| 董丽,马绍英,李晶,杨凯,杨婷,陈学英,李宝兴.腺病毒介导的hVEGF基因转染细胞修复大鼠放射性皮肤溃疡的效果研究[J].中华放射医学与防护杂志,2015,35(10):738-744 |
| 腺病毒介导的hVEGF基因转染细胞修复大鼠放射性皮肤溃疡的效果研究 |
| Effects of hVEGF-transfected fibroblasts on the repair of radiation-induced skin ulcer in rats |
| 投稿时间：2015-04-16 |
| DOI：10.3760/cma.j.issn.0254-5098.2015.10.005 |
| 中文关键词: 放射性皮肤溃疡 人血管内皮细胞生长因子 转染 损伤修复 |
| 英文关键词:Irradiated skin ulcers hVEGF Transfection Wound repair |
| 基金项目:山西省青年科技研究基金(2012021033-1) |
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| 中文摘要: |
| 目的 评价转染人血管内皮生长因子165(hVEGF 165)重组腺病毒载体的成纤维细胞体外表达状态及转染细胞修复放射性皮肤溃疡的效果。方法 构建hVEGF165基因过表达重组腺病毒载体,转染大鼠原代成纤维细胞,通过荧光定量PCR、免疫细胞化学和Western blot观察hVEGF165体外过表达状态;清洁级SD雄性大鼠24只,50 Gy γ射线局部照射,照后7 d于受照局部注射转染hVEGF165细胞,通过荧光定量PCR,分析转染基因在注射部位的表达状况,外观观察和组织学观察动物体内转基因细胞对放射性皮肤溃疡愈合效果的影响。结果 hVEGF转染组细胞体外实验中hVEGFmRNA表达显著上调,约为空转染组的88 373倍,胞质中出现hVEGF蛋白强阳性表达,且于相对分子质量23 000附近出现VEGF蛋白的特异性条带;受照后约2周局部出现溃疡,hVEGF转染组平均溃疡面积为40.2 mm2,比空转染对照组减少57%,愈合时间缩短约6 d,处理后3 d、1周皮肤组织中hVEGF mRNA相对表达量分别为空转染组的5.15和4.15倍(t=3.385、3.220,P<0.05)。结论 转染hVEGF165重组腺病毒载体的大鼠成纤维细胞能够在体外过量表达VEGF,且该转染细胞注射体内能够缩短放射性皮肤溃疡的愈合时间,增强损伤的愈合效果。 |
| 英文摘要: |
| Objective To evaluate the expression of human vascular endothelial cell growth factors 165 (hVEGF165) gene transfected into fibroblasts by recombinant adenovirus and study the repairing effect of this cells on radiated skin ulcer in rats. Methods The recombinant adenovirus with hVEGF165 was established and transfected to rat primary fibroblasts, and its expression of hVEGF165 in fibroblasts was identified with real-time PCR, immunocytochemistry and Western blot. Twenty four clean grade SD rats of were irradiated locally with 50 Gy γ rays to generate an animal model of radiation skin injury. The hVEGF165-transfected cells were injected to the irradiated site under rat skin 7 d post-irradiation. The therapeutic effects on the irradiated skin wound were evaluated through general observation as well as histological staining of HE. The expression of hVEGF in the irradiated skin tissue with fibroblasts injection was analyzed by Real-time PCR. Results The hVEGF165 gene was overexpressed in the transfected cells and approached to 88 373-fold bigger compared to controls transfected with blank vectors, and an extensive expression of VEGF in the cytoplasm of transfected cells was observed by immunohistochemistry. VEGF protein with the relative molecular mass of 23 000 was also detected in cell lysate by Western blot. The local skin ulcers in rats occurred about two weeks after irradiation. In the hVEGF165-transfected group, the average area of radiation-injured skin was 40.2 mm2, about 57% less than that of the control group transfected with blank vectors so that the healing time was shorten by 6 days. The relative concentration of hVEGF mRNA in the skin tissue of rats injected with hVEGF165-transfected cells were 5.15-fold and 4.15-fold bigger compared to that of controls (t=3.385,3.220,P<0.05) at 3 and 7 d after administration. Conclusions The primary fibroblasts transfected with hVEGF165 gene could efficiently release VEGF to the irradiated skin tissue and promote the recovery of irradiation skin ulcers by shortening the healing time and thus enhanced the therapeutic effect on skin wounds. |
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