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Lipid nanoparticle loaded with Olig2 messenger RNA (Ribonucleic Acid) and application of lipid nanoparticle

A lipid nanoparticle and messenger technology, applied in nanotechnology, nanotechnology, nanomedicine, etc., can solve problems such as insufficient differentiation, and achieve the effect of promoting remyelination, targeting and effectiveness, and promoting differentiation and maturation.

Pending Publication Date: 2022-08-05
中山大学深圳 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the ability of such resident OPCs to differentiate into mature oligodendrocytes and promote myelination is clearly insufficient

Method used

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  • Lipid nanoparticle loaded with Olig2 messenger RNA (Ribonucleic Acid) and application of lipid nanoparticle
  • Lipid nanoparticle loaded with Olig2 messenger RNA (Ribonucleic Acid) and application of lipid nanoparticle
  • Lipid nanoparticle loaded with Olig2 messenger RNA (Ribonucleic Acid) and application of lipid nanoparticle

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Preparation and Characterization of Cationic Lipid Nanoparticles Loaded with Olig2 mRNA Targeting Antibody Modification.

[0054] 1. Preparation of LNPs modified with Olig2 mRNA targeting antibody.

[0055] preparation process such as figure 1 shown.

[0056] (1) With reference to the patent of application No. 202110566034.7, a plasmid expressing mouse Olig2 was constructed, and Olig2 mRNA was transcribed in vitro to synthesize Olig2 mRNA. The sequence of the Olig2-loaded messenger RNA is shown in SEQ ID NO: 1.

[0057] (2) The lipid phase DLink-MC3-DMA: DSPC: cholesterol: DMG-PEG2000 (molar ratio of 55:5:38.5:1.5) was dissolved in 90% ethanol; Volume ratio = 3:1; citric acid adjusted pH = 4), in which the molar ratio of loaded Olig2 messenger RNA and DLink-MC3-DMA is 1:10, which is injected and mixed by Nano Assemblr Benchtop microfluidic device to form nanometers of about 100 nm. Particle solution; then use a 3000 molecular weight dialysis bag to dialyze against DE...

Embodiment 2

[0070] The effect of LNPs modified with Olig2 mRNA targeting antibody on the uptake of mouse neural stem cell-derived OPCs in vitro was verified.

[0071] In order to explore the transfection effect of different concentrations of Olig2 mRNA / LNPs, OPCs were 5×10 per well. 5 seeded in 12-well plates. Use 1, 2, 4, 6 and 8 μg mL the next day -1 A final concentration of 5-carboxyfluorescein FAM-labeled C-Olig2 was incubated with OPCs for 6 h. Uptake efficiency was measured by flow cytometry. The result is as Figure 5 As shown in A-B, it can be seen that 6μg mL -1 C-Olig2 can achieve over 92% transfection effect.

Embodiment 3

[0073] The OPCs uptake effect of LNPs modified with Olig2 mRNA targeting antibody was verified in mice.

[0074] In order to investigate the intracranial injection of Olig2 mRNA / LNPs into mouse brain, in CD140a + Targeted expression levels in OPCs cells, mice received intracranial injection of C-Olig2 (0.05 mg / kg). After 24 hours, the mice were sacrificed, and the CD140a of the brain tissue was isolated + , CD140a - OPCs ( Image 6 ). The expression of Olig2 protein was detected by Western Blot, and the results were as follows Figure 7 shown, C-Olig2 successfully targets CD140a in the mouse brain + OPC, and overexpressed Olig2 protein.

[0075] The specific operations are as follows:

[0076] 1. Middle cerebral artery occlusion (MCAO) model mice received (0.05 mg kg -1 ) C-Olig2 was injected intracranially, and 24 hours later, the mouse brains were removed. Using Adult Brain Dissociation Reagent, after dissociation, red blood cells and cell debris are removed to make...

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Abstract

The invention relates to lipid nanoparticles loaded with Olig2 messenger RNA (Ribonucleic Acid), and relates to the technical field of lipid nanoparticles. The lipid nanoparticle comprises messenger RNA (Ribonucleic Acid) loaded with Olig2, the lipid nanoparticle is modified with a CD140a monoclonal antibody, and the lipid nanoparticle delivers the messenger RNA to the surface of a cell membrane to express OPCs of the CD140a. According to the lipid nanoparticles, Olig2 can be instantly overexpressed by OPCs, the differentiation maturation and myelination capacity of the OPCs residing in the brain can be improved, and the treatment effect on cerebral arterial thrombosis can be effectively improved.

Description

technical field [0001] The present invention relates to the technical field of lipid nanoparticles, in particular to a lipid nanoparticle loaded with Olig2 messenger RNA and its application. Background technique [0002] Ischemic stroke is one of the most common cardiovascular and cerebrovascular diseases in the world. lost. Insufficient blood and oxygen supply to the brain can lead to the death of oligodendrocytes in the central nervous system, which in turn leads to myelin disintegration. Myelin is the main component of white matter in the central nervous system and plays an important role in the transmission of nerve signals and the communication of information between different regions of the brain. Although most of the pathological studies of ischemic stroke in the past focused on the death of neurons in the gray matter, the importance of myelin in the white matter of the brain in the long-term recovery of ischemic stroke has received more and more attention. Endogen...

Claims

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Application Information

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IPC IPC(8): A61K48/00A61K47/69A61K47/68A61P9/10A61P25/00B82Y5/00
CPCA61K48/0025A61K48/0066A61K47/6929A61K47/6849A61P9/10A61P25/00B82Y5/00Y02A50/30
Inventor 邓文斌徐健聂怡初赵景新谢沅
Owner 中山大学深圳
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