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Adipose tissue source exosome gel, preparation method and application

A technology of adipose tissue and exosomes, which is applied in the preparation and preparation of exosome glue derived from adipose tissue, and the application in promoting fat regeneration in vivo. In the field of exosomes derived from adipose tissue, it can solve the problems of difficulty, protein and lipid pollution, Safe and harmless, to avoid time-consuming, improve extraction efficiency, and overcome immunogenicity

Active Publication Date: 2017-05-17
成都世联康健生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, there are various extraction methods of adipose tissue exosomes, such as ultracentrifugation, membrane filtration, immunomagnetic beads and other methods. During the extraction process, there will always be technical problems and difficulties of contamination by proteins and lipids from other sources.
[0006] There are also many difficulties in the application of adipose tissue exosomes. For example, the first is the screening of exosome scaffold materials. The ideal scaffold material must first be safe and harmless, and be able to adhere to exosomes. Does not affect the microenvironment of the host in vivo; secondly, because there will be a series of physical and chemical operations in the extraction process of exosomes, it is difficult to ensure their safety and harmlessness

Method used

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  • Adipose tissue source exosome gel, preparation method and application
  • Adipose tissue source exosome gel, preparation method and application
  • Adipose tissue source exosome gel, preparation method and application

Examples

Experimental program
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example 1

[0046] Example 1 The acquisition of adipose tissue exosomes ( figure 1 )

[0047] Step 1. Processing of adipose tissue

[0048] The bodies of 4-week-old SD rats were sacrificed, sterilized with 75% ethanol Ⅰ for 30 minutes, and 75% ethanol Ⅱ for 30 minutes. Transfer to the aseptic operating table, make an inguinal incision to obtain the bilateral inguinal adipose tissue of SD rats, rinse with sterile PBS, and cut the adipose tissue into small pieces (1-2mm) with tissue scissors. 3 ), it is advisable that the normal extraction of the Pasteur tube can be performed.

[0049] Step 2. Acquisition of Adipose Tissue Extract

[0050] Transfer the shredded adipose tissue obtained in step 1 to a Celstir suspension culture bottle, add serum-free α-MEM medium, the volume ratio of adipose tissue to medium is 1:3.5, keep the speed of suspension culture at 100rpm, store At 37°C, 5% CO 2 Three days later, filter the adipose tissue through a 150-mesh cell sieve to collect the supernatant....

example 2

[0053] Example 2 Identification of adipose tissue-derived exosomes

[0054] Step 1. Morphological Identification by Transmission Electron Microscopy

[0055] Exosomes were fixed with 1% glutaraldehyde, overnight at 4°C. After washing off the fixative, the mixed exosomes were dropped onto the sample-loading copper grid, and then water phosphotungstic acid was added dropwise to negatively stain the exosomes for 1 min. Transmission electron microscopy (Hitachi H-7650) imaging and taking pictures, a circular vesicle-like structure with a diameter of about 100nm can be seen ( Figure 2A ).

[0056] Step 2. Particle size distribution identification

[0057] Take 50 microliters of sample and inject it into a dedicated channel to measure the particle size distribution of exosomes. It can be seen that the peak of the particle size distribution is around 100nm, and the particle size is mainly distributed between 60-150nm ( Figure 2B ). The results proved that the obtained samples w...

example 3

[0058] Example 3 Preparation of matrigel composite adipose tissue-derived exosome implants

[0059] Put the standard Matrigel on ice and thaw overnight in a refrigerator at 4°C. At the same time, pre-cool the required pipette tip and 1ml disposable medical syringe. 200 micrograms of exosome pellet obtained by fresh centrifugation were resuspended in 200 microliters of melted liquid Matrigel (final concentration of exosomes was 1 mg / ml). The obtained liquid matrigel mixed with adipose tissue-derived exosomes was transferred into a pre-cooled 1ml disposable medical syringe, and placed on ice for later use.

[0060] The standard type of matrigel refers to the basic type of matrigel that does not interfere with the proteins and growth factors contained in the matrigel, simulating the structure, composition, physical properties and functions of the cell basement membrane in vivo.

[0061] The volume dilution of Matrigel should not be too high, for example, it should not be greater...

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Abstract

The invention belongs to the technical field of tissue engineering and particularly relates to adipose tissue source exosome gel, a preparation method and application. The adipose tissue source exosome gel is characterized by comprising an adipose tissue exosome and a solvent in a mass-volume ratio of (1-3):1, wherein an aquogel is served as the solvent. The preparation method comprises the following steps: treating an adipose tissue, acquiring an adipose tissue extract, acquiring the adipose tissue source exosome and preparing the adipose tissue source exosome gel. The adipose tissue source exosome gel is mainly used for promoting the fat regeneration. The invention particularly relates to the exosome extracted from the adipose tissue. When the exosome is applied to the nude mouse body, a new mature adipose tissue can be generated. The defects of insufficient seed cell resource, potential malignant transformation and tumor formation of the traditional tissue engineering can be overcome; the exosome has the advantage of easiness in storage and transportation; a new thought is supplied for acquiring the fat for the tissue engineering.

Description

technical field [0001] The invention belongs to the technical field of tissue engineering, and in particular relates to adipose tissue-derived exosome glue and its preparation method and application, in particular to the application of adipose tissue-derived exosomes in promoting fat regeneration in vivo. Background technique [0002] Soft tissue defects often occur in trauma, tumor resection, congenital malformation, etc. In the field of plastic surgery, a large amount of soft tissue repair and reconstruction of the patient's appearance and function are required. The emergence of tissue engineering technology has opened a new milestone for plastic and cosmetic repair. Traditional tissue engineering believes that tissue regeneration requires three key elements, namely, seed cells, scaffold materials, and the microenvironment required for regeneration. Due to the wide range of sources of adipose tissue, the convenience of obtaining and not being restricted by medical ethics,...

Claims

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Application Information

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IPC IPC(8): C12N5/0775C12N5/071A61L27/36A61L27/52
CPCA61L27/3604A61L27/3683A61L27/52A61L2430/34C12N5/0653C12N5/0656C12N5/0667
Inventor 田卫东于湄戴敏佳张岩
Owner 成都世联康健生物科技有限公司
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