CN115462370A - Gel solution for preserving umbilical cord near totipotent stem cells and application thereof - Google Patents
Gel solution for preserving umbilical cord near totipotent stem cells and application thereof Download PDFInfo
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- CN115462370A CN115462370A CN202211368066.7A CN202211368066A CN115462370A CN 115462370 A CN115462370 A CN 115462370A CN 202211368066 A CN202211368066 A CN 202211368066A CN 115462370 A CN115462370 A CN 115462370A
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- umbilical cord
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- 210000003954 umbilical cord Anatomy 0.000 title claims abstract description 28
- 210000003014 totipotent stem cell Anatomy 0.000 title claims abstract description 23
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims abstract description 39
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 18
- 239000008367 deionised water Substances 0.000 claims abstract description 16
- 229910021641 deionized water Inorganic materials 0.000 claims abstract description 16
- 239000002184 metal Substances 0.000 claims abstract description 16
- 229910052751 metal Inorganic materials 0.000 claims abstract description 16
- 239000000843 powder Substances 0.000 claims abstract description 16
- IXPNQXFRVYWDDI-UHFFFAOYSA-N 1-methyl-2,4-dioxo-1,3-diazinane-5-carboximidamide Chemical compound CN1CC(C(N)=N)C(=O)NC1=O IXPNQXFRVYWDDI-UHFFFAOYSA-N 0.000 claims abstract description 14
- 241001312219 Amorphophallus konjac Species 0.000 claims abstract description 14
- 235000001206 Amorphophallus rivieri Nutrition 0.000 claims abstract description 14
- 108091003079 Bovine Serum Albumin Proteins 0.000 claims abstract description 14
- 229920002752 Konjac Polymers 0.000 claims abstract description 14
- 239000000252 konjac Substances 0.000 claims abstract description 14
- 235000010485 konjac Nutrition 0.000 claims abstract description 14
- 239000000661 sodium alginate Substances 0.000 claims abstract description 14
- 235000010413 sodium alginate Nutrition 0.000 claims abstract description 14
- 229940005550 sodium alginate Drugs 0.000 claims abstract description 14
- LUEWUZLMQUOBSB-FSKGGBMCSA-N (2s,3s,4s,5s,6r)-2-[(2r,3s,4r,5r,6s)-6-[(2r,3s,4r,5s,6s)-4,5-dihydroxy-2-(hydroxymethyl)-6-[(2r,4r,5s,6r)-4,5,6-trihydroxy-2-(hydroxymethyl)oxan-3-yl]oxyoxan-3-yl]oxy-4,5-dihydroxy-2-(hydroxymethyl)oxan-3-yl]oxy-6-(hydroxymethyl)oxane-3,4,5-triol Chemical compound O[C@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@@H](O[C@@H]2[C@H](O[C@@H](OC3[C@H](O[C@@H](O)[C@@H](O)[C@H]3O)CO)[C@@H](O)[C@H]2O)CO)[C@H](O)[C@H]1O LUEWUZLMQUOBSB-FSKGGBMCSA-N 0.000 claims abstract description 10
- 229920002581 Glucomannan Polymers 0.000 claims abstract description 10
- 239000012894 fetal calf serum Substances 0.000 claims abstract description 10
- 229940046240 glucomannan Drugs 0.000 claims abstract description 10
- 210000001778 pluripotent stem cell Anatomy 0.000 claims description 12
- 239000007788 liquid Substances 0.000 claims description 11
- 238000003756 stirring Methods 0.000 claims description 10
- BQCADISMDOOEFD-UHFFFAOYSA-N Silver Chemical compound [Ag] BQCADISMDOOEFD-UHFFFAOYSA-N 0.000 claims description 8
- 238000000034 method Methods 0.000 claims description 8
- 238000002360 preparation method Methods 0.000 claims description 5
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 claims description 4
- 238000005138 cryopreservation Methods 0.000 claims description 4
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 claims description 4
- 238000004321 preservation Methods 0.000 claims description 4
- 238000002156 mixing Methods 0.000 claims description 3
- 230000004083 survival effect Effects 0.000 abstract description 7
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical group CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 6
- 210000004027 cell Anatomy 0.000 description 6
- 239000002577 cryoprotective agent Substances 0.000 description 4
- 239000012091 fetal bovine serum Substances 0.000 description 4
- 238000007710 freezing Methods 0.000 description 4
- 230000008014 freezing Effects 0.000 description 4
- 210000000130 stem cell Anatomy 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 239000013078 crystal Substances 0.000 description 2
- 230000006378 damage Effects 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 238000010257 thawing Methods 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 210000001671 embryonic stem cell Anatomy 0.000 description 1
- 210000004700 fetal blood Anatomy 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 230000003834 intracellular effect Effects 0.000 description 1
- 230000003902 lesion Effects 0.000 description 1
- 230000037323 metabolic rate Effects 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000002107 myocardial effect Effects 0.000 description 1
- 210000001178 neural stem cell Anatomy 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 210000004409 osteocyte Anatomy 0.000 description 1
- 210000002826 placenta Anatomy 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 239000003223 protective agent Substances 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 230000008929 regeneration Effects 0.000 description 1
- 238000011069 regeneration method Methods 0.000 description 1
- 230000008439 repair process Effects 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 238000004017 vitrification Methods 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N1/00—Preservation of bodies of humans or animals, or parts thereof
- A01N1/02—Preservation of living parts
- A01N1/0205—Chemical aspects
- A01N1/021—Preservation or perfusion media, liquids, solids or gases used in the preservation of cells, tissue, organs or bodily fluids
- A01N1/0221—Freeze-process protecting agents, i.e. substances protecting cells from effects of the physical process, e.g. cryoprotectants, osmolarity regulators like oncotic agents
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N1/00—Preservation of bodies of humans or animals, or parts thereof
- A01N1/02—Preservation of living parts
- A01N1/0205—Chemical aspects
- A01N1/021—Preservation or perfusion media, liquids, solids or gases used in the preservation of cells, tissue, organs or bodily fluids
- A01N1/0226—Physiologically active agents, i.e. substances affecting physiological processes of cells and tissue to be preserved, e.g. anti-oxidants or nutrients
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N1/00—Preservation of bodies of humans or animals, or parts thereof
- A01N1/02—Preservation of living parts
- A01N1/0205—Chemical aspects
- A01N1/0231—Chemically defined matrices, e.g. alginate gels, for immobilising, holding or storing cells, tissue or organs for preservation purposes; Chemically altering or fixing cells, tissue or organs, e.g. by cross-linking, for preservation purposes
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Dentistry (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Environmental Sciences (AREA)
- Dispersion Chemistry (AREA)
- Chemical & Material Sciences (AREA)
- Biophysics (AREA)
- Physiology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
The invention discloses a gel solution for storing umbilical cord near totipotent stem cells and application thereof, wherein the gel solution comprises the following components: 20 to 40 parts of sodium alginate, 100 to 150 parts of glycerol, 10 to 20 parts of konjac glucomannan, 10 to 20 parts of heat-conducting metal powder, 700 to 1000 parts of fetal calf serum and 100 to 200 parts of deionized water. The invention can ensure that the near totipotent stem cells of the umbilical cord can be stored in a low-temperature environment for a long time, and can obtain a larger survival rate after the near totipotent stem cells of the umbilical cord are thawed.
Description
Technical Field
The invention relates to the technical field of stem cell preservation, in particular to a gel solution for preserving umbilical cord near totipotent stem cells and application thereof.
Background
The near totipotent stem cells are sub totipotent stem cells, the near totipotent stem cells extracted from placenta, umbilical cord and umbilical cord blood are close to embryonic stem cells, have the biological characteristics of the totipotent stem cells, can be directionally differentiated into epithelial stem cells, neural stem cells, myocardial cells, osteocytes and the like, realize the regeneration repair and functional reconstruction of various tissue lesions or injuries of organisms, and expand a new treatment technical scheme for the treatment of diseases.
At present, the preservation mode of the totipotent stem cells generally adopts a cryoprotectant for low-temperature preservation, the cryoprotectant is a certain micromolecule or high molecular compound used for preventing the cells from forming ice crystals to cause damage in the freezing and thawing processes, the commonly used cryoprotectant is DMSO, but the DMSO has certain toxicity to the cells, intracellular proteins can be denatured at normal temperature, and the totipotent stem cells can be damaged before freezing and after thawing to influence the cell survival rate. Therefore, there is a need for a cryoprotectant solution that can maintain high survival rates during cryopreservation of pluripotent stem cells.
Disclosure of Invention
The invention aims to provide a gel solution for storing umbilical cord pluripotent stem cells and application thereof, and the gel solution can solve the problem of low survival rate of the umbilical cord pluripotent stem cells after low-temperature storage.
The technical purpose of the invention is realized by the following technical scheme:
a gel solution for preserving the near totipotent stem cells of the umbilical cord comprises the following components: 20 to 40 parts of sodium alginate, 100 to 150 parts of glycerol, 10 to 20 parts of konjac glucomannan, 10 to 20 parts of heat-conducting metal powder, 700 to 1000 parts of fetal calf serum and 100 to 200 parts of deionized water.
Preferably, the composition comprises the following components: 30 parts of sodium alginate, 120 parts of glycerol, 15 parts of konjac glucomannan, 15 parts of heat-conducting metal powder, 850 parts of fetal calf serum and 150 parts of deionized water.
Preferably, the heat conducting metal powder is one of silver powder, copper powder and gold powder.
Preferably, the average diameter of the thermally conductive metal powder is in the range of 50 to 200 μm.
The invention further provides application of the gel solution for preserving the umbilical cord near totipotent stem cells in cryopreservation of the umbilical cord near totipotent stem cells.
In another aspect, the present invention provides a method for preparing a gel solution for preserving umbilical cord near totipotent stem cells, comprising: the method comprises the following steps:
step one, dissolving konjac sugar in partial deionized water to obtain transparent viscous liquid, mixing the transparent viscous liquid with fetal calf serum, stirring uniformly, and storing for 12-24h in a sterile environment at 4 ℃.
Step two, dissolving sodium alginate in the rest deionized water, adding the obtained viscous liquid into the solution obtained in the step one, and stirring uniformly.
And step three, pouring heat-conducting metal powder and glycerol into the solution obtained in the step two, and uniformly stirring.
In conclusion, the invention has the following beneficial effects:
the konjac sugar is dissolved to form gel, can play a role of non-permeable freezing protective agent, can delay and reduce the absorption of the near totipotent stem cells to glucose, reduces the metabolic rate of the near totipotent stem cells before freezing, can stabilize the physiological balance of the near totipotent stem cells by fetal calf serum, can ensure that heat-conducting metal powder is uniformly dispersed in the solution after sodium alginate is gelatinized, can accelerate the speed of temperature reduction and temperature rise by means of good heat conductivity of the heat-conducting metal powder, and realizes vitrification during temperature reduction and inhibition of ice crystal formation during temperature rise.
Detailed Description
The present invention will be described in further detail below. In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the embodiments of the present invention, and it is obvious that the described embodiments are some embodiments of the present invention, but not all embodiments. The technical solution of the present invention is not limited to the following embodiments, and includes any combination of the embodiments based on the embodiments of the present invention, and all other embodiments obtained by those skilled in the art without creative efforts shall fall within the protection scope of the present invention.
Example 1:
a gel solution for preserving the near totipotent stem cells of the umbilical cord comprises the following components:
20 parts of sodium alginate, 100 parts of glycerol, 10 parts of konjac glucomannan, 10 parts of silver powder, 700 parts of fetal calf serum and 100 parts of deionized water, wherein the diameter of the silver powder is 50 micrometers.
The preparation method comprises the following steps:
dissolving konjac sugar in part of deionized water to obtain transparent viscous liquid, mixing the transparent viscous liquid with fetal calf serum, uniformly stirring, and storing for 12-24h at 4 ℃ in an aseptic environment.
And step two, dissolving sodium alginate in the residual deionized water to obtain viscous liquid, adding the viscous liquid into the solution obtained in the step one, and stirring uniformly.
And step three, pouring heat-conducting metal powder and glycerol into the solution obtained in the step two, and uniformly stirring.
Example 2:
a gel solution for preserving umbilical cord pluripotent stem cells comprises the following components:
30 parts of sodium alginate, 120 parts of glycerol, 15 parts of konjac glucomannan, 15 parts of silver powder, 850 parts of fetal bovine serum and 150 parts of deionized water, wherein the diameter of the silver powder is 120 microns.
The procedure is as in example 1.
Example 3:
a gel solution for preserving umbilical cord pluripotent stem cells comprises the following components:
40 parts of sodium alginate, 150 parts of glycerol, 20 parts of konjac glucomannan, 20 parts of silver powder, 1000 parts of fetal bovine serum and 200 parts of deionized water, wherein the diameter of the silver powder is 200 mu m.
The procedure of the preparation method is the same as in example 1.
Example 4:
a gel solution for preserving the near totipotent stem cells of the umbilical cord comprises the following components:
40 parts of sodium alginate, 150 parts of glycerol, 20 parts of konjac glucomannan, 20 parts of copper powder, 1000 parts of fetal bovine serum and 200 parts of deionized water, wherein the diameter of the copper powder is 200 microns.
The procedure of the preparation method is the same as in example 1.
Example 5:
a gel solution for preserving umbilical cord pluripotent stem cells comprises the following components:
40 parts of sodium alginate, 150 parts of glycerol, 20 parts of konjac glucomannan, 20 parts of gold powder, 1000 parts of fetal bovine serum and 200 parts of deionized water, wherein the diameter of the gold powder is 200 mu m.
The procedure of the preparation method is the same as in example 1.
Precooling the solution prepared in the examples 1-5 to 4 ℃, quantitatively adding the precooled solution into the near-totipotent stem cells of the umbilical cord, uniformly stirring to obtain cell sap, storing the cell sap in liquid nitrogen for 3 months, 6 months, 9 months and 12 months, taking out, and unfreezing the near-totipotent stem cells of the umbilical cord to calculate the survival rate, wherein the survival rate is shown in the following table:
as can be seen from the above table, the solution prepared in the embodiment of the present application can ensure a high survival rate of the umbilical cord pluripotent stem cells after long-time cryopreservation, and in the experimental process, the smaller diameter of the heat-conducting metal powder can be uniformly dispersed, so that heat conduction is facilitated, but later-stage recovery is not facilitated, while the larger diameter of the heat-conducting metal powder can be non-uniformly dispersed, so that heat conduction is not facilitated, but later-stage recovery is facilitated.
The present embodiment is only for explaining the present invention, and it is not limited to the present invention, and those skilled in the art can make modifications of the present embodiment without inventive contribution as needed after reading the present specification, but all of them are protected by patent law within the scope of the claims of the present invention.
Claims (6)
1. A gel solution for preserving the near totipotent stem cells of the umbilical cord, which is characterized in that: comprises the following components: 20 to 40 parts of sodium alginate, 100 to 150 parts of glycerol, 10 to 20 parts of konjac glucomannan, 10 to 20 parts of heat-conducting metal powder, 700 to 1000 parts of fetal calf serum and 100 to 200 parts of deionized water.
2. The gel solution for preserving umbilical cord pluripotent stem cells according to claim 1, wherein the gel solution comprises: comprises the following components: 30 parts of sodium alginate, 120 parts of glycerol, 15 parts of konjac glucomannan, 15 parts of heat-conducting metal powder, 850 parts of fetal calf serum and 150 parts of deionized water.
3. The gel solution for preserving umbilical cord pluripotent stem cells according to any one of claims 1 or 2, wherein: the heat-conducting metal powder is one of silver powder, copper powder and gold powder.
4. The gel solution for preserving umbilical cord pluripotent stem cells according to claim 3, wherein the gel solution comprises: the average diameter of the heat-conducting metal powder is within the range of 50-200 mu m.
5. Use of the gel solution for the preservation of umbilical cord pluripotent stem cells according to any one of claims 1 to 4 for the cryopreservation of umbilical cord pluripotent stem cells.
6. A preparation method of a gel solution for preserving umbilical cord near totipotent stem cells is characterized by comprising the following steps: the method comprises the following steps:
dissolving konjac sugar in part of deionized water to obtain transparent viscous liquid, mixing the transparent viscous liquid with fetal calf serum, uniformly stirring, and storing for 12-24h at 4 ℃ in an aseptic environment.
Step two, dissolving sodium alginate in the rest deionized water, adding the obtained viscous liquid into the solution obtained in the step one, and stirring uniformly.
And step three, pouring heat-conducting metal powder and glycerol into the solution obtained in the step two, and uniformly stirring.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211368066.7A CN115462370A (en) | 2022-11-03 | 2022-11-03 | Gel solution for preserving umbilical cord near totipotent stem cells and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211368066.7A CN115462370A (en) | 2022-11-03 | 2022-11-03 | Gel solution for preserving umbilical cord near totipotent stem cells and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN115462370A true CN115462370A (en) | 2022-12-13 |
Family
ID=84338273
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202211368066.7A Withdrawn CN115462370A (en) | 2022-11-03 | 2022-11-03 | Gel solution for preserving umbilical cord near totipotent stem cells and application thereof |
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| CN (1) | CN115462370A (en) |
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| WO2004083287A1 (en) * | 2003-03-13 | 2004-09-30 | Vector Usa Inc. | A film having a liquid absorbed therein |
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Application publication date: 20221213 |