Comparative Analysis for In Vitro Differentiation Potential of Induced Pluripotent Stem Cells, Embryonic Stem Cells, and Multipotent Spermatogonial Stem Cells into Germ-lineage Cells

In the present study, embryoid bodies (EBs) obtained from induced pluripotent stem cells (iPSCs) were induced to differentiate into germ lineage cells by treatment with bone morphogenetic protein 4 (BMP4) and retinoic acid (RA). The results were compared to the results for embryonic stem cells (ESCs) and multipotent spermatogonial stem cells (mSSCs) and quantified using immunocytochemical analysis of germ cell-specific markers (integrin-α6, GFR-α1, CD90/ Thy1), fluorescence activating cell sorting (FACS), and real time-RT-PCR. We show that the highest levels of germ cell marker-expressing cells were obtained from groups treated with 10 ng/㎖ BMP4 or 0.01 ㎛ RA. In the BMP4-treated group, GFR-α1 and CD90/Thy-1 were highly expressed in the EBs of iPSCs and ESCs compared to EBs of mSSCs. The expression of Nanog was much lower in iPSCs compared to ESCs and mSSCs. In the RA treated group, the level of GFR-α1 and CD90/Thy-1 expression in the EBs of mSSCs was much higher than the levels found in the EBs of iPSCs and similar to the levels found in the EBs of ESCs. FACS analysis using integrin-α6, GFR-α1, CD90/Thy1 and immunocytochemistry using GFR-α1 antibody showed similar gene expression results. Therefore our results show that iPSC has the potential to differentiate into germ cells and suggest that a protocol optimizing germ cell induction from iPSC should be developed because of their potential usefulness in clinical applications requiring patient-specific cells.