ABSTRACT
Objective To explore the possibility of the inner vertical outer spiral complex tubular urethra scaffold vascularization in repairing long segment of urethral defect.Methods From August 2014 to October 2015,27 clean male New Zealand white rabbits were divided into 3 groups,S1 group was transfected recombinant vascular endothelial growth factor(VEGF) gene lentiviral vector group.S2 group was vascular pedicle transfer tube group.C group was simple stent group.A 3.0 cm inner vertical outer spiral complex scaffold was constructed by using the small intestine acellular matrix (SIS) and polylactic acid copolymer (PLGA) modified by type Ⅰ collagen surface,and adipose-derived mesenchymal stem cells (ADSC) and smooth muscle cells after transformation from New Zealand white rabbits.In S1 group,the seed cells were transfected by recombinant vascular endothelial growth factor (VEGF) gene lentivirus,which express VEGF protein.The complex scaffold was used to repair 3.0 cm rabbit urethral defect In S2 group,the untransfected cells were seeded into the scaffold and embedded in the skin near the groin artery 3 weeks for repairing urethral defect with vascular pedicle transfer tube.In group C,the unseeded scaffold was used to repair the urethral defect alone.Postoperative observation and urethrography were followed 4,8 and 24 weeks after implantation.The HE staining,fluorescence tracing,immunohistochemical and scanning electron microscopy were evaluated at the same phase.Results In S1 group,there were one urinary fistula and one urethral stricture-related death,respectively.The urethra was smooth and patent,histological examination showed active hyperplasia of urethral capillary.In S2 group,there were one urinary fistula and two urethral stricture-related deaths,respectively.The urethral was rough,local thinning or dilated.Fat accumulation and mucosal contraction were found in the urethral submucosal,respctively.In C group,there were one urinary fistula,three hypospadias,and three urethral stricture-related deaths.The thickness of the urethra was uneven and stricture bending.The urethral mucosa was poorly repaired and the scar was narrow.HE and CD31 staining showed that S1 and S2 groups were active in the proliferation of urethral capillaries,and the angiogenesis was abundant.VEGF staining showed that the cytoplasm of endothelial cell layer,smooth muscle layer of vascular wall and the urothelial epithelial cell layer were fully expressed at 24 weeks,especially in epithelial cell layer.CKpan staining showed that the epithelium of S1 and S2 group developed to stratified epithelium,and the morphology of urethra was similar to normal urethra at 24 weeks.The urethral epithelial in C group of grew poor as single-level,irregular arrangement,24 weeks is still a lack of effective stratified epithelium.HE and oα-SMA staining showed that the smooth muscle and actin gradually increased in group S1 and S2,α-SMA staining in group C was scarce and increased at 24 weeks.PLGA was encapsulated by the surrounding tissue and the structure of electrospinning was clear after 4 weeks,absorbed and degraded after 8 weeks and absorbed after 24 weeks.Conclusions The inner vertical outer spiral comnplex tubular urethra scaffold maybe a reasonable method in repairing long segment urethral defects,and the methods of tubular urethra scaffold vascularization by transfected VEGF gene recombinant lentiviral vector and vascular flap deserve more research.
ABSTRACT
ObjectiveTo explore the effects of myoblast formation around the urethra of stress urinary incontinence (SUI) rats after treated with bone marrow mesenchymal stem cells(BMSCs) or musclelike cells/calcium alginate composite gel injection therapy.MethodsIsolation,cultivation and identification of Sprague-Dawley rat bone marrow mesenchymal stem cell were performed.5-azacytidine was introduced to induce muscle-like cells.Calcium alginate gel was initially prepared by 2％ sodium alginate and 1％ calcium chloride solution at a volume ratio of 5∶1.Compounds of stem cells or muscle-like cells were mixed with gel,respectively,and were prepared for microinjection.SUI was produced in 72 6-week-old female Sprague-Dawley rats.The rats were then divided into 4 groups:Gel group,stem cell-gel group,muscle-like cell-gel group and mock control group.Each group was further divided into 3 groups.Submucosal injection of gel was performed at urethra and bladder neck.After preparation of cross sections of rat urinary tract at 4 weeks and 8 weeks after injection,HE staining,fluorescent tracing,staining of Desmin and α-skeletal muscle actin (α-SMA) were performed.OD values of positive rates were compared.ResultsAt 4 weeks and 8 weeks after injection in stem cell-gel group and muscle-like cell-gel group,growth of blood vessels gradually increased at gel edge,BMSCs and muscle-like cells gathered around the new blood vessels observed by fl(u)orescence tracer,muscle-like cells grew into elongated spindle-like cells.Desmin and α-SMA staining were positive in these groups,and the OD values in the stem cell-gel group and muscle-like cell-gel group was significantly higher than that from the gel only group and control group,but no difference was found between stem cell-gel group and muscle-like cell-gel group.ConclusionsCompound of BMSCs,muscle-like cells and calcium alginate composite gel has the potential to differentiate into muscle cells in the microenvironment of SUI rat model.In short term,the myoblast formation potential is the same whether the BMSCs was introduced into the micro-environment in vivo directly,or the BMSCs was implanted into microenvironment after the formation of the muscles cells induced by 5-azacytidine in vitro.
ABSTRACT
BACKGROUND:Although drug treatment.physics-behavior therapy,and postoperative therapy have been commonly used to treat stress urinary incontinence(SUI),there is still no satisfactory treatment at present.OBJECTlVE:To build a stable animal model simulating stress urinary incontinence(SUI)by bilateral transaction of pudendal nerve and nerves innervating pelvic floor muscles,including iliococcygeous muscle and pubococcygeous muscle.METHODS:A total of 18 6-week-old female SD rats weighing(1 99.44±8.41)g were randomly divided into 3 groups:normal,model,and sham-surgery groups,with 6 rats in each group.Rats in the model group underwent bilateral transaction of pudendal nerves and nerves innervating iliococcygeous/pubococcygeous muscles,while rats in the sham-surgery group had same procedures except nerve transaction.The normal group did not undergo any operation.Each rat was subjected to measure leak point pressure(LPP)at 2 weeks after the operation.After the measurement of LPP,cross sections of connection area of bladder and urethra were sent to histology.RESULTS AND CONCLUSION:One rat in the sham-surgery group died at 1 week after the operation.The LPP of model group decreased significantly by approximately 33%compared with the normal group(P＜0 05):however,there was no significant difference in LPP between sham-surgery and normal groups(P＞0.05).The results of histology showed loosely arrangement and atrophy of urethral sttriated muscle fibers in rats of the model group.Bilateral transaction of pudendal nerves and nerves innervating to iliococcygeous/pubococcygeous muscles resulted in SUI in rats stably.