Ultrastructural insights in the interface between generated renal tubules and a polyester interstitium

Abstract

In regenerative medicine, stem/progenitor cells are emerging as potential candidates for the treatment of renal failure. However, the mechanism of regeneration of renal tubules from stem/progenitor cells is not well-elucidated. In this study, a new method was developed for the generation of tubules replacing coating by extracellular matrix proteins. Renal stem/progenitor cells are mounted between layers of polyester fleece. This artificial interstitium supports spatial development of tubules within 13 days of perfusion culture in chemically defined Iscove's modified Dulbecco's medium (IMDM) containing aldosterone as the tubulogenic factor. Whole mount label by soybean agglutinin (SBA) showed that generated tubules exhibited a lumen and a continuously developed basal lamina. Immuno-labeling for cytokeratin Endo-A demonstrated the presence of isoprismatic epithelial cells, and laminin gamma1, occludin, and Na/K-ATPase alpha5 labeling revealed typical features of a polarized epithelium. To get first insight in the interface between tubules and polyester interstitium, transmission electron microscopy (TEM) was performed. The results showed that the generated tubules exhibited polar differentiation with a continuously developed basal lamina consisting of a lamina rara interna, lamina densa, and lamina rara externa. Collagen type III was found to be the linking molecule between the basal lamina and the surrounding polyester fibers by immuno labeling studies. Thus, the findings demonstrate that the spatial development involves the interface between the tubular basal lamina and the polyester interstitium of tubules and is not restricted to the epithelial portion.