含鼠胚胎成纤维细胞的组织工程皮肤体外构建及大鼠移植研究

Construction and transplantation of tissue-engineered skin with mouse embryonic fibroblasts in SD mice

目的 利用鼠胚胎成纤维细胞（mouse embryonic fibrobalsts,MEFs）三维培养修复大鼠全层皮肤缺损,并初步探讨其机制.方法 培养MEFs和普通成纤维细胞（fibroblasts,FBs）,复合鼠尾胶原形成三维构建,ELASA法测定其分泌功能.制作大鼠全层皮肤缺损,按移植物不同分为3组：MEFs组（实验组）、FBs组（对照组1）和空白胶原组（对照组2）.观察愈合时间并计算愈合面积比率,定期取创面组织行CD31、波形蛋白免疫组化检测,并以Hoechst33342荧光标记MEFs,示踪其转归.结果 与FBs组相比,MEFs组的IL-6分泌更加旺盛,而TGF-β1分泌量少（P＜0.05）.实验组创面愈合快（P＜0.05）,微血管密度高（P＜0.05）,创面中FBs排列有序,移植的MEFs随时间减少.结论 MEFs三维构建可加速创面愈合,减轻瘢痕形成,其机制可能与胚胎细胞对创面愈合的诱导有关.

Objective To investigate the application and mechanism of tissue-engineered skin with mouse embryonic fibroblasts（MEFs）for the full-thickness skin defects on mice.Methods The MEFs and fibroblasts were cultured and seeded in scaffold made of rat tail collage.ELISA method was used for detection of secretory function.The full-thickness skin defects were created on mice and covered by MEFsscaffold complex（experimental group）,or FBs-scaffold complex（control group 1）,or scaffold only（control group 2）.The process of wound healing was evaluated by observation of the re-epithelization rate.Microvessal density（MVD）and vimentin within the wound sites were also detected with immunohistochemistry staining technique to describe the characteristics of wound healing.Hoechst 33342 staining was performed to trace MEFs＇fate.Results MEFs scaffold group had higher level secretion of IL-6 and lower of TGF-β1 than FBs scaffold group（P〈0.05）.Compared with wounds in control groups,the wounds in MEFs group healed markedly fast（P〈0.05）and the MVD was significantly higher（P〈0.05）.The fibroblasts in the wounds of MEFs group were arranged regularly and the MEFs decreased during the healing process.Conclusions The MEFs-scaffold complex can promote wound healing with less scar formation.MEFs may have an inducing effect on the wound healing.