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Self-assembled Nanocarriers for Gene Delivery

Author: XuChunHong
Tutor: ShenYouQing
School: Zhejiang University
Course: Biochemical Engineering
Keywords: Gene therapy Gene delivery Non - viral vectors Polyethylenimine Gene transfection
CLC: Q782
Type: Master's thesis
Year: 2011
Downloads: 65
Quote: 0
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Abstract


Gene therapy refers exogenous normal gene into target cells in order to correct or compensate for genetic defects and diseases caused by abnormal, so as to achieve the therapeutic purposes. However, the gene therapy encounter many challenges, one of the biggest challenges is how to get efficient and safe gene delivery vehicle. Gene vector into viral vectors and non-viral vectors. Relative to viral vectors, non-viral vectors with high safety performance and the advantages of large-scale application, but the low efficiency of gene expression, and in vitro and in vivo gene transfection efficiency inconsistent. The objectives of this paper is prepared suitable for in vivo applications, and has a novel gene delivery systems in vivo high transfection efficiency. Polyethylene imine (PEI) has a high transfection efficiency in vitro, has been widely used for in vitro gene delivery. PEI with the formation of DNA complexes (polyplexes) with a positive charge, have non-specific interactions with blood proteins resulting in concentration in blood half-life is short. By electrostatic interactions wrapped the surface of the composite layer with the negatively charged polymer can reduce this non-specific effects, but with a negative charge of the polymer would be with DNA competition envelope and PEI, resulting in deterioration of the stability of the complexes, complexes blood is likely to unravel. Meanwhile, a strong electrostatic interaction between PEI and DNA, so that their composite not easily solved, therefore the DNA does not easily be expressed in the nucleus. Design ideas for the work: PEI DNA complexes after breaking bond by intracellular highly cross-linked, and then wrapped polyanionic polymer. Therefore, even if the role of polyanion polymer with PEI, DNA will not be released; inside the cell, the cross-linking bond is broken, since the poly competition envelope of the anion with PEI, and the DNA is likely to be released, so that the DNA easily be expressed to improve the efficiency of gene transfection. This thesis to PEI crosslinked of PEI / DNA composite objects is designed and a polyanion of PEI / DNA composite object surface carried modified to form a \. The first job is embedded heparin (HP) reversible cross-linked PEI / DNA complex as a targeted gene in vivo delivery system. Preparation of PEI / DNA complexes, and complexes were crosslinked with a disulfide bond that, after cross-linking the complex more stable. Then heparin embedding the composite formed HP / PEI / DNA complex, the nano particle size of about 100 nm in serum and the high salt solution, has good stability and low cytotoxicity. The effects of the gene delivery systems in vivo tumor gene transfection efficiency with breast cancer in nude mice EGFP reporter gene plasmid transfection vaccination someone source the results detected nude mice tumor is highly expressed green fluorescent protein, and by the immune staining experiments confirmed that the gene delivery system has the potential for in vivo applications. The second work is the embedded with hyaluronic acid (HA) PEI-Dexa/DNA composite. HA has good performance, the \Some tumor cells expressed high HA receptor, therefore, the HA-wrapped composite can be through the cells of the receptor-mediated endocytosis into tumor cells. Dexamethasone (Dexa) has a nuclear targeting ability, and therefore the introduction of Dexa up can further improve the nuclear capability to the complex. Therefore, the prepared HA embedded PEI-Dexa/DNA composite. First Dexa covalently connected to PEI (MW25 kDa) chain investigated cytotoxicity and transfection efficiency of the introduction of dexamethasone. The results show that the modified dexamethasone, cytotoxicity decreased, but the transfection efficiency may be due to of Dexa the shielding effect than PEI (MW25 kDa) slightly reduced. Secondly, hyaluronic acid embedding formed HA / PEI-Dexa / DNA complexes for high expression of the transfection efficiency of the cell strain of the HA receptor higher than the HA receptor-negative cell lines, indicate that the introduction of the HA receptor on the HA positive cells transfected with a catalytic role. The third is entrapped in anionic liposomes PEI / DNA complexes. The purpose of this work is a preliminary study to take advantage of the ability of the preparation of the liposome membrane fusion simulation viral gene delivery systems. Initially prepared anionic liposome entrapped of Lipid / PEI / DNA complex found that the particle surface carries a negative charge, with the potential of in vivo applications, but because of the surface of the gene delivery system with a strong negative charge, even if the targeting group. folic acid at the cellular level, the efficiency of transfection is still low. Therefore, the need for the kind of anionic liposomes optimized to obtain a high transfection efficiency of the gene delivery system.

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CLC: > Biological Sciences > Molecular Biology > Genetic engineering (genetic engineering) > Gene vector
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