CN107080753A - A kind of cosmetic formulation of human umbilical cord mesenchymal stem cells source excretion body - Google Patents
A kind of cosmetic formulation of human umbilical cord mesenchymal stem cells source excretion body Download PDFInfo
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- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
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Abstract
The invention belongs to biomedicine field, and in particular to a kind of cosmetic formulation of human umbilical cord mesenchymal stem cells source excretion body, and the preparation includes following component：Human mesenchymal stem cell source excretion body freeze-dried powder, glycerine, asparagus juice；The weight ratio of three is 1:1:10‑20；The preparation method of the preparation is：The human mesenchymal stem cell source excretion body freeze-dried powder, glycerine, asparagus juice are weighed in proportion, mixes to transparent, and then the filtering with microporous membrane through 0.45 μm, 0.22 μm successively, that is, obtain the cosmetic formulation.The present invention as active component, can be prepared a kind of cosmetic formulation for having a positive effect, can solve the problems, such as the skin aging of people using a kind of new human mesenchymal stem cell source excretion body；With great market prospects and application value.
Description
Technical field
The invention belongs to biomedicine field, and in particular to a kind of beauty system of human umbilical cord mesenchymal stem cells source excretion body
Agent.
Background technology
2013, the regulation mechanism for the cell vesicle transport that the U.S., 3 scientists of Germany are had found by them was won
Nobel prize's soul.Excretion body (exosomes) is also begun to by increasingly as the important vesica of a class in human body
Many concerns.Scientists have found that excretion, which is known from experience, participates in immune response, apoptosis, angiogenesis, inflammatory reaction, condensation etc.
In important bioprocess, some signaling molecules can be secreted into tissue or cell farther out by cell by secreting excretion body,
To play regulating and controlling effect.
2016, Jiangsu Province, China University Medical College was permitted literary honor et al. research discovery, human umbilical cord mesenchymal stem cells source excretion
The protein that body is included, can be cooperated with the signaling molecule during skin regeneration.(document：Zhang,B.,et al.
(2016)."HucMSC Exosome-Delivered 14-3-3zeta Orchestrates Self-control of the
Wnt Response via Modulation of YAP during Cutaneous Regeneration."Stem
Cells.) first, the effect of rat deepⅱdegreeburn, finder are treated by assessing human umbilical cord mesenchymal stem cells source excretion body
Umbilical cord mesenchymal stem cells source excretion body promotes self adjusting for Wnt/ β-catenin signal paths in the regeneration remodelling phase of skin
Section.Secondly, the proteome analysis of human umbilical cord mesenchymal stem cells source excretion body shows, relevant albumen can pass through excretion body
Transport.Comprehensive research result shows that human umbilical cord mesenchymal stem cells source excretion body can not only be used as Wnt/ β-catenin signals
" accelerator " can control skin regeneration to be used as the signal to promote the Regeneration and Repair of damaged skin tissue by adjusting YAP
" brake ".
Therefore, in terms of aging skin/damaged skin is repaired, with wide market prospects, but the production in the field at present
Product anti-aging effects are unsatisfactory, and finding a kind of anti-aging, good product is the very urgent thing in the current field.
The content of the invention
Therefore, the technical problems to be solved by the invention are the clear area for overcoming prior art, so as to propose a kind of anti-
The cosmetic formulation of the good human umbilical cord mesenchymal stem cells source excretion body of aging effects.
In order to solve the above technical problems, the invention discloses a kind of beauty system of human umbilical cord mesenchymal stem cells source excretion body
Agent, the preparation includes following component：Human mesenchymal stem cell source excretion body freeze-dried powder, glycerine, asparagus juice；The weight ratio of three
For 1:1:10-20.
It is preferred that, the preparation method of the preparation is：The human mesenchymal stem cell source excretion body is weighed in proportion to freeze
Powder, glycerine, asparagus juice, mix to transparent, and then the filtering with microporous membrane through 0.45 μm, 0.22 μm successively, that is, obtain institute
State cosmetic formulation.
It is preferred that, the outer membrane of the excretion body is to contain micro-RNA, institute inside phophoslipid bilayer structure, the excretion body
State at least one of mRNA, DNA and nucleic acid of excretion body source cell.
It is preferred that, the acquisition methods of described excretion body are as follows：
A) human mesenchymal stem cell is cultivated, cultivation stage added in cell culture medium interferon, EPO and
PDGF-BB；
B) stem cell after culture is pre-processed；
C) the human umbilical cord mesenchymal stem cells source excretion body is extracted in the supernatant of culture medium after the pre-treatment.
It is preferred that, in the asparagus juice, containing aloe polysaccharide, content is 700-800mg/L.
It is preferred that, the excretion body differentiates for MSC.
It is preferred that, the mescenchymal stem cell is human umbilical cord mesenchymal stem cells.
It is preferred that, the pretreatment is concretely comprised the following steps：
1) people's passage MSC is subjected to serum free suspension culture, and it is 1.0 × 10 to adjust cell concentration6/ml；
2) according to 106The big culture dish inoculations of/150mm, are cultivated to cell fusion degree to 70-80%, incubation time is 2-3
My god；
3) liquid in culture dish is absorbed, and culture dish is washed with alpha-MEM fluid nutrient mediums；
4) 20ml alpha-MEM is added again in 150mm culture dishes；
5) recombinant human interferon alpha 2, PDGF-BB and EPO are sequentially added；
7) at 37 DEG C, 5% CO2Under the conditions of cultivate 96 hours, collection obtain MSC nutrient solutions.
It is preferred that, the concentration of interferon is that 10U/ml~600U/ml, PDGF-BB concentration are in the MSC nutrient solutions
2ng/ml~60ng/ml, EPO concentration are 0.1IU/ml~5IU/ml.
It is preferred that, the step 2) pretreatment be specially：
1) supernatant of the MSC nutrient solutions is taken, centrifugal treating is carried out, cell fragment is removed；
2) and then by supernatant membrane filtration handle, remove apoptotic body, obtain pretreated supernatant.
It is preferred that, the step 3) be specially：
1) centrifuge tube of the product of trying to please for 5 parts by volume, adds the excretion body separation agent of 1 parts by volume；
2) the pretreated supernatant of process of 4 parts by volume is added, is fully mixed；
3) after stewing process, then centrifugal treating twice is carried out, obtains the excretion body.
The above-mentioned technical proposal of the present invention has advantages below compared with prior art：The present invention uses a kind of new human world
Mesenchymal stem cells source excretion body can prepare a kind of cosmetic formulation for having a positive effect as active component, can be in solution
The certainly skin aging problem of people；Preparation method is easy, be adapted to it is instant match somebody with somebody, the bioactive ingredients of excretion body are utilized well；
Prepare cost relatively low, be adapted to be used for multiple times, user can be allowed to keep cosmetic result for a long time；Material used is by national medicine
The pharmaceutic adjuvant of prison department approval, can reduce side effect, it is ensured that the skin safe of user；With great market prospects and
Application value.
Embodiment
Present embodiment discloses a kind of cosmetic formulation of human umbilical cord mesenchymal stem cells source excretion body, the system for embodiment 1
Agent includes following component：Human mesenchymal stem cell source excretion body freeze-dried powder, glycerine, asparagus juice；The weight ratio of three is 1:1:15；
In the asparagus juice, containing aloe polysaccharide, content is 750mg/L.
The preparation method of the preparation is：Weigh in proportion the human mesenchymal stem cell source excretion body freeze-dried powder, glycerine,
Asparagus juice, mixes to transparent, and then the filtering with microporous membrane through 0.45 μm, 0.22 μm successively, that is, obtain the beauty system
Agent.
Present embodiment discloses a kind of cosmetic formulation of human umbilical cord mesenchymal stem cells source excretion body, the system for embodiment 2
Agent includes following component：Human mesenchymal stem cell source excretion body freeze-dried powder, glycerine, asparagus juice；The weight ratio of three is 1:1:10；
In the asparagus juice, containing aloe polysaccharide, content is 800mg/L.
The preparation method of the preparation is：Weigh in proportion the human mesenchymal stem cell source excretion body freeze-dried powder, glycerine,
Asparagus juice, mixes to transparent, and then the filtering with microporous membrane through 0.45 μm, 0.22 μm successively, that is, obtain the beauty system
Agent.
Present embodiment discloses a kind of cosmetic formulation of human umbilical cord mesenchymal stem cells source excretion body, the system for embodiment 3
Agent includes following component：Human mesenchymal stem cell source excretion body freeze-dried powder, glycerine, asparagus juice；The weight ratio of three is 1:1:20；
In the asparagus juice, containing aloe polysaccharide, content is 700mg/L.
The preparation method of the preparation is：Weigh in proportion the human mesenchymal stem cell source excretion body freeze-dried powder, glycerine,
Asparagus juice, mixes to transparent, and then the filtering with microporous membrane through 0.45 μm, 0.22 μm successively, that is, obtain the beauty system
Agent.
The acquisition methods of excretion body mentioned in the embodiment 1 of embodiment 4 are as follows：
A. people MSC culture and pretreatment
1. material and reagent
1) people's umbilical cord, placenta, marrow, the MSC in fatty or other sources, passage number is within 6 generations.
2) people MSC serum free mediums
3)alpha-MEM
4) recombinant human interferon alpha 2
5) recombinant human epo
6) rhPDGF-BB-BB
7) culture dish (diameter 150mm)
8) other consumptive materials
2. cell culture
1) it will count, suspended using MSC Serum-free complete mediums, adjustment cell concentration is 1.0 after people's passage MSC digestion
×106/ml。
2) according to 106The big culture dish inoculations of/150mm, culture to cell fusion degree to 70-80%.Generally require 2-3 days.
3) liquid in culture dish is absorbed, and culture dish is washed with alpha-MEM fluid nutrient mediums.
4) alpha-MEM 20ml/150mm culture dishes are added.
5) recombinant human interferon alpha 2, PDGF-BB are firstly added, in 37 DEG C, 5% CO2Under the conditions of cultivate 24 hours；Then,
EPO is added, in 37 DEG C, 5% CO2Under the conditions of cultivate 72 hours.
Recombinant human interferon alpha 2 (final concentration of 10U/ml~600U/ml), PDGF-BB (2ng/ml~60ng/ml) and EPO
(0.1IU/ml~5IU/ml).
7) 37 DEG C, 5%CO2Under the conditions of cultivate 96 hours.
8) collect supernatant and carry out following handle.It such as can not in time handle, supernatant is placed in -80 DEG C of preservations.
B. the pretreatment of people MSC culture supernatants
1. material and reagent
1) consumptive material such as centrifuge tube.
2) aperture is 1 μm, 0.45 μm and 0.22 μm of filter.
2. operating procedure
1) collect and centrifuge 2000g under the conditions of cells and supernatant, room temperature or 4 DEG C, 10 minutes, remove cell fragment.
2) by culture supernatant or body fluid respectively through a diameter of 1 μm, 0.45 μm and 0.22 μm membrane filtration, to remove wherein
Apoptotic body that may be present.
C. the harvest of excretion body
1) 50ml centrifuge tubes are taken, excretion body separation agent 10ml is added.Or 250ml centrifuge tubes are taken, add excretion body reagent
50ml。
2) treated cells and supernatant 40ml is added, is fully mixed.Or add culture supernatant 200ml.
3) 4 DEG C are placed in overnight, or at least 6 hours.
4) centrifuge, 2000g, 30 minutes.
5) liquid is removed, is centrifuged again, 2000g, 5 minutes.
6) residual liquid is carefully absorbed with pipette.
7) the PBS or μ l to 1ml of physiological saline 500 is added.Or add PBS or physiological saline 2.5ml to 5ml.
8) protein concentration is determined, or carries out the Instrumental Analysis such as flow cyctometry.
D. prepared by the freeze-dried powder of gained excretion body
Gained excretion body is taken, mannitol (1%~15%) is added, mixed, is filtered (0.22 μm), packing, then be positioned over super
Low temperature refrigerator (- 80 DEG C) 10h.Then, lyophilized (- 50 DEG C, 24h) are carried out under 10Pa vacuum, obtains human mesenchyme dry thin
Born of the same parents source excretion body freeze-dried powder.
Experimental example
First, stability test
Gained cosmetic formulation is preserved under the conditions of being positioned over -20 DEG C.Observed after six months, no significant change.
2nd, validity test
Ten Mies are selected in beauty parlor, are divided into two groups, one of which tries out gained cosmetic formulation, and another group using conventional
Cosmetics.After a course for the treatment of (30 days) is completed, assay result is listed in table 1, table 2.
Feel and sensory evaluation after the use of the excretion body cosmetic formulation of table 1 group on probation
| Sequence number | Skin type and situation | Sex | Age bracket | It is invalid | Typically | Well | It is effective |
| 01 | Oiliness microgroove | Female | 30~35 | √ | |||
| 02 | Neutrality wrinkle | Female | 40~45 | √ | |||
| 03 | Neutral spot | Female | 25~30 | √ | |||
| 04 | Dryness microgroove | Female | 25~30 | √ | |||
| 05 | Dryness wrinkle spot | Female | 40~45 | √ |
Feel and sensory evaluation after the use of the conventional cosmetic preparations. Control group of table 2
| Sequence number | Skin type and situation | Sex | Age bracket | It is invalid | Typically | Well | It is effective |
| 01 | Oiliness microgroove | Female | 30~35 | √ | |||
| 02 | Neutrality wrinkle | Female | 40~45 | √ | |||
| 03 | Neutral spot | Female | 25~30 | √ | |||
| 04 | Dryness microgroove | Female | 25~30 | √ | |||
| 05 | Dryness wrinkle spot | Female | 40~45 | √ |
Consolidated statement 1, the result of table 2 are it can be found that the cosmetic formulation of human umbilical cord mesenchymal stem cells source excretion body, for each
The skin of type has good or effective effect.And compared to conventional cosmetic preparation, human umbilical cord mesenchymal stem cells
The cosmetic formulation of source excretion body has more preferable effect for the skin problem of neutral wrinkle and dryness wrinkle spot, and can releive ladies face
To wrinkle problem.
Obviously, above-described embodiment is only intended to clearly illustrate example, and the not restriction to embodiment.It is right
For those of ordinary skill in the art, can also make on the basis of the above description it is other it is various forms of change or
Change.There is no necessity and possibility to exhaust all the enbodiments.And the obvious change thus extended out or
Among changing still in the protection domain of the invention.
Claims (10)
1. a kind of cosmetic formulation of human umbilical cord mesenchymal stem cells source excretion body, it is characterised in that the preparation includes such as the following group
Point：Human mesenchymal stem cell source excretion body freeze-dried powder, glycerine, asparagus juice；The weight ratio of three is 1:1:10-20.
2. cosmetic formulation as claimed in claim 1, it is characterised in that the preparation method of the preparation is：Institute is weighed in proportion
State human mesenchymal stem cell source excretion body freeze-dried powder, glycerine, asparagus juice, mix to it is transparent, then successively through 0.45 μm,
0.22 μm of filtering with microporous membrane, that is, obtain the cosmetic formulation.
3. cosmetic formulation as claimed in claim 1 or 2, it is characterised in that the outer membrane of the excretion body is phophoslipid bilayer structure,
Contain at least one of micro-RNA, mRNA, DNA of the excretion body source cell and nucleic acid inside the excretion body.
4. cosmetic formulation as claimed in claim 3, it is characterised in that the acquisition methods of described excretion body are as follows：
A) human mesenchymal stem cell is cultivated, cultivation stage adds interferon, EPO and PDGF-BB in cell culture medium；
B) stem cell after culture is pre-processed；
C) the human umbilical cord mesenchymal stem cells source excretion body is extracted in the supernatant of culture medium after the pre-treatment.
5. the acquisition methods of excretion body as claimed in claim 4, it is characterised in that in the asparagus juice, containing aloe polysaccharide,
Content is 700-800mg/L.
6. the acquisition methods of excretion body as claimed in claim 5, it is characterised in that between the mescenchymal stem cell behaviour umbilical cord
Mesenchymal stem cells；The excretion body differentiates for MSC.
7. the acquisition methods of excretion body as claimed in claim 6, it is characterised in that the specific steps of the pretreatment
For：
1) people's passage MSC is subjected to serum free suspension culture, and it is 1.0 × 10 to adjust cell concentration6/ml；
2) according to 106The big culture dish inoculations of/150mm, culture to cell fusion degree to 70-80%, incubation time is 2-3 days；
3) liquid in culture dish is absorbed, and culture dish is washed with alpha-MEM fluid nutrient mediums；
4) 20ml alpha-MEM is added again in 150mm culture dishes；
5) recombinant human interferon alpha 2, PDGF-BB and EPO are sequentially added；
7) at 37 DEG C, 5% CO2Under the conditions of cultivate 96 hours, collection obtain MSC nutrient solutions.
8. the acquisition methods of excretion body as claimed in claim 7, it is characterised in that interferon is dense in the MSC nutrient solutions
Degree is that 10U/ml~600U/ml, PDGF-BB concentration are 2ng/ml~60ng/ml, and EPO concentration is 0.1IU/ml~5IU/
ml。
9. the acquisition methods of excretion body as claimed in claim 8, it is characterised in that the step 2) pretreatment it is specific
For：
1) supernatant of the MSC nutrient solutions is taken, centrifugal treating is carried out, cell fragment is removed；
2) and then by supernatant membrane filtration handle, remove apoptotic body, obtain pretreated supernatant.
10. the acquisition methods of excretion body as claimed in claim 9, it is characterised in that the step 3) be specially：
1) centrifuge tube of the product of trying to please for 5 parts by volume, adds the excretion body separation agent of 1 parts by volume；
2) the pretreated supernatant of process of 4 parts by volume is added, is fully mixed；
3) after stewing process, then centrifugal treating twice is carried out, obtains the excretion body.
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Cited By (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108823159A (en) * | 2018-07-13 | 2018-11-16 | 李璇 | The influence for the excretion body that PDGF-BB discharges mescenchymal stem cell |
| CN108823159B (en) * | 2018-07-13 | 2022-08-09 | 上海齐昔生物科技集团有限公司 | Effect of PDGF-BB on exosomes released by mesenchymal Stem cells |
| CN109865033A (en) * | 2019-03-21 | 2019-06-11 | 北京隆祺生物科技有限公司 | A kind of external application medical ointment and its application in reparation diabetes wound |
| CN110403898A (en) * | 2019-09-06 | 2019-11-05 | 天津珂芬迪斯生物医药科技有限公司 | A kind of jelly peptide facial mask and preparation method thereof |
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