RESUMEN
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.
RESUMEN
BACKGROUND: The aim of this study is to investigate the co-culture effects of human endometrial mesenchymal stem cells (EnMSCs) with mouse oocytes to enhance their maturation and development by using the hanging drop and sodium alginate hydrogel methods. MATERIALS AND METHODS: In this experimental study, we prepared human EnMSCs (2.5×105 cells/mL) and co-cultured them with partially denuded mouse oocytes by the hanging drop (n=120) and sodium alginate hydrogel (n=120) methods. Control oocytes (n=230, total) were cultured in both systems in the absence of human EnMSCs for 18 hours. Both survival and maturation rates of the oocytes were analysed morphologically. After insemination with capacitated sperm, the fertilization and development of the embryos up to the blastocyst stage were assessed and compared statistically for all of the study groups via one-way ANOVA and the t tests. RESULTS: Oocytes cultured in the hanging drop method had a significantly higher survival rate than their control group (92.60 ± 4.36% vs. 84.20 ± 3.12%, P=0.018). There were no significant differences between the two experimental groups in terms of survival. The mean percent of oocytes that reached the metaphase II (MII) stage was 64.35 ± 3.19% and fertilised was 62.25 ± 4.43% in the hanging drop method; these rates were 63.43 ± 1.92% and 58.14 ± 4.14 in sodium alginate hydrogel method, respectively. These rates were higher than their controls (P<0.050), but there were no statistical differences between the two experimental groups (P>0.050). Among the studied groups, the highest significant blastocyst rate (32.55 ± 2.18%) was observed in the hanging drop experimental group (P=0.0017). CONCLUSION: The results of this study show that human EnMSCs improve the survival, maturation, and development rates of oocytes and they could have future clinical applications.
RESUMEN
Purpose: Given the antiandrogenic effects of spearmint, in this study we evaluated the effects of its essential oil on polycystic ovarian syndrome in a rat model. Methods: Female rats were treated as follows: Control, normal rats which received 150 mg/kg spearmint oil or 300 mg/kg spearmint oil, or sesame oil; and PCOS-induced rats which received 150 mg/kg spearmint oil or 300 mg/kg spearmint oil, or sesame oil. Then the animals were killed and the levels of LH, FSH, testosterone and ovarian folliculogenesis were evaluated. Results: Spearmint oil reduced body weight, testosterone level, ovarian cysts and atretic follicles and increased Graafian follicles in PCOS rats. Conclusion: Spearmint has treatment potential on PCOS through inhibition of testosterone and restoration of follicular development in ovarian tissue.