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Non-viral gene transfection in vitro using endosomal pH-sensitive reversibly hydrophobilized polyethylenimine

Paper ID Volume ID Publish Year Pages File Format Full-Text
7132 535 2011 11 PDF Available
Title
Non-viral gene transfection in vitro using endosomal pH-sensitive reversibly hydrophobilized polyethylenimine
Abstract

Reversibly hydrophobilized 10 kDa polyethylenimine (PEI) based on rapidly acid-degradable acetal-containing hydrophobe was designed for nontoxic and highly efficient non-viral gene transfer. Water soluble PEI derivatives with average 5, 9 and 14 units of pH-sensitive 2,4,6-trimethoxybenzylidene-tris(hydroxymethyl)ethane (TMB-THME) hydrophobe per molecule, denoted as PEI-g-(TMB-THME)n, were readily obtained by treating 10 kDa PEI with varying amounts of TMB-THME-nitrophenyl chloroformate. Gel retardation assays showed that all PEI-g-(TMB-THME)n derivatives could effectively condense DNA at an N/P ratio of 5/1. Notably, polyplexes of PEI-g-(TMB-THME)n derivatives had smaller sizes (about 100∼170 nm) and higher surface charges (+25 ∼ +43 mV) than the parent 10 kDa PEI at the same N/P ratios ranging from 10/1 to 40/1. MTT assays revealed that these PEI-g-(TMB-THME)n derivatives were practically non-toxic at polymer concentrations used in transfection experiments. The acetal degradation of PEI-g-(TMB-THME)9 was shown to be highly pH dependent in which half lives of 1.3, 2.8 and 11 h were determined for pH 4.0, 5.0 and 6.0, respectively, while negligible hydrolysis (<12%) was observed after 24 h at pH 7.4. Gel electrophoresis, dynamic light scattering (DLS) and zeta potential analyses indicated that polyplexes formed at an N/P ratio of 10/1 were dissociated following 5 h incubation at pH 5.0, highlighting the importance of hydrophobic TMB-THME moieties in DNA condensation and supporting that acetal hydrolysis in endosomes would facilitate DNA release. Notably, in vitro transfection experiments performed at N/P ratios of 10/1 and 20/1 in HeLa, 293T, HepG2 and KB cells using plasmid pGL3 expressing luciferase as the reporter gene showed that reversibly hydrophobilized PEIs had superior transfection activity to 25 kDa PEI control. For example, polyplexes of PEI-g-(TMB-THME)14 showed about 235-fold and 175-fold higher transfection efficiency as compared to 10 kDa PEI in HeLa cells in serum-free and 10% serum media, respectively, which were approximately 7-fold and 16-fold higher than 25 kDa PEI formulation at its optimal N/P ratio under otherwise the same conditions. Confocal laser scanning microscope (CLSM) studies confirmed that PEI-g-(TMB-THME)14 efficiently delivered Cy5-labeled DNA to the nuclei of HeLa cells. These endosomal pH-sensitive reversibly hydrophobilized PEIs have great potentials for safe and efficient non-viral gene transfection.

Keywords
pH-sensitive; Cationic polymers; Polyethylenimine; Gene delivery; Plasmid DNA; Degradable polymers
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Non-viral gene transfection in vitro using endosomal pH-sensitive reversibly hydrophobilized polyethylenimine
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Publisher
Database: Elsevier - ScienceDirect
Journal: Biomaterials - Volume 32, Issue 34, December 2011, Pages 9109–9119
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us