In vitro and in vivo gene delivery using chitosan/hyaluronic acid nanoparticles: Influences of molecular mass of hyaluronic acid and lyophilization on transfection efficiency

被引:22
作者
Sato, Toshinori [1 ]
Nakata, Mitsuhiro [1 ]
Yang, Zhihong [1 ]
Torizuka, Yu [1 ]
Kishimoto, Satoko [2 ]
Ishihara, Masayuki [3 ]
机构
[1] Keio Univ, Dept Biosci & Informat, Yokohama, Kanagawa 2238522, Japan
[2] Dokkyo Med Univ, Res Support Ctr, Shimotsuga, Tochigi, Japan
[3] Natl Def Med Coll, Res Inst, Tokorozawa, Saitama, Japan
基金
日本科学技术振兴机构;
关键词
anti-tumor effect; chitosan; gene transfer; hyaluronic acid; lyophilization; suicide gene therapy; CHONDROITIN SULFATE; INTRACELLULAR TRAFFICKING; PDNA/CHITOSAN COMPLEXES; CHITOSAN NANOPARTICLES; CD44; DOXORUBICIN; MECHANISMS; EXPRESSION; PROTECTION; OLIGOMERS;
D O I
10.1002/jgm.2968
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
BackgroundLyophilization is an effective method for preserving nonviral gene vectors. To improve the stability and transgene expression of lyophilized plasmid DNA (pDNA) complexes, we coated the surfaces of pDNA/chitosan complexes with hyaluronic acid (HA) of varying molecular masses. The transgene expression of pDNA/chitosan/HA ternary complexes was characterized in vitro and in vivo. MethodspDNA complexes were lyophilized overnight and the resultant products with spongy, porous consistencies were stored at -30, 4 or 25 degrees C for 2weeks. Rehydrated complexes were characterized using gel retardation assays, aiming to confirm complex formation, measure particle size and evaluate zeta potential, as well as conduct luciferase gene reporter assays. The anti-tumor effects of pDNA ternary complexes were evaluated using suicide gene (pTK) coding thymidine kinase in Huh7-implanted mice. ResultsTransfection efficiencies of pDNA/chitosan/HA ternary complexes were dependent on the average molecular masses of HA. The coating of pDNA/chitosan complexes with HA maintained the cellular transfection efficiencies of lyophilized pDNA ternary complexes. Furthermore, intratumoral injection of lyophilized, rehydrated pDNA ternary complexes into tumor-bearing mice showed a significant suppression of tumor growth. ConclusionsThe coating of pDNA/chitosan complexes with high-molecular-weight HA augmented the stability and cellular transfection ability of the complexes after lyophilization-rehydration.
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页数:10
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