RRC ID 65105
Author Sato T, Nakata M, Yang Z, Torizuka Y, Kishimoto S, Ishihara M.
Title In vitro and in vivo gene delivery using chitosan/hyaluronic acid nanoparticles: Influences of molecular mass of hyaluronic acid and lyophilization on transfection efficiency.
Journal J Gene Med
Abstract BACKGROUND:Lyophilization 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.
METHODS:pDNA complexes were lyophilized overnight and the resultant products with spongy, porous consistencies were stored at -30, 4 or 25°C for 2 weeks. 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.
RESULTS:Transfection 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.
CONCLUSIONS:The coating of pDNA/chitosan complexes with high-molecular-weight HA augmented the stability and cellular transfection ability of the complexes after lyophilization-rehydration.
Volume 19(8)
Published 2017-8-1
DOI 10.1002/jgm.2968
PMID 28667693
MeSH Animals Chitosan / therapeutic use* DNA / genetics Electrophoretic Mobility Shift Assay Freeze Drying Gene Transfer Techniques Genes, Reporter Genetic Therapy / methods* Genetic Vectors Humans Hyaluronic Acid / therapeutic use* Luciferases / genetics Mice Nanoparticles Particle Size Plasmids Thymidine Kinase / genetics Transfection
IF 3.258
Resource
Human and Animal Cells HuH-7(RCB1366)