Establishment of functional germ cell precursor cell lines

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The aim of this research is to derive cell lines from mouse primordial germ cells (PGCs) and spermatogonia and to differentiate them into functional gametes. For this purpose, Ifirst examined whether PGC-derived pluripotential EG cells could redifferentiate into PGCs when culturing them with somatic cells from embryonic gonads. After culture, I stained them with PGC-specific vasa antibody, and found that EG cells differentiated to vasa-positive cells. In addition, EGF could also induced vasa-positive cells from EG cells without gonadal somatic cells. I also attempted to establish spermatogonial cell lines form a transgenic mice overexpressing bcl-2 in spermatogonia. Dissociated transgenic testicular cells were cultured on a feeder layer of STO cells and I found that TRA98-positive germ cells determined by imnmunostaining could be detected up to 45 days after culture, while wild type testicular cells survived up to only 30 days in culture. In addition, I found that the transgenic germ cells differentiated intoTRA369-positive spermatocytes and/or spermatid after 10 days in culture.