchymal stem cells (EnSCs) derived from menstrual blood are a unique stem cell source. Evidence suggests that EnSCs exhibit a multi-lineage potential and have attracted extensive attention in regenerative medicine. However, the potential of EnSCs to differentiate into germline cells in vitro remains unclear. In this study, EnSCs were induced to differentiate into germ cells in a differentiation medium supplemented with 20% human follicular fluid. Our results demonstrated that EnSCs derived from human menstrual blood form oocyte-like cells and express germ cell markers. The induced cell aggregates contained not only oocyte-like structures but also cells expressing follicle stimulating hormone receptor and luteotropic hormone receptor, and produced estrogen and progesterone regulated by gonodatropin, suggesting that granulosa-like and theca-like cells were also induced. We further found that granulosa cells promote the development of oocyte-like cells and activate the induction of blastocyst-like structures derived from EnSCs. In conclusion, EnSCs may potentially represent an in vitro system for the investigation of human folliculogenesis.

Formation of ovarian organoid by co-culture of human endometrial mesenchymal stem cells and mouse oocyte in 3-dimensional culture system.

The purpose of this study was to compare the formation of organoid structures by co-culturing of human endometrial mesenchymal stem cells (hEnMSCs) and mouse germinal vesicle (GV) oocytes in hanging drop and sodium alginate hydrogel co-culture methods. Following the preparation of hEnMSCs and partially denuded mouse germinal vesicle oocytes, they were co-cultured in hanging drop and sodium alginate hydrogel systems as two experimental groups. In respected control groups the hEnMSCs were cultured without oocytes. The organoid formation was evaluated under the inverted microscope in all studied groups during the culture period. The hematoxylin and eosin, alcian blue, periodic acid Schiff, and Masson's trichrome methods, were applied for morphological evaluation and extracellular matrix components staining such as glycosaminoglycan, carbohydrate, and collagen fibers. In addition, the germ cell-like characteristics within the organoid structures were investigated via alkaline phosphatase activity immunocytochemistry for DEAD-box polypeptide 4 (DDX4), and the expression of octamer-binding transcription factor 4 (OCT4), DDX4, and synaptonemal complex protein 3 (SYCP3) genes by real-time RT-PCR. The culturing of hEnMSCs in the hanging drop method led to the formation of organoid structures while this structure was not seen in sodium alginate hydrogel culture. The mean diameter of organoid structures was increased during 4 days of culture in both the experimental and control groups in the hanging drop method, reaching 675.50 ± 18.55 µm and 670.25 ± 21.40 µm, respectively (P < 0.05). Morphological staining indicated some large ovoid cells with euchromatin nuclei in the experimental group, whereas, in the control group cells showed dark and dense nuclei. The extracellular matrix components were deposited in organoid structures in both control and experimental groups. The positive alkaline phosphatase activity and immunocytochemistry for DDX4 confirmed the presence of germ cell-like in the experimental group. Real-time RT-PCR showed a significant increase in the expression of DDX4 and SYCP3 genes and a decrease in the level of OCT4 expression in the experimental group compared with its controls. This study successfully generated organoid structures by co-culture of hEnMSCs and oocytes in the hanging drop method and the hEnMSCs could be differentiated into germ cell-like. This organoid structure has potential applications in regenerative medicine and reproductive biology. Supplementary Information: The online version contains supplementary material available at 10.1007/s10616-024-00639-w.