Differentiation of Human Menstrual Blood Stem Cells into Female Germ Cells in Indirect Co-culture with Human Granulosa Cells

Background: Infertility is a genital disease that defines a couple's inability to conceive after twelve months of regular and unprotected sex. Infertility affects approximately 10 to 15 percent of couples worldwide. The evaluation of female factors has traditionally been the trigger for infertility studies. Causes of ovulation are the most common contributing factor to infertility. Despite advances in assisted reproductive technologies, infertility remains a serious problem. In recent years, significant advances in cell therapy have emerged as an emerging approach to infertility treatment. This study aimed to differentiate human menstrual blood stem cells (MenSCs) from female germ cells in indirect co-culture with human granulosa cells (GCs), which is a type of human in vitro gametogenesis that can transform reproductive medicine. Methods: In this study, MenSCs and GCs after isolation and culture were examined for expression of mesenchymal (CD105, CD29, CD44, CD73, CD90, CD10, and CD9) and hematopoietic (CD45, CD34, CD38, C and D133) markers by flow cytometry. Subsequently, the cells were divided into two groups, including the control group (culture of MenSCs without co-culture with GCs) and the co-culture group (culture of MenSCs in co-culture with GCs). After two weeks, the expression of specific markers of female gametes (Vasa, SCP3, and GDF9) at both molecular and protein levels were examined by Real time-PCR and immunocytochemistry techniques, respectively. Results: Results from real-time PCR of the co-culture group showed the activation of germ cell-specific genes, including GDF9, Vasa, and SCP3, after two weeks, indicating cell differentiation. The expression of the above-mentioned genes was not observed in the control group. Immunocytochemical analysis used for the expression of oocyte markers showed that the mentioned genes were expressed in the co-culture group, while the control group did not express these genes. Conclusion: Co-culture of GCs with MenSCs is an effective inducer for the differentiation of MenSCs into oocyte-like cells. Keywords * Cell differentiation * Peripheral blood stem cells * Germ cells * Coculture techniques * Granulosa cells * Infertility, female