title: Tuning the buffering capacity of polyethylenimine with glycerol molecules for efficient gene delivery: staying in or out of the endosomes.
abstract: Endosomal escape is a major bottleneck for efficient non-viral gene delivery. This paper presents the development of two novel non-viral vectors by cross-linking glycerol molecules with low molecular weight polyethylenimine (PEI). The vectors, namely, HG-PEI (45 mol% glycerol content) and LG-PEI (9 mol% glycerol content) have apparently similar DNA binding, DNA unpacking and cellular uptake abilities but differ in buffering capacity. The cellular uptake and subsequent transfection efficiency of LG-PEI is superior to commercially available PEI 25 k. Interestingly, although the cellular uptake of HG-PEI is higher than that of PEI 25 k, the transgene expression by HG-PEI-mediated transfection is very low. Inhibitor and co-localization studies demonstrate the mechanism of endocytosis and formation of endosomes prone to lysosomal lysis of HG-PEI polyplexes as a consequence of its weak buffering capacity. Importantly, when the lysosomal lysis is inhibited, the transgene expression of HG-PEI-mediated transfection increases by 9-fold of its initial capacity which is comparable to the transfection efficiency of PEI 25 k. These results indicated that the buffering capacity of the polymers primarily impacts endosomal escape and subsequent transfection efficiency. Furthermore, this study highlights the significance of cross-linkers in optimizing the buffering capacity when designing polymers for gene delivery.
authors.affiliation: Department of Agricultural Biotechnology, Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul, 151-921, Korea.
authors.forename: Bijay
authors.initials: B
authors.lastname: Singh
authors.affiliation: Department of Agricultural Biotechnology, Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul, 151-921, Korea.
authors.forename: Sushila
authors.initials: S
authors.lastname: Maharjan
authors.affiliation: Department of Agricultural Biotechnology, Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul, 151-921, Korea.
authors.forename: Tae-Eun
authors.initials: TE
authors.lastname: Park
authors.affiliation: Department of Agricultural Biotechnology, Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul, 151-921, Korea.
authors.forename: Tao
authors.initials: T
authors.lastname: Jiang
authors.affiliation: Institute of Green-Bio Science and Technology, Seoul National University, Kangwon-Do, 232-916, Korea.
authors.forename: Sang-Kee
authors.initials: SK
authors.lastname: Kang
authors.affiliation: Department of Agricultural Biotechnology, Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul, 151-921, Korea. cyjcow@snu.ac.kr.
authors.forename: Yun-Jaie
authors.initials: YJ
authors.lastname: Choi
authors.affiliation: Department of Agricultural Biotechnology, Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul, 151-921, Korea. chocs@snu.ac.kr.
authors.forename: Chong-Su
authors.initials: CS
authors.lastname: Cho
mesh_terms: D002021:Buffers; D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D002738:Chloroquine / Q000494:pharmacology; D004247:DNA / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects; D011992:Endosomes / Q000378:metabolism*; D005434:Flow Cytometry; D015870:Gene Expression / Q000187:drug effects; D018014:Gene Transfer Techniques; D005990:Glycerol / Q000737:chemistry*; D006801:Humans; D018942:Macrolides / Q000494:pharmacology; D018613:Microscopy, Confocal; D010316:Particle Size; D011094:Polyethyleneimine / Q000138:chemical synthesis / Q000737:chemistry* / Q000633:toxicity; D054328:Proton Pump Inhibitors / Q000494:pharmacology; D055672:Static Electricity; D014162:Transfection
chemical_list: D002021:Buffers; D018942:Macrolides; D054328:Proton Pump Inhibitors; D002738:Chloroquine; C040929:bafilomycin A1; D011094:Polyethyleneimine; D004247:DNA; D005990:Glycerol
keywords: buffering capacity; endosomal escape; glycerol; polyethylenimine; proton sponge