膀胱上皮细胞和膀胱脱细胞基质构建组织工程管状移植物行兔膀胱切除术后尿流改道

Tissue-engineered conduit using bladder acellular matrix and bladder epithelial cells for urinary diversioninrabbits

目的利用膀胱脱细胞基质(BAM)和膀胱上皮细胞构建组织工程管状移植物(TETG), 在兔模型上验证其进行尿流改道的可行性。方法选取雄性新西兰大白兔48只, 将其分为对照组和实验组, 各24只, 首先制作兔的BAM, 体外培养扩增兔膀胱上皮细胞, 观察上皮细胞在BAM上的生长情况。将复合物包绕尿管制作成长为4 cm、长径为0.8 cm的TETG, 使膀胱上皮细胞位于管腔内面, 将TETG移植到兔模型上验证其进行尿流改道的可行性。分别于尿流改道术后1、2、4、8周进行组织学、免疫组化检查, 术后12周进行静脉尿路造影(IVU)检查流出道的通畅情况。结果 BAM行HE检测染色显示未见细胞生长;膀胱上皮细胞种植到BAM上的生长曲线结果显示细胞生长良好。将TETG移植到动物体内进行尿流改道, 结果显示, 实验组均存活, 取材时发现移植物与盆腔脂肪轻度粘连, 未见明显尿瘘、狭窄及严重的肾积水。组织学检查揭示了有管腔上皮的生长。免疫组化检查也证实了管腔内上皮的再生情况, TETG移植到体内, 随时间增长, 内腔覆盖多层的成熟上皮细胞, 并可见新生血管形成。术后12周行IVU和膀胱镜检未见梗阻发生。相反, 对照组术后1个月内均死亡, 尸体解剖见流出道有瘢痕形成、闭锁及双肾重度积水。结论利用种植有尿路上皮细胞的BAM制作成TETG, 并用于动物体内进行尿流改道是可行的, 能使流出道的内腔有成熟上皮覆盖, 具有防止尿外渗的功能。

Objective To verify the feasibility of urine diversion in rabbit models by construction of tissue-engineered tubular grafts(TETG)using bladder acellular matrix(BAM)and bladder epithelial cells.Methods A total of 48 male New Zealand white rabbits(24 in the control group and 24 in the experimental group)were selected to prepare rabbit BAM,and the growth of rabbit bladder epithelial cells on BAM was observed.The compound was wrapped around the urinary tube as TETG with a length of 4 cm and a diameter of 0.8 cm,so that the bladder epithelial cells were located in the inner surface of the lumen.The TETG was transplanted into the rabbit model to verify the feasibility of urine flow diversion.Histological and immunohistochemical tests were performed at 1 week,2,4,and 8 weeks after the operation,and intraudio videoenous urography(IVU)was performed at 12 weeks after the operation to check the patency of the outlow tract.Results BAM staining showed no cell growth.The growth curve of bladder epithelial cells implanted on BAM showed good cell growth.TETG was transplanted into animals for urine flow diversion.The results showed that all 24 rabbits in the experimental group survived.Mild adhesion between the grafts and pelvic fat was found during sampling,and no obvious urinary fistula,stenosis,or severe hydronphrosis was observed.Histological examination revealed the growth of luminal epithelium.Immunohistochemical examination also confirmed the regeneration of lumen epithelium.When TETG was transplanted into the body,the lumen covered multiple layers of mature epithelial cells with the increase of time,and neovascularization was observed.No obstruction was found by intravenous urography and cystoscopy 12 weeks after operation.On the contrary,24 rabbits in the control group died within 1 month after operation,and the autopsy showed outflow tract scar formation,atresia,and severe hydronephrosis in both kidneys.Conclusions It is feasible to use BAM with urothelial cells to produce TETG for urine flow diversion in animals.The outflow tract is covered by mature epithelium and has the function of preventing urine extravasation.