Lee Minhyung, Choi Donghoon, Choi Min Ji, Jeong Ji Hoon, Kim Won Jong, Oh Seungjoon, Kim Yong-Hee, Bull David A, Kim Sung Wan
Department of Bioengineering, College of Engineering, Hanyang University, Seoul, Korea.
J Control Release. 2006 Sep 28;115(1):113-9. doi: 10.1016/j.jconrel.2006.07.010. Epub 2006 Jul 18.
Gene therapy with the vascular endothelial growth factor (VEGF) gene is a potential treatment for many disorders or injuries with ischemia. However, unregulated expression of VEGF may induce pathological angiogenesis, promoting tumor growth, diabetic proliferative retinopathy and rupture of atherosclerotic plaque. Therefore, the effective regulation of the gene expression is one of the requirements for the VEGF gene therapy. In this research, we evaluated the hypoxia-inducible gene expression system with the erythropoietin (Epo) enhancer and the Epo 3'-untranslated region (UTR). The luciferase plasmids were constructed with the Epo enhancer (pEpo-SV-Luc), the Epo 3'-UTR (pSV-Luc-EpoUTR) or both (pEpo-SV-Luc-EpoUTR). The polyethylenimine/plasmid complexes were transfected to 293 or A7R5 cells and the cells were incubated under normoxia or hypoxia. The results showed that the Epo enhancer or Epo 3'-UTR increased the target gene expression under hypoxia. pEpo-SV-Luc-EpoUTR showed the highest luciferase expression. The VEGF expression plasmid with the Epo enhancer and 3'-UTR was also constructed. The VEGF expression by pEpo-SV-VEGF-EpoUTR showed the highest specificity of the gene expression in the hypoxic cells. The results suggest that the VEGF plasmid with the Epo enhancer and the Epo 3'-UTR may be useful for gene therapy for ischemic diseases.
采用血管内皮生长因子（VEGF）基因进行基因治疗是针对许多缺血性疾病或损伤的一种潜在治疗方法。然而，VEGF的无节制表达可能会诱导病理性血管生成，促进肿瘤生长、糖尿病性增殖性视网膜病变以及动脉粥样硬化斑块破裂。因此，有效调控基因表达是VEGF基因治疗的必要条件之一。在本研究中，我们评估了带有促红细胞生成素（Epo）增强子和Epo 3'非翻译区（UTR）的缺氧诱导基因表达系统。构建了带有Epo增强子的荧光素酶质粒（pEpo-SV-Luc）、带有Epo 3'-UTR的质粒（pSV-Luc-EpoUTR）或两者兼具的质粒（pEpo-SV-Luc-EpoUTR）。将聚乙烯亚胺/质粒复合物转染至293或A7R5细胞，并在常氧或缺氧条件下培养这些细胞。结果表明，Epo增强子或Epo 3'-UTR在缺氧条件下可增加靶基因表达。pEpo-SV-Luc-EpoUTR显示出最高的荧光素酶表达。还构建了带有Epo增强子和3'-UTR的VEGF表达质粒。pEpo-SV-VEGF-EpoUTR介导的VEGF表达在缺氧细胞中显示出最高的基因表达特异性。结果表明，带有Epo增强子和Epo 3'-UTR的VEGF质粒可能对缺血性疾病的基因治疗有用。
