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目的以大鼠深Ⅱ度烧伤为模型,观察工程细胞即腺病毒介导肝细胞生长因子(hepatocyte growth factor,HGF)基因修饰的骨髓间充质干细胞(mesenchymal stem cells,MSCs)对烧伤创面愈合中皮肤附属器再生的作用。方法体外分离、培养雄性Wistar大鼠骨髓MSCs,转染携带HGF基因的重组腺病毒(100 pfu/cell)后备用。16只雌性Wistar大鼠复制深Ⅱ度烧伤模型,随机分为观察组与对照组各8只。在造模同时,观察组将携带HGF基因的重组腺病毒修饰的骨髓MSCs悬液,对照组将同体积磷酸盐缓冲液注射于创面真皮下。分别于细胞移植后第3,5,7,14,21天计算创口愈合率,第7,14,21天光镜下观察组织愈合及皮肤附属器情况,常规PCR法检测细胞移植后第21天创面组织中性别决定基因Sry片段。结果携带HGF基因的重组腺病毒可有效转染MSCs,修饰的MSCs移植后第7,14天,观察组创口收缩率与对照组比较,差异有统计学意义(P<0.01)。HE染色结果显示,第7天观察组表皮化优于对照组,第21天观察组再生表皮明显厚于对照组,并可见再生毛囊和皮脂腺。观察组雌性大鼠创面组织中扩增到Sry基因片段。结论 MSCs联合携带HGF基因的重组腺病毒异体移植可促进烧伤创面愈合修复过程中皮肤附属器的形成,且移植的细胞也参与了烧伤创面修复过程中附属器的形成。
OBJECTIVE: To investigate the effect of mesenchymal stem cells (MSCs) modified by adenovirus-mediated hepatocyte growth factor (HGF) on the healing of burn wounds in rats with deep second degree burn in rats. The role of skin appendage regeneration. Methods MSCs from male Wistar rats were isolated and cultured in vitro. Recombinant adenovirus (100 pfu / cell) carrying HGF gene was transfected into MSCs. Sixteen female Wistar rats were duplicated deep second degree burn model and randomly divided into observation group and control group. At the same time of modeling, the observation group will carry the recombinant adenovirus-modified bone marrow MSCs suspension containing HGF gene, while the control group will inject the same volume of phosphate buffer under the wound dermis. The wound healing rate was calculated on the 3rd, 5th, 7th, 14th and 21st days after the cell transplantation. The wound healing and the skin appendages were observed on the 7th, 14th and 21st day under light microscope. Tissue genotyping Sry fragments. Results The recombinant adenovirus carrying HGF gene was successfully transfected into MSCs. On the 7th and 14th day after transplantation, the shrinkage rate of wounds in the observation group was significantly lower than that in the control group (P <0.01). The results of HE staining showed that on the 7th day, the epidermis of the observation group was better than that of the control group. On the 21st day, the regenerated epidermis of the observation group was thicker than that of the control group, and the regenerating hair follicles and sebaceous glands were seen. In the observation group, the Sry gene fragment was amplified from the wound surface of female rats. Conclusions MSCs combined with recombinant adenovirus carrying HGF gene can promote the formation of skin appendages during the healing process of burn wounds, and the implanted cells also participate in the formation of appendages during the repair of burn wounds.