Genetic engineering with endothelial nitric oxide synthase improves functional properties of endothelial progenitor cells from patients with coronary artery disease: an in vitro study
Genetic engineering with endothelial nitric oxide synthase improves functional properties of endothelial progenitor cells from patients with coronary artery disease: an in vitro study
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DOI：
10.1007/s00395-009-0039-x
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发表时间：
2009-11-01
影响因子：
9.5
通讯作者：
Kartha, Chandrasekharan Cheranellore
中科院分区：
文献类型：
--
作者：
Kaur, Savneet;Kumar, T. R. Santhosh;Kartha, Chandrasekharan Cheranellore
最近的研究报道了冠状动脉疾病（CAD）患者内皮祖细胞（EPCs）的数量和功能明显受损。鉴于eNOS在血管生成中的重要作用，在本研究中，我们评估了eNOS基因转移对从CAD患者中分离的离体扩增的EPCs的影响。用脂质体法将含有人eNOS全长基因的真核表达载体pcDNA3.1-eNOS转染扩增的EPCs。与未转染的EPCs相比，约35-40%的eNOS-EPCs具有更高的eNOS表达。转染pcDNA3.0-EGFP（表达绿色荧光蛋白的质粒载体）的EPCs作为对照。比较了未转染、GFP转染和eNOS转染的EPCs在血管生成的重要功能属性方面的差异，如增殖、迁移、分化和粘附/整合成体外管状结构。功能研究表明，在特定的生长条件下，与未转染和GFP转染的细胞相比，来自CAD患者的eNOS-EPCs的[H-3]胸苷标记的DNA显著增加（P < 0.01），迁移（14.6 +/- A 1.8和16.5 +/- A 1.9对比23.5 +/- A 3.4细胞/视野，P < 0.01），分化为内皮样梭形细胞的能力（46 +/- A 4.5和56.5 +/- A 2.1对93.2 +/- A 6.6细胞/视野，P < 0.001），并且还掺入基质胶上的管状结构中。（GFP-EPCs：21.25 +/-Δ 2.9对比GFP-eNOS-EPCs：34.5 +/-Δ 5.5细胞/视野，P < 0.05）。我们的结论是，eNOS基因转染是一种有价值的方法，以增强血管生成特性的离体扩增的EPCs和eNOS修饰的EPCs可能提供显着的优势，比EPCs单独在其临床应用中与心肌缺血的患者。
Recent studies have reported a marked impairment in the number and functions of endothelial progenitor cells (EPCs) in patients with coronary artery disease (CAD). In view of an important role of eNOS in angiogenesis, in the present study, we evaluated the effects of eNOS gene transfer in ex vivo expanded EPCs isolated from patients with CAD. The expanded EPCs were transfected with mammalian expression vector pcDNA3.1-eNOS containing the full-length human eNOS gene using lipofectamine. About 35-40% of the eNOS-EPCs had higher expression of eNOS as compared to untransfected EPCs. EPCs transfected with pcDNA3.0-EGFP, the plasmid vector expressing green fluorescent protein (GFP) were used as control. The untransfected, GFP-transfected and eNOS-transfected EPCs were compared in terms of important functional attributes of angiogenesis such as proliferation, migration, differentiation and adhesion/integration into tube-like structures in vitro. Functional studies revealed that in the presence of defined growth conditions, compared to the untransfected and GFP-transfected cells, eNOS-EPCs from patients with CAD have a significant increase in [H-3] thymidine-labeled DNA (P < 0.01), migration (14.6 +/- A 1.8 and 16.5 +/- A 1.9 vs. 23.5 +/- A 3.4 cells/field, P < 0.01), ability to differentiate into endothelial-like spindle-shaped cells (46 +/- A 4.5 and 56.5 +/- A 2.1 vs. 93.2 +/- A 6.6 cells/field, P < 0.001) and also incorporation into tube-like structures on the matrigel (GFP-EPCs: 21.25 +/- A 2.9 vs. GFP-eNOS-EPCs: 34.5 +/- A 5.5 cells/field, P < 0.05). We conclude that eNOS gene transfection is a valuable approach to augment angiogenic properties of ex vivo expanded EPCs and eNOS-modified EPCs may offer significant advantages than EPCs alone in terms of their clinical use in patients with myocardial ischemia.