聚苯丙氨酸修饰低分子量聚乙烯亚胺制备高效基因载体

林琳; 郭兆培; 陈杰; 田华雨; 陈学思

doi:10.11777/j.issn1000-3304.2017.16277

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聚苯丙氨酸修饰低分子量聚乙烯亚胺制备高效基因载体

研究论文 | 更新时间：2021-05-18

- 聚苯丙氨酸修饰低分子量聚乙烯亚胺制备高效基因载体
- Synthesis and Characterization of Polyphenylalanine Grafted Low Molecular Weight PEI as Efficient Gene Carriers
- 高分子学报 2017年第2期页码：321-328
- 作者机构：
  
  中国科学院长春应用化学研究所中国科学院生态环境高分子材料重点实验室长春 130022
- 作者简介：
  
  田华雨, E-mail: thy@ciac.ac.cn Tian Hua-yu, E-mail: thy@ciac.ac.cn
  陈学思, E-mail: xschen@ciac.ac.cn Chen Xue-si, E-mail: xschen@ciac.ac.cn
- 基金信息：
  
  国家自然科学基金(51390484);国家自然科学基金(51520105004);国家自然科学基金(21474104);国家自然科学基金(51233004);中共中央组织部“万人计划”青年拔尖人才项目和吉林省重点科技攻关项目(20160204032GX)
- DOI：10.11777/j.issn1000-3304.2017.16277
  中图分类号：
- 纸质出版日期：2017-2-20，
  
  收稿日期：2016-9-11，
  
  修回日期：2016-10-24，
扫描看全文
林琳, 郭兆培, 陈杰, 田华雨, 陈学思. 聚苯丙氨酸修饰低分子量聚乙烯亚胺制备高效基因载体[J]. 高分子学报, 2017,(2):321-328.

Lin Lin, Zhao-pei Guo, Jie Chen, Hua-yu Tian, Xue-si Chen. Synthesis and Characterization of Polyphenylalanine Grafted Low Molecular Weight PEI as Efficient Gene Carriers[J]. Acta Polymerica Sinica, 2017,(2):321-328.
林琳, 郭兆培, 陈杰, 田华雨, 陈学思. 聚苯丙氨酸修饰低分子量聚乙烯亚胺制备高效基因载体[J]. 高分子学报, 2017,(2):321-328. DOI： 10.11777/j.issn1000-3304.2017.16277.

Lin Lin, Zhao-pei Guo, Jie Chen, Hua-yu Tian, Xue-si Chen. Synthesis and Characterization of Polyphenylalanine Grafted Low Molecular Weight PEI as Efficient Gene Carriers[J]. Acta Polymerica Sinica, 2017,(2):321-328. DOI： 10.11777/j.issn1000-3304.2017.16277.

摘要

分别制备了以支化小分子量聚乙烯亚胺（PEI-1.8k）为引发剂，引发苯丙氨酸-NCA开环聚合得到聚乙烯亚胺-聚苯丙氨酸（PEI1.8k-

-PPhe）以及聚乙烯亚胺接枝苯丙氨酸单体（PEI1.8k-

-Phe）的系列基因载体材料.利用核磁、粒度、zeta电位仪、荧光光度计、流式细胞仪以及激光共聚焦显微镜对PEI1.8k-

-PPhe，PEI1.8k-

-Phe以及PEI1.8k-

-PPhe/DNA和PEI1.8k-

-Phe/DNA复合物颗粒进行了系统的表征.研究结果表明，最佳转染条件下，PEI1.8k-

-PPhe10/DNA复合物颗粒的粒径约为150 nm，表面电位约为16 mV.在人源宫颈癌（HeLa）和人源乳腺癌（MCF-7）2种细胞系中均具有较高的基因转染效率，且最佳转染效率可达到PEI-25k的12倍.MTT细胞毒性实验分别比较了PEI1.8k-

-PPhe和PEI1.8k-

-Phe对HeLa细胞毒性的大小.从实验结果可见，苯丙氨酸引入的方式及数量决定着其细胞毒性的大小.PEI1.8k-

-PPhe和PEI1.8k-

-Phe都具有较低的细胞毒性（材料在较高浓度1 mg/mL时的细胞存活率大于70%）.内吞实验结果表明，PEI1.8k-

-PPhe由于接入了具有规则聚合链的聚苯丙氨酸，而易于被HeLa细胞内吞.PEI1.8k-

-PPhe10/DNA复合物颗粒相比于PEI-25k/DNA，PEI-1.8k/DNA和PEI1.8k-

-PPhe/DNA具有更高的细胞内吞效率.

Abstract

To search safe and evaluate non-viral nucleic acids carrier

a series of polyphenylalanine and phenylalanine grafted low molecular weight PEI with a molecular weight of 1.8×10

(PEI1.8k-

-PPhe and PEI1.8k-

-Phe) were prepared by NCA ring opening polymerization initiated by PEI-1.8k and phenylalanines conjugation to PEI-1.8k

respectively. The polymers and the complexes of PEI1.8k-

-PPhe/DNA and PEI1.8k-

-Phe/DNA were characterized by nuclear magnetic resonance (NMR) analysis

particle size analysis

zeta potential analysis

luciferase analysis

flow cytometry (FCM) analysis and confocal laser scan microscopy (CLSM). In order to compare the property for compacting DNA into nanoparticles

the particle size and zeta potential analysis were carried out. Both PEI1.8k-

-PPhe and PEI1.8k-

-Phe showed suitable particle size and zeta potential for gene delivery. The particle size of PEI1.8k-

-PPhe10/DNA complexes were about 150 nm and the zeta potentials were about +16 mV

which were suitable for the

in vitro

experiments. Moreover

cell viability

after treating with different copolymers at various concentrations

was studied by an MTT assay. The reduced cytotoxicity of PEI1.8k-

-PPhe and PEI1.8k-

-Phe may be because of the introduction of neutral hydrophobic phenylalanine moieties. And both of the PEI1.8k-

-PPhe and PEI1.8k-

-Phe had lower cytotoxicity (above 70% viability at a higher concentration 1 mg/mL) than that of PEI with a molecular weight of 2.5×10

(PEI-25k) in HeLa cells. The

in vitro

gene transfection of PEI1.8k-

-PPhe10 and PEI1.8k-

-Phe10 was conducted in human cervical cancer (HeLa) and breast cancer (MCF-7) cells. In both of the cells

PEI1.8k-

-PPhe10 exhibited much higher gene transfection efficiency. PEI1.8k-

-PPhe10/DNA complexes showed remarkable gene transfection efficiency

which was about twelve times higher than that of PEI-25k. The endocytosis efficiency of PEI1.8k-

-PPhe10/DNA and PEI1.8k-

-Phe10/DNA were quantified using flow cytometry. Due to their regular polymer chain

PEI1.8k-

-PPhe10/Cy5-DNA showed better internalization efficiency than PEI1.8k-

-Phe10/Cy5-DNA

PEI-25k/Cy5-DNA and PEI-1.8k/Cy5-DNA. The CLSM assay was carried out to verify the internalization efficiency together which also indicated that PEI1.8k-

-PPhe10/Cy5-DNA induced higher intracellular uptake efficiency than the others.

关键词

聚苯丙氨酸低分子量聚乙烯亚胺基因载体基因转染

Keywords

PolyphenylalanineLow molecular weight PEIGene carriersGene transfection

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