Gene expression profiles in mouse urethral development.

OBJECTIVE To analyse the gene expression profiles of the mouse genital tubercle (GT) during urethral tube development at embryonic (E) days E14, E15, E16 and E17, as the aetiology of hypospadias, one of the most common congenital anomalies, remains unknown. MATERIALS AND METHODS During GT development the urethral folds fuse to form an epithelial seam; subsequently, the epithelial seam disappears, resulting in the normal tubular urethra. Abnormalities in urethral seam formation and remodelling might explain hypospadias, and elucidating the molecular developmental mechanisms underlying normal penile development might provide the basis for understanding hypospadias. Total RNA was isolated from the genital tubercle at embryonic days E14, E15, E16, and E17. Together with reference RNA, sample RNA was labelled with Cy-3 and Cy-5 respectively and hybridized to a 16 000-mouse gene array that included the Incyte GEM2.1 and NIA 7k sets. Candidate genes were analysed by immunohistochemistry and real-time polymerase chain reaction. RESULTS Using cDNA microarrays, we identified the up-regulation of genes involved in the transforming growth factor (TGF)-β and Wnt-Frizzled pathways, and of thrombospondin (TSP) 4, a member of a cell-migration molecule family, all candidates for involvement in urethral tube formation. Immunohistochemistry showed TGFβ1, TGFβ receptor III, and Frizzled1 were expressed exclusively in E14–E17 urethral epithelium. TSP4 was expressed in the mesenchymal basal layer underlying E17 GT skin epidermis. CONCLUSIONS Many signalling pathways are involved in late GT development, and cell migration molecules might have an important role in urethral tube formation.