Centro de Investigación Cardiovascular, CSIC-ICCC, IIB-Sant Pau, Barcelona, Spain.
Hum Mol Genet. 2013 May 15;22(10):1949-59. doi: 10.1093/hmg/ddt042. Epub 2013 Feb 5.
We have previously shown that NOR-1 (NR4A3) modulates the proliferation and survival of vascular cells in culture. However, in genetically modified animal models, somewhat conflicting results have been reported concerning the involvement of NOR-1 in neointimal formation after vascular injury. The aim of this study was to generate a transgenic mouse model over-expressing NOR-1 in smooth muscle cells (SMCs) and assess the consequence of a gain of function of this receptor on intimal hyperplasia after vascular injury. The transgene construct (SM22-NOR1) was prepared by ligating the full-length human NOR-1 cDNA (hNOR-1) and a mouse SM22α minimal promoter able to drive NOR-1 expression to SMC. Two founders were generated and two stable transgenic mouse lines (TgNOR-1) were established by backcrossing the transgene-carrying founders with C57BL/6J mice. Real-time PCR and immunohistochemistry confirmed that hNOR-1 was mainly targeted to vascular beds such as aorta and carotid arteries, and was similar in both transgenic lines. Vascular SMC from transgenic animals exhibit increased NOR-1 transcriptional activity (assessed by electrophoretic mobility shift assay and luciferase assays), increased mitogenic activity (determined by [(3)H]-thymidine incorporation; 1.58-fold induction, P < 0.001) and increased expression of embryonic smooth muscle myosin heavy chain (SMemb) than wild-type cells from control littermates. Using the carotid artery ligation model, we show that neointima formation was increased in transgenic versus wild-type mice (2.36-fold induction, P < 0.01). Our in vivo data support a role for NOR-1 in VSMC proliferation and vascular remodelling. This NOR-1 transgenic mouse could be a useful model to study fibroproliferative vascular diseases.
我们之前的研究表明，NOR-1（NR4A3）调节培养中的血管细胞的增殖和存活。然而，在基因修饰的动物模型中，关于 NOR-1 在血管损伤后的新生内膜形成中的参与，报告了一些相互矛盾的结果。本研究的目的是生成一种在平滑肌细胞（SMC）中过表达 NOR-1 的转基因小鼠模型，并评估该受体功能获得对血管损伤后内膜增生的影响。转基因构建体（SM22-NOR1）通过连接全长人 NOR-1 cDNA（hNOR-1）和一个能够驱动 NOR-1 表达到 SMC 的小鼠 SM22α 最小启动子来制备。生成了两个启动子，并通过将携带转基因的启动子与 C57BL/6J 小鼠回交，建立了两个稳定的转基因小鼠系（TgNOR-1）。实时 PCR 和免疫组织化学证实，hNOR-1 主要靶向血管床，如主动脉和颈动脉，并且在两个转基因系中相似。来自转基因动物的血管 SMC 显示出增加的 NOR-1 转录活性（通过电泳迁移率变动分析和荧光素酶测定评估）、增加的有丝分裂活性（通过[(3)H]-胸苷掺入确定；1.58 倍诱导，P < 0.001）和胚胎平滑肌肌球蛋白重链（SMemb）的表达增加，与来自对照同窝仔的野生型细胞相比。使用颈动脉结扎模型，我们表明，与野生型小鼠相比，转基因小鼠的新生内膜形成增加（2.36 倍诱导，P < 0.01）。我们的体内数据支持 NOR-1 在 VSMC 增殖和血管重塑中的作用。这种 NOR-1 转基因小鼠可能是研究纤维增殖性血管疾病的有用模型。
