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背景:近年研究表明,生长因子作为一种分子信号调控细胞的增殖、分化、迁移与代谢,其表达与调控在慢性创面愈合中起着很重要的作用。目的:观察血管内皮生长因子对创伤组织血管内皮生长因子受体,碱性成纤维细胞生长因子和血小板源性生长因子mRNA表达的影响,分析其促进损伤组织修复的作用途径。设计:对照动物实验。单位:解放军第四军医大学西京医院整形科。材料:实验用新西兰白兔6只,兔龄48～60个月。纳米微囊包裹的重组质粒DNA真核表达载体pcDNA3.1/myc-hisA-血管内皮生长因子165(VEGF165)由第四军医大学西京医院整形外科贾宁硕士惠赠。方法:实验于2004-10/2005-06在西京医院整形外科实验室完成。①以血管内皮生长因子165为目的基因,构建真核表达载体pcDNA3.1/myc-hisA-VEGF165,利用纳米微囊包裹制成纳米微囊-血管内皮生长因子165复合体。兔经麻醉后,在其耳腹侧制成直径为6mm的3个圆形创面,暴露软骨。以明胶海棉薄片蘸取纳米微囊-血管内皮生长因子165复合体100μL覆于每只兔一侧耳创面,外层覆盖无菌密封薄膜,作为血管内皮生长因子组;对侧每一创面以明胶海棉薄片蘸取100μL纳米微囊-空载质粒覆于创面作为对照组;在环切部位近端的兔耳皮肤作为正常皮肤组。②利用反转录聚合酶链反应方法观察术后14d损伤组织血管内皮生长因子受体,碱性成纤维细胞生长因子和血小板源性生长因子mRNA的表达变化。③术后14d,以创面为中心,切取1cm×1cm正方形组织块(含兔耳全层),制成标本,苏木精-伊红染色,光镜下观察新生肉芽组织的生长情况。主要观察指标:创伤组织血管内皮生长因子受体、碱性成纤维细胞生长因子和血小板源性生长因子表达。结果:①苏木精-伊红染色显示,血管内皮生长因子组创面肉芽生长及上皮爬行速度明显快于对照组。②反转录聚合酶链反应显示,血管内皮生长因子组损伤组织内血管内皮生长因子受体,碱性成纤维细胞生长因子和血小板源性生长因子mRNA的表达水平明显高于对照组(P<0.05)及正常组(P<0.01)。结论:外源性血管内皮生长因子可以上调创伤组织中血管内皮生长因子受体,碱性成纤维细胞生长因子和血小板源性生长因子的表达,血管内皮生长因子可能作用于其受体,通过与其他细胞因子的协同作用来实现其对伤口愈合的促进作用。
BACKGROUND: Recent studies have shown that growth factor acts as a molecular signal to regulate the proliferation, differentiation, migration and metabolism of cells. Its expression and regulation play an important role in the healing of chronic wounds. OBJECTIVE: To observe the effect of vascular endothelial growth factor (VEGF) on the expression of vascular endothelial growth factor receptor, basic fibroblast growth factor and platelet-derived growth factor mRNA in wound tissues and to explore the pathways that promote vascular endothelial growth factor repair. Design: Controlled animal experiments. Unit: Fourth Military Medical University Xijing Hospital Plastic Surgery. MATERIALS: Six New Zealand white rabbits were used for experiment, 48 to 60 months old. The eukaryotic expression vector pcDNA3.1 / myc-hisA-Vascular Endothelial Growth Factor 165 (VEGF165), a recombinant plasmid DNA encapsulated by nanocapsules, was kindly donated by Dr. Justin, a professor of plastic surgery at Xijing Hospital, Fourth Military Medical University. Methods: The experiment was performed in Plastic Surgery Laboratory of Xijing Hospital from October 2004 to June 2005. (1) The eukaryotic expression vector pcDNA3.1 / myc-hisA-VEGF165 was constructed by using vascular endothelial growth factor 165 as a target gene, and the nano-microencapsulated-vascular endothelial growth factor 165 complex was prepared by encapsulation with nano-microencapsulated cells. Rabbits were anesthetized and made into 3 circular wounds 6 mm in diameter on the ventral side of the ear to expose the cartilage. 100 microliters of nano-microvessel-vascular endothelial growth factor 165 complex was dipped in gelatin sponge slice on the ear wound surface of each rabbit, and the outer layer was covered with aseptic sealing film as vascular endothelial growth factor. On the opposite side of each wound, gelatin Sponge chip dipped 100μL nano-microcapsules - empty plasmid overlying the wound as a control group; the rabbit ear skin near the circumcision site as a normal skin group. ② The expression of vascular endothelial growth factor receptor, basic fibroblast growth factor and platelet-derived growth factor mRNA were detected by reverse transcription-polymerase chain reaction 14 days after operation. ③ At 14 days after operation, the wounds were taken as the center and 1 cm × 1 cm square tissue blocks (including the whole ear of rabbit) were excised and stained with hematoxylin-eosin. The growth of new granulation tissue was observed under light microscope. MAIN OUTCOME MEASURES: Expression of vascular endothelial growth factor receptor, basic fibroblast growth factor and platelet derived growth factor in wound tissues. Results: ① Hematoxylin-eosin staining showed that the growth of granulation tissue and the rate of epithelial creep in vascular endothelial growth factor group were significantly faster than those in control group. ② Reverse transcriptase-polymerase chain reaction showed that the expression of vascular endothelial growth factor receptor, basic fibroblast growth factor and platelet-derived growth factor mRNA in vascular endothelial growth factor group was significantly higher than that in the control group (P < 0.05) and normal group (P <0.01). Conclusion: Exogenous vascular endothelial growth factor can up-regulate the expression of vascular endothelial growth factor receptor, basic fibroblast growth factor and platelet-derived growth factor in wound tissue. Vascular endothelial growth factor may act on its receptor, The synergistic effect of other cytokines to achieve its role in promoting wound healing.