尿道细胞外基质修复尿道缺损的研究

目的 寻求一种较理想的尿道修复材料。方法 20只家兔尿道制成尿道细胞外基质(ECM)；另40只随机分3组：尿道ECM移植组(实验组)，对照组Ⅰ及对照组Ⅱ。实验组切除尿道1．O～1．5cm后用尿道ECM修复并于术后10d、3周、6周及24周行组织再生情况观察；另于术后10周、24周各取4只行膀胱尿道造影；24周时实验组及对照组Ⅰ各取4只行尿流动力学检测；24周实验组取4只行尿道镜检查。结果 缺损修复术后10d，基质中见单层上皮细胞且有血管长入ECM；3周时尿道ECM管腔已完全被上皮细胞覆盖；6周时可见平滑肌细胞再生，炎性细胞消失；24周后其组织结构与正常组织相比差异无统计学意义。膀胱尿道造影无尿液外渗，无梗阻及结石形成。尿流动力学检测结果实验组与对照组Ⅰ差异无统计学意义；尿道镜检查证实尿道黏膜完整光滑，尿道内径及其形态正常。结论 尿道细胞外基质是一种理想的尿道修复材料。     

异体真皮细胞外基质重建尿道的实验和临床研究

目的：寻求理想的尿道修复材料。方法: 2只犬真皮脱细胞处理后制成真皮细胞外基质框架;18只犬分为2组;异体真皮细胞外基质（ECM）移植组（实验组）15只,异体真皮移植组（对照组）3只,全麻下将尿道中段切除5cm,将同样长度真皮ECM或异体真皮缝成直径0．3cm管状替代缺损尿道,8F硅塑管留置4周,实验组分别于术后1,3,6,12,24周每次3只取材行光镜,电镜及免疫组化检查。结果：组织学显示：1周时移植物边缘腔内面有尿上皮,管壁可见少量肌束,6周时移植物区尿路上皮达4－5层,肌层分布均匀,24周后移植物已无法与宿主尿道辨别,对照组术后8－10d移植物坏死出现尿瘘,在实验基础上用于临床2例均获成功。结论ECM为理想的尿道修复材料。     

小肠黏膜下层修复尿道的实验研究

目的探讨小肠黏膜下层(small intestinal submucosa,SIS)在尿道修复重建中的应用价值.方法 24只日本雄性大耳白兔,随机分为A、B、C及D组(n=6).A、B组切除前尿道2.0 cm,A组,用管状SIS修复尿道缺损;B组将其断端与周围组织直接缝合作为对照.C、D组仅切除2.0 cm尿道前壁,保留一半尿道壁为底板,C组用片状SIS修复尿道缺损;D组将其残端与周围组织直接缝合作为对照.均于修复后6、12周行组织学观察;12周行尿道膀胱造影及尿动力学检查.结果术后6周,A、C组修复的尿道有单层上皮细胞覆盖,基层组织中可见SIS的微小碎片包裹,出现不规则紊乱的平滑肌细胞生长,A组较C组的炎性反应重,有白细胞及淋巴细胞浸润,C组出现新生血管.术后12周,C组的上皮组织及基层下组织与D组无明显差别,平滑肌排列规则,血管数目进一步增多,炎性反应消失,未见SIS组织;A组中仍可见少数SIS的微小碎片;B组1只、D组2只尿道自行修复,余可见尿道闭塞,大量结缔组织生长,炎性细胞浸润,无正常上皮结构.术后12周尿道膀胱造影,A、C组可见尿道完整、光滑,无尿液外渗、尿道憩室等形成;尿动力学检查示A、C组的膀胱容量、最大尿道压分别与术前比较,差异无统计学意义(P＞0.05),而B、D组不能置入测压管检测.结论 SIS可作为兔尿道修复重建的良好支架材料,片状SIS修复优于管状SIS修复.     

血管细胞外基质和尿道细胞外基质修复兔尿道缺损的比较研究

目的　寻求较理想的尿道修复材料。　方法　切取 10只家兔的腹主动脉和尿道各 3cm ,制备成血管细胞外基质 (VECM)和尿道细胞外基质 (UECM)。另 2 0只分别切除尿道 2 .5cm并随机分为VECM修复组和UECM修复组。两组均于修复术后 10d、3周、6周及 2 4周行组织再生情况研究 ;另于术后 10周、2 4周各取 2只行膀胱尿道造影 ;2 4周两组各取 2只行尿动力学检测和尿道镜检查。　结果　制备的VECM和UECM均为白色透明状 ,但VECM较UECM弹性和机械强度好。缺损修复术后10d ,基质中见单层上皮细胞且有血管长入ECM ,基质和受体尿道连接处有炎性细胞浸润 ;3周时基质管腔已完全被上皮细胞覆盖 ;6周时可见平滑肌细胞再生 ,炎性细胞消失 ;2 4周后其组织结构与正常尿道组织结构一致。VECM修复组和UECM修复组相比其组织再生过程无差异。尿动力学检测VECM修复组和UECM修复组的膀胱容量分别为 (30 .2± 1.6 )ml和 (32 .1± 1.4 )ml、尿道最高压分别为(15 .2 7± 1.36 )mmHg和 (14 .6 8± 1.6 5 )mmHg、尿道最低压分别为 (12 .4 9± 1.2 3)mmHg和 (11.96±0 .98)mmHg ,差异无统计学意义 (P >0 .0 5 )。膀胱尿道造影可见尿道壁完整光滑通畅 ,不能分辨移植区与正常组织 ,无尿液外渗 ,无梗阻及结石形成 ;尿道镜检查证实VECM修复组和U     

应用异体脱细胞尿道基质修复尿道缺损

目的探讨应用同种脱细胞尿道基质修复尿道缺损的可行性。方法将14只雄性新西兰兔分为两组，切除实验组长约1.0～1．5cm的尿道，用相应长度脱细胞尿道基质修复；对照组行假手术。术后行尿道造影并取尿道标本作病理检查。结果12只实验兔的脱细胞基质移植物没有移位。除2例狭窄、2例尿瘘外，其余满意效果。病理检测示，术后3周尿道管腔上皮化，6个月基质中平滑肌及血管再生明显。结论同种脱细胞尿道基质材料可以修复兔尿道部分缺损。     

脱细胞膀胱粘膜下筋膜修复尿道的实验研究

目的探索组织工程方法在尿道修复中的应用。方法将12只雄兔作为实验对象,切除部分尿道的全周粘膜,长度分1cm以内(Ⅰ组)和2cm以上(Ⅱ组)两组,将制备完成的同种家兔膀胱粘膜下脱细胞筋膜用可吸收线,缝合到尿道缺损处,术后2周、6周分别处死一批实验对象,通过尿道造影,大体观察,修复段组织切片HE染色光镜研究和免疫组化等来研究修复段尿道的解剖和组织学变化。结果Ⅰ组仅1只(1/6)发生尿瘘,其余尿道造影显示尿道基本通畅；而Ⅱ组4只(4/6)发生狭窄(不伴尿瘘),1只(1/6)发生尿瘘；术后2周两组植入筋膜依然存在；术后6周时,尿道植入筋膜已降解,Ⅰ组的修复段尿道已全部被宿主自身粘膜所覆盖,粘膜光洁完整,管腔无明显狭小；Ⅱ组的修复段尿道中间苍白增厚,致密坚硬且管腔狭小。结论单纯脱细胞膀胱粘膜下筋膜能修复短距离兔尿道缺损,而对长距离缺损修复效果不理想。     

前尿道替代膜部尿道治疗男性后尿道狭窄52例报告

目的：探讨长段后尿道狭窄手术治疗方法。方法：同顾性总结前尿道替代膜部尿道端端吻合治疗狭窄段〉2cm的后尿道狭窄患者52例。结果：一次手术成功49例,成功率94．2％,3例术后尿线细,排尿不畅经内窥镜切除0．3～0．5cm瘢痕后排尿通畅。随访5～20年,全部排尿通畅,最大尿流率20～25ml／s,平均22ml／s。结论：切除狭窄段瘢痕前尿道替代膜部尿道端一端吻合治疗后尿道狭窄成功率高,远期效果好。术中彻底切除瘢痕,满意的无张力外翻端一端吻合及术后预防感染是手术成功的关键。     

前尿道代后尿道成形术治疗男性复杂性后尿道狭窄

目的探讨长段复杂性后尿道狭窄治疗新方法。方法采用分期前尿道代后尿道成形术治疗3例复杂性后尿道长段狭窄（6．5—10．0cm）患者。第一期行阴茎转位尿道端端吻合术，术后3—6个月行二期阴茎伸直、尿道会阴造口术，6个月后行第三期前尿道成形术（Johanson Ⅱ期尿道成形术）。结果例1术后排尿通畅，膀胱尿道造影检查示尿道通畅，双侧输尿管返流近消失，最大尿流率18．8ml／s，随访2年，最大尿流率18ml／s，无剩余尿。例2术后排尿通畅，最大尿流率19．5ml／s，无剩余尿，尿道扩张可顺利通过22F尿道探子。例3经会阴一耻骨联合径路行第一期阴茎转位尿道端端吻合术、尿道直肠瘘、尿道会阴瘘切除、修补术，术后尿道直肠瘘及尿道会阴瘘治愈，但因耻骨联合切口感染致吻合口狭窄，有待进一步治疗。结论分期前尿道代后尿道加前尿道重建方法是治疗男性长段复杂性尿道狭窄的有效方法。     

结肠粘膜重建尿道治疗复杂性尿道狭窄的探讨

目的 探讨结肠粘膜替代尿道治疗复杂性较长段尿道狭窄或闭锁的可能性。方法 采用结肠粘膜重建尿道一期尿道成形术治疗6例复杂性较长段尿道狭窄，患者术前有平均3次不成功的尿道修复史，用结肠粘膜重建的尿道长10－15cm，平均12.7cm。术后随访时分别行逆行尿道造影，尿道镜和尿流率检查。结果 术后随访3－14个月，平均7.8个月。1例在术后3个月并发尿道外口狭窄性，经手术矫正后排尿通畅，术后1年随访时最大尿流率28.7ml/s。余5例术后排尿通畅，最大尿流率大于15ml/s。术后6个月4例经尿道镜检查，肉眼较难将尿道的结肠粘膜与正常的尿道粘膜相区别。结论 结肠粘膜重建尿道治疗较长段尿道狭窄或闭锁是一种可行而有效的方法，适合阴茎皮肤或膀胱粘膜利用有困难时的尿道重建。     

尿道细胞外基质在兔尿道重建中的应用

目的 评价尿道细胞外基质作为一种生物材料重建尿道的效果. 方法 切取新西兰兔的尿道制备尿道细胞外基质.手术切除实验组1～1.5 cm的尿道片段后用细胞外基质修复缺损,采用ELISA法检测术前、术后12、24及48 h血清TNFα的水平,评估术后兔的免疫反应状态.术后10d和3、6、24周取修复段尿道,行组织学观察并做尿道造影、尿道镜及尿流动力学检查. 结果 术后实验组血清TNFα水平较对照组略有升高,但无统计学意义(P>0.05).术后10 d,上皮细胞开始从边缘向细胞外基质移行并出现新生小血管;3周上皮细胞覆盖细胞外基质的整个管腔;6周出现排列不规则的平滑肌纤维;24周平滑肌数量明显增多,成束状排列.尿道造影、尿道镜及尿流动力学检测检查显示,尿道基质管壁光滑,排尿通畅. 结论 尿道细胞外基质是一种安全有效的尿道重建材料.     

Urethral Reconstruction Using Bone Marrow Mesenchymal Stem Cell– and Smooth Muscle Cell–Seeded Bladder Acellular Matrix

Background

Congenital or acquired abnormalities may lead to a urethral defect that often requires surgical reconstruction. The traditional methods often lead to complications, including urethrocutaneous fistulae and strictures. In this study, we proposed to construct a tissue-engineered sheet graft (TESG) using a bone marrow mesenchymal stem cell (BMSC)– and smooth muscle cell (SMC)–seeded bladder acellular matrix (BAM) for urethral reconstruction.

Methods

Rabbit BMSCs and SMCs were isolated, expanded, and identified in vitro before seeding into BAM as the experimental group, whereas BAM-only was the control group. The graft was used to construct TESG for implantation into the rabbit omentum for 2 weeks before urethral reconstruction. We divided 24 male rabbits into four experimental groups six each, and six other were the control group. Histological analysis was performed at 2 weeks, 4 weeks, 8 weeks, and 16 weeks postoperatively. Retrograde urethrography was performed at 16 weeks postoperatively.

Results

All experimental rabbits survived to they were humanly killed. At 8 weeks, there was no difference between the graft and the normal urethra with no severe shrinkage. At 8 and 16 weeks after TESG grafting in vivo, multilayer urothelium covered the graft, neovascularization was visible within the center of TESG, and organized smooth muscle bundles were present. Retrograde urethrography failed to demonstrate diverticula formation or urethral stricture. Three control rabbits died within 4 weeks postoperatively. Autopsy showed their urethras to be almost completely blocked whereas another three hosts displays urethral strictures.

Conclusion

A TESG was constructed using a BMSC- and SMC-seeded BAM for urethral reconstruction.     

犬舌黏膜与颊黏膜组合移植重建尿道的实验研究

目的 探讨犬舌黏膜与颊黏膜组合移植替代尿道的可行性和有效性. 方法杂种雌犬7只,平均体质量13 kg,剥离尿道黏膜4 cm×1 cm,取2 cm×1 cm舌侧面黏膜和颊黏膜各1条,拼接后移植至尿道内,恢复尿道黏膜的连续性.术后留置硅胶导尿管1周,拔除尿管后观察排尿情况.1.5～12.0个月后行逆行尿道造影检测实验犬尿道通畅情况,并用10 F尿管证实有无狭窄.随后处死实验犬,测定移植物长度并行病理组织学检查,观察舌黏膜与颊黏膜组合移植至尿道后组织学改变情况. 结果7只实验犬均存活,排尿通畅6只,发生尿道狭窄1只;无尿瘘.移植手术前后舌黏膜和颊黏膜拼接长度分别为(4.00±0.15)和(3.75±0.23)cm,差异有统计学意义(P＜0.05).排尿通畅犬移植黏膜存活良好;舌黏膜与颊黏膜交界处愈合良好,无狭窄发生;光镜下见鳞状上皮呈连续分布,拼接移植黏膜的鳞状上皮与尿道的移行上皮区分明显,移植黏膜的鳞状上:皮有渐被尿道移行上皮替代的趋势.结论 犬舌黏膜和颊黏膜组合移植可替代尿道黏膜并恢复尿道的连续性.     

Urethral stricture repair with an off-the-shelf collagen matrix

PURPOSE: In select patients with urethral strictures in whom genital skin is insufficient alternative tissues are needed for urethral reconstruction. We explored the feasibility of using a bladder submucosa collagen based inert matrix as a free graft substitute for urethral stricture repair. MATERIALS AND METHODS: A total of 28 patients 22 to 61 years old with a diagnosis of urethral stricture underwent reconstructive surgery using a collagen based inert matrix for urethral repair. The inert collagen matrix was trimmed to size as needed for each patient and the neourethra was created by anastomosing the matrix in an onlay fashion to the urethral plate with continuous 6-zero absorbable sutures. The size of the created neourethra ranged from 1.5 to 16 cm. A voiding history, physical examination, retrograde urethrography, uroflowmetry and cystoscopic examinations were performed preoperatively and postoperatively. Random urethral biopsies were also performed. RESULTS: After a 36 to 48-month followup (mean 37) 24 of the 28 patients had a successful outcome. The remaining 4 patients had a slight caliber decrease at the anastomotic sites on urethrography. A subcoronal fistula developed in 1 patient which closed spontaneously 1 year after repair. Mean maximum urine flow rate increased from the preoperative value of 9 +/- 1.29 to 19.7 +/- 3.07 ml. per second postoperatively. Cystoscopic studies revealed adequate caliber conduits and normal appearing urethral tissues. Histological examination of the biopsy specimens showed the typical urethral stratified epithelium. CONCLUSIONS: Use of an off-the-shelf collagen inert matrix appears to be beneficial for patients with urethral strictures and obviates the need for obtaining an autologous graft, thus eliminating donor site morbidity.     

Urethral replacement using epidermal cell-seeded tubular acellular bladder collagen matrix

OBJECTIVES: To investigate the feasibility of replacing urinary epithelium cells with foreskin epidermal cells to reconstruct engineered anterior urethra with an acellular collagen matrix. MATERIALS AND METHODS: Acellular collagen matrices were generated from allogeneic rabbit bladder submucosa. In nine rabbits, autologous foreskin epidermal cells were isolated, expanded in vitro, and labelled with 5-bromo2'-deoxy-uridine (BrdU) before seeding onto a tubular acellular collagen matrix (1.5x1 cm). In male rabbits, a urethral mucosal defect was created, and urethroplasty performed with a tubular acellular collagen matrix seeded with epidermal cells (nine rabbits) or with a matrix with no cell seeding (nine rabbits; control group). Urethrography was done at 1, 2 and 6 months after grafting. The urethral grafts were harvested and analysed grossly and histologically. RESULTS: In the control group, gross views and urethrography revealed stricture of repaired defects at the different sample times. In the experimental group, a wide urethral calibre was maintained with no sign of strictures. Histology in the control group showed a single layer of epithelium cells with disorganized muscle fibre bundles in the submucosa layer at 1 month after grafting, and a transitional cell layer surrounded by disorganized muscle fibre bundles at 2 and at 6 months. Grafts seeded with epidermal cells formed a single-layer structure by 1 month, and at 2 and 6 months there were several layers of epidermal cells with abundant vessels in the submucosa. There was an evident margin between graft epidermal cells and host epithelium at 6 months. The implanted cells expressed keratin, shown by staining with anti-pancytokeratins. Immunofluorescence for BrdU confirmed the presence of implanted epidermal cells at 1 month after grafting; there were fewer positive cells at the implantation site at 2 months. At 6 months, there were several layers of epidermal cells with no signs of BrdU staining. CONCLUSIONS: Urethral reconstruction was better with an acellular collagen matrix seeded with epidermal cells than with the acellular collagen matrix alone. Foreskin epidermal cells seem adequate in replacing urethral epithelium cells for urethral reconstruction.     

Acellular matrix tube for canine urethral replacement: is it fact or fiction?

PURPOSE: We evaluated the results of acellular matrix used as a tube for replacement of a relatively long segment of the canine urethra. MATERIALS AND METHODS: The study included 9 female and 5 male mongrel dogs in which a 3 cm segment of the whole urethral circumference was excised and replaced by a tube of acellular matrix of the same length and width. The acellular matrix was obtained by excision of the whole urethra of donor female dogs that were sacrificed and not included in the study group. The retrieved urethra was treated to have complete cell lysis with maintenance of the fiber framework. In all dogs the urethra was stented for 4 weeks. Ascending urethrogram was done after stent removal every month until the dogs were sacrificed. The dogs were sacrificed at a rate of 1 weekly for 4 weeks and then monthly. If urine retention occurred, ascending urethrogram was performed and the dog was sacrificed. Before sacrifice the urethra was exposed and carefully examined. The whole specimen was then excised for histopathological examination. RESULTS: All dogs survived surgery and no postoperative complications were seen during urethral stenting. After stent removal all dogs had a urethral fistula and/or stricture, which increased in severity until urine retention occurred by the end of month 3. Exploration at 1, 2, 3 and 4 weeks showed an intact graft, although there was progressive shrinkage in length and urethral calibration also demonstrated progressive narrowing of the lumen. At 3 months there was marked shrinkage (length equaled 0.3 cm) and complete obliteration of the lumen. Histopathological examination showed progressive fibrosis, which was extensive by week 4. CONCLUSIONS: Acellular matrix tube is not able to replace a 3 cm segment of canine urethra.     

Long-term study of male rabbit urethral mucosa reconstruction using epidermal cell

Aim： To investigate the transformation of characteristics of epidermal cells from foreskin which were used to reconstruct male rabbit anterior urethra in combination with acellular collagen matrices. Methods： In three rabbits, autologous foreskin epidermal cells were isolated, expanded in vitro, and seeded （inoculated） onto a tubular acellular collagen matrix, acquired from allogeneic rabbit bladder submucosa. A urethral mucosal defect was created, and urethral reconstruction was performed with the tubular acellular collagen matrix seeded with epidermal cells. Results： On gross examination at 12 months following the procedure, the mucosa of the urethral grafts appeared lubricous and smooth. Urethrography showed that a wide urethral caliber had been maintained without any sign of strictures. Histological examination showed a transitional cell layer in the graft without evidence of a margin between the graft and the host tissue at 12 months postoperatively. Conclusion： Epidermal cells seeded onto acellular collagen matrices can be successfully used to reconstruct urethras that have defects and are transformed to transitional epithelial cells.     

Laparoscopic replacement of urinary tract segments using biodegradable materials in a large-animal model.

OBJECTIVE: We elected to evaluate laparoscopic segmental bladder and ureteral replacement with free biodegradable graft materials in a large-animal model. MATERIALS AND METHODS: In 18 Yucatan minipigs, a 1.5- to 2.8-cm segment of the upper ureter was excised. In 15 study animals, the ureter was laparoscopically replaced: by a stinted (6F double-J stent) tube graft made of acellular matrix (AMX) prepared from minipig ureters (MUMX) in 6 animals, acellular matrix prepared from domestic pig ureters (DUMX) in 3, and small-intestinal submucosa (SIS) in 6. In 3 control animals, the ureteral gap was bridged only by an indwelling stent. The stent was removed at 6 weeks, and retrograde ureteropyelography was performed preoperatively and at 8 and 12 weeks postoperatively, when animals were sacrificed. In 18 Yucatan minipigs, 3 x 3-cm bladder dome segments were laparoscopically replaced: by acellular matrix prepared from minipig small bowel (MBMX) in 6 animals, and SIS in 6 animals. The bladder was closed primarily in 6 control animals. Bladder capacity was assessed preoperatively and at 6 and at 12 weeks, when the animals were sacrificed. RESULTS: The average operating time for ureteral replacement was 187 (range 105-360) minutes. At 12 weeks, all animals had complete obstruction at the level of the replacement, with fibrosis +/- bone formation at the level of the stricture. For the bladder replacement groups, the average operating time was 147 (range 85-200) minutes. At 12 weeks, the bladder capacity was 60% of the preoperative capacity in the control group, 118% in the MBMX group, and 142% in the SIS group. Histologic examination showed regeneration of urothelium and some muscle with both MBMX and SIS. CONCLUSIONS: We were able to develop a reliable laparoscopic technique for both segmental ureteral and partial bladder replacement in a porcine model. With AMX and SIS replacement, regeneration of urothelium occurred in both ureter and bladder. However, functional replacement was successful only in the bladder.