预血管化BAM水凝胶／丝素蛋白复合支架修复尿道缺损的实验研究

At present, complex urethral repair and reconstruction using tissue engineering technology is one of the most promising clinical application research. There are three key issues to should be solved before clinical application: choosing ideal biological materials, combining materials with cells for constructing ideal grafts and rapid neovascularization on the grafts in vivo. In the study, it is the first to introduce thermosensitive bladder acellular matrix (BAM) hydorgel into urethral reconstruction area, and modify the hydogel by a variety of methods. The BAM hydorgel not only mimic the native extracellular matix environation which is essential for cell to grow, but also release angiogenic growth factors slowly, which are contributed to constructing a new urethra through combining the hydrogel with keratincocytes and vascular endothelial cells. Based on prevascularization theory in the tissue enginerring area, the BAM solution is added onto the cell surface of silk fibroin scaffold seeded with vascular endothelial cells and form prevascularized BAM hydrogel/silk fibroin composited scaffold in vitro. In vivo, the rapid anastomosis between prevascularized networks on the composited scaffold and the host vasculature contribute to rapid formation of new blood supply in the new urethral tissue, thereby resulting in better effect of urethral reconstruction. Based on these better effects, the study will give the complex urethral reconstruction new idea and approach.

利用组织工程技术进行复杂性尿道修复重建是目前最具临床应用前景的研究方向之一。选择理想生物材料、细胞与材料构建组织工程移植物以及移植物回植体内快速有效的获得新生血供是将此研究从实验推向临床过程中急需解决的三个关键性问题。本研究首次将温敏型膀胱脱细胞基质（BAM）水凝胶引入尿道修复重建领域，并通过多种方法对其理化特性进行改进。BAM水凝胶不仅最大限度地模拟细胞生长所必需的天然细胞外基质微环境，而且缓慢地释放其固有的促血管化生长因子，使其有利于与上皮细胞和血管内皮细胞共同构建新尿道。本研究依据组织工程预血管化理论，将BAM凝胶水溶液覆盖于复合血管内皮细胞的丝素蛋白支架的细胞面，体外培养、构建富含微血管网的BAM水凝胶/丝素蛋白复合支架，使新生血管网在复合支架回植体内后快速有效地与宿主血管吻合，以促进新尿道新生血供快速形成，最终达到有效修复尿道缺损的目的，为复杂性尿道修复重建提供新的思路与方法。

利用组织工程技术进行复杂性尿道修复重建是目前最具临床应用前景的研究方向之一。选择理想生物材料、细胞与材料构建组织工程移植物以及移植物回植体内快速有效的获得新生血供是将此研究从实验推向临床过程中急需解决的三个关键性问题。本研究首次将温敏型膀胱脱细胞基质（BAM）水凝胶引入尿道修复重建领域，制备了不同浓度的BAM水凝胶，通过BAM水凝胶孔径的测定、成胶时间、固有生长因子的释放以及血管内皮细胞生长情况等评估，发现在体外浓度为6mg/ml的BAM水凝胶更适合血管内皮细胞的生长。将包含血管内皮细胞的BAM水凝胶种植于裸鼠体内，发现BAM水凝胶在体内能明显促进血管内皮细胞的再生。所以认为BAM水凝胶是一种理想的天然温敏水凝胶。制备BAM水凝胶/丝素蛋白复合支架，将复合支架种植于兔大网膜内，2周后发现BAM水凝胶利用其促血管化作用在大网膜内形成预血管化的BAM水凝胶／丝素蛋白复合支架。将预血管化的BAM水凝胶／丝素蛋白复合支架修复兔2.5cm尿道缺损模型，术后评估尿道修复部位的组织再生情况，发现预血管化的BAM水凝胶／丝素蛋白能明显促进尿道组织的再生，最终达到有效修复尿道缺损的目的，为复杂性尿道修复重建提供新的思路与方法。