利用pcDNA 3.0裸质粒探讨创伤愈合动物模型中转入基因的分布规律

Approach of transgenic disposition of wound healing models with the naked plasmid pcDNA 3.0 in vivo

目的 通过在Wistar大鼠皮肤伤口局部应用含半乳糖苷酶（LacZ）报告基因的裸peDNA3．0重组质粒，建立创伤愈合转基因治疗动物模型并评价其应用价值。方法 通过建立大鼠全层皮肤切割伤模型，采用自身对照，在治疗侧伤口周围局部注射重组转染裸pcDNA3．0-LacZ质粒，通过B-gal原位染色检测注射3、30、100μg剂量后1、3、6、15天及愈合时的伤口组织标本中报告基因LacZ的表达，确定目标基因表达效率。结果 报告基因LacZ主要表达于皮下成纤维细胞中，100μg剂量有较高表达，表达效率为10．3％，表达时间主要在注射后3-6天，15天时仍可检测到少量表达细胞，愈合时及对照侧各个时相点均未检测到表达。结论 重组的裸pcDNA3．0-LacZ质粒能成功转染伤口的成纤维细胞等修复细胞，表达效率及时间适用于动物创伤愈合的转基因治疗。

Objective To establish wound healing models of transgenic therapy in vivo, and evaluate its applications. Methods The rats were given a full-thickness wound and received local subcutaneous injection of 3, 30,100μg naked plasmid Lac Z respectively. Then,the rats were executed on 1,3,6,15d and healing day respectively,and the transfection was confirmed by histochemical assays of the tissue of the wound for 13-gal. Results The blue reaction product of the 13-gal reaction was predominantly present in the fibroblasts underlying the wound, they were efficiently expressed in 100μg group with an expressing efficiency of 10.3% ,and the peak time of report gene expression was between 3-6d, but they were not expressed in the tissue of the control side and at healing stage. Conclusion The report gene can be efficiently expressed in the fibroblasts underlying the wound and have a short expressing time. The result indicated that this animal model was ideal for transgenic therapy of wound healing in vivo.