Generation and ex vivo characterization of a full-thickness substitute of the human urethra by tissue engineering

Tissue engineering may offer efficient alternatives for the surgical repair of severe conditions affecting the human urethra. However, development of tubular full-thickness substitutes is challenging. In this work, we have generated and evaluated ex vivo a novel full-thickness human urethra substitute (FHUS) containing its three main layers: the urethral mucosa (UM), the spongy layer (SP), and the tunica albuginea (AL). Results first showed that the generation of a FHUS significantly improved the biomechanical properties of this artificial tissue as compared to the individual layers, although the resistance of the native urethra was not reached. At the structural level, we found that FHUS shared important histological similarities with the native urethra. Analysis of the individual layers showed that UM had a stratified epithelium that expressed several epithelial markers, including cytokeratins CK7 and CK14, uroplakin 1b, and the intercellular junction proteins desmoplakin, tight junction protein 1, and claudin. At the stromal level, UM tended to increase the presence of collagen fibers and versican with time. The SP layer displayed abundant CD31 and CD34-positive blood vessels, but small amounts of collagen and proteoglycans. The AL layer showed scattered smooth muscle cells expressing α-smooth muscle actin, smoothelin, and desmin cell markers, and contained low amounts of collagen and proteoglycans. Analysis of the basement membrane components collagen IV and laminin revealed their progressive development with time, especially collagen IV. These results confirm the possibility of developing a partially biomimetic full-thickness substitute of human urethra that might have potential clinical usefulness for the clinical repair of severe urethral lesions.