OBJECTIVE: To investigate the microscopic and molecular structure of the urethral plate in hypospadias patients compared to normal urethral tissue, focusing on epithelial characteristics, cytokeratin expression, vascular patterns, and extracellular matrix components, including elastin fibers. METHODS: We analyzed 8 full-thickness mid urethral plate samples from pediatric hypospadias patients using immunohistochemistry to evaluate epithelial structure, cytokeratin expression (PanCK, CK7, CK10, CK13, CK14), vascular patterns (von Willebrand Factor, CD31), and extracellular matrix components (elastin). Control tissues included fetal urethras and penile urethras from gender confirmation surgery. RESULTS: The hypospadias urethral plate exhibited a multi-layered epithelium, contrasting with the single-layered normal urethral epithelium. Cytokeratin expression differed significantly: CK7 was predominantly negative in hypospadias samples but strongly positive in controls
CK10 showed variable expression
CK13 and CK14 were consistently positive in hypospadias samples, with CK14 exhibiting strong expression in the basal layer. Vascular analysis revealed fewer sub-epithelial capillaries and smaller vascular spaces in hypospadias tissue. Elastin expression was positive in hypospadias samples, while fetal urethra showed lower expression and adult urethra displayed distinct elastin fiber bundles. Minimal smooth muscle tissue was observed in hypospadias and fetal samples compared to adult controls. CONCLUSION: The urethral plate in hypospadias patients demonstrates significant structural and molecular differences from normal urethral tissue, suggesting a developmental delay or arrest. These differences may contribute to surgical challenges and post-operative complications. Understanding these unique tissue characteristics could inform the development of novel surgical techniques or tissue engineering approaches to improve hypospadias repair outcomes.