Development of Endothelial-Specific Single Inducible Lentiviral Vectors for Genetic Engineering of Endothelial Progenitor Cells.
Development of Endothelial-Specific Single Inducible Lentiviral Vectors for Genetic Engineering of Endothelial Progenitor Cells.
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DOI：
10.1038/srep17166
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发表时间：
2015-11-27
影响因子：
4.6
通讯作者：
Qian C
中科院分区：
文献类型：
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作者：
Yang G;Kramer MG;Fernandez-Ruiz V;Kawa MP;Huang X;Liu Z;Prieto J;Qian C
内皮祖细胞（Endothelial progenitor cells， EPC）能够迁移到肿瘤血管。这些细胞，如果经过基因改造，可以用作向肿瘤输送有毒物质或表达抗癌蛋白的载体。为了验证这一假设，我们开发了几种单一的、内皮特异性的、强西环素诱导的自我灭活（SIN）慢病毒载体。将两个不同的表达盒插入到sin载体中：一个由内皮细胞特异性的小鼠血管内皮钙粘蛋白（mVEcad）启动子控制，用于表达反激活子rtTA2S-M2；另一个由萤火虫荧光素酶报告基因的诱导启动子TREalb驱动。我们比较了荧光素酶在不同载体结构中的表达水平，包含与载体序列相同或相反的方向。结果表明，将这两个表达盒放置在相反方向的载体是最佳的，其特点是在强力霉素存在的情况下，在几种内皮细胞系中诱导转基因表达（17.7- 73倍），但在未诱导的情况下没有泄漏。综上所述，我们已经建立了内皮细胞特异性的单诱导SIN慢病毒载体。这种慢病毒载体可以用来赋予内皮祖细胞抗肿瘤的特性。
Endothelial progenitor cells (EPC) are able to migrate to tumor vasculature. These cells, if genetically modified, can be used as vehicles to deliver toxic material to, or express anticancer proteins in tumor. To test this hypothesis, we developed several single, endothelial-specific, and doxycycline-inducible self-inactivating (SIN) lentiviral vectors. Two distinct expression cassettes were inserted into a SIN-vector: one controlled by an endothelial lineage-specific, murine vascular endothelial cadherin (mVEcad) promoter for the expression of a transactivator, rtTA2S-M2; and the other driven by an inducible promoter, TREalb, for a firefly luciferase reporter gene. We compared the expression levels of luciferase in different vector constructs, containing either the same or opposite orientation with respect to the vector sequence. The results showed that the vector with these two expression cassettes placed in opposite directions was optimal, characterized by a robust induction of the transgene expression (17.7- to 73-fold) in the presence of doxycycline in several endothelial cell lines, but without leakiness when uninduced. In conclusion, an endothelial lineage-specific single inducible SIN lentiviral vector has been developed. Such a lentiviral vector can be used to endow endothelial progenitor cells with anti-tumor properties.