CN113930393A - Method for extracting exosome from umbilical cord stem cells and preparing hydrogel - Google Patents
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- 210000001808 exosome Anatomy 0.000 title claims abstract description 57
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- 238000000034 method Methods 0.000 title claims abstract description 18
- 210000000130 stem cell Anatomy 0.000 title abstract description 10
- 210000002901 mesenchymal stem cell Anatomy 0.000 claims abstract description 61
- 239000006228 supernatant Substances 0.000 claims abstract description 25
- 238000002360 preparation method Methods 0.000 claims abstract description 19
- 239000000661 sodium alginate Substances 0.000 claims abstract description 16
- 235000010413 sodium alginate Nutrition 0.000 claims abstract description 16
- 229940005550 sodium alginate Drugs 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 15
- 239000001963 growth medium Substances 0.000 claims abstract description 13
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 claims abstract description 6
- 238000000108 ultra-filtration Methods 0.000 claims abstract description 6
- 235000010443 alginic acid Nutrition 0.000 claims abstract description 5
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- 238000001914 filtration Methods 0.000 claims abstract description 5
- FHVDTGUDJYJELY-UHFFFAOYSA-N 6-{[2-carboxy-4,5-dihydroxy-6-(phosphanyloxy)oxan-3-yl]oxy}-4,5-dihydroxy-3-phosphanyloxane-2-carboxylic acid Chemical compound O1C(C(O)=O)C(P)C(O)C(O)C1OC1C(C(O)=O)OC(OP)C(O)C1O FHVDTGUDJYJELY-UHFFFAOYSA-N 0.000 claims abstract description 4
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- 239000007640 basal medium Substances 0.000 claims description 4
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- 125000003172 aldehyde group Chemical group 0.000 description 1
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- 210000002966 serum Anatomy 0.000 description 1
- 210000004927 skin cell Anatomy 0.000 description 1
- -1 sodium alginate hyaluronic acid derivative Chemical class 0.000 description 1
- 230000029663 wound healing Effects 0.000 description 1
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Abstract
The invention discloses an extraction method of umbilical cord stem cell-derived exosomes and a preparation method of hydrogel, belongs to the technical field of stem cell processing, solves the problems of complicated separation operation of existing umbilical cord mesenchymal stem cells and preparation process of the exosome hydrogel, and comprises the following steps: primary culture of umbilical cord mesenchymal stem cells; collecting culture medium of umbilical cord mesenchymal stem cells in sections, obtaining supernatant after centrifugal filtration, and obtaining exosome of human umbilical cord mesenchymal stem cells after centrifugal ultrafiltration of the supernatant; the method comprises the steps of adding the extracted human umbilical cord mesenchymal stem cell exosomes and a calcium chloride solution into a sodium alginate solution, standing and incubating to form the alginate hydrogel of the human umbilical cord mesenchymal stem cell exosomes.
Description
Technical Field
The invention relates to the technical field of stem cell processing, in particular to the technical field of extraction of exosomes derived from umbilical cord stem cells and hydrogel preparation.
Background
Umbilical cord Mesenchymal Stem Cells (Umbilical cord Mesenchyl Stem Cells, ucMSCs) refer to a multifunctional Stem cell existing in Umbilical cord tissues of newborns, can be differentiated into a plurality of tissue Cells, and has wide clinical application prospects.
At present, methods for extracting umbilical cord mesenchymal stem cell exosomes and applying exosomes appear, for example, a chinese patent with application number of 202011184764.2 discloses a method for preparing an extracting solution of human umbilical cord mesenchymal stem cell exosomes and a method for preparing exosome cream, and the preparation method comprises the following steps: placing the human umbilical cord mesenchymal stem cells into an incubator, and culturing with DMEM/F12 culture solution; until the cells are fused by 60-70%, culturing for 12-16h by using a serum-free starvation culture medium; changing a serum-free starvation culture solution into a DMEM/F12 culture solution containing epidermal growth factor EGF, fibroblast growth factor FGF and keratinocyte growth factor KGF, culturing until cells are in a logarithmic phase, collecting cell culture supernatant, carrying out gradient centrifugation, and filtering the supernatant obtained by centrifugation by using a filter; and preparing the obtained human umbilical cord mesenchymal stem cell exosome extract and a cream base formula to obtain the cream. The prepared cream can improve cell division and secrete a large amount of extracellular matrix to help complete the renewal of skin cells and improve the cell state.
Further, as disclosed in the Chinese patent application No. 202010225566.X, "an exosome hydrogel wound dressing and a preparation method thereof", includes mesenchymal stem cell exosomes and hydrogels, wherein the hydrogel is composed of pluronic F127 and pluronic F68; respectively preparing physiological saline solution of pluronic F127 and pluronic F68 and mesenchymal stem cell exosome physiological saline resuspension, and mixing the solutions at 4 ℃ to obtain the exosome hydrogel wound dressing. Compared with the method that the mesenchymal stem cell exosome and hydrogel are independently adopted, the wound dressing prepared by the method can better promote the regeneration of skin wound cells and shorten the wound healing time.
In addition, the Chinese patent with application number 201910623484.8 discloses an injectable bone repair hydrogel containing human umbilical cord mesenchymal stem cell exosomes and a preparation method thereof, wherein the hydrogel material contains human umbilical cord mesenchymal stem cell exosomes, nano-hydroxyapatite, a hyaluronic acid-adipic acid dihydrazide derivative and an alginic acid derivative with an aldehyde group on a main chain, the preparation method comprises the steps of separation of human umbilical cord mesenchymal cells, extraction of exosomes, preparation of nano-hydroxyapatite, preparation of sodium alginate hyaluronic acid derivative, preparation of composite hydrogel and the like, and the technical scheme provides a new repair method for bone defect treatment and has wide application prospect and ideal repair effect.
The above patent documents all have the following problems: 1. the separation operation steps of the umbilical cord mesenchymal stem cells are complex, and the concentration of the extracted exosomes is low; 2. the preparation process of the exosome hydrogel is complex, has more components, and is not beneficial to the formation of a subsequent composite preparation. Therefore, the method for extracting exosomes from umbilical cord stem cells and preparing hydrogel by optimizing and improving the existing method has very important significance for obtaining umbilical cord mesenchymal stem cells, establishing a mesenchymal stem cell bank and clinically applying the umbilical cord mesenchymal stem cells.
Disclosure of Invention
The invention aims to: in order to solve the technical problems, the invention provides a method for extracting exosomes derived from umbilical cord stem cells and preparing hydrogel.
The invention specifically adopts the following technical scheme for realizing the purpose:
a preparation method of a human umbilical cord mesenchymal stem cell exosome hydrogel comprises the following steps:
s1: dissolving sodium alginate powder in PBS buffer solution to prepare sodium alginate solution with mass concentration of 2%;
s2: adding human umbilical cord mesenchymal stem cell exosome into a sodium alginate solution, and uniformly stirring;
s3: adding a calcium chloride solution with the mass concentration of 1% into a sodium alginate solution, and incubating for 10min at 37 ℃ to form the alginate hydrogel of the human umbilical cord mesenchymal stem cell exosome with the concentration of 0.2 mug/muL.
The volume ratio of the calcium chloride solution to the sodium alginate solution is 1: 4.
The extraction method of the human umbilical cord mesenchymal stem cell exosomes in the step S2 comprises the following steps:
s201: placing human umbilical cord mesenchymal stem cells into an incubator, and using DMEM/F12 basal medium at 37 ℃ and 5% CO2Culturing under the condition;
s202: when the cells are 80% confluent, culturing the human umbilical cord mesenchymal stem cells in a serum-free culture medium for 24-72 h, and collecting the culture medium in sections;
s203: centrifuging the collected culture medium at 4 deg.C for 15min at 1000 Xg, removing cell debris and dead cells, collecting supernatant, centrifuging the supernatant at 4 deg.C for 30min at 12000 Xg for further removing cell debris, collecting supernatant, and filtering;
s204: centrifuging the filtered supernatant at 150000 Xg for 1.5h at 4 ℃, immediately sucking out the supernatant after centrifugation, washing the precipitate with PBS, centrifuging again at 150000 Xg and 4 ℃ for 1.5h, collecting the supernatant, combining the supernatants collected after two times of centrifugation, and performing ultrafiltration to obtain the human umbilical cord mesenchymal stem cell exosome;
s205: and newly suspending the obtained exosome of the human umbilical cord mesenchymal stem cells in PBS to obtain an exosome extracting solution of the human umbilical cord mesenchymal stem cells.
In the step S201, the human umbilical cord mesenchymal stem cells are primary umbilical cord tissue blocks, the umbilical cord tissue is washed by sterile PBS and 75% alcohol, cut into 1-3 mm-sized pasty tissue, and then cultured on a DMEM/F12 basic culture medium.
The number of passages of the human umbilical cord mesenchymal stem cells in the step S202 is less than 5.
The culture time for collecting culture medium in the step S202 in sections is 24h, 48h and 72h respectively.
The invention has the following beneficial effects: the method optimizes the cell extraction process in the extraction method of the exosomes derived from the umbilical cord mesenchymal stem cells, so that the extraction efficiency of the umbilical cord mesenchymal stem cells is higher, the purity is higher, the acquisition amount of the exosomes can be ensured on the premise of acquiring more cells, the process is simple, good economic benefits and wide application prospects are realized, and the method has very important significance for the acquisition of the umbilical cord mesenchymal stem cells, the establishment of a mesenchymal stem cell bank and clinical application.
Drawings
FIG. 1 is a schematic diagram of the particle size analysis of umbilical cord mesenchymal stem cell-derived exosomes of the present invention;
FIG. 2 is a diagram of identification of an umbilical cord mesenchymal stem cell-derived exosome marker of the present invention;
FIG. 3 is a transmission electron microscope image of the exosome derived from umbilical cord mesenchymal stem cell of the present invention.
Detailed Description
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.
Thus, the following detailed description of the embodiments of the present invention is not intended to limit the scope of the invention as claimed, but is merely representative of selected embodiments of the invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1
The embodiment provides a preparation method of a human umbilical cord mesenchymal stem cell exosome hydrogel, which specifically comprises the following steps:
s1: primary culture of umbilical cord mesenchymal stem cells
S101: fresh umbilical cords of healthy lying-in women without diseases after full-term cesarean section are taken, washed by sterile PBS for a plurality of times, washed by 75% alcohol, and cut into pasty tissues with the size of 1-3mm by sterile ophthalmology after being washed clean by sterile PBS. Placed in a basal Medium containing DMEM/F12(Gibco Dulbecco's Modified Eagle Medium: F-12) at 37 ℃ with 5% CO2Culturing under the condition;
s102: when 80% confluence was reached, cells were trypsinized for passage;
s103: culturing the human umbilical cord mesenchymal stem cells with the passage times less than 5 in a serum-free culture medium for 24-72 h, and collecting the culture medium in sections;
s2: hUCMSC exosome extraction
S201: culturing the human umbilical cord mesenchymal stem cells with the passage times less than 5 times in a culture medium without exosome serum, and continuously collecting the culture medium for 24h, 48h and 72 h;
s202: centrifuging the collected culture medium at 4 deg.C for 15min at 1000 Xg, removing cell debris and dead cells, collecting supernatant, centrifuging the supernatant at 4 deg.C for 30min at 12000 Xg for further removing cell debris, collecting supernatant, and filtering the collected supernatant with 0.2 μm filter;
s203: centrifuging the filtered supernatant at 150000 Xg at 4 ℃ for 1.5h, immediately sucking out the supernatant after centrifugation, washing the precipitate with PBS, centrifuging again at 150000 Xg and 4 ℃ for 1.5h, separating the supernatant and the precipitate, combining the supernatants collected after two times of centrifugation, performing ultrafiltration with a 3KD ultrafiltration tube, collecting the attachments on the 3KD ultrafiltration tube, combining the attachments and the precipitate to obtain the human umbilical cord mesenchymal stem cell exosomes, and suspending the obtained human umbilical cord mesenchymal stem cell exosomes in PBS again and extracting for subsequent use;
s3: preparation of human umbilical cord mesenchymal stem cell exosome hydrogel
S301: dissolving sodium alginate powder in PBS buffer solution at room temperature to prepare sodium alginate solution with mass concentration of 2%;
s302: adding 200 mu g of hUCMSC exosomes into sodium alginate solution and stirring uniformly;
s303: adding a calcium chloride solution with the mass concentration of 1% into a sodium alginate solution in a volume ratio of 1:4, and incubating at 37 ℃ for 10min to form the alginate hydrogel of the hUCMSC exosomes with the final concentration of 0.2 mug/muL.
Claims (6)
1. A preparation method of a human umbilical cord mesenchymal stem cell exosome hydrogel is characterized by comprising the following steps:
s1: dissolving sodium alginate powder in PBS buffer solution to prepare sodium alginate solution with mass concentration of 2%;
s2: adding human umbilical cord mesenchymal stem cell exosome into a sodium alginate solution, and uniformly stirring;
s3: adding a calcium chloride solution with the mass concentration of 1% into a sodium alginate solution, and incubating for 10min at 37 ℃ to form the alginate hydrogel of the human umbilical cord mesenchymal stem cell exosome with the concentration of 0.2 mug/muL.
2. The preparation method of the human umbilical cord mesenchymal stem cell exosome hydrogel according to claim 1, which is characterized in that: the volume ratio of the calcium chloride solution to the sodium alginate solution is 1: 4.
3. The method for preparing the hydrogel of human umbilical cord mesenchymal stem cell exosomes according to claim 1, wherein the method for extracting human umbilical cord mesenchymal stem cell exosomes in the step S2 specifically comprises the following steps:
s201: placing human umbilical cord mesenchymal stem cells into an incubator, and using DMEM/F12 basal medium at 37 ℃ and 5% CO2Culturing under the condition;
s202: when the cells are 80% confluent, culturing the human umbilical cord mesenchymal stem cells in a serum-free culture medium for 24-72 h, and collecting the culture medium in sections;
s203: centrifuging the collected culture medium at 4 deg.C for 15min at 1000 Xg, removing cell debris and dead cells, collecting supernatant, centrifuging the supernatant at 4 deg.C for 30min at 12000 Xg for further removing cell debris, collecting supernatant, and filtering;
s204: centrifuging the filtered supernatant at 150000 Xg for 1.5h at 4 ℃, immediately sucking out the supernatant after centrifugation, washing the precipitate with PBS, centrifuging again at 150000 Xg and 4 ℃ for 1.5h, collecting the supernatant, combining the supernatants collected after two times of centrifugation, and performing ultrafiltration to obtain the human umbilical cord mesenchymal stem cell exosome;
s205: and newly suspending the obtained exosome of the human umbilical cord mesenchymal stem cells in PBS to obtain an exosome extracting solution of the human umbilical cord mesenchymal stem cells.
4. The method for preparing the human umbilical cord mesenchymal stem cell exosome hydrogel according to claim 3, wherein the human umbilical cord mesenchymal stem cells in the step S201 are obtained by using an umbilical cord tissue block as a primary product, washing the umbilical cord tissue with sterile PBS and 75% alcohol, cutting the umbilical cord tissue into 1-3 mm-sized pasty tissue, and culturing the umbilical cord tissue on a DMEM/F12 basal medium.
5. The preparation method of the human umbilical cord mesenchymal stem cell exosome hydrogel according to claim 3, which is characterized in that: the number of passages of the human umbilical cord mesenchymal stem cells in the step S202 is less than 5.
6. The preparation method of the human umbilical cord mesenchymal stem cell exosome hydrogel according to claim 3, which is characterized in that: the culture time for collecting culture medium in the step S202 in sections is 24h, 48h and 72h respectively.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114939098A (en) * | 2022-05-19 | 2022-08-26 | 明德南加(成都)生物技术有限公司 | Exosome-loaded hydrogel and preparation method and application thereof |
| CN115463154A (en) * | 2022-10-21 | 2022-12-13 | 上海君益禾生物医学科技有限公司 | Preparation method and application of platelet lysate and exosome composite gel |
| CN115581810A (en) * | 2022-10-26 | 2023-01-10 | 济宁医学院附属医院 | Hydrogel rich in exosomes and preparation method and application thereof |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105349487A (en) * | 2015-11-16 | 2016-02-24 | 中国人民解放军第二军医大学 | Method for promoting human bone mesenchymal stem cell proliferation based on exosome |
| US20170121685A1 (en) * | 2015-11-02 | 2017-05-04 | Tigenix S.A.U. | Mesenchymal stem cell-derived exosomes and their uses |
| CN109880797A (en) * | 2019-04-08 | 2019-06-14 | 济南磐升生物技术有限公司 | A method of preparing human umbilical cord mesenchymal stem cells excretion body |
| CN112641803A (en) * | 2019-09-25 | 2021-04-13 | 深圳光彩生命工程技术有限公司 | Stem cell exosome preparation and preparation method thereof |
-
2021
- 2021-09-27 CN CN202111135546.4A patent/CN113930393A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20170121685A1 (en) * | 2015-11-02 | 2017-05-04 | Tigenix S.A.U. | Mesenchymal stem cell-derived exosomes and their uses |
| CN105349487A (en) * | 2015-11-16 | 2016-02-24 | 中国人民解放军第二军医大学 | Method for promoting human bone mesenchymal stem cell proliferation based on exosome |
| CN109880797A (en) * | 2019-04-08 | 2019-06-14 | 济南磐升生物技术有限公司 | A method of preparing human umbilical cord mesenchymal stem cells excretion body |
| CN112641803A (en) * | 2019-09-25 | 2021-04-13 | 深圳光彩生命工程技术有限公司 | Stem cell exosome preparation and preparation method thereof |
Non-Patent Citations (3)
| Title |
|---|
| JONG-MING HSU等: "Locally Applied Stem Cell Exosome-Scaffold Attenuates Nerve Injury-Induced Pain in Rats", JOURNAL OF PAIN RESEARCH * |
| SHUO YANG等: "Integration of Human Umbilical Cord Mesenchymal Stem Cells-Derived Exosomes with Hydroxyapatite-Embedded Hyaluronic Acid-Alginate Hydrogel for Bone Regeneration", ACS BIOMATER. SCI. ENG. * |
| 郭尚春: "外泌体对组织修复的作用及其机制研究", 中国博士学位论文全文数据库医药卫生科技辑 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114939098A (en) * | 2022-05-19 | 2022-08-26 | 明德南加(成都)生物技术有限公司 | Exosome-loaded hydrogel and preparation method and application thereof |
| CN114939098B (en) * | 2022-05-19 | 2023-09-26 | 明德南加(成都)生物技术有限公司 | Exosome-loaded hydrogel and preparation method and application thereof |
| CN115463154A (en) * | 2022-10-21 | 2022-12-13 | 上海君益禾生物医学科技有限公司 | Preparation method and application of platelet lysate and exosome composite gel |
| CN115581810A (en) * | 2022-10-26 | 2023-01-10 | 济宁医学院附属医院 | Hydrogel rich in exosomes and preparation method and application thereof |
| CN115581810B (en) * | 2022-10-26 | 2023-12-22 | 济宁医学院附属医院 | Hydrogel rich in exosomes as well as preparation method and application thereof |
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Application publication date: 20220114 |