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. 2014 Mar 11;9(3):e90088. doi: 10.1371/journal.pone.0090088

Figure 4. Spermatogonial stem cells were identified in co-cultures by immunocytochemistry and transplantation assay, and these cells were supported by endogenous GDNF secretion.

Figure 4

A) Image of 1-week-old co-culture showing proliferating, phosphorylated Histone-3 (PH3) positive cells expressing spermatogonial stem cell marker PLZF. B) A high proportion of LIN28 positive cells also expressed PH3 after 10 days of culture. DAPI stains the nuclei of cells. C) Section of a wild-type SSC-depleted seminiferous tubule that has been colonized by transplanted SSCs. Samples were prepared five weeks after transplantation of GFP-expressing SSCs and show that spermatogenic differentiation of the transplanted SSCs had advanced to meiotic prophase. Arrows indicate pachytene spermatocytes. D) GDNF concentration in co-cultures. Analysis of GDNF levels in the culture medium showed that approximately 5–15 ng/ml were present in 1–5-week co-cultures.