利用载有角质细胞生长因子微囊的组织工程皮肤修复裸鼠皮肤缺损

摘要

Abstract BACKGROUND: As an emerging technology, tissue-engineered skin has great application prospects. Keratinocyte growth factor (KGF) is proved to promote the proliferation of epidermal cels. OBJECTIVE: To evaluate the effect and characteristics of tissue-engineered skin carrying KGF nanocapsules in repairing skin defects of nude mice. METHODS:(1) The acelular dermal matrix loading KGF (KGF-ADM) was constructed. The human epidermal stem cel population and fibroblasts were captured and cultivated, and then identified. Epidermal stem cels were cultivated on the KGF-ADM and their growth was observed. The tissue-engineered skin loading KGF nanocapsules was transplanted onto the ful-skin defects on the back of nude mice compared with a blank group without keratinocyte growth factor nanocapsules and a control group with skin autograft. In 2, 4 and 6 weeks after transplantation, the contraction and histological healing of the skin were observed respectively. Then anti-human keratin 10-FITC and β1-integrin-Cy3 immunofluorescence were applied to detect the origin, growth and differentiation of stem cels in the epidermis and dermis. RESULTS AND CONCLUSION: The epidermal stem cel population grew wel on the surface of KGF-ADM and attached tightly. There were smal round epidermal stem cels and polygonal terminaly-differentiated cels, which presented with partly cloning growth and a tendency of merging into pieces. The results of tissue-engineered skin with KGF nanocapsules in repairing the skin defects were better than those of the blank group and the control group in 2, 4 and 6 weeks after transplantation. The transplanted skin could fuse with adjacent skin completely, but stil showed some contraction. Under the microscope, they showed good epidermis with layers and normal keratose stratum, and meanwhile, there were stil some β1-integrin+ cels at 8 and 10 weeks, which were epidermal stem cels or transient amplifying cels identified by immunofluorescence. These findings indicate that the tissue-engineered skin carrying KGF nanocapsules has good outcomes in repairing skin defects of nude mice, which is better than common tissue-engineered skin without KGF nanocapsules and autogeneous skin transplantation.%背景：组织工程皮肤作为一项新兴技术拥有良好的应用前景。有研究表明，角质细胞生长因子可以促进表皮细胞增殖。目的：观察荷载角质细胞生长因子纳米微囊的新型组织工程皮肤修复裸鼠皮肤缺损的效果和特点。方法：构建荷载角质细胞生长因子的脱细胞真皮基质复合物；将人表皮干细胞群和成纤维细胞分离、培养，并且进行鉴定；将表皮干细胞群接种于复合物之上，观察其生长状况；将荷载角质细胞生长因子纳米微囊的组织工程皮肤移植于裸鼠皮肤缺损处，将无角质细胞生长因子纳米微囊的组织工程皮肤作为空白组，将其自体皮肤移植修复缺损组作为对照组，于移植后2，4，6周时观察皮片挛缩及组织学愈合情况，并应用抗人角蛋白10及β1-整合素免疫荧光检测修复区表皮和真皮层细胞来源、分化及生长情况。结果与结论：表皮干细胞在复合物表面生长良好，黏贴紧密，可见有连接成片趋势的小圆形的表皮干细胞及多角形的终末表皮细胞，部分形成克隆团块。移植后第2，4，6周，荷载角质细胞生长因子纳米微囊组织工程皮肤修复裸鼠皮肤缺损的结果均优于空白组及对照组，移植的皮肤边缘与邻近皮肤完全融合，但存在一定程度的挛缩。修复区组织工程皮肤的表皮细胞分层良好并能产生角质层，同时，移植后8，10周，组织工程皮肤切片免疫荧光染色可以鉴别出少量β1-整合素阳性细胞，均为表皮干细胞或短暂扩充细胞。结果证实，荷载角质细胞生长因子纳米胶囊的新型组织工程皮肤修复裸鼠皮肤缺损的效果较好，优于普通组织工程皮肤及自体全厚皮片移植修复。