Regenerating Tubules for Kidney Repair

Abstract

Stem/progenitor cells are in the focus of regenerative medicine for a future therapy of acute and chronic renal failure. However, broad knowledge about parenchymal regeneration in kidney is lacking. For that reason developmental pathways leading from stem/progenitor cells to newly formed tubules have to be investigated. A new technique promotes renal stem/progenitor cells to form numerous tubules between layers of polyester fleeces. This artificial interstitium replaces coating by extracellular matrix proteins, supports spatial extension of renal tubules, and can be used with chemically defined Iscove’s modified Dulbecco’s medium (IMDM) during a long-term culture period of 13 days. The development of tubules is stimulated by aldosterone and depends on the applied hormone concentration. The tubulogenic effect cannot be mimicked by precursors of the aldosterone synthesis pathway or by other steroid hormones. Antagonists such as spironolactone or canrenoate prevent the development of tubules, which indicates that the mineralocorticoid receptor (MR) is involved. Administration of geldanamycin, radicicol, quercetin, or KNK 437 in combination with aldosterone blocks development of tubules by disturbing the contact between MR and heat-shock proteins. Transmission electron microscopy (TEM) further demonstrates that generated tubules exhibit a junctional complex between the apical and the lateral plasma membrane. At the basal aspect a continuously developed basal lamina is present. Immuno-label for Troma I (cytokeratin Endo-A) shows isoprismatic cells, while label for laminin γ1, occludin, and Na/K-ATPase α5 confirms typical features of a polarized epithelium. Finally, the introduced system makes it possible to pile and pave renal stem/progenitor cells, so that the spatial development of tubules can be systematically investigated.