Stimulus-responsive Diselenide-crosslinked Polyethyleneimine as Gene Vector†

doi:10.7503/cjcu20130926

Abstract

Abstract:

Four different diselenide-crosslinked polyethylenimine(PEISeSe) were successfully prepared by changing the dosage of cross-linker and reaction time. The influence of cross-linking level on the DNA condensation, proton buffer capacity and transfection efficiency were investigated. The results indicated that the proton buffer capacity of PEISeSe was reduced with cross-linking level. All of the polycations could efficiently condense DNA into spherical particles with diameter of 150 nm at m(PEISeSe)/m(DNA)=8. Moreover, diselenide bonds were easily cleaved in the presence of DTT. Cell culture results indicated that by adding chloroquine to exclude the limitation of lysosome escape, PEISeSe showed decreased cell cytotoxicity and efficient transfection. The proper reason was that the intracellular cleavage of diselenide bonds facilitated DNA release and nuclear entry. The diselenide-crosslinked polycations had promising potential in redox-sensitive gene delivery.

Key words: Diselenide crosslinked, Polyethylenimine, Stimulus-response, Non-viral gene vector

CLC Number:

O631.3
Q782

TrendMD:

KONG Yunna, LI Wenyu, DU Jianwei, TANG Jianguo, HU Qiaoling, WANG Youxiang. Stimulus-responsive Diselenide-crosslinked Polyethyleneimine as Gene Vector^†[J]. Chem. J. Chinese Universities, 2014, 35(4): 881.

Figures/Tables 10

Scheme 1 Synthesis routes of PEISeSe

Fig.1 1H NMR spectrum of PEISeSe2.7 in D2O

Fig.2 Buffer capacity(BC) of different polymer by acid-base titration Polymer concentration was fixed at 0.2 mg/mL; c(HCl)=0.1 mol/L.

Fig.3 Agarose gel electrophoresis retardation of different polyplexes with various m(PEISeSe)/m(DNA)

Fig.4 TEM images of different polyplexes at m(polymer)/m(DNA)=16 after treatment with 10 mmol/L DTT (A) PEI1800/DNA; (B) PEISeSe2.7/DNA; (C) PEISeSe2.7/DNA after treament with 10 mmol/L DTT.

Fig.5 Zeta potential of different polyplexes with various m(polymer)/m(DNA)Error bars represent mean±SD for n=3. a.PEI1800; b. PEISeSe2.3; c. PEISeSe2.7; d. PEISeSe3.4; e. PEISeSe4.1.

Fig.6 Ethidium bromide exclusion assay of PEISeSe2.7/DNA polyplexes with various m(PEISeSe)/m(DNA)

Fig.7 Cytotoxicity of HEK293T cells exposed to different polymer complexed with 1 μg DNA after incubation for 48 h Error bars represent mean±SD for n=5. a. Control; b.PEI25000; c. PEI1800; d. PEISeSe2.3; e. PEISeSe2.7; f. PEISeSe3.4; g. PEISeSe4.1.

Fig.8 Transfection efficiency of HEK293T cells exposed to different polyplexes in the presence of chloroquine detected by flow cytometryPEI25000 at N/P ratio of 10 is used as compared. Error bars represent mean±SD for n=3.a.PEI25000; b. PEI1800; c. PEISeSe2.3; d. PEISeSe2.7; e. PEISeSe3.4; f. PEISeSe4.1.

Fig.9 Intracellular distribution of PEISeSe2.7/Cy3-DNA polyplexes at m(PEISeSe)/m(DNA)=16The polyplexes were incubated with HEK293T cells for 4 h in the absence(A) and presence of chloroquine(B), followed by incubation in the absence of polyplexes and chloroquine for another 12 h(A3, B3). Cy3-DNA presented as red fluorescence(A1, B1) and DAPI-stained nuclei showed blue color(A2, B2).

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