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Genetic modification of embryonic stem cells with VEGF enhances cell survival and improves cardiac function

lizongjin 添加于 2012-4-1 19:49 | 1131 次阅读 | 0 个评论

作者
Xie X, Cao F, Sheikh AY, Li Z, Connolly AJ, Pei X, Li R-K, Robbins RC, Wu JC
摘要
Cardiac stem cell therapy remains hampered by acute donor cell death posttransplantation and the lack of reliable methods for tracking cell survival in vivo. We hypothesize that cells transfected with inducible vascular endothelial growth factor 165 (VEGF(165)) can improve their survival as monitored by novel molecular imaging techniques. Mouse embryonic stem (ES) cells were transfected with an inducible, bidirectional tetracycline (Bi-Tet) promoter driving VEGF(165) and renilla luciferase (Rluc). Addition of doxycycline induced Bi-Tet expression of VEGF(165) and Rluc significantly compared to baseline (p<0.05). Expression of VEGF(165) enhanced ES cell proliferation and inhibited apoptosis as determined by Annexin-V staining. For noninvasive imaging, ES cells were transduced with a double fusion (DF) reporter gene consisting of firefly luciferase and enhanced green fluorescence protein (Fluc-eGFP). There was a robust correlation between cell number and Fluc activity (R(2)=0.99). Analysis by immunostaining, histology, and RT-PCR confirmed that expression of Bi-Tet and DF systems did not affect ES cell self-renewal or pluripotency. ES cells were differentiated into beating embryoid bodies expressing cardiac markers such as troponin, Nkx2.5, and beta-MHC. Afterward, 5 x 10(5) cells obtained from these beating embryoid bodies or saline were injected into the myocardium of SV129 mice (n=36) following ligation of the left anterior descending (LAD) artery. Bioluminescence imaging (BLI) and echocardiography showed that VEGF(165) induction led to significant improvements in both transplanted cell survival and cardiac function (p<0.05). This is the first study to demonstrate imaging of embryonic stem cell-mediated gene therapy targeting cardiovascular disease. With further validation, this platform may have broad applications for current basic research and further clinical studies.
详细资料
- 关键词: Animals; Apoptosis; Cell Proliferation; Cell Survival; Doxycycline/pharmacology; Echocardiography/methods; Embryonic Stem Cells/*cytology; Gene Therapy/*methods; *Genetic Techniques; Green Fluorescent Proteins/metabolism; Heart/*physiology; Mice; Myocardium/*pathology; Promoter Regions, Genetic; Vascular Endothelial Growth Factor A/*metabolism
- 文献种类:期刊
- 期刊名称: Cloning and Stem Cells
- 期刊缩写: Cloning Stem Cells
- 期卷页: 2007年第9卷第4期 549-563页
- 地址: The Department of Radiology and Molecular Imaging Program at Stanford, Stanford University, Stanford, California, USA
- ISBN: 1536-2302
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