Choi Ung Hyune, Ha Yoon, Huang Xian, Park So Ra, Chung Joonho, Hyun Dong Keun, Park Hyeonseon, Park Hyung Chun, Kim Sung Wan, Lee Minhyung
Inha Research Institute for Medical Sciences, Inha University College of Medicine, Incheon, Korea.
J Neurosurg Spine. 2007 Jul;7(1):54-60. doi: 10.3171/SPI-07/07/054.
Vascular endothelial growth factor (VEGF) has been investigated as a therapy for many disorders and injuries involving ischemia. In this report, we constructed and evaluated a hypoxia-inducible VEGF expression system as a treatment for spinal cord injury (SCI).
The hypoxia-inducible VEGF plasmid was constructed using the erythropoietin (Epo) enhancer with the Simian virus 40 (SV40) promoter (pEpo-SV-VEGF) or the RTP801 promoter (pRTP801-VEGF). The expression of VEGF in vitro was evaluated after transfection into N2A cells. The plasmids were then injected into rat spinal cords with contusion injuries. The expression of VEGF in vivo was measured using reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. Locomotor recovery in the rats was evaluated using the Basso, Beattie and Bresnahan (BBB) scale for locomotor analysis.
In vitro transfection showed that pEpo-SV-VEGF or pRTP801-VEGF induced VEGF expression under hypoxic conditions, whereas pSV-VEGF did not. The VEGF level was higher in the pEpo-SV-VEGF and pRTP801-VEGF groups than in the control group. The VEGF expression was detected in neurons and astrocytes of the spinal cord. Locomotor recovery was improved in the pEpo-SV-VEGF and pRTP801-VEGF groups, and BBB scores were higher than in the control group. Staining using terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling showed that the number of apoptotic cells decreased in the plasmid-injected groups compared with the control group, and significant differences were observed between the hypoxia-responsive groups and the pSV-VEGF group.
These results suggest that the hypoxia-inducible VEGF expression system may be useful for gene therapy of SCI.
血管内皮生长因子（VEGF）已被研究用于治疗多种涉及缺血的疾病和损伤。在本报告中，我们构建并评估了一种缺氧诱导型VEGF表达系统用于治疗脊髓损伤（SCI）。
使用促红细胞生成素（Epo）增强子与猿猴病毒40（SV40）启动子（pEpo-SV-VEGF）或RTP801启动子（pRTP801-VEGF）构建缺氧诱导型VEGF质粒。将其转染到N2A细胞后评估VEGF在体外的表达。然后将质粒注射到有挫伤损伤的大鼠脊髓中。使用逆转录-聚合酶链反应和酶联免疫吸附测定法测量体内VEGF的表达。使用Basso、Beattie和Bresnahan（BBB）运动分析量表评估大鼠的运动恢复情况。
体外转染显示，pEpo-SV-VEGF或pRTP801-VEGF在缺氧条件下诱导VEGF表达，而pSV-VEGF则不能。pEpo-SV-VEGF和pRTP801-VEGF组的VEGF水平高于对照组。在脊髓的神经元和星形胶质细胞中检测到VEGF表达。pEpo-SV-VEGF和pRTP801-VEGF组的运动恢复得到改善，BBB评分高于对照组。使用末端脱氧核苷酸转移酶介导的脱氧尿苷三磷酸缺口末端标记染色显示，与对照组相比，质粒注射组的凋亡细胞数量减少，缺氧反应组与pSV-VEGF组之间观察到显著差异。
这些结果表明，缺氧诱导型VEGF表达系统可能对SCI的基因治疗有用。
