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Nano artificial red blood cell and application thereof in preparation of medicine for treating bacterial infection

A technology for erythrocytes and erythrocyte membranes, which can be used in nano-drugs, anti-bacterial drugs, nanotechnology, etc., and can solve the problems of complex preparation process and difficulty in maintaining the complete structure of erythrocyte membranes.

Inactive Publication Date: 2021-06-29
FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

According to the interaction mechanism between pore-forming toxins and red blood cell membranes, a series of red blood cell-like nano-medicines have been designed for the removal of pore-forming toxins and the treatment of bacterial infections. Polymeric nanoparticles (aka nanosponges) for the clearance of various pore-forming toxins and the treatment of bacterial infections (Hu, C.J.; Fang, R.H.; Copp, J.; Luk, B.T.; Zhang, L., Abiomimetic nanosponge that absorbs pore-forming toxins.Nature Nanotechnology2013,8,336-340.); Pang Zhiqing’s research group fused red blood cell membranes with artificial lipid membranes to construct red blood cell membrane fusion liposomes, which are used for efficient removal of pore-forming toxins and bacterial infection (He, Y.; Li, R.; Li, H.; Zhang, S.; Dai, W.; Wu, Q.; Jiang, L.; Zheng, Z.; Shen, S.; Chen, X.; Zhu, Y.; Wang, J.; Pang, Z., Erythroliposomes: Integrated Hybrid Nanovesicles Composed of Erythrocyte Membranes and Artificial Lipid Membranes for Pore-Forming Toxin Clearance. ACS Nano 2019, 13, 4148-4159.); Both nanosponge and erythrocyte membrane fusion liposome can effectively remove pore-forming toxins and treat bacterial infections, but the preparation process is complicated, it is difficult to maintain the original intact structure of erythrocyte membrane, and it is easy to be cleared by mononuclear macrophages in vivo, and clinical Applications are limited by lack of red blood cell resources and blood type pairing requirements

Method used

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  • Nano artificial red blood cell and application thereof in preparation of medicine for treating bacterial infection
  • Nano artificial red blood cell and application thereof in preparation of medicine for treating bacterial infection
  • Nano artificial red blood cell and application thereof in preparation of medicine for treating bacterial infection

Examples

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Embodiment 1

[0054] Embodiment 1: the construction of erythrocyte membrane protein liposome

[0055] In this embodiment, red blood cells are firstly extracted from whole blood, and red blood cell membranes are separated and purified by hypotonic and centrifugation methods, and then red blood cell membrane proteins are extracted. Specific operation: remove the eyeballs of six-week-old male ICR mice to take blood, anticoagulate the whole blood with heparin sodium, centrifuge at 700g for 10 minutes at 4°C, discard the upper plasma and buffy coat (white blood cells and platelets), and then use Resuspend the bottom erythrocytes in 1 mM EDTA in PBS, and repeat the washing three times. Collect the red blood cells at the bottom and add an equal volume of PBS solution containing 1mM EDTA to make a red blood cell suspension, add each 250μL to a 1.5mL EP tube, then add 950μL 0.2mM EDTA aqueous solution, mix well, vortex to break the red blood cells, and then add 50μL 20× Adjust to isotonic with PBS....

Embodiment 2

[0060] Example 2: Membrane protein integration analysis in erythrocyte membrane protein liposome

[0061] In this embodiment, first adopt differential scanning calorimetry to carry out thermal analysis to RP-PLs ( figure 2 A); The results show that, compared with PLs, the phase transition temperature (transition temperature, T of RP-PLs and RMVs m ) obviously shifted to the right, which indirectly proves that the membrane protein is integrated in the liposome bilayer membrane. The infrared spectra of RP-PLs were measured by Fourier transform infrared spectrometer; the results showed that both RP-PLs and RMVs had protein characteristic peaks (1,700-1,600 cm -1 and 1,600-1,500cm -1 ) ( figure 2 B), showing that membrane proteins in RP-PLs are integrated on liposomes.

Embodiment 3

[0062] Example 3: Membrane protein composition and orientation analysis in erythrocyte membrane protein liposome

[0063] Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was used to determine the protein components of RP-PLs; the samples were prepared with SDS sample buffer, and the membrane protein concentration of the samples was determined by the Bradford method, and the protein samples were stored at 90 °C Heating for 5 minutes, loading on 10% SDS polyacrylamide gel (Bio-Rad), running at 120V for 1 hour, then staining and imaging with Coomassie brilliant blue; Western blotting to detect certain specific proteins in RP-PLs, Such as CD47 and ADAM10; for samples 1 × sample buffer preparation, SDS-PAGE separation as above, the protein was transferred to nitrocellulose membrane (PALL), respectively, with CD47 (ABCAM, USA) and ADAM10 (R&D, USA) specific antibodies and the corresponding horseradish Peroxidase (HRP)-conjugated secondary antibody (1:200 dilut...

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Abstract

The invention belongs to the technical field of biology, and relates to a nano artificial red blood cell and application thereof in preparation of a medicine for treating bacterial infection. The nano medicine is a red blood cell membrane protein liposome formed by integrating red blood cell membrane protein into an artificial lipid membrane, wherein the lipid membrane is composed of phosphatidylcholine, polyethylene glycol distearoyl phosphatidyl ethanolamine and / or cholesterol. The nano medicine adsorbs pore-forming toxins on the surface by simulating the red blood cell, neutralizes the toxicity of the toxins, and realizes anti-virulence factor treatment, thereby enhancing the effect of anti-bacterial infection treatment, especially drug-resistant bacterial infection.

Description

technical field [0001] The invention belongs to the field of biological technology, and relates to a nano-artificial red blood cell and its application in the preparation of a drug for treating bacterial infection, in particular to a nano-artificial red blood cell with bacterial pore-forming toxin adsorption performance and its use in the preparation of a drug for treating bacterial infection. Uses: The nanomedicine absorbs pore-forming toxins on its surface by simulating red blood cells, neutralizes the toxicity of the toxins, and realizes anti-virulence factor treatment, thereby improving the effect of anti-bacterial infection treatment, especially drug-resistant bacterial infection. Background technique [0002] The prior art discloses that blood is a kind of liquid connective tissue in the body, which plays an extremely important role in maintaining body homeostasis and regulating immunity. Red blood cells, also known as erythrocytes, are the most common cell type in blo...

Claims

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Application Information

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IPC IPC(8): A61K9/127A61K38/17A61K45/00A61K47/24A61K47/28A61K47/42A61P31/04B82Y5/00
CPCA61K9/127A61K47/28A61K47/24A61K47/42A61K45/00A61K38/1709A61P31/04B82Y5/00
Inventor 王建新庞志清何雨薇李瑞翔李海春
Owner FUDAN UNIV
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