乙酰普鲁兰纳米粒子BeWo b30细胞毒性及摄取研究

doi:10.3969/j.issn.1006-7795.2016.04.002

首都医科大学学报 ›› 2016, Vol. 37 ›› Issue (4): 418-423.doi: 10.3969/j.issn.1006-7795.2016.04.002

乙酰普鲁兰纳米粒子BeWo b30细胞毒性及摄取研究

蒋子雯¹, 周志敏², 唐红波¹, 杜博², 张其清², 代荫梅¹

1. 首都医科大学附属北京妇产医院妇科, 北京 100026;
2. 中国医学科学院北京协和医学院生物医学工程研究所, 天津市生物医学材料重点实验室, 天津 300192

收稿日期:2016-06-15 出版日期:2016-08-21 发布日期:2016-07-18
通讯作者: 代荫梅 E-mail:fcyydym@163.com
基金资助:
国家自然科学基金（81301322），天津市应用基础与前沿技术研究计划（15JCQNJC14400）。

Cell toxicity and cell uptake study of pullulan acetate nanoparticles to BeWo b30 cells

Jiang Ziwen¹, Zhou Zhimin², Tang Hongbo¹, Du bo², Zhang Qiqing², Dai Yinmei¹

1. Department of Gynecology, Beijing Obstetrics and Gynecology Hospital, Capital Medical University, Beijing 100026, China;
2. Tianjin Key Laboratory of Biomedical Materials, Institute of Biomedical Engineering, Chinese Academy of Medical Sciences & Peking Union Medical College, Tianjin 300192, China

Received:2016-06-15 Online:2016-08-21 Published:2016-07-18
Supported by:
This study was supported by National Natural Science Foundation of China(81301322),Tianjin Research Program of Application Foundation and Advanced Technology(15JCQNJC14400).

摘要/Abstract

摘要： 目的通过乙酰普鲁兰纳米粒子对人绒毛膜癌细胞（BeWo b30）的细胞毒性以及细胞摄取研究，为普鲁兰基纳米药物跨胎盘屏障机制及妊娠期用药提供科学依据。方法以透析法制备异硫氰酸荧光素标记的乙酰普鲁兰纳米粒子（fluorescein isothiocyanate labelled pullulan acetate nanoparticles， PA-FITC NPs），分别利用激光粒度分析仪、扫描电镜、透射电镜考察乙酰普鲁兰纳米粒子粒径、Zeta电位、形貌及结构；用细胞计数试剂盒-8（cell counting kit-8， CCK-8）检测细胞生长曲线，考察不同浓度纳米粒子（0.4～2.0 mg/mL）对BeWo b30细胞的细胞毒性；考察纳米粒子浓度、孵育时间、孵育温度对BeWo b30细胞摄取纳米粒子的影响。结果 PA-FITC纳米粒子水合直径为（348.0±114.3）nm，Zeta电位为（-26.0±5.1）mV，呈表面光滑、内部结构规整的球形。PA-FITC纳米粒子在0.4～2.0 mg/mL 浓度范围内细胞存活率为95.0%～100.5%，差异无统计学意义（P>0.05）；BeWo b30细胞摄取实验表明:细胞吞噬纳米粒子的量随着PA-FITC NPs的浓度升高和孵育时间延长而增大；当孵育温度从37℃降低至4℃，PA-FITC NPs的细胞摄取量显著降低（P<0.05）。结论乙酰普鲁兰纳米粒子呈球形，对BeWo b30细胞无明显细胞毒性；BeWo b30细胞摄取PA-FITC 纳米粒子与纳米粒子浓度、孵育时间和孵育温度呈正相关。

关键词: 普鲁兰, BeWo b30细胞, 纳米药物, 妊娠期用药

Abstract: Objective To provide evidence for application in obstetrics field of drugs during pregnancy and the mechanism of pullulan-base nanoparticles across the placental barrier by investigating the cell uptake of pullulan acetate nanoparticles and the cell toxicity of pullulan acetate nanoparticles to BeWo b30 cells. Methods The fluorescein isothiocyanate labelled pullulan acetate nanoparticles (PA-FITC NPs) were prepared by using dialysis method. The methods to investigate the characteristics involved particle size and distribution, zeta potential and morphology of nanoparticles using dynamic light scattering (DLS) and transmission electron microscopy. The growth curve of BeWo b30 and the cell toxicity of different concentration of PA-FITC NPs were studied by cell counting kit-8. The influence of different NPs concentration, incubation time and incubation temperature on cell up-taking were observed in this study.Results The prepared pullulan acetate nanoparticles are solid and spherical in shape. The mean diameter of PA-FITC NPs by DLS were (348.0±114.3) nm and the Zeta potential was (-26.0±5.1) mV. Cell viability in all concentration groups have no significant difference compared with the control group (P>0.05); higher PA-FITC NPs concentration and longer incubation time showed an increased cell uptake; cell uptake was decreased significantly after the temperature was reduced to 4℃. Conclusion The results demonstrated that there was no obvious toxic effect of pullulan acetate nanoparticles on BeWo b30 cells; There is a positive correlation between cell uptake and the concentration, incubation time and environment temperature of PA-FITC NPs.

Key words: pullulan, BeWo b30, nanomedicine, drug delivery in pregnancy

中图分类号:

R318

蒋子雯, 周志敏, 唐红波, 杜博, 张其清, 代荫梅. 乙酰普鲁兰纳米粒子BeWo b30细胞毒性及摄取研究[J]. 首都医科大学学报, 2016, 37(4): 418-423.

Jiang Ziwen, Zhou Zhimin, Tang Hongbo, Du bo, Zhang Qiqing, Dai Yinmei. Cell toxicity and cell uptake study of pullulan acetate nanoparticles to BeWo b30 cells[J]. Journal of Capital Medical University, 2016, 37(4): 418-423.

参考文献

[1] Tang H, Feng X, Zhang T, et al. Stability, pharmacokinetics, biodistributionand safety assessment of folate-conjugated pullulan acetate nanoparticles as cervical cancer targeted drug carriers [J]. J Nanosci Nanotechnol, 2015,15(9):6405-6412.
[2] Zhou Z, Anselmo A C, Mitragotri S. Synthesis of protein-based, rod-shaped particles from spherical templates using layer-by-layer assembly [J]. Adv Mater,2013, 25(19):2723-2727.
[3] Singh R S, Kaur N, Kennedy J F. Pullulan and pullulan derivatives as promising biomolecules for drug and gene targeting [J]. Carbohydr Polym, 2015,123:190-207.
[4] Tang H B, Li L, Chen H. Stability and in vivo evaluation of pullulan acetate as a drug nanocarrier [J]. Drug Deliv, 2010,17(7):552-558.
[5] Wang Y, Chen H, Liu Y, et al. pH-sensitive pullulan-based nanoparticle carrier of methotrexate and combretastatin A4 for the combination therapy against hepatocellular carcinoma [J]. Biomaterials, 2013,34(29):7181-7190.
[6] Zhang C, An T, Wang D, et al. Stepwise pH-responsive nanoparticles containing charge-reversible pullulan-based shells and poly (β-amino ester)/poly(lactic-co-glycolic acid) cores as carriers of anticancer drugs for combination therapy on hepatocellular carcinoma [J]. J Control Release, 2016,226:193-204.
[7] Avery M L, Meek C E, Audus K L. The presence of inducible cytochrome P450 types 1A1 and 1A2 in the BeWo cell line [J]. Placenta, 2003,24(1):45-52.
[8] Bode C J, Jin H, Rytting E, et al. In vitro models for studying trophoblast transcellular transport [J]. Methods Mol Med, 2006,122:225-239.
[9] 唐红波,周志敏,闫素英,等. 纳米递药系统胚胎毒性研究进展 [J]. 药物不良反应杂志, 2015,17(1):44-45.
[10] Fujitani T, Ohyama K, Hirose A, et al. Teratogenicity of multiwall carbon nanotube (MWCNT) in ICR mice [J]. J Toxicol Sci, 2012,37(1):81-89.
[11] Yamashita K, Yoshioka Y, Higashisaka K, et al. Silica and titanium dioxide nanoparticles cause pregnancy complications in mice [J]. Nat Nanotechnol, 2011,6(5):321-328.
[12] Kloet S K, Walczak A P, Louisse J, et al. Translocation of positively and negatively charged polystyrene nanoparticles in an in vitro placental model [J]. Toxicol In Vitro, 2015,29(7):1701-1710.
[13] Gitrowski C, Al-Jubory A R, Handy R D. Uptake of different crystal structures of TiO2 nanoparticles by Caco-2 intestinal cells [J]. Toxicol Lett, 2014,226(3):264-276.
[14] Benfer M, Kissel T. Cellular uptake mechanism and knockdown activity of siRNA-loaded biodegradable DEAPA-PVA-g-PLGA nanoparticles [J]. Eur J Pharm Biopharm, 2012,80(2):247-256.
[15] 黄乾鹏,朱立新,许小亮,等. 免疫金纳米笼子介导热疗诱导人乳腺癌细胞凋亡研究[J]. 中华肿瘤防治杂志,2014,21(15):1129-1133.
[16] 郏亚静,陈红丽,唐红波,等. 普鲁兰基肿瘤靶向纳米粒的体外抑瘤效应和细胞摄取途径研究 [J]. 医学研究生学报, 2015(2):127-130.

乙酰普鲁兰纳米粒子BeWo b30细胞毒性及摄取研究

Cell toxicity and cell uptake study of pullulan acetate nanoparticles to BeWo b30 cells

PDF

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 1

编辑推荐

Metrics

本文评价