Adipose tissue-derived composite cell group and preparation method and application thereof

An adipose tissue and mature adipocyte technology, applied in the field of stem cell therapy, can solve the problems of high cost, complicated operation, damage to cell structure, etc., and achieve the effects of easy cleaning, disinfection and sterilization, improved therapeutic effect, and high biological safety.

Pending Publication Date: 2019-02-19
瑞梦德医药科技(北京)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0002] The prior art uses the enzyme digestion method to prepare composite cell populations derived from adipose tissue, but there are many shortcomings in the enzyme digestion method, which limits the clinical application of multicellular components derived from adipose tissue, as follows:
[0003] 1. Collagenase (bovine origin) and collagenase combined with trypsin (pig origin) are commonly used for enzymatic digestion to digest collagen and protein between fat cells to obtain the cell components of the compound cell group derived from adipose tissue, and use bovine Serum albumin (BSA) terminates the reaction of trypsin and uses animal-derived digestive enzymes, which is complicated to operate, high in cost, and poses a biological safety hazard
[0004] 2. Enzyme digestion method needs to add human serum albumin during separation in order to ensure cell viability, using drugs from scarce human resources, and the cost is high
[0005] 3. A large number of red clumps can be seen in the cells separated by enzymatic digestion, and a large number of round cells with uniform shape can be seen under the microscope. The current method to remove red blood cell contamination is to use red blood cell lysate and use the principle of hypotonicity to lyse red blood cells, or use lymphocytes The separation liquid is passed through the density gradient centrifugation method, the reagents used in the two methods are not medical products, the red blood cell pollution is serious, and the cost is high
Similarly, ultrasonic cavitation destroys cells indiscriminately, damages target products such as mesenchymal stem cells, destroys cell structure, and affects the therapeutic effect of products

Method used

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  • Adipose tissue-derived composite cell group and preparation method and application thereof
  • Adipose tissue-derived composite cell group and preparation method and application thereof
  • Adipose tissue-derived composite cell group and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] Example 1, Preparation of Adipose Tissue-derived Compound Cell Population

[0074] 1. Washing of fatty tissue

[0075] (1) In the ultra-clean bench, push the fat mixture obtained from clinical liposuction into sterilized 50ml centrifuge tubes, 40ml in each tube, and gently invert 3 times to mix; the fat mixture is obtained by using a 2.5mm ultra-fine liposuction needle;

[0076] (2) Centrifuge at room temperature, the speed of centrifugation is 700rpm / min, and the time is 1min;

[0077] (3) Take the adipose tissue from the upper layer in the ultra-clean bench and transfer it to another sterile 50ml centrifuge tube, add 20ml of adipose tissue to each tube, and add an equal amount of normal saline to 40ml;

[0078] (4) Centrifuge at room temperature, the speed of centrifugal washing is 800rpm / min, and the time is 1min;

[0079] (5) After centrifugation, if the physiological saline is not clarified, repeat steps 3 and 4, and wash the adipose tissue in the centrifuge tube...

Embodiment 2

[0097] Example 2, Preparation of Adipose Tissue-derived Compound Cell Population

[0098] 1. Washing of fatty tissue

[0099] (1) In the ultra-clean bench, push the fat mixture obtained from clinical liposuction into sterilized 50ml centrifuge tubes, 40ml in each tube, and gently invert 3 times to mix; the fat mixture is obtained by using a 2.5mm ultra-fine liposuction needle;

[0100] (2) centrifugation at room temperature, the speed of centrifugation is 1200rpm / min, and the time is 5min;

[0101] (3) Take the adipose tissue from the upper layer in the ultra-clean bench and transfer it to another sterile 50ml centrifuge tube, add 20ml of adipose tissue to each tube, and add an equal amount of normal saline to 40ml;

[0102](4) Centrifuge at room temperature, the speed of centrifugal washing is 1500rpm / min, and the time is 5min;

[0103] (5) After centrifugation, if the physiological saline is not clarified, repeat steps 3 and 4, and wash the adipose tissue in the centrifuge...

Embodiment 3

[0121] Example 3, Preparation of Adipose Tissue-derived Compound Cell Population

[0122] 1. Cleanse fatty tissue

[0123] (1) In the ultra-clean bench, push the fat mixture obtained from clinical liposuction into sterilized 50ml centrifuge tubes, 40ml in each tube, and gently invert 3 times to mix; the fat mixture is obtained by using a 2.5mm ultra-fine liposuction needle;

[0124] (2) centrifugation at room temperature, the speed of centrifugation is 1000rpm / min, and the time is 3min;

[0125] (3) Take the adipose tissue from the upper layer in the ultra-clean bench and transfer it to another sterile 50ml centrifuge tube, add 20ml of adipose tissue to each tube, and add an equal amount of normal saline to 40ml;

[0126] (4) Centrifuge at room temperature, the speed of centrifugal washing is 1200rpm / min, and the time is 3min;

[0127] (5) After centrifugation, if the physiological saline is not clarified, repeat steps 3 and 4, and wash the adipose tissue in the centrifuge t...

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Abstract

The invention discloses an adipose tissue-derived composite cell group and a preparation method and an application thereof. The preparation method of the adipose tissue-derived composite cell group includes the following steps: (1) washing adipose tissue; (2) separating the adipose tissue, wherein the washed adipose tissue sequentially passes through three blade-type nano adipose transplantation emulsifiers with different pore diameters of 1.5 mm-2.0 mm, 0.8 mm-1.0 mm and 0.5 mm-0.7 mm, removing an oil drop layer, a mature adipose cell layer and a physiological saline layer after centrifugation, and collecting the composite cell group layer at the bottom; filtering the composite cell group layer with a cell sieve to remove cell fragments and base substance components in the composite cellgroup layer, centrifuging, and discarding supernatant to obtain the adipose tissue-derived composite cell group. The adipose tissue is crushed under the cutting action, the acting force on the adiposetissue is reduced, and damage to target products such as mesenchymal stem cells and the like is avoided, so that the cell structure is protected, and the treatment effect of the products is improved.

Description

technical field [0001] The invention belongs to the field of stem cell therapy, and in particular relates to a compound cell group derived from adipose tissue and its preparation method and application. Background technique [0002] In the prior art, adipose tissue-derived composite cell populations are prepared by enzymatic digestion, but there are many deficiencies in the enzymatic digestion method, which limits the clinical application of multicellular components derived from adipose tissue, as follows: [0003] 1. Collagenase (bovine origin) and collagenase combined with trypsin (pig origin) are commonly used for enzymatic digestion to digest collagen and protein between fat cells to obtain the cell components of the adipose tissue-derived compound cell group, and use bovine Serum albumin (BSA) terminates the reaction of trypsin and uses animal-derived digestive enzymes, which is complicated to operate, high in cost, and poses a biological safety hazard. [0004] 2. In ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/077A61K35/35A61P19/02A61P29/00A61P19/08A61P19/04
CPCA61K35/35A61P19/02A61P19/04A61P19/08A61P29/00C12N5/0653C12N2509/00
Inventor 蔡哲
Owner 瑞梦德医药科技(北京)有限公司
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