Augmentation of erythropoietin enhancer-mediated hypoxia-inducible gene expression by co-transfection of a plasmid encoding hypoxia-inducible factor 1alpha for ischemic tissue targeting gene therapy.
Therapeutic angiogenesis with gene encoding vascular endothelial growth factor (VEGF) is a potential treatment for ischemic diseases. However, VEGF expression should be tightly regulated to avoid side effects such as tumor growth. Previously, our group developed the erythropoietin (Epo) enhancer-SV40 promoter system for hypoxia-specific gene expression. In the present study, the activity of the Epo enhancer-SV40 promoter system was further enhanced without significant decrease in its specificity by co-transfection of the hypoxia-inducible factor 1alpha (HIF1alpha) gene. pSV-HIF1alpha was constructed by the insertion of the HIF1alpha cDNA into pSI. At a 1:1 ratio, co-transfection of pSV-HIF1alpha and pEpo-SV-Luc increased the promoter activity of the Epo enhancer-SV40 promoter system, showing at least three times higher gene expression under hypoxia as compared with the pEpo-SV-Luc single-plasmid transfection. Furthermore, co-transfection showed significant hypoxia specificity. Also, co-transfection of pEpo-SV-VEGF with pSV-HIF1alpha showed the enhanced VEGF expression without loss of hypoxia specificity, as compared with pEpo-SV-VEGF single-plasmid transfection. Furthermore, pSV-HIF1alpha induced the endogenous hypoxia-responsive genes such as angiopoietin-1, which would be beneficial for therapeutic angiogenesis. Therefore, with hypoxia specificity and higher gene expression, co-transfection of pSV-HIF1alpha and pEpo-SV-VEGF may be useful for ischemia targeting gene therapy.
Lee
Suyeon
S
Department of Bioengineering, College of Engineering, Hanyang University, 17 Haengdang-dong, Seongdong-gu, Seoul, South Korea.
Kim
Kyunghwa
K
Kim
Hyun Ah
HA
Kim
Sung Wan
SW
Lee
Minhyung
M
D000199:Actins / Q000096:biosynthesis; D000803:Angiotensin I / Q000096:biosynthesis; D002460:Cell Line; D004797:Enzyme-Linked Immunosorbent Assay; D004921:Erythropoietin / Q000502:physiology*; D015316:Genetic Therapy / Q000379:methods*; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis* / Q000235:genetics; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008156:Luciferases / Q000235:genetics; D010957:Plasmids / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013539:Simian virus 40 / Q000235:genetics; D014158:Transcription, Genetic / Q000235:genetics; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology
D000199:Actins; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D042461:Vascular Endothelial Growth Factor A; D004921:Erythropoietin; D000803:Angiotensin I; D008156:Luciferases