Mechanisms of cellular uptake and intracellular trafficking with chitosan/DNA/poly(γ-glutamic acid) complexes as a gene delivery vector
Autor: | Ming-Cheng Wei, Shu-Fen Peng, Zi-Xian Liao, Michael T. Tseng, Yi Cheng Ho, Hsing-Wen Sung |
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Rok vydání: | 2011 |
Předmět: |
Cell Survival
Endocytosis Inhibition media_common.quotation_subject Intracellular Space Biophysics Bioengineering Molecular Dynamics Simulation Biology Transfection Endocytosis Models Biological Biomaterials chemistry.chemical_compound Cell Line Tumor Humans Luciferases Internalization media_common Chitosan Pinocytosis Polyglutamic acid Gene Transfer Techniques DNA Cell biology Kinetics Protein Transport Polyglutamic Acid chemistry Mechanics of Materials Ceramics and Composites Intracellular |
Zdroj: | Biomaterials. 32:239-248 |
ISSN: | 0142-9612 |
DOI: | 10.1016/j.biomaterials.2010.08.081 |
Popis: | Chitosan (CS)-based complexes have been considered as a vector for DNA delivery; nonetheless, their transfection efficiency is relatively low. An approach by incorporating poly(γ-glutamic acid) (γ-PGA) in CS/DNA complexes was developed in our previous study to enhance their gene expression level; however, the detailed mechanisms remain to be understood. The study was designed to investigate the mechanisms in cellular uptake and intracellular trafficking of CS/DNA/γ-PGA complexes. The results of our molecular dynamic simulations suggest that after forming complexes with CS, γ-PGA displays a free γ-glutamic acid in its N-terminal end and thus may be recognized by γ-glutamyl transpeptidase in the cell membrane, resulting in a significant increase in their cellular uptake. In the endocytosis inhibition study, we found that the internalization of CS/DNA complexes took place via macropinocytosis and caveolae-mediated pathway; by incorporating γ-PGA in complexes, both uptake pathways were further enhanced but the caveolae-mediated pathway played a major role. TEM was used to gain directly understanding of the internalization mechanism of test complexes and confirmed our findings obtained in the inhibition experiments. After internalization, a less percentage of co-localization of CS/DNA/γ-PGA complexes with lysosomes was observed when compared with their CS/DNA counterparts. A greater cellular uptake together with a less entry into lysosomes might thus explain the promotion of transfection efficiency of CS/DNA/γ-PGA complexes. Knowledge of these mechanisms involving CS-based complexes containing γ-PGA is critical for the development of an efficient vector for DNA transfection. |
Databáze: | OpenAIRE |
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