Engineering of vascular grafts with genetically modified bone marrow mesenchymal stem cells on poly (propylene carbonate) graft.
Engineering of vascular grafts with genetically modified bone marrow mesenchymal stem cells on poly (propylene carbonate) graft.
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DOI：
10.1111/j.1525-1594.2006.00322.x
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发表时间：
2006-12
影响因子：
2.4
通讯作者：
Jun Zhang;Hongxu Qi;Hongjun Wang;P. Hu;L. Ou;Shuhua Guo;Jing Li;Yongzhe Che;Yao-ting Yu;D. Kong
中科院分区：
文献类型：
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作者：
Jun Zhang;Hongxu Qi;Hongjun Wang;P. Hu;L. Ou;Shuhua Guo;Jing Li;Yongzhe Che;Yao-ting Yu;D. Kong
骨髓间充质干细胞（MSCs）具有多向分化潜能，有望成为血管组织工程的另一种细胞来源。本研究的目的是通过将遗传修饰的MSC接种和培养到由静电纺丝技术产生的合成聚合物支架上来创建小直径血管。通过静电纺丝聚（碳酸亚丙酯）（PPC）的管状支架（直径为2 mm）的非织造纤维的微观结构产生。将从骨髓中获得的大鼠MSC在培养中扩增并用血管保护基因内皮一氧化氮合酶（eNOS）或标记基因绿色荧光蛋白（GFP）修饰。将这些MSC接种到电纺纤维移植物（内径= 2 mm）上，并在37 ℃下在5%CO（2）中培养。用扫描电镜（SEM）和苏木精-伊红（H&E）染色分析支架中MSC的生长。采用荧光激活细胞分选（FACS）、免疫组化、逆转录-聚合酶链反应（RT-PCR）和western blot检测转基因细胞的基因转移和转基因表达。用NO检测试剂盒测定工程化血管产生的一氧化氮（NO）。我们的数据显示，种子细胞与支架的微纤维整合形成三维细胞网络，表明这种合成PPC支架与MSC之间存在有利的相互作用。用浓缩的逆转录病毒进行基因转染，获得了较高的转染效率。RT-PCR和免疫组化染色检测eNOS基因转录产物和eNOS蛋白表达。与天然血管相比，植入eNOS修饰的MSC的移植物产生的NO量相当，并且显著高于植入未修饰的MSC的移植物。综上所述，eNOS基因修饰的MSC在电纺管状支架上培养产生的血管移植物在功能方面显示出有希望的结果。在血管组织工程中使用MSCs和治疗基因可能有助于改善血管再生和通畅性。
Bone marrow mesenchymal stem cells (MSCs) have demonstrated their pluripotency to differentiate into different cell lineages and may be an alternative cell source for vascular tissue engineering. The objective of this study is to create small diameter vessels by seeding and culture of genetically modified MSCs onto a synthetic polymer scaffold produced by an electrospinning technique. A tubular scaffold (2 mm in diameter) with a microstructure of nonwoven fibers was produced by electrospinning of poly (propylene carbonate) (PPC). Rat MSCs obtained from bone marrow were expanded in culture and modified with vasculoprotective gene endothelial nitric oxide synthase (eNOS) or marker gene green fluorescent protein (GFP). These MSCs were seeded onto the electrospun fibrous grafts (internal diameter = 2 mm), and cultured in 5% CO(2) at 37 degrees C. The growth of MSCs in the scaffold was analyzed with scanning electron microscopy (SEM) and hematoxylin and eosin (H&E) staining. The gene transfer and transgenic gene expression were examined with fluorescence-activated cell sorting (FACS), immunochemical staining, reverse transcriptase-polymerase chain reaction (RT-PCR), and western blot. The production of nitric oxide (NO) by the engineered vessels was measured with an NO detection kit. Our data showed that the seeded cells integrated with the microfibers of the scaffold to form a three-dimensional cellular network, indicating a favorable interaction between this synthetic PPC scaffold with MSCs. High transduction efficiency was obtained with the use of concentrated retrovirus in the gene transfection of MSCs. The eNOS gene transcripts and protein were detected in the grafts seeded with eNOS-modified MSCs by RT-PCR and immunochemical staining. The amount of NO produced by grafts seeded with eNOS-modified MSCs was comparable to that produced by native blood vessels, and it was significantly higher than that in the grafts seeded with nonmodified MSCs. In summary, the vascular graft produced by culture of eNOS gene-modified MSCs onto the electrospun tubular scaffolds shows promising results in terms of function. The use of MSCs and therapeutic genes in tissue engineering of blood vessels could be helpful in improving vessel regeneration and patency.