Mechanisms of cellular uptake and intracellular trafficking with chitosan/DNA/poly(g-glutamic acid) complexes as a gene delivery vector

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Title:	Mechanisms of cellular uptake and intracellular trafficking with chitosan/DNA/poly(g-glutamic acid) complexes as a gene delivery vector
Authors:	彭淑芬(Shu-Fen Peng);(Michael T Tseng);(Yi-Cheng Ho);(Ming-Cheng Wei);(Zi-Xian Liao);(Hsing-Wen Sung)*
Contributors:	中國附醫醫學研究部遺傳中心
Keywords:	lysosome;macropinocytosis;caveolae-mediated pathway: transfection
Date:	2011-01
Issue Date:	2011-09-07 09:34:50 (UTC+8)
Abstract:	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.
Relation:	BIOMATERIALS 32(1):239-248
Appears in Collections:	[China Medical University Hospital] Jurnal articles

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