Urethral reconstruction with autologous urine-derived stem cells seeded in three-dimensional porous small intestinal submucosa in a rabbit model

Abstract

BackgroundUrethral reconstruction is one of the great surgical challenges for urologists. A cell-based tissue-engineered urethra may be an alternative for patients who have complicated long strictures and need urethral reconstruction. Here, we demonstrated the feasibility of using autologous urine-derived stem cells (USCs) seeded on small intestinal submucosa (SIS) to repair a urethral defect in a rabbit model.MethodsAutologous USCs were obtained and characterized, and their capacity to differentiate into urothelial cells (UCs) and smooth muscle cells (SMCs) was tested. Then, USCs were labeled with PKH67, seeded on SIS, and transplanted to repair a urethral defect. The urethral defect model was surgically established in New Zealand white male rabbits. A ventral urethral gap was created, and the urethral mucosa was completely removed, with a mean rabbit penile urethra length of 2 cm. The urethral mucosal defect was repaired with a SIS scaffold (control group: SIS with no USCs; experimental group: autologous USC-seeded SIS; n = 12 for each group). A series of tests, including a retrograde urethrogram, histological analysis, and immunofluorescence, was undertaken 2, 3, 4, and 12 weeks after the operation to evaluate the effect of the autologous USCs on urethral reconstruction.ResultsAutologous USCs could be easily collected and induced to differentiate into UCs and SMCs. In addition, the urethral caliber, speed of urothelial regeneration, content of smooth muscle, and vessel density were significantly improved in the group with autologous USC-seeded SIS. Moreover, inflammatory cell infiltration and fibrosis were found in the control group with only SIS, but not in the experimental autologous USC-seeded SIS group. Furthermore, immunofluorescence staining demonstrated that the transplanted USCs differentiated into UCs and SMCs in vivo.ConclusionsAutologous USCs can be used as an alternative cell source for cell-based tissue engineering for urethral reconstruction.

Reno-protection of Urine-derived Stem Cells in A Chronic Kidney Disease Rat Model Induced by Renal Ischemia and Nephrotoxicity

“…USC at passage 3-5 were strong positive for glomerular parietal epithelial cells (CD24, CD29 and CD44), and also classical cell surface markers for MSC (CD73, CD90, CD105, CD146 and SSEA-4), but lack of the expression of hematopoietic markers (CD 31, CD34 and CD45). These results were consistent with our previous reports (Supplementary Figure 1) [12][13][14][15][16][17][18][19][20] . In addition, USC differentiated into osteogenic and adipogenic cell lineages, respectively (Figure 2).…”

“…Collection of human urine samples for USC isolation in this study was approved by the Wake Forest University Health Sciences Institutional Review Board. USC culture and characterization have been described previously [12][13][14][15][16][17][18][19][20] . In brief, USC were collected from healthy adult males (n=6) and expanded in culture media comprised of keratinocyte serum-free medium (Gibco, Gaithersburg, MD) and Dulbecco's Modified Eagle's medium (DMEM, high glucose, Gibco, Gaithersburg, MD) with 5% fetal bovine serum (FBS) (Gibco, Gaithersburg, MD) containing supplements 12 , see Table 1.…”

“…Recently, we demonstrated that stem cells exist in human urine and possess the potential for clinical applications [12][13][14][15][16][17][18][19][20] . Urine derived stem cells (USC) are thought to originate from the parietal cells in renal glomeruli 14 and have the potential for tissue regenerative effects, including robust proliferative potential, multi-potential differentiation, and the ability to exert regenerative effects via paracrine factors 14,21,22 .…”

Urine as a Main Effector in Urological Tissue Engineering—A Double-Edged Sword

“…[40] On the other hand, autologous rabbit UDSCs harvested via bladder irrigation were labeled with PKH67 and seeded into SIS by Wu et al [40] Cell-loaded scaffolds implanted into rabbits to repair ventral urethral defect showed differentiation of the cells and regeneration of the urethral defect. [68] (Figure 2 Yang et al also seeded UDSCs with dynamic culture on bladder submucosa, which significantly promoted cell-matrix penetration in vitro, as well as cell growth in vivo [69]. Bodin et al seeded UDSCs on microporous bacterial cellulose (BC), obtaining layered urothelial cells and SMCs with excellent cell-matrix infiltration [55].…”

Current Status of Tissue Engineering in the Management of Severe Hypospadias

“…For example, urothelial cells have been obtained from bladder washings, a method that has shown to be safe and highly reproducible in adults and children ( 45 , 46 ). Urine-derived stem cells have also shown the ability to expand and to differentiate into both urothelial and smooth muscle phenotypes ( 47 , 48 ). Cell proliferation is markedly influenced by the mechanical properties of the polymer scaffolds, and coculture of several cell types was shown to be superior to culture of individual cell types ( 38 ), perhaps because the former represents a more physiological condition, involving paracrine signaling among different cell types.…”