The development of new bladder augmentation technique using cultured bladder epithelial cell sheets.

胃肠瓣膀胱重建可引起严重的并发症。大多数并发症与肠黏膜有关。我们假设用尿路上皮细胞替代胃肠道黏膜可以减少临床并发症。本研究的目的是研究未经蛋白水解酶处理(1)的新型温度反应性细胞培养系统获得的完整的尿路上皮细胞片能否在狗模型中成功地自体移植到去黏膜的胃瓣上，以促进天然样尿路上皮的再生。通过部分膀胱切除术从beagle犬获得活的尿路上皮（n=8）。将收获的尿路上皮细胞接种于用热敏聚合物聚n -异丙基丙烯酰胺修饰的温度敏感培养皿中。培养三周的尿路上皮细胞产生的尿路上皮细胞片是无创收获的，无需从这些培养皿中进行酶处理。将尿路上皮细胞片自体移植到手术去黏膜的胃瓣上。自体移植3周后处死狗，观察带尿路上皮细胞片的胃瓣。比较了胃瓣移植的尿路上皮细胞与天然的尿路上皮细胞的细胞和组织特性。超细结构也进行了检测。在没有任何蛋白水解酶的情况下，通过降低培养温度从温度敏感表面获得完整的尿路上皮细胞片，显示出独特的移植特性。8个移植物中有5个显示出可再生的尿路上皮细胞。尿路上皮细胞片自发地附着在脱粘膜组织表面，完全不需要任何缝合或固定，并发展成与天然尿路上皮高度相似的分层活上皮。再生的尿路上皮未被抗质子泵抗体染色，而抗质子泵抗体通常对胃粘膜上皮细胞呈阳性染色。我们成功地将尿路上皮细胞片自体移植到狗去黏膜胃瓣上，无需缝合或固定，在体内生成了多层尿路上皮。完整的细胞片移植方法在体内快速产生原生样上皮。这种多用途的技术在泌尿道组织工程和外科重建中应该是有用的。少

Bladder reconstruction using gastrointestinal flaps may induce severe complications. The majority of the complications are related to intestinal mucosa. We have hypothesized that the replacement of gastrointestinal mucosa with urothelial cells could reduce clinical complications. The purpose of this study was to examine whether intact urothelial cell sheets harvested from new temperature-responsive cell culture systems without proteolytic enzyme treatment (1) would be autografted successfully onto demucosalized gastric flaps in a dog model to promote improved regeneration of native-like urothelium. Viable urothelium was obtained from beagle dogs by partial cystectomy (n=8). Harvested urothelial cells were seeded on temperature-responsive culture dishes modified with the thermally sensitive polymer, poly(N-isopropylacrylamide). Urothelial cells cultured for three weeks generated urothelial cell sheets that were non-invasively harvested without enzymatic treatment from these dishes simpl … More y by reducing culture temperature. Urothelial cell sheets were autografted onto surgically demucosalized gastric flaps. Three weeks after autografting, dogs were sacrificed and the gastric flaps with the urothelial cell sheets were examined. Cell and tissue characteristics were compared between these urothelial cell sheet-grafted gastric flaps and native urothelium. Ultrafine structures were also examined. Intact urothelial cell sheets harvested from temperature-responsive surfaces by reducing culture temperature without any proteolytic enzymes exhibit unique properties for transplantation. Five of the eight urothelial cell sheet-grafted flaps showed viable urothelial regeneration. Urothelial cell sheets attached spontaneously to demucosalized tissue surfaces completely without any suture or fixing, and developed into a stratified viable epithelium highly similar to native urothelium. Regenerated urothelium remained unstained by anti-proton pump antibody, which typically stains epithelial cells positively in gastric mucosal layers. Urothelial cell sheets were autografted onto dog demucosalized gastric flaps successfully without suture or fixation requirements, generating a multilayered urothelium in vivo. The intact cell sheet grafting method rapidly produces native-like epithelium in vivo. This versatile technology should prove useful in urinary tract tissue engineering and surgical reconstructions. Less