Objective To observe whether umbilical cord mesenchymal stem cells (UCMSCs) can differentiate into the smooth muscle cells (SMCs) induced by bladder SMCs (BSMCs) conditioned medium so as to seek an alternative seed cells for the repair and reconstruction of the urology system. Methods UCMSCs and BSMCs were harvested from umbilical cord of full-term births and bladder tissues which were obtained from patients who underwent a radical cystectomy. BSMCs conditioned medium was prepared by mixing supernatant of BSMCs at passages 1-5 with complete medium at ratio of 1 ∶ 1. UCMSCs at passage 3 were cultured with BSMCs conditioned medium (induced group, group A) and complete medium (control group, group B), respectively; simple BSMCs served as positive control group (group C). The morphological changes of co-cultured UCMSCs were observed by inverted phase microscope, the expressions of α-smooth muscle actin (α-SMA), Calponin, and smooth muscle myosin heavy chain (SM-MHC) of UCMSCs were tested by immunofluorescence staining and Western blot at 7 and 14 days. Results The morphology of UCMSCs in group A started to change from a polygonal and short spindle shape to a large and spindle shape after co-culture, which was similar to BSMCs morphology; but the morphology of UCMSCs did not change obviously in group B. Immunofluorescence staining showed that the expressions of α-SMA, Calponin, and SM-MHC were positive in group C. At 7 days, the expression of α-SMA could be observed in groups A and B; at 14 days, the positive expression of α-SMA increased gradually in group A, but it did not increase in group B. At 7 days, a positive expression of Calponin could be observed in group A, and positive expression increased obviously at 14 days; the expression of Calponin could not be observed at 7 and 14 days in group B. However, the expression of SM-MHC could not be observed in groups A and B. The results of Western blot showed the expressions of α-SMA, Calponin, and SM-MHC protein were consistent with the results of immunofluorescence staining. Conclusion UCMSCs have the potential of differentiation into SMCs and may be a potential seed cells for bladder tissue engineering.
Objective To sum up the common mode in urinary diversion after radical cystectomy. Methods The recent original articles about the common mode in urinary diversion after radical cystectomy were extensively reviewed. Results Urinary diversion includes no continent ureterocutaneostomy, continent ureterocutaneostomy and orthotopic neobladder. Ileal conduit, an ideal procedure of urinary diversion, has been widely used in patients after radical cystectomy and it is uncertain whether the health related quality of life in patients undergoing orthotopic ileal neobladder is superior to those undergoing ileal conduit. A series of basic researches of tissue engineering show a wide prospect of clinical application in the future. Conclusion Intestinal segment will remain the main material for urinary diversion and bladder reconstruction in a long time. Tissue engineering materials may be ideal for the substitution of bladder, and tissue engineering becomes the ultimate approach to solve the problem of missing bladder.
Objective To evaluate tissue regeneration, body reaction, and biological safety of xenogeneous bladder acellular matrix (BAM) that can be used to repair rabbit bladder. Methods Porcine BAM was prepared through physical, chemical, and enzymatic methods, and the effects of acellularization and the structure were observed with HE staining and scanning electron microscope (SEM). Eighteen New Zealand white rabbits (weighing, 2.5-3.0 kg) undergoing partial cystectomy were randomly divided into 2 groups. After partial (about 30%) cystectomy, the porcine BAM was used to replace partial rabbit bladder in the experimental group (n=12), and the incision was directly sutured as control group (n=6). The survival condition of animals was observed after operation. At 15 days, 1, 2, 3, and 6 months after operation, the blood routine, renal function, and electrolyte were tested by collecting the blood samples. At 1, 2, 3, and 6 months after operation, maximum bladder capacity, bladder leak point pressure, and bladder compliance were measured through urodynamic studies. Then gross observation was performed for regeneration of bladder, and the specimens of the bladder were harvested for HE staining and immunohistochemical staining. The surrounding organs and local lymphoid tissues were harvested for gross observation and HE staining. Results Cell components were completely removed in the porcine BAM, showing three-dimensional porous structure under SEM. All the animals survived during the experiment. At 15 days after operation, white blood cell count increased, and then returned to normal level in 2 groups, showing no significant difference between 2 groups (P gt; 0.05). The tests of renal function and electrolyte suggested no significant difference between 2 groups (P gt; 0.05). The level of serum creatinine showed a tendency of increase, but it remained within normal range at 6 months after operation. The maximum bladder capacity and compliance in experimental group were significantly higher than those in control group at 3 and 6 months after operation (P lt; 0.05), but no significant difference in bladder leak point pressure at each time point between 2 groups (P gt; 0.05). The urothelial regeneration, smooth muscle regeneration, and blood vessel regeneration were seen by histological observation in 2 groups. In the 2 groups, chronic inflammatory cells infiltration could be observed at 1 month postoperatively, and then chronic inflammatory cells decreased significantly (P lt; 0.05), until complete disappearance. There was no significant difference in score of chronic inflammatory cell infiltration between 2 groups at 3 and 6 months after operation (P gt; 0.05). The α-smooth muscle actin expression was significantly increased with time passing in 2 groups (P lt; 0.05), and it was significantly higher in control group than in experimental group at each time point (P lt; 0.05). In addition, gross and HE staining observations showed no abnormalities in surrounding organs and local lymphoid tissues. Conclusion No immune rejection response occurs when porcine BAM is used for xenotransplantation. It is indicated that porcine BAM is relative safety for xenotransplantation.
Objective To study the microstructural change of detrusor muscle and neuromuscular junction (NMJ) after bladder functional reconstruction for atonic bladder caused by medullary cone injury and to discuss the feasibility of bladder functional reconstruction for improving the detrusor muscle degeneration. Methods A total of 104 adult female Sprague-Dawley rats (weighing, 200-250 g) were randomized divided into 3 groups: normal group (n=8), control group (n=48), and experimental group (n=48). No treatment was given in normal group; the medullary cone injury was established by sharp transection of spinal cord at L4, 5 levels in control group; and the anastomosis of bilateral L5 ventral root (VR)-S2 VR and L5 dorsal root (DR)-S2 DR was performed for bladder functional reconstruction after modeling of medullary cone injury in experimental group. After operation, the survival condition of rats was observed. At 3 days and 3 consecutive days before 1, 2, 3, 4, 5, and 6 months after operation, the residual urine volume was measured; at 1, 2, 3, 4, 5, and 6 months after operation, the detrusor muscle was harvested to measure the muscle fiber cross-sectional area by HE staining, to calculate the percentage of connective tissue by Masson trichrome staining, and to observe the ultrastructure of the detrusor muscle and the NMJ by transmission electron microscope (TEM). Results Eleven rats were supplemented because of death after operation. In control group, a significant increase of the residual urine volume was observed with the extension of time (P lt; 0.05); in experimental group, an increase was observed at the first 3 months after operation, and then gradually decreased, showing significant differences between the other time point (P lt; 0.05) except between at 3 days and at 5 months after operation (P gt; 0.05); there was significant difference between control and experimental groups at other each time point (P lt; 0.05) except at 3 days, 1 month, and 2 months (P gt; 0.05). HE staining and Masson trichrome staining indicated that the muscle fibers arranged in disorder with gradually aggravated atrophy and gradually increased connective tissue in control group, while the shape of the detrusor muscle recovered with no increased connective tissue at 4, 5, and 6 months after operation in experimental group; there was significant difference in cross-sectional area of detrusor muscle and percentage of connective tissue between normal group and experimental group, and between normal group and control group at each time point (P lt; 0.05). In control group, the cross-sectional area of detrusor muscle decreased and the percentage of connective tissue increased with the extension of time (P lt; 0.05). In experimental group, the cross-sectional area of detrusor muscle decreased at the first 3 months and then increased, and the percentage of connective tissue increased slowly with the extension of time. There was no significant difference of cross-sectional area of detrusor muscle at the first 3 months between control and experimental groups (P gt; 0.05), but the values in experimental group were significantly higher than those in control group at 4, 5, and 6 months after operation (P lt; 0.05). There were significant differences of the percentage of connective tissue between control and experimental groups at each time point (P lt; 0.05). In control group, the amount of synaptic vesicles decreased in the NMJ with time passing; vacuole like structure was observed in NMJ at 3 months; there was almost no nerve ending at 6 months. In experimental group, the amount of synaptic vesicles decreased at 1 and 3 months after operation, but obviously increased at 6 months. Conclusion The reconstruction of bladder function with L5 nerve roots above the paraplegic plane can effectively inhibit the degeneration of detrusor muscle and improve its microstructural changes after medullary cone injury.
Objective To review the study on adi pose derived stem cells (ADSCs) in the therapy of urological diseases. Methods The recent l iterature concerning ADSCs in bladder repair, urethral reconstruction, incontinence treatment, and erectile dysfunction treatment was reviewed. Results The appl ication of tissue engineering using ADSCs has made significant achievements in the treatment of urological diseases and in animal studies, and has been initially used in cl inicaland has achieved a good therapeutic effect. Conclusion Tissue engineering using ADSCs has good prospects in the study on urological diseases, and is expected to widely used in the treatment of urological diseases.
Objective It is a thorny problem to reconstruct long ureteral defect in urinary surgery. To investigate the feasibil ity of intestinal sero-muscular segment with autograft of bladder mucosa as a replacement material for reconstructionof long ureteral defect. Methods Twelve adult Beagle dogs (weighing 6.5-9.3 kg and being male or female) were randomlydivided into 3 groups, each group including 4 dogs. In group A, lower segment of ureter was reconstructed by autograft of bladder mucosa to the intestinal sero-muscular segment; furthermore, the proximal and distal reconstructed ureter were anastomosed to the bladder and the upper ureter, respectively. In group B, upper segment of ureter was reconstructed by the same method as that of group A, the proximal and distal reconstructed ureter anastomosised with pelvic and lower ureter, respectively. In group C, whole ureter was reconstructed by the same method as that of group A, the proximal and distal reconstructed ureter were anastomosised with pelvic and bladder, respectively. Blood urea nitrogen, Cr2+, K+, Na+, Cl-, Ca2+ and carbon dioxide combining power were detected before operation, the general state, drainage volume, heal ing of wound, and compl ications were observed after operation. At 6 weeks, the blood biochemical indexes and intravenous urography (IVU) were detected, and the gross and histological observations of ureter were done. Results In group B, urine leakeage and infection occurred in 1 dog 2 days after operation because ureter stent prolapsed; other dogs had no complications. There was no significant difference in the biochemical indexes between before operation and 6 weeks after operation. IVU showed: in group A, hydronepherosis and ureterectasia occurred on the operation side of 1 dog; in group B, anastomotic stricture between the reconstructed ureter and lower ureter and hydronepherosis occurred in 1 dog; and in other dogs of all groups, renal function was good and the reconstructed ureter had peristalsis function. The histopathological observation showed that the reconstructed ureter had similar structure to normal ureterat 6 weeks in 3 groups; the inflammatory cells infiltrating of the reconstructed ureter was observed in 1 dog of groups A and C, respectively. Conclusion Reconstruction of ureter by intestinal sero-muscular segment with autograft of bladder mucosa has similar structure and function to the normal ureter. The results might provide an experimental basis for cl inical use.
To introduce a micturition alert device dedicated to neurogenic bladders. Methods The design and mechanism of the micturition alert device were explained, the effectiveness was tested in a cranine experiment. Results The micturition alert device consisted of a permanent magnet sutured on the anterior bladder wall and a warning unit sutured on theinferior abdominal wall. The warning unit was assembled with a compass-l ike switch, a power supply, a buzzer and a power switch. Bladder volume determined the position of the magnet which determined the magnetic field at the point of the warning unit. The change of magnetic field was read by the warning unit. With increasing bladder volume from initial state to 200 mL in 8 dogs, the magnet moved cranially 32.8 mm averagely (from 31.3 mm to 34.1 mm) and the hand of warning unit turned 52° (from 47° to 57°). The value of the warning unit was correlated positively to the bladder volume (r =1.0, P lt; 0.01）. If the desired bladder volume was determined as 150 mL to activate the warning unit to alarm in advance, the fullness of bladder was 147.6 mL averagely from135 mL to 160 mL, with an error less than 15 mL (10%). Conclusion The micturition alert device including a warning unit and permanent magnet could monitor bladder volume continuously and alarm in time for the patients with loss of micturition desire. It is simple, easily-made, cheap and conveniently used. It is worth of further study.
【Abstract】 Objective To establ ish an artificial physiological reflex arc with reconstruction of the sensory and themotorial functions of atonic bladder simultaneously after the conus medullary injury in rats. Methods Twenty 3-month-oldmale SD rats, with the weight of 250 to 300 g, were included. The right side was the experimental side, while the left side served as a control. Intradural microanastomosis of the right L5 ventral root to S2 ventral root and L5 dorsal root to S2 dorsal root wasperformed to reconstruct the sensory and the motorial functions of atonic bladder. After axonal regeneration, the new motor-tomotor and sensory-to-sensory artificial bladder reflex pathway was establ ished. At 5 months postoperatively, the early function of the reflex arc was observed by electrophysiological examinations, and the bladder pressure was tested. Results Eighteen rats survived for 5 months after the operation. Single stimul i (3 mA, 0.3 ms) of the S2 dorsal root of the experimental side resulted in evoked potentials recorded from the right vesical plexus before and after the spinal cord was destroyed horizontally between L6 and S4 segmental levels. The ampl itudes of the evoked potentials were (0.10 ± 0.02) mV and (0.11 ± 0.03) mV, respectively, before and after paraplegia, and there was no statistically significant difference (P gt; 0.05). The figures of the evoked potentials were similar to those of the control side. Bladder contraction was initiated by trains of stimul i (3 mA, 20 Hz, 5 s) of the S2 dorsal root of the experimental side. The bladder pressures were (6.55 ± 1.33) cmH2O and (6.11 ± 2.01) cmH2O, respectively, and the ampl itudes of bladder smooth muscle complex action potential were (0.11 ± 0.02) mV and (0.11 ± 0.03) mV, respectively, beforeand after paraplegia. There was no significant difference (P gt; 0.05). These figures were similar to those of the control side before paraplegia. Before paraplegia, when the S2 dorsal root of the control side was stimulated, the ampl itude of the evoked potential was (0.14 ± 0.02) mV, the bladder pressures was (10.77 ± 1.78) cmH2O and the ampl itude of bladder smooth muscle complex action potential was (0.17 ± 0.02) mV. There was statistically significant difference bewteen the experimental side and the control side (P lt; 0.01). All the results of electrophysiological examinations and bladder pressure were negative when the left S2 dorsal root was stimulated after paraplegia. Conclusion Suprasacral nerve motor-to-motor and sensory-to-sensory transfers after the spinal cord injury to reconstruct the bladder autonomic reflex arc by intradural microanastomosis of ventral root and the dorsal root between L5 and S2 simultaneously is practical in a rat model and may have potential in cl inical appl ication.
Objective To study major influential factors of the micturition alert device dedicated to neurogenic bladders for the product design and cl inical appl ication of the device. Methods One ferrite permanent magnet with thickness and diameter of 3 mm and 10 mm, respectively, and three NdFeB permanent magnets with the thickness of 3 mm and diameter of 10, 15 and 20 mm, respectively, were used. The effects of thickness of the abdominal wall as well as the position and type of permanent magnets on the micturition alert device dedicated to neurogenic bladders were measured in vitro simulated test, when the abdominal wall was set to 2, 3, 4, 5, 6, 7, 8 and 9 cm, respectively, and the position of permanent magnets was 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 and 12 cm, respectively. The effect of the geomagnetic field on the device was measured under the condition that the thickness of the simulated abdominal wall was set to 2, 3, 4 and 5 cm, respectively,and the position of permanent magnets was 2, 3, 4, 5, 6, 7, 8, 9 and 10 cm, respectively. Results The value showed inthe warning unit was positively correlated with the position of the ferrite permanent magnet only when the thickness ofthe simulated abdominal wall was 2 cm (r=0.632, P lt; 0.05). The correlation between the value of the warning unit andthe position of NdFeB permanent magnets was significant (r gt; 0.622, P lt; 0.05), which was intensified with the increasingdiameter of NdFeB permanent magnets, but weakened with the increasing thickness of the simulated abdominal wall. The effect of the geomagnetic field was correlated with the exposition of the body, the position of the permanent magnet and the thickness of the abdominal wall. Conclusion The major influential factors of the micturition alert device dedicated to neurogenic bladder include the magnetism and location of the permanent magnet, the thickness of the abdominal wall and the geomagnetic field. These factors are correlated with and affect each other. Reasonable allocation of these factors may optimize the device.
Objective To investigate the biocompatibil ity of sil ica gel embedded permanent magnets of themicturition alert device dedicated to neurogenic bladder. Methods According to the national standards of biologicalevaluation of medical equipment (GB/T 16886), Shanghai Biomaterial Research and Test Center was confided to evaluate the biocompatibil ity of sil ica gel embedded permanent magnets both in vitro and in vivo, including cytotoxicity test, sensitization test, primary skin irritant test and acute general toxicity test. The cytotoxicity test was performed according to the agar diffusion method. The L929 cell discoloration index and cell lysis index were counted at 24 hours after the action of the specimen. The sensitization test was performed according to the maximal dose method. The skin response was evaluated in 30 male albino guinea-pigs at 24 and 48 hours after the routine induction and provocation of leaching l iquors of the specimen. The primary skin irritant test was evaluated in 2 male healthy New Zealand rabbits according to the local tissue response at 24, 48 and 72 hours after intradermal injection of leaching l iquors of the specimen. The acute general toxicity test was evaluated in 10 male Kumming mice musculus albus according to animal condition at 4, 24, 48 and 72 hours after injection of leaching l iquors of the specimen through the caudal vein. Both the general reaction of canines and the pathology of the local bladder walls were observed at 2, 4 and 8 weeks after a permanent magnet was fixed on the anterior wall of urinary bladder in three canines. Results No sensitization, no stimulation and no acute general toxicity were observed except sl ight cytotoxicity to sil ica gel embeddedpermanent magnets. After implantation of a permanent magnet, the canines showed excellent tolerace, which manifested as no abnormal ity in spirit, appetite, urine and stool, healed wounds and no infection. Adhesions occurred between the epiploon and the bladder wall around the permanent magnet in two canines at 2 and 4 weeks, and between the lower abdominal wall and the bladder wall around the permanent magnet in the other canine at 8 weeks. The local bladder wall below permanent magnet was thickened, the fibrous capsule around the permanent magnet was thin, but the bladder mucosa was normal. Inflammatory reaction such as congestion, edema and inflammatory cells lessened from the serosa layer to the mucosa layer microscopically. Conclusion Sil ica gel embedded permanent magnets used in the micturition alert device dedicated to neurogenic bladde has excellent biocompatibil ity and meet the criteria for cl inical appl ication．