[1]余勇波,唐健,张桂龙,等.载BDNF基因PEG-PLGA纳米微球的制备及其转染外源性神经干细胞的研究[J].临床神经外科杂志,2019,16(2):109-114.[doi:10.3969/j.issn.1672-7770.2019.02.004]
 YU Yong-bo,TANG Jian,ZHANG Gui-long,et al.Preparation of PEG-PLGA-BDNF nanoparticles and exploratory study on transfection of exogenous neural stem cells[J].Journal of Clinical Neurosurgery,2019,16(2):109-114.[doi:10.3969/j.issn.1672-7770.2019.02.004]
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载BDNF基因PEG-PLGA纳米微球的制备及其转染外源性神经干细胞的研究()
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《临床神经外科杂志》[ISSN:1672-7770/CN:32-1727/R]

卷:
16
期数:
2019年第2期
页码:
109-114
栏目:
转化医学专题
出版日期:
2019-04-17

文章信息/Info

Title:
Preparation of PEG-PLGA-BDNF nanoparticles and exploratory study on transfection of exogenous neural stem cells
作者:
余勇波唐健张桂龙朱治翰杨芳陈陆馗
210009南京,东南大学附属中大医院神经外科(余勇波,张桂龙,陈陆馗);东南大学医学院(朱治翰);东南大学生物科学与医学工程学院,生物电子国家重点实验室,江苏省生物材料与器件重点实验室(唐健,杨芳)
Author(s):
YU Yong-bo TANG Jian ZHANG Gui-long et al.
Department of Neurosurgery, Zhongda Hospital, Southeast University, Nanjing 210009, China
关键词:
纳米微球神经干细胞基因治疗转染
Keywords:
nanoparticleneural stem cellsgene therapytransfection
分类号:
RQ78;Q291
DOI:
10.3969/j.issn.1672-7770.2019.02.004
文献标志码:
A
摘要:
【摘要】目的 构建转染后能表达神经修复功能蛋白的载脑源性神经营养因子(brain-derived neurotrophic factor,BDNF)基因的聚乙二醇-聚乳酸-羟基乙酸纳米微球[poly(ethylene glycol)-poly(lactin-co-glycolic acid)-BDNF-nanoparticles,PEG-PLGA-BDNF-NPs];探讨其转染外源性神经干细胞(neural stem cells,NSCs)后对细胞生物学功能的影响。方法采用W/O/W复乳溶剂挥发法制备装载BDNF基因质粒的PEG-PLGA纳米微球转染复合体。用扫描电镜和透射电镜表征纳米微球的形貌,粒度电位分析仪测量纳米微球的粒径及Zeta电位。测定纳米微球对BDNF基因质粒的包封率和载药量;检测PEG-PLGA纳米微球转染复合体的体外缓释质粒的情况。倒置显微镜观察纳米微球转染复合体对NSCs形态的影响;并用CCK-8法检测其对NSCs增殖的影响。荧光倒置显微镜观察NSCs胞吞纳米微球转染复合体后的分泌作用;Western-blot及RT-PCR检测其BDNF的分泌能力。结果 (1)制备了包载BDNF基因的PEG-PLGA纳米微球,微球的粒径约为200 nm,粒径均一;包封率及载药量均较高,具有一定的缓释效果。(2)分离培养出了具有不断增殖能力,表达神经巢蛋白的NSCs;并能经过诱导分化为神经元和神经胶质细胞。(3)载BDNF基因的PEG-PLGA纳米微球高效转染NSCs,且转染不会引起显著细胞毒性;Western-blot及RT-PCR检测显示其大量表达BDNF。结论采用W/O/W复乳溶剂挥发法制备得到安全高效的PEG-PLGA-BDNF纳米微球。并首次构建了装载PEG-PLGA-BDNFNPs,能表达具有神经修复功能的BDNF的外源性NSCs系。
Abstract:
Abstract: ObjectiveTo construct nano-gene microspheres(PEG-PLGA-BDNF-NPs) with the function of express nerve repair protein, and explore the effects of transfection on exogenous neural stem cells(NSCs) on their biological functions. MethodsPEG-PLGA nanoparticles transfected with BDNF gene plasmids were prepared by double emulsionsolvent evaporation method. The morphology of nanoparticles was observed by scanning electron microscopy and transmission electron microscope. The particle size and Zeta potential of nanoparticles were measured by laser particle size analyzer. The entrapment efficiency of PEG-PLGA nanoparticles and the drug loading of BDNF gene plasmids were examined. The in vitro release plasmid of PEG-PLGA nanoparticle transfection complex was tested. The biocompatibility of the nanoparticle transfection complex and the effect of the nanoparticle complex on the morphology of NSCs were detected by CCK-8. The secretion of neural stem cells after transfection of nanoparticles transfected complex was observed by inverted fluorescence microscope. Western blot and quantitative RT-PCR were used to detect the secretion of BDNF. Results (1) The gene-loaded PEG-PLGA nanoparticles were prepared. The encapsulation efficiency and drug loading were high, which had a certain sustained release effect. (2) The neural stem cells with the ability to proliferate and express the nestin protein are isolated and cultured, and can be induced to differentiate into neurons and glial cells. (3) PEG-PLGA nanoparticles containing BDNF gene were highly transfected into neural stem cells, and transfection did not cause significant cytotoxicity. Western-blot and RT-PCR showed that BDNF was abundantly expressed in these stem cells. ConclusionsThe rat neural stem cells were successfully extracted and cultured, and the PEG-PLGA-BDNF-NPs were prepared. An exogenous neural stem cell line loaded with PEG-PLGA-BDNF-NPs was first constructed to express BDNF protein with neurorestorative function.

相似文献/References:

[1]刘友,刘希光,李爱民,等.悬浮培养与贴壁培养方式神经干细胞分化表型特点[J].临床神经外科杂志,2017,14(05):385.
 LIU You,LIU Xi-guang,LI Ai-min..Neural stem cells and phenotypic characteristics of suspension culture and adherent culture[J].Journal of Clinical Neurosurgery,2017,14(2):385.
[2]李世安,刘友,刘希光,等.神经干细胞与神经元差异表达基因的生物信息学分析[J].临床神经外科杂志,2017,14(06):406.
 LI Shi-an,LIU You,LIU Xi-guang,et al.Bioinformatics analysis of differentially expressed genes in neural stem cells and neurons[J].Journal of Clinical Neurosurgery,2017,14(2):406.

更新日期/Last Update: 2019-04-17