Thy-1+ cells isolated from adult human testicular tissues express human embryonic stem cell genes OCT3/4 and NANOG and may include spermatogonial stem cells.

Abstract

Spermatogonial stem cells (SSCs) are self-renewing cells whose progeny are committed to differentiate into spermatozoa; this is a life-long process in male mammals. There are several methods for obtaining enriched populations of mouse SSCs, and immunological separation using surface antigens is a commonly used technique. The study of human SSCs is much less advanced. We used biopsied human testicular tissues [obstructive azoospermia patients (n = 5) and patients who underwent a testis biopsy as part of an evaluation for infertility (n = 7)] to obtain Thy-1+ cells. Thy-1-a glycosyl phosphatidylinositol-anchored surface antigen-is a marker uniquely expressed on SSCs that is used to isolate SSC-enriched cell populations in mice. The Thy-1+ cells from human testicular tissues were cultured in a basic system consisting of serum-free medium and mitotically inactivated STO (SIM mouse embryo-derived thioguanine- and ouabain-resistant) cell feeders with added growth factors: glial cell line-derived neurotrophic factor (GDNF), basic fibroblast growth factor (bFGF), and GDNF-family receptor α1 (GFRα-1). The Thy-1+ cells were maintained in vitro using this system for 1 week. The Thy-1+ cells expressed OCT3/4 and alkaline phosphatase, like mouse SSCs. They also expressed NANOG. Thy-1+ cells injected into nude mice did not cause tumor formation over a period of at least 6 months. These results support the possibility that the Thy-1+ cell population included human SSCs, and that Thy-1 may be a marker for human SSCs.