title: Elucidating the role of free polycationic chains in polycation gene carriers by free chains of polyethylenimine or N,N,N-trimethyl chitosan plus a certain polyplex.
abstract: Polycations as gene carriers have attracted considerable attention over the past decade. Generally, polyplexes between polycations and deoxyribonucleic acid (DNA) are formed at low N/P ratios (the ratios of the numbers of nitrogen atoms in a polycation to the numbers of phosphorus atoms in DNA), but high transfection efficiency can only be obtained at much higher N/P ratios. Thus, many polycationic chains are still free in solution. In this study, we investigated the detailed functions of free polyethylenimine chains (PEI-F) and free N,N,N-trimethyl chitosan chains (TMC-F) using the same polyplex, ie, TMC polyplex (TMC-P), which has high stability in Dulbecco's Modified Eagle's Medium (DMEM). Meanwhile, PEI polyplex (PEI-P)/PEI-F was also evaluated rather than PEI-P/TMC-F because the stability of PEI-P is low in DMEM and, in the latter case, the TMC-F may replace the bound PEI chain in PEI-P to form TMC-P. The transfection results show that both TMC-F and PEI-F can significantly increase the transfection efficiency of TMC-P; however, PEI-F can upregulate the gene expression of TMC-P more efficiently than TMC-F. Further investigations on the endocytosis and intracellular trafficking show that PEI-P/PEI-F, TMC-P/PEI-F, and TMC-P/TMC-F exhibit similar cellular uptake efficiency. However, by shutting down the clathrin-mediated endocytosis or vacuolar proton pump, the transfection efficiency decreases in the order of PEI-P/PEI-F > TMC-P/PEI-F > TMC-P/TMC-F. These findings indicate that PEI-F and TMC-F may promote the transfection efficiency of the polyplex by affecting its cellular uptake pathway and intracellular trafficking.
authors: State Key Laboratory of Molecular Engineering of Polymer, Advanced Materials Laboratory, Department of Macromolecular Science, Fudan University, Shanghai, People's Republic of China.
authors: Tao
authors: T
authors: Xu
authors: Laboratory for Reproductive Immunology, Hospital and Institute of Obstetrics and Gynecology, Fudan University, Shanghai, People's Republic of China.
authors: Wei
authors: W
authors: Liu
authors: State Key Laboratory of Molecular Engineering of Polymer, Advanced Materials Laboratory, Department of Macromolecular Science, Fudan University, Shanghai, People's Republic of China.
authors: Suhang
authors: S
authors: Wang
authors: State Key Laboratory of Molecular Engineering of Polymer, Advanced Materials Laboratory, Department of Macromolecular Science, Fudan University, Shanghai, People's Republic of China.
authors: Zhengzhong
authors: Z
authors: Shao
mesh_terms: D000818:Animals; D019556:COS Cells; D048271:Chitosan / Q000737:chemistry* / Q000493:pharmacokinetics / Q000494:pharmacology; D002522:Chlorocebus aethiops; D004705:Endocytosis; D015870:Gene Expression / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D053758:Nanoparticles / Q000737:chemistry*; D011073:Polyamines / Q000737:chemistry* / Q000493:pharmacokinetics / Q000494:pharmacology; D000071228:Polyelectrolytes; D011094:Polyethyleneimine / Q000737:chemistry* / Q000493:pharmacokinetics / Q000494:pharmacology; D014162:Transfection / Q000379:methods*
chemical_list: D048271:Chitosan; D049452:Green Fluorescent Proteins; D011073:Polyamines; D000071228:Polyelectrolytes; D011094:Polyethyleneimine; C118071:N-trimethyl chitosan chloride; C485184:enhanced green fluorescent protein
keywords: cellular uptake pathway; gene delivery; intracellular trafficking; mechanism; transfection efficiency