Abstract
In mammals, the pre-Sertoli cell of the male genital ridge is the first cell type to display sex specific differentiation and differential gene expression. The genetic cascade driving the differentiation of pre-Sertoli cells and ultimately testis formation is beginning to be unravelled, but many questions remain. A better understanding of the transcriptome of pre-Sertoli cells immediately after sex determination is essential in order to further understand this differentiation process. A mouse model expressing Red Fluorescent Protein (RFP) under the control of a hybrid mouse/pig SRY promoter (HybSRYp-RFP) was used to purify cells from embryonic day 12.0 (e12.0) male genital ridges. To compare the transcriptomes of HybSRYp-RFP cell populations versus age matched whole female genital ridges, RNA was extracted and used to generate molecular probes that were hybridized onto Affymetrix Mouse Genome 430 2.0 micro-arrays. The expression of genes considered markers for pre-Sertoli cells, including Sox9, Mis, Dhh and Fgf9 were identified within the HybSRYp-RFP expressing cell population, while markers for germ cells (Oct4, SSEA-1) and endothelial cells (Ntrk3) were not identified. In contrast, markers for ovarian somatic cell expression, including Fst and Bmp2, were identified as overexpressed within the ovarian cell population. In a general fashion, genes identified as 2.5-fold over expressed in HybSRYp-RFP expressing cells coded notably for cell signalling and extra cellular proteins. The expression of Sox10, Stc2, Fgf18, Fgf13 and Wnt6 were further characterized via whole mount in situ hybridization (WISH) on male and female genital ridges between e11.5 and e14.5. Sox10, Fgf18, Fgf13 and Stc2 gene expression was detected within the male genital ridges while Wnt6 was found diffusely within both the male and female genital ridges. These data represent the earliest comprehensive microarray expression analysis of purified presumptive pre-Sertoli cells available to date.
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