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[用新型阳离子基因载体聚乙烯亚胺-聚(γ-苄基-L-谷氨酸酯)进行转染]

[Transfection with a novel cationic gene carrier: PEI-PBLG].

作者信息

Wei Ji-Zheng, Lin Lei, Xiong Wei, Zhu Qing-Yu

机构信息

State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, China.

出版信息

Sheng Wu Gong Cheng Xue Bao. 2007 Mar;23(2):229-34.

Abstract

This study mainly deals with cell transfection and cytotoxicity for PEI(10kD)-PBLG, a novel cationic copolymer, to observe its potential as a gene carrier. Size measurement and SEM were used to show the modality of the PEI-PBLG/pDNA complexes. Cytotoxicity of PEI (10kD)-PBLG was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay and compared with PEI(25kD)-PBLG, PEI(10kD), and PEI(25kD). Furthermore, pEGFP that can express the enhanced green fluorescent protein was chosen as a reporter to observe the transfection efficiency directly. Then, PEI (10kD)-PBLG/pEGFP complexes were transfected into several cell lines, such as Hela, COS-7, Vero-E6, and ECV-304, and effects of the transfection conditions were evaluated. The efficiencies were measured by FACS. Size measurement of complex particles indicated that PEI-PBLG/pDNA tended to form smaller nanoparticles compared with PEI/pDNA. The representative size of the PEI(10kD)-PBLG/pDNA complex was approximately 100 - 200 nm. SEM images showed that the particles were condense and compact. This can be suitable for their entry into cells. Cytotoxicity studies suggested that PEI (10kD)-PBLG had considerably lower toxicity than the other three materials. In the transfection tests, PEI (10kD)-PBLG/pDNA complexes could be transfected into all the cell lines that were tested. These provided the highest level of EGFP expression (45.02%) in Hela cells, which was considerably higher than that of PEI(10kD)/pEGFP (29.16%). Being less affected by the serum during transfection, PEI-PBLG/pDNA complexes offered greater biocompatibility than PEI. PEI-PBLG copolymer reduces the cytotoxicity of PEI, improves the transfection efficiency, and offers greater biocompatibility than PEI. It shows considerable potential as an efficient nonviral carrier for gene delivery.

摘要

本研究主要探讨新型阳离子共聚物PEI(10kD)-PBLG的细胞转染及细胞毒性,以观察其作为基因载体的潜力。通过尺寸测量和扫描电子显微镜(SEM)来展示PEI-PBLG/pDNA复合物的形态。采用3-(4,5-二甲基噻唑-2)-2,5-二苯基溴化四氮唑(MTT)比色法评估PEI(10kD)-PBLG的细胞毒性,并与PEI(25kD)-PBLG、PEI(10kD)和PEI(25kD)进行比较。此外,选择可表达增强型绿色荧光蛋白的pEGFP作为报告基因,直接观察转染效率。然后,将PEI(10kD)-PBLG/pEGFP复合物转染至多种细胞系,如Hela、COS-7、Vero-E6和ECV-304,并评估转染条件的影响。通过流式细胞仪(FACS)测定效率。复合物颗粒的尺寸测量表明,与PEI/pDNA相比,PEI-PBLG/pDNA倾向于形成更小的纳米颗粒。PEI(10kD)-PBLG/pDNA复合物的典型尺寸约为100 - 200 nm。SEM图像显示颗粒凝聚且致密。这有利于它们进入细胞。细胞毒性研究表明,PEI(10kD)-PBLG的毒性远低于其他三种材料。在转染试验中,PEI(10kD)-PBLG/pDNA复合物可转染至所有测试的细胞系。这些复合物在Hela细胞中提供了最高水平的EGFP表达(45.02%),显著高于PEI(10kD)/pEGFP(29.16%)。在转染过程中受血清影响较小,PEI-PBLG/pDNA复合物比PEI具有更高的生物相容性。PEI-PBLG共聚物降低了PEI的细胞毒性,提高了转染效率,并且比PEI具有更高的生物相容性。它作为一种高效的非病毒基因递送载体显示出相当大的潜力。

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