Human umbilical cord mesenchymal stem cell membrane granules, preparation and applications thereof

A technology of stem cells and human umbilical cord, applied in the field of synthetic biology, can solve problems such as the limitation of clinical application of stem cells, and achieve the effect of avoiding immune response

Inactive Publication Date: 2017-12-08
SHANTOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] It has been reported that stem cells may become cancerous, so concerns about the biological safety of stem cells have severely limited the clinical application of stem cells

Method used

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  • Human umbilical cord mesenchymal stem cell membrane granules, preparation and applications thereof
  • Human umbilical cord mesenchymal stem cell membrane granules, preparation and applications thereof
  • Human umbilical cord mesenchymal stem cell membrane granules, preparation and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] The preparation method of human umbilical cord mesenchymal stem cell membrane particles mainly comprises the following steps:

[0037] (1) Healthy full-term umbilical cord and amniotic membrane delivered by caesarean section were obtained from the Department of Obstetrics and Gynecology of the Second Affiliated Hospital of Shantou University School of Medicine with the consent of the family members and the approval of the school ethics committee;

[0038] (2) After washing with PBS to remove red blood cells, cut Wharton's jelly into small pieces of 3-4cm, remove blood vessels, cut into smaller pieces, and transfer to a 10cm culture plate, and incubate for 30 minutes until the tissue can be attached on the culture plate;

[0039] (3) Carefully add 5ml of high-sugar DMEM (10% fetal bovine serum, 100mg / ml penicillin, 100mg / ml streptomycin) culture solution, cultivate in a 5% CO2 humidified incubator at 37 degrees, replace the culture solution every two days, 5 - Fibrous c...

Embodiment 2

[0045] Collect 2x10 according to the method of embodiment 1 6 Human umbilical cord mesenchymal stem cells were resuspended in 200 μl of culture medium, and then added 1:50 diluted PE fluorescently labeled antibodies, including CD13, CD29, CD44, CD73, HLA-ABC, CD45, CD117, CD31, CD34 and HLA-DR antibody was incubated at 4°C for 1-2h, fixed with 1% paraformaldehyde, and then analyzed by flow cytometry. Analysis results such as figure 2 As shown, the human umbilical cord mesenchymal stem cells cultured in Example 1 express typical undifferentiated cell surface marker proteins, such as CD13, CD29, CD44, CD73 and HLA-ABC, but do not express cell differentiation coding marker proteins, such as CD45 , CD117, CD31, CD34 and HLA-DR.

Embodiment 3

[0047] Collect 2x10 6 Human umbilical cord mesenchymal stem cells were resuspended in 200 μl of culture medium, and then added 1:50 diluted PE fluorescently labeled antibodies, including CD13, CD29, CD44, CD73, HLA-ABC, CD45, CD117, CD31, CD34 and HLA-DR antibody, incubated at 4°C for 2h, washed 3 times, and then resuspended in 3mL culture medium. Place the polycarbonate membrane with a pore size of 3 μm in a 25mm replaceable membrane syringe filter, and tighten the filter, draw the cell suspension with a 10ml syringe, and forcefully push the cells from one side of the filter to the other. Collect in a 15ml centrifuge tube.

[0048] Collect the obtained human umbilical cord mesenchymal stem cell membrane particles, centrifuge at 1000g for 10min, gently absorb the supernatant, leave about 200μl of culture medium, and gently shake the centrifuge tube to obtain human umbilical cord mesenchymal stem cells with a size of about 1-5μm. Stem cell membrane granules. The obtained hum...

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Abstract

The invention relates to human umbilical cord mesenchymal stem cell membrane granules, preparation and applications thereof. The surfaces of the cell membrane granules express surface markers of human umbilical cord mesenchymal stem cells and wrap biomacromolecules and organelles; the biomacromolecules comprise mRNA products of dry genes of the human umbilical cord mesenchymal stem cells; the organelles comprise mitochondria, proteasome, lysosome and autophagosome; and the cell membrane granules does not contain cell nucleuses, cell nucleuse DNA and nucleoproteins as well as virus glycoproteins VSV-G. 1-5mum cell membrane granules can be formed by a filtering membrane with the pore diameter of 3mum in mechanical extrusion, are larger, can be used for delivering the biomacromolecules and the organelles in vitro and delivering the biomacromolecules and the organelles to the liver and the spleen, and has a function of repairing aged cells and injured cells. The human umbilical cord mesenchymal stem cell membrane granules, the preparation and the applications have the advantages that due to no containing of the VSV-G, the generation of immunoreactions for viral proteins is avoided; and due to no containing of nuclear DNA, the probability of inducing canceration of target cells is almost zero.

Description

technical field [0001] The invention relates to synthetic biology technology, in particular to human umbilical cord mesenchymal stem cell membrane particles and their preparation and application in delivering biological macromolecules and organelles. Background technique [0002] Recently, a controversial mitochondrial replacement therapy was approved by the UK government. The method, based on nuclear transfer technology, is used to treat infertility problems in elderly women and can also be used to solve the dilemma caused by eggs carrying lethal mitochondrial DNA mutations. In addition, diseases associated with aging, such as atherosclerosis, type 2 diabetes, Alzheimer's disease, and cancer, are often multifaceted. Studies have shown that lipid accumulation, amyloid deposition, retention of misfolded proteins in the endoplasmic reticulum, and damaged proteasomes, autophagosomes, and mitochondria all contribute to the onset and progression of aging diseases. At present, t...

Claims

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Application Information

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IPC IPC(8): C12N5/0775A61K47/46
CPCA61K47/46C12N5/0665C12N2509/00
Inventor 魏炽炬杨旅军李爽林美嘉胥立群
Owner SHANTOU UNIV
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