Identifizierung und funktionelle Untersuchung von humanen adulten spermatogonalen und germalen Stammzellen

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Our in vitro and in vivo experiments have shown that we are able to generate cultures of human adult pluripotent germline-derived stem cells (haGSCs) from enriched spermatogonia (Conrad et al., 2008). These haGSCs possess expression profiles which resemble those of pluripotent embryonic stem cells (hES). haGSCs can be differentiated in vitro into cell of all three germ layers, and they also form teratomas. These are the first pluripotent stem cells from an adult source which are suitable for use in humans while at the same time being ethically non-controversial. At present, however, only a very few surface markers are available for the prospective isolation of human and/or murine adult spermatogonial stem cells (hSSCs/mSSCs) and haGSCs, including c-kit, Thy-1, α6-integrin, β1-integrin, CD9 and GFRa1, most of which have only been tested in the mouse. In addition, these markers are not highly selective for spermatogonia. Currently employed methods cause substantial loss of cells and enrich not only hSSCs, but other cells involved in spermatogenesis as well. In the proposed project, we plan to develop new methods for the isolation and characterization of highly purified hSSCs and haGSCs, taking advantage of our large library of stem cell-specific antibodies which are promising candidates for a more efficient isolation of these cells. Finally, we will use single cell sorting to establish clones with ES cell-like growth behavior. A central goal is to optimize the culture and selection methods for the generation of hSSCs to obtain sufficient cell numbers from small biopsies for therapeutic applications.