title: The molecular signature of spermatogonial stem/progenitor cells in the 6-day-old mouse testis.
abstract: To characterize the molecular phenotype of spermatogonial stem cells (SSCs), we examined genes that are differentially expressed in the stem/progenitor spermatogonia compared to nonstem spermatogonia. We isolated type A spermatogonia (stem and nonstem type A) from 6-day-old mice using sedimentation velocity at unit gravity and further selected the stem/progenitor cell subpopulation by magnetic activated cell sorting with an antibody to GDNF-receptor-alpha-1 (GFRA1). It has been previously shown that GFRA1 is expressed in SSCs and is required for their stemness. The purity of the isolated cells was approximately 95% to 99% as indicated by immunocytochemistry using anti-GFRA1. Comparison of GFRA1-positive and GFRA1-negative spermatogonia by microarray analysis revealed 99 known genes and 12 uncharacterized transcripts that are overexpressed in the former cell population with a >2-fold change. Interestingly, the highest level of overexpression was observed for Csf1r, encoding the receptor for macrophage colony-stimulating factor (M-CSF, official symbol CSF1), which has a well-established role in the regulation of myeloid progenitor cells. Analysis of our microarray data with a bioinformatics software program (Ingenuity Systems) revealed the potential role of various signaling pathways in stem/progenitor spermatogonia and suggested a common pathway for GFRA1 and CSF1R that may lead to their proliferation. Further investigation to test this hypothesis has shown that CSF1 promotes cell proliferation in primary cultures of the isolated type A spermatogonia and in the spermatogonial-derived stem cell line C18-4. Semiquantitative RT-PCR and immunohistochemistry confirmed the previously mentioned microarray data. Collectively, this study provides novel molecular signatures for stem/progenitor spermatogonia and demonstrates a role for CSF1/CSF1R signaling in regulating their proliferation.
authors.affiliation: Department of Biochemistry and Molecular and Cellular Biology, Georgetown University Medical Center, Washington, District of Columbia 20057, USA.
authors.forename: Maria
authors.initials: M
authors.lastname: Kokkinaki
authors.forename: Tin-Lap
authors.initials: TL
authors.lastname: Lee
authors.forename: Zuping
authors.initials: Z
authors.lastname: He
authors.forename: Jiji
authors.initials: J
authors.lastname: Jiang
authors.forename: Nady
authors.initials: N
authors.lastname: Golestaneh
authors.forename: Marie-Claude
authors.initials: MC
authors.lastname: Hofmann
authors.forename: Wai-Yee
authors.initials: WY
authors.lastname: Chan
authors.forename: Martin
authors.initials: M
authors.lastname: Dym
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D049109:Cell Proliferation; D002478:Cells, Cultured; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D053263:Gene Regulatory Networks; D051097:Glial Cell Line-Derived Neurotrophic Factor Receptors / Q000378:metabolism; D016173:Macrophage Colony-Stimulating Factor / Q000235:genetics / Q000378:metabolism; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008954:Models, Biological; D020411:Oligonucleotide Array Sequence Analysis; D016186:Receptor, Macrophage Colony-Stimulating Factor / Q000235:genetics / Q000378:metabolism; D013091:Spermatogenesis / Q000235:genetics; D013093:Spermatogonia / Q000378:metabolism* / Q000502:physiology; D013234:Stem Cells / Q000378:metabolism* / Q000502:physiology; D013737:Testis / Q000254:growth & development / Q000378:metabolism*
chemical_list: C494988:Gfra1 protein, mouse; D051097:Glial Cell Line-Derived Neurotrophic Factor Receptors; D016173:Macrophage Colony-Stimulating Factor; D016186:Receptor, Macrophage Colony-Stimulating Factor