权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Regulation of endothelial cell functions by gene transfer using viral vectors

使用病毒载体进行基因转移调节内皮细胞功能

基本信息

批准号：
12670687
负责人：
MIMURO Jun
金额：
$ 2.5万
依托单位：
Jichi Medical School
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12670687/
关键词：
endothelial cell thrombosis thrombomodulin adeno-associated virus vector gene therapy plasminogen activator inhibitor 1 AAV vector Thrombosis

项目摘要

We could facilitate plasminogen activator inhibitor 1 (PAI-1) promoter activity approximately by 20-fold using an enhancer element. This enhanced PAI- 1 promoter has a strong basal activity, comparable to CAG promoter activity, and has a response similar to the PAI-1 promoter with respect to TGF β1 and TNF α stimulation. The characteristic of the enhanced PAI-1 promoter is thought to be suited to timely and tissue specific expression of anticoagulant molecules in the vascular cells. Thus, we developed recombinant adeno-associated virus (rAAV) vectors using the enhanced PAI-1 promoter and were successful in transducing vascular endothelial cells to express the thrombomodulin transgene under the regulation of the enhanced PAI-1 promoter in vitro. Thromobomodulin transgene expression driven by the enhanced PAI- 1 promoter in rAAV vector-transduced cultured endothelial cells was 600〜1000-fold higher than constitutive thrombomodulin gene expression in cultured human umbilical vein endothelial cells and was up-regulated by TGFβ1 and TNFα stimulation which may down-regulate endogenous thrombomodulin gene expression in endothelial cells. The brain vascular endothelial cells of Mongolian gerbils also could be transduced by the same rAAV vector in vivo. Transition of endothelial cells by rAAV vectors to express enhanced PAI- 1 promoter-driven transgenes may be a useful gene therapy approach for vascular diseases.

我们可以使用增强元件将纤溶酶原激活物抑制剂1 （PAI-1）的启动子活性提高约20倍。这种增强的PAI-1启动子具有很强的基础活性，与CAG启动子活性相当，并且对TGF β1和TNF α的刺激具有与PAI-1启动子相似的反应。PAI-1启动子增强的特性被认为适合于血管细胞中抗凝血分子的及时和组织特异性表达。因此，我们利用增强型PAI-1启动子构建了重组腺相关病毒（rAAV）载体，并成功地在体外诱导血管内皮细胞在增强型PAI-1启动子的调控下表达血栓调节蛋白转基因。在rAAV载体介导的体外培养的内皮细胞中，PAI- 1启动子增强驱动的凝血调节蛋白转基因表达量比体外培养的人脐静脉内皮细胞中的组成型凝血调节蛋白基因表达量高600 ~ 1000倍，TGFβ1和TNFα刺激可上调凝血调节蛋白基因表达，这可能下调内皮细胞内源性凝血调节蛋白基因的表达。同样的rAAV载体也能在体内转染蒙古沙鼠的脑血管内皮细胞。rAAV载体转染内皮细胞表达PAI- 1启动子驱动的基因可能是一种有效的血管疾病基因治疗方法。