Preparation method of human adipose mesenchymal stem cell exosome for treating myocardial infarction

A technology of mesenchymal stem cells and exosomes, applied in the field of preparation of exosomes from human adipose-derived mesenchymal stem cells, to achieve stable effects, enhance heart function, and reduce compensatory cardiac load

Pending Publication Date: 2020-11-17
FUJIAN MEDICAL UNIV UNION HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the great differences in the biologically active substances in exosomes expressed by stem cells from different sources, more research

Method used

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  • Preparation method of human adipose mesenchymal stem cell exosome for treating myocardial infarction
  • Preparation method of human adipose mesenchymal stem cell exosome for treating myocardial infarction
  • Preparation method of human adipose mesenchymal stem cell exosome for treating myocardial infarction

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0032] Example 1

[0033] The object of this embodiment is to separate the culture of human fatty mesenchymal stem cells.

[0034] Material

[0035] Preparing human fat is normal human fat after surgery (signed informed consent), PBS, BSA, IV collagenase, fetal bovine serum, CD29 + CD44 + , CD90 + CD34 - And CD45 - Monoclonal antibody.

[0036] 2. Method

[0037] 2.1 Separation of human fat mesenchymal stem cells

[0038] (1) The adipose tissue is collected under sterile conditions, and the connective tissue envelope and blood vessels of the surface of the adipose tissue are removed.

[0039] (2) Quickly cut the untraped to the paste, add 3 ml of medium containing medium, containing a medium containing a final concentration of 0.06 to 0.16% IV type collagenase and 0.015 ~ 0.02% BSA At 37 ° C constant temperature 120R / min oscillation digestion 30 ~ 35min, observe the tissue digestion at any time, during which the mix is ​​removed every 5 to 8 minutes, and then use 100 mesh and 20...

Example Embodiment

[0057] Example 2

[0058] The object of this embodiment is to separate and purify the human fatty mixed stem cells of human fatty mesenchymal stem cells, including the following steps:

[0059] Material method

[0060] Preparing human fat (normal human fat after surgery, signed an informed consent), PBS, BSA, IV collagenase, fetal bovine serum, CD9, CD63, monoclonal antibody of CD81, α-Actinin-4 and CD40 Dimethylene deargylglycine.

[0061] 2. Collect human fat mesenchymal stem cell exosomes

[0062] (1) Collect human fat mesenchymal stem cells, inoculate, serum medium with no external body at 37 ° C, 5% CO 2 Cultivate under conditions;

[0063] (2) After 12 h, the serum medium containing 300 μmol / L dimethylactylglycine-containing immobilized stem cell exosomeric body was exchanged, and 36 h was continued to collect cell culture fluid;

[0064] (3) The cell culture liquid obtained in step (2) was discarded at 4 ° C, i.e., at 1500 g of centrifugation for 10 min, then centrifuged ...

Example Embodiment

[0079] Example 3

[0080] The object of this embodiment is to cultivate H-MSCS cell lines.

[0081] Material

[0082] Prepare human fat (normal human fat after surgery, have signed informed consent), PBS, BSA, IV collagenase, fetal bovine serum, CD29 + CD44 + , CD90 + CD34 - And CD45 - Monoclonal antibody.

[0083] 2.H-MSCS pass

[0084] (1) After the method shown in Example 1 was collected, the cells were cultured in the method to 80% to 90% in accordance with the method described in Example 1 in Example 1. At 0.25% trunca-0.02% EDTA, the cell precipitate was obtained after centrifugation;

[0085] (2) Cell precipitation with fresh non-subcontinent medium retransmitted, in accordance with 1: 2 to 4 passage, cultured to P10, each generation of cells named P1-H-MSCS, P2-H-MSCS, P3- H-MSCS, P4-H-MSCs, P5-H-MSCs, P6-H-MSCs, P7-H-MSCs, P8-H-MSCs, P9-H-MSCs and P10-H-MSCs;

[0086] (3) collecting P2-h-MSCs, P5-h-MSCs, P8-h-MSCs and P10-h-MSCs, were digested into single cell suspensions w...

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Abstract

The invention provides a preparation method of human adipose mesenchymal stem cell exosome for treating myocardial infarction, and relates to the technical field of biology. The preparation method ofthe human mesenchymal stem cell exosome for treating myocardial infarction provided by the invention comprises the following steps: culturing human adipose mesenchymal stem cells, and collecting a cell culture solution; and carrying out differential centrifugation on the cell culture solution, collecting the supernatant, carrying out ultracentrifugation on the supernatant to obtain target microspheres, and then filtering, purifying and sterilizing the target microspheres to obtain the human adipose mesenchymal stem cell exosome for treating myocardial infarction. According to the present invention, the advantage is that the human adipose mesenchymal stem cell exosome is prepared by culturing human adipose mesenchymal stem cells, and the human adipose mesenchymal stem cell exosome containsvarious RNAs and cytokines, so that infarcted myocardial cells are effectively repaired, the cardiac function is enhanced, compensatory myocardial load is reduced, and heart failure is relieved; and atreatment medicine or a treatment strategy is provided for myocardial infarction.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for preparing human adipose-derived mesenchymal stem cell exosomes for treating myocardial infarction. Background technique [0002] Coronary atherosclerotic heart disease is due to abnormal lipid metabolism. The lipids in the blood deposit on the smooth arterial intima, and gradually accumulate to form white plaques, causing vascular stenosis or blockage, which can lead to severe Coronary artery occlusion, blood flow interruption, partial necrosis of myocardium due to severe persistent ischemia, resulting in myocardial infarction. After myocardial infarction, some myocardial cells are difficult to repair due to ischemia and hypoxia, resulting in reduced cardiac function and increased compensatory myocardial load, which will lead to serious complications such as heart failure over time. During the onset of coronary heart disease, the coronary artery will be blocked for 20-3...

Claims

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Application Information

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IPC IPC(8): C12N5/0775A61P9/10
CPCC12N5/0667A61P9/10C12N2509/00C12N2509/10
Inventor 陈良万柴天赐
Owner FUJIAN MEDICAL UNIV UNION HOSPITAL
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