PEG-PLGA nanosphere loading BDNF gene plasmid as well as preparation method and application thereof

A nano-microsphere and gene technology, applied in the fields of material biology and neuroscience, can solve the problems of low gene transfer efficiency of neural stem cells, limited application of cytotoxicity, unstable properties, etc., and achieve good monodispersity and biocompatibility Good performance and good encapsulation rate

Active Publication Date: 2019-01-08
SOUTHEAST UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, liposomes are the most researched, but the gene transfer efficiency of liposomes to neural stem cells is not high, their properties are unstable, easily digested by hydrolytic enzymes inside and outside the cells, the cost is high, and their cytotoxicity limits their application.

Method used

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  • PEG-PLGA nanosphere loading BDNF gene plasmid as well as preparation method and application thereof
  • PEG-PLGA nanosphere loading BDNF gene plasmid as well as preparation method and application thereof
  • PEG-PLGA nanosphere loading BDNF gene plasmid as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] Preparation of PEG-PLGA nanospheres carrying BDNF gene plasmid

[0056] (1) Weigh 200 mg of polyethylene glycol-modified polylactic acid / glycolic acid copolymer (PEG-PLGA, the weight average molecular weight of polyethylene glycol is 2000Da, the weight average molecular weight of polylactic acid glycolic acid segment is 11500Da or can adopt The weight average molecular weight is 10000Da, the molar ratio of two monomers of polylactic acid and glycolic acid is 50:50) is dissolved in 2ml of dichloromethane (DCM, 10%wt / vol) as the organic oil phase, 200μl of BDNF gene plasmid solution (Plasmid concentration 1000ng / μl) was added dropwise to the organic phase as the aqueous phase, that is, the mass ratio of PEG-PLGA to the BDNF gene plasmid solution was 1:1, and the ultrasonic cell disruptor was used to sonicate in an ice bath at 100W ultrasonic power for 30s to prepare Obtain solution colostrum (W / O); add colostrum dropwise to 5mL 5% polyvinyl alcohol (PVA) in the external a...

Embodiment 1

[0059] The physical and chemical properties of the PEG-PLGA nanospheres carrying the BDNF gene plasmid prepared in Example 1 were detected as follows:

[0060] (1) Scanning electron microscope and transmission electron microscope to observe the physical morphology of nanoparticles

[0061] The morphology of nano-microspheres was observed by transmission electron microscope and scanning electron microscope.

[0062] The result is as figure 1 As shown, observing the PEG-PLGA nanospheres loaded with BDNF plasmids under scanning electron microscope and transmission electron microscope, it can be found that the appearance of the microspheres is spherical, the surface is smooth and complete, the size distribution is uniform, the monodispersity is good, and there is little mutual fusion phenomenon. See obvious cracks.

[0063] (2) Malvern ZS90 particle size analyzer measures the particle size and Zeta potential of nanospheres

[0064] The results show that the particle size of the...

Embodiment 3

[0069] Detection of Biological Properties of PEG-PLGA Nanospheres Carrying BDNF Gene Plasmid

[0070] The biological characteristics of the PEG-PLGA nanospheres carrying the BDNF gene plasmid prepared in Example 1 were detected as follows:

[0071] (1) Effects of PEG-PLGA nanospheres carrying BDNF gene plasmid on the morphology and proliferation of neural stem cells

[0072] Take the NSCs passaged in vitro, plant them in a 12-well plate, take them out after incubation for 4 hours, add 300 μl of complete medium containing 6 mg / ml PEG-PLGA nanospheres containing the BDNF gene plasmid, 37 ° C, 5% CO 2 Observed with an inverted microscope after incubation for 3 days.

[0073] Take the NSCs of the third generation, digest them into single cells, adjust them to 200,000 cells / ml with medium, add them to a 96-well plate, set 27 experimental wells in each plate, and divide them into 3 groups, 50 μl per hole (about 10,000 cells) cells); after incubation for 4 h, take them out, add 50 ...

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Abstract

The invention discloses a PEG-PLGA nanosphere loading a BDNF gene plasmid as well as a preparation method and application thereof. The nanosphere is obtained after the BDNF gene plasmid is loaded by apoly(ethylene glyco)-b-poly(lactic-co-glycolic acid) (PEG-PLGA) nanoparticle. The nanosphere provided by the invention, which has the advantages of uniform particle size distribution, good dispersion, excellent ability in transfecting neural stem cells and high expression of brain derived neurotrophic factor (BDNF) functional protein after transfecting the neural stem cells, is applicable to protection and repairing of the function of nerve cells and can play an important role in the medical field as a transfection material for the neural stem cells, thereby having a broad application prospect.

Description

technical field [0001] The invention belongs to the technical fields of material biology and neuroscience, and specifically relates to a PEG-PLGA nanometer microsphere carrying a BDNF gene plasmid and a preparation method and application thereof. Background technique [0002] In recent years, neural stem cells have been proved to have the ability to cross the blood-brain barrier and the potential to migrate to pathological sites. They can be used as seed cells for cell transplantation and at the same time as carriers to deliver gene drugs to play the role of gene therapy. At present, the focus of research on the gene carrier delivery system of neural stem cells has shifted from the viral vector system to the non-viral vector system. Among them, liposomes are the most researched, but the gene transfer efficiency of liposomes to neural stem cells is not high, their properties are unstable, easily digested by hydrolytic enzymes inside and outside the cells, the cost is high, an...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K9/51A61K38/18A61K48/00A61K47/34A61P25/00
CPCA61K9/5146A61K38/185A61K48/0008A61K48/005A61P25/00
Inventor 陈陆馗余勇波
Owner SOUTHEAST UNIV
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