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BOR - Papers in Press, published online ahead of print July 13, 2005.
Biol Reprod 2005, 10.1095/biolreprod.105.043810
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BIOLOGY OF REPRODUCTION 73, 1011–1016 (2005)
DOI: 10.1095/biolreprod.105.043810
© 2005 by the Society for the Study of Reproduction, Inc.

GDNF Family Receptor alpha1 Phenotype of Spermatogonial Stem Cells in Immature Mouse Testes1

Anyanee Buageaw 3,4,5  , Meena Sukhwani 6, Ahmi Ben-Yehudah 6, Jens Ehmcke 5, Vanesa Y. Rawe 6, Chumpol Pholpramool 3,4 , Kyle E. Orwig 6, and Stefan Schlatt 2, 5

Department of Physiology,3 Faculty of Science, Mahidol University, Bangkok 10400, Thailand Institute of Science and Technology for Research and Development,4 Mahidol University, Salaya Campus, Nakorn Pathom 73170, Thailand University of Pittsburgh School of Medicine,5 Center for Research in Reproductive Physiology, Department of Cell Biology and Physiology, Pittsburgh, Pennsylvania 15261 University of Pittsburgh School of Medicine,6 Departments of Obstetrics/Gynecology & Reproductive Sciences and Molecular Genetics & Biochemistry, Pittsburgh Development Center of Magee-Womens Research Institute, Pittsburgh, Pennsylvania 15213

Spermatogonial stem cells (SSCs) are essential for spermatogenesis, and these adult tissue stem cells balance self-renewal and differentiation to meet the biological demand of the testis. The developmental dynamics of SSCs are controlled, in part, by factors in the stem cell niche, which is located on the basement membrane of seminiferous tubules situated among Sertoli cells. Sertoli cells produce glial cell line-derived neurotrophic factor (GDNF), and disruption of GDNF expression results in spermatogenic defects and infertility. The GDNF signals through a receptor complex that includes GDNF family receptor {alpha}1 (GFRA1), which is thought to be expressed by SSCs. However, expression of GFRA1 on SSCs has not been confirmed by in vivo functional assay, which is the only method that allows definitive identification of SSCs. Therefore, we fractionated mouse pup testis cells based on GFRA1 expression using magnetic activated cell sorting. The sorted and depleted fractions of GFRA1 were characterized for germ cell markers by immunocytochemistry and for stem cell activity by germ cell transplantation. The GFRA1-positive cell fraction coeluted with other markers of SSCs, including ITGA6 and CD9, and was significantly depleted of KIT-positive cells. The transplantation results confirmed that a subpopulation of SSCs expresses GFRA1, but also that the stem cell pool is heterogeneous with respect to the level of GFRA1 expression. Interestingly, POU5F1-positive cells were enriched nearly 15-fold in the GFRA1-selected fraction, possibly suggesting heterogeneity of developmental potential within the stem cell pool.

gamete biology, spermatogenesis, testis

1 Supported by a fellowship from the Commission on Higher Education Staff Development Project for the joint Ph.D. program in physiology at Mahidol University, Bangkok, Thailand (to A.B.). Startup funds from the University of Pittsburgh School of Medicine, a grant from the German-Israeli Foundation (GIF I-760-205.2/2002), and an NIA/NIH grant were available to S.S. (1R21 AG024914-01). K.E.O. was supported by startup funds from the Magee-Womens Health Corporation, NCRR/NIH (1R01 RR018500-02), NIA/NIH (1R01 AG024992-01), and the Pennsylvania Department of Health formula funds. V.Y.R. was supported by the Americas Fellowship RSANET, NICHD, NIH.

2 Correspondence: Stefan Schlatt, University of Pittsburgh School of Medicine, Department of Cell Biology and Physiology, W952 Biomedical Science Tower, 3500 Terrace Street, Pittsburgh, PA 15261. FAX: 412 648 8315; schlatt{at}pitt.edu




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