Establishment of cell lines with rat spermatogonial stem cell characteristics

Spermatogonial cell lines were established by transfecting a mixed population of purified rat A(s) (stem cells), A(pr) and A(al) spermatogonia with SV40 large T antigen. Two cell lines were characterized and found to express Hsp90alpha and oct-4, specific markers for germ cells and A spermatogonia, respectively. Expression of c-kit, normally expressed in A spermatogonia from late A(al) spermatogonia onwards, could not be detected in either cell line. Furthermore, no expression of vimentin (Sertoli cell marker) and alpha-smooth muscle actin (peritubular cell marker) could be found. Upon transplantation of these cell lines into recipient mice, the cells were found to be able to migrate to the basement membrane and to colonize seminiferous tubules. Taken together, we conclude that our cell lines have spermatogonial stem cell characteristics. These first spermatogonial cell lines with stem cell characteristics can now be used to study spermatogonial gene expression in comparison with more advanced germ cells.