Daidzein activates Akt pathway to promote the proliferation of female germline stem cells through upregulating Clec11a

Female germline stem cells (FGSCs) have been successfully isolated from postnatal mammalian and human ovarian tissues. However, the effects and mechanisms of action of natural small-molecule compounds on FGSCs are largely unknown. Here, we determined the effects of daidzein, a phytoestrogen present in soybean , on cell viability and proliferation using Cell Counting Kit-8 (CCK8) and 5-ethynyl-2-deoxyuridine (EdU). We found that daidzein promoted the viability and proliferation of FGSCs in a concentration-dependent manner. To elucidate the mechanism underlying this, we performed RNA-Seq in daidzein-treated FGSCs and controls. The results showed that there were 153 upregulated and 156 downregulated genes after daidzein treatment. We confirmed the expression of some genes related to cell proliferation in the sequencing results by RT-PCR, such as Type C lectin domain family 11 member a (Clec11a), Mucin1 (Muc1), Glutathione peroxidase 3 (Gpx3), and Tet methylcytosine dioxygenase 1(Tet1). The high expression of Clec11a at the protein level after daidzein treatment was also confirmed by western blotting. Furthermore, recombinant mouse Clec11a (rmClec11a) protein was shown to promote the viability and proliferation of FGSCs. However, knockdown of Clec11a inhibited the viability and proliferation of FGSCs, which could not be rescued by the administration of daidzein. These results indicate that daidzein promoted the viability and proliferation of FGSCs through Clec11a. In addition, both daidzein and rmClec11a activated the Akt signaling pathway in FGSCs. However, Clec11a knockdown inhibited this pathway, which could not be rescued by daidzein administration. Taken together, our findings revealed that daidzein activates the Akt signaling pathway to promote cell viability and proliferation through upregulating Clec11a. This study should deepen our understanding of the developmental mechanism of FGSCs and female infertility.