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Application of AMSC-MALAT1-Exo to preparation of medicine for treating hepatic diseases and preparation method of AMSC-MALAT1-Exo

A liver disease, amsc-malat1-exo technology, applied in the application of liver macrophage targeting biological preparations, the preparation of liver disease treatment biological preparations, the construction of adipose-derived mesenchymal stem cells and the field of exosome preparation, can Solve the problems of difficult mass preparation and limited preparation technology

Active Publication Date: 2019-08-02
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the limitations of the current MSC-Exo preparation technology, it is still difficult to prepare a large amount of MSC-Exo that meets the clinical therapeutic dose.
Therefore, the promotion of its clinical application still depends on improving its efficacy through modification.

Method used

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  • Application of AMSC-MALAT1-Exo to preparation of medicine for treating hepatic diseases and preparation method of AMSC-MALAT1-Exo
  • Application of AMSC-MALAT1-Exo to preparation of medicine for treating hepatic diseases and preparation method of AMSC-MALAT1-Exo
  • Application of AMSC-MALAT1-Exo to preparation of medicine for treating hepatic diseases and preparation method of AMSC-MALAT1-Exo

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] Example 1 AMSC-MALAT1-Exo can mediate the transfer of MALAT1 to macrophages

[0071] qRT-PCR analysis of MALAT1 was performed as described above. Such as figure 1 As shown on the left, AMSC-MALAT1-Exo contains high levels of MALAT1.

[0072] AMSC-MALAT1-Exo, AMSC-Ctrl-Exo, and native AMSC-Exo were added to macrophage RAW264.7, and the cells were collected after 24 hours for qPCR analysis of MALAT1 levels.

[0073] qPCR results showed that AMSC-MALAT1-Exo could significantly up-regulate the MALAT1 level of macrophages compared with AMSC-Ctrl-Exo and native AMSC-Exo ( figure 1 right).

Embodiment 2

[0074] Example 2 AMSC-MALAT1-Exo regulates macrophage phenotype

[0075] Add gradient concentrations of AMSC-MALAT1-Exo, AMSC-Ctrl-Exo and native AMSC-Exo (0.2 μg / mL, 4 μg / mL, 80 μg / mL) to macrophage RAW264.7, and add corresponding volume of PBS as In the control group (Vehicle), the cells were collected after 24 hours for qPCR analysis of macrophage phenotype-related molecules.

[0076] Compared with AMSC-Ctrl-Exo and natural AMSC-Exo, AMSC-MALAT1-Exo can more significantly up-regulate the expression of molecules related to M2 phenotype of macrophages Arg1 and Fizz-1, while down-regulate the expression of M1 phenotype under the same concentration conditions Expression of related molecules CD86 and iNOS. AMSC-MALAT1-Exo can significantly affect the expression of M1 / 2 phenotype-related molecules at a concentration of 0.2 μg / mL, while AMSC-Ctrl-Exo and native AMSC-Exo can significantly regulate the M1 / 2 phenotype at a concentration of 80 μg / mL Expression of related molecules (...

Embodiment 3

[0077] Example 3 AMSC-MALAT1-Exo regulates the secretion of macrophage inflammatory factors

[0078] Macrophages RAW264.7 were treated with gradient concentrations of AMSC-MALAT1-Exo, AMSC-Ctrl-Exo and natural AMSC-Exo (0.2 μg / mL, 4 μg / mL, 80 μg / mL) for 24 hours, and corresponding volumes of PBS was used as the control group (Vehicle); and then stimulated with LPS (100ng / mL) for 12h, and then ELISA was used to detect the levels of IL-1β, IL-33 and HMGB1 in the cell supernatant.

[0079] Under the same concentration conditions, AMSC-MALAT1-Exo can more significantly reduce the secretion of LPS-induced macrophage inflammatory factors IL-1β, IL-33 and HMGB1 than AMSC-Ctrl-Exo and natural AMSC-Exo. AMSC-MALAT1-Exo can significantly inhibit the secretion of macrophage inflammatory factors at a concentration of 0.2 μg / mL, and its inhibitory effect is comparable to that of AMSC-Ctrl-Exo and natural AMSC-Exo at a concentration of 80 μg / mL ( image 3 ).

[0080] Moreover, the AMSC-Ex...

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Abstract

The invention belongs to the technical field of biology, and relates to novel application of mesenchymal stem cell source exosomes, in particular to building of lncRNA MALAT1 modified fat source mesenchymal stem cells and preparation of the exosome of the mesenchymal stem cells, and application of the exosome to preparation of a hepatic disease treatment biological agent and a liver macrophage targeted biological agent. According to the thought, MALAT1 modified AMSCs are built, and the exosome is prepared; through experiments such as ELISA, WB and streaming in the cell level, the regulation and control advantages of the AMSC-MALAT1-Exo on the macrophage functional phenotype are defined; through in vivo experiments, the curative effect on hepatic diseases and the action advantages on the regulation and control of the liver macrophage functional phenotype of the MALAT1 modified AMSC source exosome are defined. The liver macrophage targeted biological agent or medicine with the improved curative effect is further prepared.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to the new application of exosomes derived from mesenchymal stem cells, in particular to the construction of adipose-derived mesenchymal stem cells modified by lncRNA MALAT1 and the preparation of exosomes, and its application in the preparation of biological preparations for liver disease treatment and Application of liver macrophage targeting biologics. Background technique [0002] The mechanism of liver inflammation is very complex, involving many factors such as etiology, host genetics, body immunity, and clinical intervention, among which the body's immune response plays a decisive role in the occurrence and development of liver inflammation. Various innate immune cells, such as monocytes / macrophages, natural killer cells, and dendritic cells, are recruited, activated, and proliferated in the inflamed liver, suggesting that the innate immune system plays a very important role in the...

Claims

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Application Information

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IPC IPC(8): A61K35/28A61P1/16C12N5/10C12N15/867
CPCA61K35/28A61P1/16C12N5/0667C12N15/86C12N2510/00C12N2740/15043
Inventor 楼国华刘艳宁陈智
Owner ZHEJIANG UNIV
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