Development of Tissue-Engineered Tubular Substitutions for Urinary Diversion in a Preclinical Rabbit Model
Authors: Wang Qianliang , Liu Qingling 
Published: June 2024

Abstract

Objectives
At present, the gold standard of bladder substitution is using the intestine as a neobladder for urinary diversion. However, there are many kinds of complications with this substitution, such as hematuria, stone formation, chronic urinary tract infection, tumor recurrence and secretion of mucus, which seriously affect the quality of life of the patients. for a long time, the medical community has not yet found a suitable way to replace this kind of urinary diversion.The rise of tissue engineering provides the possibility to solve this kind of problem. As an important research field of regenerative medicine, tissue engineering has made great progress in nearly half a century. In recent years, with the deepening of research, while the study of tissue-engineered bladder has made significant progress, but nowadays, tissue engineering cannot build a bladder with the complete structure and function. Because the gut as neobladder is only served as the urine drainage and does not involve complex neuromuscular regulation to modulate the contraction and relaxation of bladder, the current tissue engineering technology can construct a urine diversion device to solve the radical cystectomy patients' problems of urine drainage. In this experiment, as a starting point, using the idea of tissue engineering, we try to build a new type of urinary diversion device which was implanted in rabbits for urine diversion, and explore the feasibility and effectiveness of the device for urine drainage.
Methods
ADSCs obtained from the adipose tissue of New Zealand rabbits by the method of collagenase digestion, and ADSCs were cultured, amplified and identified by flow cytometry in vitro. At the same time, SMCs harvested from bladder tissue patch were cultured, amplified and identified by immunofluorescence. After obtaining the fish swim bladder, we prepared decellularized fish swim bladder(DFSB) by removing cells, and the effect of cell-free and property of Cell-free swim bladder were evaluated by HE, Masson, Sirius red staining and scanning electron microscope. ADSCs and SMCs were seeded onto the two sides of the DFSB respectively, and with 7 days' culture, the growth of cells on DFSB was observed by HE staining and scanning electron microscopy. Also, the cell compatibility test was performed next. The cell-seeded matrix was used to build tissue-engineered tubular substitutions, which were then implanted and wrapped into the omentum in vivo for two weeks to promote angiogenesis. In the experimental group, the bladder of 24 rabbits was totally resected, and the above tissue-engineered tubular substitutions were used for urinary diversion. In the control group, tissue-engineered tubular substitutions with unseeded cells were implanted into the omentum and were used as urinary diversion on another Six rabbits with the same process. The implants were harvested, and histological examination was conducted at 2, 4, 8, and 16 weeks after operation. Intravenous urography assessment was performed at 16 weeks postoperatively.
Results
We successfully acquired ADSCs and SMCs confirmed by flow cytometry and immunofluorescence respectively, and in this study. After cell-free processing, HE, Masson, Sirius red staining and scanning electron microscope displayed a good effect of cell-free and there were rich collagen fiber and high porosity and permeability within DFSB. Cells grow well after DFSB loaded with ADSCs and SMCs, and test showed that DFSB had good cell compatibility. After incubation of omentum, the status of vascular regeneration was obvious. After urinary diversion, histological and immunohistochemical analysis showed that the lumen of TETSs generated well differentiated and mature urothelial cells. Intravenous urography revealed that the TETSs in experimental group had a clear image, and the contrast agent flows unimpeded. There was no obvious stenosis, obstruction and leakage phenomenon. In the control group, all the rabbits were died within 3 weeks,The urinary diversion device appeared leakage of urine, contracture, obvious inflammatory reaction and scar formation, and there were a lot of stones in the outflow tract.
Conclusions
In this study, the constructed new TETSs was close to the normal bladder tissue, which can prevent urinary extravasation, protect renal function, and void urine efficiently. This research provided a preliminary experimental basis for further research of tissue-engineered bladder of complete structure and function, and offered a theoretical possibility for the clinical application of this new TETSs.