The Interface Between Generating Renal Tubules and a Polyester Fleece in Comparison to the Interstitium of the Developing Kidney

Abstract

Abstract-An increasing number of investigations is dealing with the repair of acute and chronic renal failure by the application of stem/progenitor cells. However, accurate data concerning the cell biological mechanisms controlling the process of regeneration are scarce. For that reason new implantation techniques, advanced biomaterials and morphogens supporting regeneration of renal parenchyma are under research. Special focus is directed to structural and functional features of the interface between generating tubules and the surrounding interstitial space. The aim of the present experiments was to investigate structural features of the interstitium during generation of tubules. Stem/ progenitor cells were isolated from neonatal rabbit kidney and mounted between layers of a polyester fleece to create an artificial interstitium. Perfusion culture was performed for 13 days in chemically defined Iscove's Modified Dulbecco's Medium containing aldosterone (1 9 10 À7 M) as tubulogenic factor. Recordings of the artificial interstitium in comparison to the developing kidney were performed by morphometric analysis, scanning and transmission electron microscopy. The degree of differentiation was registered by immunohistochemistry. The data reveal that generated tubules are embedded in a complex network of fibers consisting of newly synthesized extracellular matrix proteins. Morphometric analysis further shows that the majority of tubules within the artificial interstitium develops in a surprisingly close distance between 5 and 25 lm to each other. The abundance of synthesized extracellular matrix acts obviously as a spacer keeping generated tubules in distance. For comparison, the same principle of construction is found in the developing parenchyma of the neonatal kidney. Most astonishingly, scanning electron microscopy reveals that the composition of interstitial matrix is not homogeneous but differs along a cortico-medullary axis of proceeding tubule development.Keywords-Tissue engineering, Perfusion culture, Kidney, Tubule, Artificial interstitium, Collagen type III. INTRODUCTIONRecovery from renal failure requires the replacement of injured tissue with new cells that restore epithelial integrity and functionality within tubules. An increasing number of papers is therefore dealing with strategies for repair of parenchyma by the help of stem/progenitor cells. 5,12 However, recent data show that an effective therapy is still far away from a widespread clinic application. Unsolved issues in renal tissue engineering are the concentration of stem/progenitor cells at the site of damage, their integration in a diseased environment, the process of differentiation into nephron-specific cells, and the spatial development of tubules within the kidney. 32 Part of actual research is focusing on cell biological mechanisms involved in the formation of tubules during regeneration. 4 Due to the spatial microarchitecture of the kidney experiments are frequently performed applying three-dimensional culture experiments in combination wi...

Illustration of extensive extracellular matrix at the epithelial-mesenchymal interface within the renal stem/progenitor cell niche

“…In contrast, previous and present experiments demonstrate that after conventional fixation by GA an astonishingly wide interstitial space separates epithelial and mesenchymal stem/progenitor cells (Figures 1, 2; asterisk) [28,31,43,53]. Further it was shown that numerous cellular protrusions from mesenchymal stem/progenitor cells are lining through the interstitial space to contact the lamina fibroreticularis at the tip of a CD ampulla (Figures 2a, 3a, 4a, 5a; arrow).…”

Polyester Fleeces used as an Artificial Interstitium Influence the Spatial Growth of Regenerating Tubules

“…These findings are in conformity with previous studies [41]. As yet the nature of these particles remains to be elaborated.…”

“…In the present study the processes do not seem to end but rather are interwoven with the lamina fibroreticularis of the generated tubules. It has been shown by transmission electron microscopy that the lamina fibroreticularis acts as connecting element to the interstitial space [41]. These adhesions and connections to the basal lamina of the generated tubules suggest that cross-talk between epithelial cell and interstitial, "fibroblast-like" cell might be mediated by mechanical forces [53,54].…”

Bridging the gap between traditional cell cultures and bioreactors applied in regenerative medicine: practical experiences with the MINUSHEET perfusion culture system

“…All up to date performed experiments yet point out that development of tubules is triggered by interactions between their basal lamina, newly synthesized fibers of the extracellular matrix and fibers of the polyester fleece (Minuth et al 2010a, b, c, d;Miess et al 2010;Glashauser et al 2011).…”