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Preparation method for lysosomal membrane coated nanoparticle

A nanoparticle and lysosomal membrane technology, applied in medical preparations with non-active ingredients, medical preparations containing active ingredients, pharmaceutical formulas, etc., can solve the problems of low loading efficiency, loss, and damage to membrane structure biologically active proteins and other problems, to achieve the effects of high loading efficiency, economical savings, and uniform appearance

Active Publication Date: 2020-01-21
BEIJING UNIV OF CHEM TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These passive coating methods have low loading efficiency and are easy to cause damage and loss to the membrane structure and biologically active proteins

Method used

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  • Preparation method for lysosomal membrane coated nanoparticle
  • Preparation method for lysosomal membrane coated nanoparticle
  • Preparation method for lysosomal membrane coated nanoparticle

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Nanoparticle PMCS TEM figure 1 As shown, the PMCS nanoparticles are 140 nm in size. Lysosome transmission electron microscope figure 2 As shown, lysosome size is 450 nm.

example 2

[0050] Observation of macrophage phagocytosis of PMCS nanoparticles and internalization in lysosomes. First, macrophages were seeded in 6-well plates at a concentration of 400,000 cells per well, and incubated in an incubator. The old medium culture in the 6-well plate was replaced with 50 μg / mL PMCS nanoparticles dispersed in DMEM. Cells were washed with PBS buffer and collected by centrifugation. Wash twice with PBS. The pellet was fixed in 2.5% glutaraldehyde and 4% paraformaldehyde in PBS. Biological transmission electron microscope Figure 4 As shown, the extracted lysosomal membrane-coated nanoparticle PMCS is a single-layer membrane structure vesicle-like particle on the outside, the particle size is 450 nm, and the interior contains PMCS nanoparticles. In addition, the three potentials of PMCS, lysosome and lysosomal membrane-coated nanoparticle PMCS are as follows: Figure 5 shown. They are: -5.6mV, -24.7mV, -23.2mV.

Embodiment 3

[0052] (1) The macrophages were cultured in DMEM containing 10% fetal bovine serum, 100 μL of cell suspension was prepared in a 96-well plate, and 5000 cells were plated per well. Pre-incubate the culture plate in the incubator. After the cells adhered, activated macrophages were stimulated with 100 μL of lipopolysaccharide with a final concentration of 10 μg / mL for 1 h. Then remove the culture medium.

[0053] (2) Add 100 μL of different concentrations (6.25, 12.5, 25, 50, 100, 200 μg / mL) of nanoparticle PMCS to the culture plate.

[0054] (3) Continue to incubate for 24h, add 100μL of CCK-8 solution to each well, incubate the culture plate in the incubator for 1.5h, measure the absorbance with a microplate reader, and process the data. like Image 6 As shown in the figure, when the PMCS concentration is greater than 50 μg / mL, the incubation time is 24 h, and the macrophage activity is lower than 60%. On the premise of ensuring the macrophage activity, the macrophages can ...

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Abstract

The invention relates to a preparation method for a lysosomal membrane coated nanoparticle. Under the function of the driving of macrophages, an active lysosomal membrane coated nanoparticle is synthesized. Specifically, the preparation method comprises the following steps of: firstly, activating the macrophages by lipopolysaccharides, stimulating the secretion of lysosomal related proteases in the macrophages, and enhancing the enzymatic activity of the lysosomal related proteases; then, utilizing the phagocytic ability of the macrophages to jointly incubate the prepared nanoparticle and themacrophages to enable the macrophages to carry out phagocytosis on the nanoparticle, and carrying out internalization on the nanoparticle in macrophage lysosome; and finally, utilizing a lysosome extraction kit to extract the lysosome to obtain the lysosomal membrane coated nanoparticle. Compared with a preparation method for a traditional lipid membrane coated nanoparticle, on one hand, the preparation method disclosed by the invention is simple, and does not need to consider damage caused for the membrane by pressure or electric shock. On the other hand, the macrophages are taken as drivingforce, and the prepared lysosomal membrane coated nanoparticle keeps the activity of hydrolase in the lysosome.

Description

Technical field: [0001] The present invention relates to the preparation technology and method of lysosome-coated functional nanometer medicine. Under the action of macrophage drive, the nanoparticle coated with active lysosome membrane is synthesized. Specifically, macrophages are first activated with lipopolysaccharide to stimulate the secretion of lysosome-related proteases in macrophages and enhance their enzymatic activity, and secondly, using the phagocytic ability of macrophages, the prepared nanoparticles are co-incubated with macrophages , macrophages phagocytose the nanoparticles, internalize the nanoparticles in the macrophage lysosomes, and finally extract the lysosomes with a lysosome extraction kit to obtain nanoparticles coated with a lysosomal membrane. Background technique: [0002] Due to the diversity, heterogeneity and recurrence of tumors, it is difficult to effectively inhibit tumor recurrence and metastasis by simply relying on traditional surgery and ...

Claims

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Application Information

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IPC IPC(8): A61K41/00A61K9/51A61K38/46A61K47/46A61P35/00
CPCA61K9/5176A61K9/5192A61K38/46A61K41/0033A61P35/00A61K2300/00
Inventor 刘惠玉魏炜李闪闪
Owner BEIJING UNIV OF CHEM TECH
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