The crucial role of Activin A on the formation of primordial germ cell-like cells from skin-derived stem cells in vitro

Primordial germ cells (PGCs) are founder cells of the germ cell lineage, and can be differentiated from stem cells in an induced system <i>in vitro</i>. However, the induction conditions need to be optimized in order to improve the differentiation efficiency. Activin A (ActA) is a member of the TGF-β super family and plays an important role in oogenesis and folliculogenesis. In the present study, we found that ActA promoted PGC-like cells (PGCLCs) formation from mouse skin-derived stem cells (SDSCs) in both embryoid body-like structure (EBLS) differentiation and the co-culture stage in a dose dependent manner. ActA treatment (100 ng/ml) during EBLS differentiation stage and further co-cultured for 6 days without ActA significantly increased PGCLCs from 53.2% to 82.8%, and as well as EBLS differentiation without ActA followed by co-cultured with 100 ng/ml ActA for 4 to 12 days with the percentage of PGCLCs increasing markedly <i>in vitro</i>. Moreover, mice treated with ActA at 100 ng/kg body weight from embryonic day (E) 5.5–12.5 led to more PGCs formation. However, the stimulating effects of ActA were interrupted by <i>Smad3</i> RNAi, and in an <i>in vitro</i> cultured Smad3^−/− mouse skin cells scenario. SMAD3 is thus likely a key effecter molecule in the ActA signaling pathway. In addition, we found that the expression of some epiblast cell markers, <i>Fgf5, Dnmt3a, Dnmt3b</i> and <i>Wnt3</i>, was increased in EBLSs cultured for 4 days or PGCLCs co-cultured for 12 days with ActA treatment. Interestingly, at 16 days of differentiation, the percentage of PGCLCs was decreased in the presence of ActA, but the expression of meiosis-relative genes, such as <i>Stra8, Dmc1, Sycp3</i> and <i>Sycp1</i>, was increased. In conclusion, our data here demonstrated that ActA can promote PGCLC formation from SDSCs <i>in vitro</i>, at early stages of differentiation, and affect meiotic initiation of PGCLCs in later stages