Although previous studies have shown that stem cells can be differentiated into Leydig cells by gene transfection, a simple, safe and effective induction method has not yet been reported. Therefore, the present study investigated novel methods for the induction of human umbilical cord mesenchymal stem cell (HUMSC) differentiation into Leydig-like, steroidogenic cells. HUMSCs were acquired using the tissue block culture attachment method, and the expression of MSC surface markers was evaluated by flow cytometry. Leydig cells were obtained by enzymatic digestion and identified by lineage-specific markers via immunofluorescence. Third-passage HUMSCs were cultured with differentiation-inducing medium (DIM) or Leydig cell-conditioned medium (LC-CM), and HUMSCs before induction were used as the control group. Following the induction of HUMSCs, Leydig cell lineage-specific markers (CYP11A1, CYP17A1 and 3β-HSD) were positively identified using immunofluorescence analysis. Additionally, reverse transcription-quantitative polymerase chain reaction and western blot analysis were performed to evaluate the expression levels of these genes and enzymes. In contrast, the control group cells did not show the characteristics of Leydig cells. Collectively, these results indicate that, under in vitro conditions, LC-CM can achieve a comparable effect to that of DIM on inducing HUMSCs differentiation into steroidogenic cells.
Keywords: Leydig cells; steroidogenesis; umbilical cord mesenchymal stem cells.