Immune cell therapeutic agent for atherosclerotic plaque as well as preparation method and application thereof

A technology for immune cell therapy and atherosclerosis, which is applied in the field of cell engineering, can solve the problem of insignificant treatment effects such as dose adoption, achieve excellent gene load and transfection ability, good application prospects, and no obvious cytotoxic effect

Pending Publication Date: 2022-08-09
CHONGQING UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In view of this, the present invention provides an immune cell therapy agent M-CD47p-GQDs-miRNA223 for atherosclerotic plaques and a preparation method, which solves the problem

Method used

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  • Immune cell therapeutic agent for atherosclerotic plaque as well as preparation method and application thereof
  • Immune cell therapeutic agent for atherosclerotic plaque as well as preparation method and application thereof
  • Immune cell therapeutic agent for atherosclerotic plaque as well as preparation method and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Preparation of immune cell therapeutic agent M-CD47p-GQDs-miRNA223 for atherosclerotic plaque (see appendix for the synthesis route). figure 1 )

[0051] (1) Dissolve 1 mg of CD47p powder in 1 ml of 75% ACN and 25% H 2 A 1 mg / mL CD47p solution was prepared in the mixed solution of O, and then 20 mg of 1-ethyl-(3-dimethylaminopropyl) carbodiimide (EDC) and 10 mg of N-hydroxysuccinimide were added in sequence. (NHS) in CD47p solution, stir magnetically until completely dissolved, adjust the pH value to be about 6.0, and react at room temperature for 30 min to obtain solution A;

[0052] (2) 1 mg / mL GQDs-NH with 5×PBS solution 2 The stock solution was diluted to 0.1mg / mL, and then 1ml of GQDs-NH was taken 2 The diluent was added to solution A, and the reaction was slightly stirred at 20-25 °C for 24 h to obtain solution B;

[0053] (3) Dialyze solution B with a dialysis bag with a molecular weight cut-off of 1kDa for 48h, and replace the dialysate every 12h to remove r...

Embodiment 2

[0061] Phagocytosis test of immune cell therapy M-CD47p-GQDs-miRNA223 for atherosclerotic plaque

[0062] (1) M-CD47p-GQDs-miRNA223, an immune cell therapy for atherosclerotic plaques, at 37°C, 5% CO2 Construct 12h under conditions;

[0063] (2) After 12h, discard the spent cell culture medium, wash it three times with PBS, and add it at a density of about 1x10 5 / ml yellow polystyrene fluorescent particles in fresh complete medium, co-cultivated with the immune cell therapeutic agent of the present invention for 6h;

[0064] (3) After 6h, discard the spent cell culture medium and wash with PBS three times. Subsequently, the nuclei were stained with DAPI with DAPI staining solution containing mounting solution for 10 min and mounted; fluorescence images were taken under a laser confocal microscope, and the results are shown in the appendix. Image 6 shown.

[0065] (4) Interpretation of the results:

[0066] attached Image 6 It can be seen that the immune cell therapeuti...

Embodiment 3

[0068] Homing ability detection of immune cell therapy M-CD47p-GQDs-miRNA223 in atherosclerotic plaques

[0069] The blank group (M+DMEM), the control group (M+MCP-1) and the experimental group (M-CD47p-GQDs-miRNA223+MCP-1) were set up respectively.

[0070] (1) Glue laying: put the transwell chamber into a 24-well plate, add 100 μL of Matrigel glue diluted with serum-free medium at a ratio of 1:8 to the upper chamber, and solidify at 37°C for 4 hours;

[0071] (2) RAW264.7 monocytes (blank group and control group) and M-CD47p-GQDs-miRNA223 cells (experimental group) were prepared to a density of 1×10 5 cells / ml single cell suspension and add 100 μL to the upper chamber of the transwell chamber, 37°C, 5% CO 2 Incubate for 6h under conditions to make cells adherent;

[0072] (3) 500 μL of monocyte chemokine MCP-1 with a concentration of 200 μg / ml was added to the lower chamber of the transwell chamber of the control and experimental groups respectively, and 500 μL of DMEM com...

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Abstract

The invention discloses an immune cell therapeutic agent for atherosclerotic plaque as well as a preparation method and application of the immune cell therapeutic agent. Relates to the technical field of cell engineering. According to the immune cell therapeutic agent for atherosclerotic plaque, mononuclear macrophages are taken as a basis, a system CD47p-GQDs-miRNA223 formed by crosslinking a membrane modification material CD47p, a nano auxiliary intermediate vector GQDs and a gene drug miRNA223 is taken as a tool, and the M-CD47p-GQDs-miRNA223 is constructed by utilizing a cell surface nano engineering technology. The immune cell therapeutic agent disclosed by the invention can be used for precisely delivering the medicine to an AS focus in a targeting manner, so that the problem that the treatment effect obtained by whole-body non-specific distribution of the medicine in AS non-invasive treatment is not remarkable is solved.

Description

technical field [0001] The present invention relates to the technical field of cell engineering, and more particularly, to an immune cell therapeutic agent M-CD47p-GQDs-miRNA223 for atherosclerotic plaque and a preparation method and application thereof. Background technique [0002] At present, in recent years, with the rapid development of cytology, immunology, molecular biology and tissue engineering technology, cell therapy therapy, as a safe and effective treatment method, plays an increasingly prominent role in clinical treatment, and is well-known It is the "third pillar of future medicine". For example, cell therapy has become the fourth treatment mode after the traditional "surgery, chemotherapy, radiotherapy" three major tumor treatment methods. Due to the three advantages of high selectivity, high local concentration, and personalized modification, cell therapy products will occupy an important share of the future disease treatment market. [0003] Atheroscleros...

Claims

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Application Information

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IPC IPC(8): A61K47/64A61K31/713C07K14/705C07K1/107A61P9/10
CPCA61K31/713A61K47/6425A61P9/10C07K14/70503
Inventor 刘非拉夏青
Owner CHONGQING UNIV OF TECH
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