CN105769911A - Hair regeneration method for mesenchymal stem cell-induced alopecia areata and application - Google Patents
Hair regeneration method for mesenchymal stem cell-induced alopecia areata and application Download PDFInfo
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Abstract
The invention relates to the technical field of stem cells and hair loss treatment, and in particular relates to a hair regeneration method for mesenchymal stem cell-induced alopecia areata and an application. The invention provides a new application of the mesenchymal stem cells, namely the application of the mesenchymal stem cells in preparation of products for hair regeneration of induced alopecia areata, the hair regeneration method for the mesenchymal stem cell-induced alopecia areata, and an animal model for hair regeneration after alopecia areata built by the method. The hair regeneration method can be applied to hair follicle regeneration research and alopecia areata treatment.
Description
Technical field
The present invention relates to stem cell and alopeciaing therapeutic technical field, be specifically related at a kind of mescenchymal stem cell induction alopecia areata
The method of hair regeneration, and the method is applied in hair follicle regeneration research and alopecia areata treatment.
Background technology
Hair has important physiology and a social function, but huge social competition's pressure, spirit constant tension, stay up late for a long time
Etc. life style, all can seriously damage the health of hair follicle and then cause alopecia.Epidemiological study finds: patient's alopecia is younger
Changing, the young and the middle aged becomes " main force " in alopecia crowd.Although at present understanding and the understanding of alopecia being on the increase and is goed deep into,
The remedy measures held or assist also gets along with, and clinic still expects to find more effectively, more fully Therapeutic Method.
Alopecia areata (alopecia areata), is the non-cicatricial alopecia of a kind of autoimmunity, often betides health hairiness
The position sent out, local skin is normal, no conscious sympton.The circular alopecia of another name, circle is bald, alopecia areata.Etiology unknown, hair follicle, ball top
And hair papilla all reduces, position is also upper to be moved, lymphocytic infiltration seen from perifollicolar, and primary disease sometimes merges other and self exempts from
Epidemic disease disease (such as vitiligo, atopic dermatitis), therefore it is now recognized that primary disease there may be autoimmune pathogenesis.
There is no effective Therapeutic Method for alopecia areata at present, common treatment measures have oral glucose corticosteroid hormone, ciclosporin etc..
Currently also have an employing hair grafting treatment alopecia areata, but exist this not enough from chaeta source in survival efficiency problem (see document:
Thakur BK,Verma S.Is hair transplantation always successful in secondary
cicatricial alopecia？Int J Trichology.2015Jan-Mar；7(1):43-4).Therefore, promote that alopecia areata is suffered from
The hair regeneration of person self has become one of tissue repair field difficult problem urgently to be resolved hurrily, and research hair regeneration has important reason
Opinion and practical significance.
In recent years, mescenchymal stem cell (mesenchymal stem cell, MSC) clinical practice is in solving multiple blood
Systemic disease, cardiovascular disease, liver cirrhosis, nervous system disease, meniscus of knee joint Partial Resection injury repairing, autoimmune
The aspects such as property disease achieve important breakthrough and (see document: Forostyak S, Jendelova P, Sykova E.The role
of mesenchymal stromal cells in spinal cord injury,regenerative medicine and
possible clinical applications.Biochimie.2013Dec；95(12):2257-70).
After the previous research of the present inventor: MSC injection nude mouse is subcutaneous, can be shown by detection anti-human mitochondrial antibody
The differentiation of track MSC with go back to the nest, its be divided into more Hair follicle epithelial cells and hair papilla cell (dermal papilla cell,
DPC), position of going back to the nest (sees document: Minjuan Wu, Qing Sun, et with common at raw coal bunker and hair papilla
al.XiaocanGuo,Houqi Liu.hMSCs possess the potential to differentiate into DP
cells in vivo and in vitro.Cell Biology International Reports.2012；19(2):
e00019)。
In the regulation and control of hair follicle growth, some hair growth associated protein play vital effect.There is experiment display,
VEGF (vascular endothelial growth factor, VEGF) is at hair follicle growth phase high expressed.
VEGF can affect cell migration by stimulating actin restructuring, raises Akt/PKB gene to prevent cell death simultaneously.Become
Fibroblast growth factor 7 (fibroblast growth factor, FGF-7) can promote that hair follicle turns to trophophase from catagen
Become, and can promote stem cell activity (see document: Greco, V., Chen, T., Rendl, M., Schober, M.,
Pasolli,H.A.,Stokes,N.,Dela Cruz-Racelis,J.,and Fuchs,E.A two-step mechanism
for stem cell activation during hair regeneration.Cell Stem Cell.2009；4,155–
169.)。
There is no document report at present and promote mice alopecia areata position hair follicle reconstitution and the research of hair regeneration about MSC subcutaneous injection
And application.
Summary of the invention
It is an object of the invention to provide the new application of mescenchymal stem cell, specifically mescenchymal stem cell in preparation induction
The method of hair regeneration, Yi Jiyong at application in the product of hair regeneration at alopecia areata, and mescenchymal stem cell induction alopecia areata
The method builds a kind of animal model.
First MSC supernatant secretions identified by we, when MSC cell amplification is to total amount 107, melt about 70%
The cell closed is replaced by serum-free medium, and 24 culture medium as a child collecting these mescenchymal stem cells carry out protein chip inspection
Survey, it is seen that it secretes substantial amounts of somatomedin, including somatomedin more closely-related with hair growth, such as VEGF, FGF-7
Deng.In view of this, it is contemplated that mescenchymal stem cell may can promote hair follicle growth, hair growth is promoted afterwards.
A first aspect of the present invention, it is provided that mescenchymal stem cell is the product of hair regeneration at preparation induction (promotion) alopecia areata
Application in product.
Present invention provides mescenchymal stem cell application in the medicine of preparation treatment alopecia areata.
Present invention provides the reagent that mescenchymal stem cell promotes VEGF (VEGF) to express in preparation
In application.
Present invention provides mescenchymal stem cell and promote what fibroblast growth factor 7 (FGF-7) was expressed in preparation
Application in reagent.
Mescenchymal stem cell in the present invention, is preferably people's embryo source property mescenchymal stem cell.
Described product, medicine, reagent, can be the culture fluid of mescenchymal stem cell, supernatant, cell suspension etc., excellent
Electing as is mesenchyma stem cell suspension, and concentration is with 5 × 104Every square centimeter is preferred.
A second aspect of the present invention, it is provided that the method for hair regeneration at a kind of mescenchymal stem cell induction alopecia areata.
At described a kind of mescenchymal stem cell induction alopecia areata, the method for hair regeneration, refers to MSC with hypodermic
Mode is transplanted in the skin histology at alopecia areata position.
Preferably, MSC is with 5 × 104The concentration of every square centimeter is injected.
Preferably, the skin histology at alopecia areata position is skin corium.
MSC at mice alopecia areata position subcutaneous injection, can be induced hair regeneration by the present invention.
The invention provides a kind of side using mescenchymal stem cell subcutaneous injection inducing mouse alopecia areata position hair regeneration
Method, comprises the following steps:
The recovery of A.MSC and qualification；
B. step A is identified the MSC obtained, to be transplanted to mice alopecia areata area skin tissue after hypodermic mode.
Above-mentioned injected in mice area skin is drawn materials, is fixed, makes section after paraffin embedding, HE dyeing, observe speckle
The structure change of bald place skin and hair follicle.Use immunohistochemical method, observe MSC differentiation direction and position of going back to the nest, measure hair
Send out the expression of growth associated protein.Found that MSC can induce hair regeneration at alopecia areata, can be at hair papilla and raw coal bunker
Go back to the nest and transdifferentiation is Hair follicle epithelial cells and hair papilla cell.
A third aspect of the present invention, it is provided that the construction method of hair regeneration animal model after a kind of alopecia areata.
Described construction method, comprises the following steps:
A, imiquimod induction alopecia areata animal model, builds the alopecia areata model of C3H/HeJ system mice napex skin；
B, MSC is transplanted in the skin histology at mice alopecia areata position in hypodermic mode.
Preferably, step B particularly as follows:
The recovery of B1, MSC and qualification；
B2, the MSC qualification of step B obtained, with 5 × 104Mice alopecia areata position it is expelled to after the concentration of every square centimeter
Skin corium.
Concrete, construction method comprises the following steps:
A. build the alopecia areata animal model of imiquimod induction, carry out recovery and the qualification of MSC simultaneously
Concrete implementation is, first uses 0.9% sodium chloride solution to moisten the skin after mouse ear, exposes its Rhizoma Imperatae
With skin, imiquimod cream (trade name: Ming Xinlidi, the bright glad Pharmaceutical in Sichuan) is used to smear, every Mus at Rhizoma Imperatae
About 25mg, smears once for every 2 days every time.After smearing for three to 4 weeks, the mice of ear back part skin alopecia areata can be obtained.
From liquid nitrogen, take out the MSC of cryopreservation, be immediately placed in 37 C water bath, be shaken gently for cryovial so that it is
Rapid rewarming.Then seed cells in complete grown cultures liquid and cultivate, change liquid after 24 hours to remove DMSO.Streaming
Cell instrument detection MSC surface marker, result be the positive rate of CD29, CD44, CD106 all more than 90%, CD34, CD45 sun
Property rate is below 1%, it is seen that MSC purity is higher.
B, by recovery the MSC that identifies with 5 × 104Individual/cm2Concentration be seeded to the cutaneous lesion of alopecia areata mice.By mice
It is divided into 2 groups, often organizes mouse subcutaneous injection 5 × 10 respectively4Individual/cm2MSC and physiological salt liquid.Perusal 4 after raising 2 weeks
Group mice alopecia areata portion hair growth situation, locates after death skin and draws materials.
C. draw materials area skin through fixing, wash and being dehydrated, transparent, waxdip, embed, cut into slices with bonding die, dewax, dye,
HE stained preparation made by dehydration, transparent, mounting, observe each group of mice draw materials portion's hair follicle from catagen to the recovery shape of trophophase
Condition, ball top is positioned at skin or hypodermic position and hair condition such as diameter, density etc..In addition, will draw materials position
Carry out SABC process, use antibody labeling VEGF, FGF-7 equimolecular and humanized's mitochondrion, detect the differentiation of MSC and return
Nest.
By experimentation, after 14 days, it is observed visually at MSC transplantation group mouse back alopecia areata skin and has hair to grow,
And mice regenerated hairs and the density of primary hair, hair color closest to, but more glossy.After 14 days, paraffin section H.E contaminates
The visible hair follicle transplanted in MSC group mice skin tissue of color is in trophophase more, and hair follicles locations substantially moves down, with subepidermal skin
Undertissue's layer is main.SABC is identified and is found, in MSC transplantation group mouse skin, the expression of VEGF with FGF-7 molecule is relatively noted
Penetrating normal saline group skin to increase, the position gone back to the nest of humanized's mitochondrial markers spike display MSC is mainly at hair papilla and true
At Intradermal new capillary vessel.
After the present invention builds the alopecia areata obtained, hair regeneration animal model can be used for clinical research hair follicle regeneration, hair again
Raw, treatment alopecia areata etc..
The present invention provides for realizing the formation of skin appendages hair in hair regeneration at alopecia areata and organization engineering skin
New thinking and mode.
Accompanying drawing explanation
Fig. 1 is the alopecia areata model of preparation, and wherein A is substantially animal picture, and wherein A1 is normal mouse, and A2 is alopecia areata model
Mice；B is skin and the change of hair follicle structure at H.E dyeing detection alopecia areata, and wherein B1 is normal mouse, and B2 is that alopecia areata model is little
Mus；Most of hair follicle at visible alopecia areata enters catagen.
Fig. 2 is hair regeneration at mescenchymal stem cell induction alopecia areata, and wherein A is substantially animal picture；B is H.E dyeing inspection
Survey skin and the change of hair follicle structure at hair regeneration；Visible hair follicle is entered trophophase by catagen.
Fig. 3 is Immunohistochemical detection humanized's mitochondrion and the expression of hair follicle growth associated protein, and wherein A is people source
Property mitochondrion；B is VEGF；C is FGF-7；Humanized's mitochondrion, the positive signal of VEGF and FGF-7 it is respectively shown in arrow.
Fig. 4 is MSC supernatant antibody chip result, and lower row's arrow indication is VEGF, and upper row's arrow indication is FGF-7.
Detailed description of the invention
In conjunction with embodiment and accompanying drawing, the invention will be further described, but the enforcement of the present invention is not limited to that.
Agents useful for same of the present invention and raw material the most maybe can be prepared by literature method.Unreceipted tool in the following example
The experimental technique of concrete conditions in the establishment of a specific crime, generally according to normal condition such as Sambrook et al. " molecular cloning: lab guide " (New
York:Cold Spring Harbor Laboratory Press, 1989) condition described in, or according to normal condition, or
According to the condition proposed by manufacturer.Unless otherwise indicated, otherwise percentage ratio and number are calculated by weight.
The qualification of embodiment 1:MSC supernatant secretions
MSC cell amplification is to total amount 107, during fusion rate 70-80%, start to change serum-free medium continuation cultivation 12 into little
Time, collect serum-free medium and carry out protein chip cytokines measurement, use Kang Cheng company RayBio human cell factor antibody
Chip, detects nearly 500 kinds of cytokines altogether, and concrete operation step provides handbook to carry out by Kang Cheng company.
After X-ray film exposure, by the image scanner scanning on film the picture file that is converted to gray scale tiff format
Preserving, as shown in Figure 4, wherein lower row's arrow indication is VEGF, and upper row's arrow indication is FGF-7.
Run ScanAlyze software, the dot matrix of gray scale tiff format picture is converted into digital data, by this original number
According to saving as Microsoft Excel file, and original value is deducted all BLANK averages away from Pos revise, use Pos
Do standardization, formation following table:
The result of MSC supernatant secretions protein chip detection: visible its secretes substantial amounts of somatomedin, including raw with hair
Long more closely-related somatomedin, such as VEGF, FGF-7 etc..
Embodiment 2: prepared by mice alopecia areata model
1. prepared by alopecia areata model
All of zoopery is all in accordance with NIH laboratory animal requirement.Laboratory animal is 6 week old C3H/HeJ mouses, male and female
Do not limit, weigh about 30g, The 2nd Army Medical College Experimental Animal Center provide.
Alopecia areata model is formed first by imiquimod induction C3H/HeJ mouse skin of back.Method is as follows: the most careful
Taking out mice, drink with cleaning medical cotton writing paper and take imiquimod cream body about 0.05 gram, uniform application is to napex skin, and area is about
1.5 × 1.5cm, two days once.The longest continuous use is less than 10 weeks.All clean local with normal saline before being administered every time, with
Remove previous left drug.In operation, action is the softst, smears uniformly, it is to avoid friction causes skin tears or hair repeatedly
Send out mechanicalness to come off.About 4 weeks, medicine-feeding part can be formed > 1 × 1cm size alopecia areata area, prompting modeling success, such as Fig. 1 institute
Show.
Stop being administered, within 5 days after obvious alopecia areata is formed, draw materials, with 75% alcohol disinfecting skin 2 times, cut with sharp keen cutter
Skin holostrome, is put immediately into 4% paraformaldehyde solution and fixes 4-5 hour.
Successful for alopecia areata modeling mice is randomly divided into 2 groups, often group 6, lumbar injection 1.5% Nembutal sodium solution
(0.1ml/20g), use hypodermic mode by skin group at a certain amount of mesenchymal stem cell transplantation to alopecia areata after anesthesia
Knit.Every day observes, and within after transplanting 3,5,7,9,14 days, draws materials.
2. alopecia areata model is identified
Equipment apparatus: microtome, calorstat, wax-dissolving box, glass jar, microscope slide, coverslip, knife blade, tweezers, embedding
Box.
Reagent: 10% formalin, ethanol, dimethylbenzene, hard paraffin, hematoxylin, sour water, ammonia, ethanol eosin stains
Dosage form, neutral gum etc..Next carry out the specimen fixed embedding and routine paraffin wax section and H.E dye.
(1) be dehydrated transparent:
Take out fixing specimen, make dehydrant with by low concentration to alcohol in high concentration, gradually slough the water in piece of tissue
Part.Again piece of tissue is placed in and is both dissolved in ethanol, be dissolved in again in the clarifier dimethylbenzene of paraffin transparent, replace out tissue with dimethylbenzene
Ethanol in block.
(2) waxdip embedding:
The most transparent piece of tissue is placed in the paraffin dissolved, puts into wax-dissolving box insulation.Treat that paraffin is completely immersed in tissue
Embedding after block: in the small paper box folded, pour the paraffin dissolved into, rapid gripping has been impregnated with the piece of tissue of paraffin and has put
Enter wherein, treat liquid paraffin cooled and solidified in bulk.On paraffin mass, carry out labelling with pencil, prepare section.
(3) section and paster:
Embedded wax stone is fixed on microtome, thinly slices, generally 5 microns thickness.The thin slice cut often wrinkles
Folding, plates in the water of heating to be put into, then is attached on microscope slide, puts in 45 DEG C of calorstats and dries.
(4) H.E dyeing
Hematoxylin-eosin staining is frequently referred to H.E dyeing, to strengthen the aberration of intracellular each ingredient in tissue, in order to
In observation.
H.E dyeing course is:
Dyeing several minutes in hematoxylin aqueous solution is put in section after entering distilled water.Color separation 5 seconds in sour water and ammonia
The clock several seconds.Flowing water enters distilled water for a moment after rinsing 1 hour.70% and 90% ethanol is dehydrated each 10 minutes.Enter ethanol Yihong
Dyeing liquor dyes 2-3 minute.It is dehydrated transparent: the section after dyeing is dehydrated through absolute alcohol, then makes section transparent through dimethylbenzene.Neutral
Can preserve for a long time after natural gum mounting.
Skin and the change of hair follicle structure at result: H.E dyeing detection alopecia areata, it is seen that at the alopecia areata of alopecia areata model mice
Major part hair follicle enters catagen.
Embodiment 3: the recovery of human mesenchymal stem cell and the recovery of Transplanted Human mescenchymal stem cell and transplanting
Human mesenchymal stem cell (MSC) can be according to documents below method separating-purifying: list of references: Minjuan Wu,
XiaocanGuo,LingYang,Yue Wang,Ying Tang,Yongji Yang,Houqi Liu.Mesenchymal Stem
Cells with Modification of JAM-AInduce Hair Formation.Stem Cells
Translational Medicine.2014Apr；3(4):481-8).MSC used by the present embodiment separates according to preceding method and carries
After pure frozen by the Histology and Embryology teaching and research room of The 2nd Army Medical College at the present inventor place.
From nitrogen storage tank, take out MSC, be placed in 37 DEG C of water-baths and rock until dissolving.It is centrifuged under the conditions of 1000rpm
5min, abandons supernatant, adds DMEM (L) culture fluid (L is the culture medium of low sugar) the 1ml suspension cell containing 20%FBS, and is transferred to
In T25 culture bottle, add DMEM (L) culture fluid of 20%FBS to 5ml.It is placed in 37 DEG C, 5%CO2Incubator is hatched, 24 hours
After change DMEM (L) culture fluid containing 10%FBS into, within the most every 3 days, change a culture fluid.
When cell grows to 70-80% fusion, peptic cell, collection cell carries out Flow cytometry mesenchyme to be done
The expression of cell specific marker molecules.
Qualification result: the surface marker of flow cytomery MSC, result is that the positive rate of CD29, CD44, CD106 is equal
More than 90%, CD34, CD45 positive rate is below 1%, it is seen that MSC purity is higher.
Mescenchymal stem cell after identifying is with 5 × 104The mode of the concentration subcutaneous injection of every square centimeter is inoculated into
Skin at alopecia areata.
Matched group is the normal saline of same volume.Observe hair regeneration situation after inoculation, inoculate observation inoculation of drawing materials latter 7 days
Place's skin and the microstructural change of hair follicle.Draw materials and the same embodiment of paraffin section step.And the routine carrying out paraffin section exempts from
Epidemic disease histochemical stain observation VEGF and FGF-7 is in experimental group and the expression of matched group, with the mechanism of preliminary analysis hair regeneration.
Experimental result:
After 14 days, it is observed visually at MSC transplantation group mouse back alopecia areata skin and has hair to grow, and mice regenerated hairs
With the density of primary hair, hair color closest to, but more glossy.As shown in Figure 2.
After 14 days, the visible hair follicle transplanted in MSC group mice skin tissue of paraffin section H.E dyeing is in trophophase more,
Hair follicles locations substantially moves down, based on subepidermal hypodermis layer.SABC is identified and is found, MSC transplantation group mouse skin
The expression of middle VEGF with FGF-7 molecule relatively injecting normal saline group skin increases, humanized's mitochondrial markers spike display MSC
The position gone back to the nest is mainly at hair papilla and at intradermal new capillary vessel.As shown in Figure 3.
The ultimate principle of the present invention, principal character and advantages of the present invention have more than been shown and described.The technology of the industry
Personnel, it should be appreciated that the present invention is not restricted to the described embodiments, simply illustrating this described in above-described embodiment and description
The principle of invention, the present invention also has various changes and modifications without departing from the spirit and scope of the present invention, and these become
Change and improvement both falls within scope of the claimed invention.Claimed scope by appending claims and
Equivalent defines.
Claims (10)
1. mescenchymal stem cell application in the product of hair regeneration at preparation induction alopecia areata.
Mescenchymal stem cell the most according to claim 1 is the application in the product of hair regeneration at preparation induction alopecia areata,
It is characterized in that, described product is the medicine for the treatment of alopecia areata.
3. mescenchymal stem cell application in preparation promotes the reagent of vascular endothelial growth factor expression.
4. mescenchymal stem cell application in the reagent that preparation promotes fibroblast growth factor 7 to express.
Mescenchymal stem cell the most according to claim 1 is the application in the product of hair regeneration at preparation induction alopecia areata,
It is characterized in that, described product is the culture fluid of mescenchymal stem cell, supernatant, or cell suspension.
6. a method of hair regeneration at mescenchymal stem cell induction alopecia areata, it is characterised in that described method is to fill
Matter stem cell is transplanted in the skin histology at alopecia areata position in hypodermic mode.
The method of hair regeneration at a kind of mescenchymal stem cell the most according to claim 6 induction alopecia areata, it is characterised in that
The method is used for inducing mouse alopecia areata position hair regeneration, comprises the following steps:
A. the recovery of mescenchymal stem cell and qualification；
B. step A is identified the mescenchymal stem cell obtained, to be transplanted to mice alopecia areata area skin after hypodermic mode
Tissue.
8. the construction method of hair regeneration animal model after an alopecia areata, it is characterised in that described construction method includes following
Step:
A, imiquimod induction alopecia areata animal model, builds the alopecia areata model of C3H/HeJ system mice napex skin；
B, mescenchymal stem cell is transplanted in the skin histology at mice alopecia areata position in hypodermic mode.
The construction method of hair regeneration animal model after a kind of alopecia areata the most according to claim 8, it is characterised in that step
B is:
B1, the recovery of mescenchymal stem cell and qualification；
B2, the mescenchymal stem cell qualification of step B obtained, with 5 × 104Mice alopecia areata it is expelled to after the concentration of every square centimeter
The skin corium at position.
The construction method of hair regeneration animal model after a kind of alopecia areata the most according to claim 8, it is characterised in that institute
The construction method stated comprises the following steps:
A, imiquimod induction alopecia areata animal model, the alopecia areata model of structure C3H/HeJ system mice napex skin:
First use 0.9% sodium chloride solution to moisten the skin after mouse ear, expose its Rhizoma Imperatae and skin, at Rhizoma Imperatae, use miaow
The not special emulsifiable paste of quinoline is smeared, every Mus about 25mg every time, within every 2 days, smears once；After smearing for three to 4 weeks, ear back can be obtained
The mice of portion's skin alopecia areata；
B, mescenchymal stem cell is transplanted in the skin histology at mice alopecia areata position in hypodermic mode:
B1, the recovery of mescenchymal stem cell and qualification；
From liquid nitrogen, take out the mescenchymal stem cell of cryopreservation, be immediately placed in 37 C water bath, be shaken gently for cryovial,
Make its rapid rewarming；Then seed cells in complete grown cultures liquid and cultivate, change liquid after 24 hours to remove DMSO；
Surface marker CD29, CD44, CD106, CD34, CD45 of flow cytomery MSC；
B2, the mescenchymal stem cell qualification of step B obtained, with 5 × 104Mice alopecia areata it is expelled to after the concentration of every square centimeter
The skin corium at position.
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Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106497886A (en) * | 2016-12-09 | 2017-03-15 | 北京恒峰铭成生物科技有限公司 | A kind of hair regeneration induction liquid and its preparation method and application |
| RU2631642C1 (en) * | 2016-11-29 | 2017-09-25 | Общество с ограниченной ответственностью "Научно-производственная компания Стемма" | Method for treating nonscarring alopecia |
| CN108236614A (en) * | 2016-12-23 | 2018-07-03 | 吟斯蔻碧株式会社 | Biological implantation for promoting hair tonic uses graft |
| CN109554454A (en) * | 2017-09-26 | 2019-04-02 | 东莞自然衡健康科技有限公司 | A method of freeze-dried powder hair regrowth is evaluated by measurement cytokine content |
| CN109718251A (en) * | 2019-01-25 | 2019-05-07 | 华子昂 | Hair growth method and application using stem cell reconstructed hair follicles |
| RU2695364C1 (en) * | 2018-11-08 | 2019-07-23 | Общество с ограниченной ответственностью "Научно-производственная компания Стемма" | Method of treating non-scarring alopecia |
| CN110551692A (en) * | 2019-07-09 | 2019-12-10 | 吕贯廷 | hFGF9 gene modified mesenchymal stem cell and preparation method and application thereof |
| CN110878287A (en) * | 2019-12-05 | 2020-03-13 | 伯仕利生物科技发展（盐城）有限公司 | Preparation method and application of stem cell supernatant over-expressing TAT-KGF |
| CN110917334A (en) * | 2019-11-27 | 2020-03-27 | 浙江卫未生物医药科技有限公司 | Hair regenerating agent based on hair follicle cell dryness maintenance |
| CN114748507A (en) * | 2022-02-17 | 2022-07-15 | 浙江卫未生物医药科技有限公司 | Human hair follicle mesenchymal stem cell and CGF composition and preparation method and application thereof |
| CN115918610A (en) * | 2023-02-24 | 2023-04-07 | 中日友好医院(中日友好临床医学研究所) | Preparation method and application of stress liver depression and spleen deficiency type alopecia areata-like mouse model |
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| RU2631642C1 (en) * | 2016-11-29 | 2017-09-25 | Общество с ограниченной ответственностью "Научно-производственная компания Стемма" | Method for treating nonscarring alopecia |
| CN106497886A (en) * | 2016-12-09 | 2017-03-15 | 北京恒峰铭成生物科技有限公司 | A kind of hair regeneration induction liquid and its preparation method and application |
| CN108236614A (en) * | 2016-12-23 | 2018-07-03 | 吟斯蔻碧株式会社 | Biological implantation for promoting hair tonic uses graft |
| CN108236614B (en) * | 2016-12-23 | 2021-07-20 | 吟斯蔻碧株式会社 | Implant for biological transplantation for promoting hair growth |
| CN109554454A (en) * | 2017-09-26 | 2019-04-02 | 东莞自然衡健康科技有限公司 | A method of freeze-dried powder hair regrowth is evaluated by measurement cytokine content |
| RU2695364C1 (en) * | 2018-11-08 | 2019-07-23 | Общество с ограниченной ответственностью "Научно-производственная компания Стемма" | Method of treating non-scarring alopecia |
| CN109718251A (en) * | 2019-01-25 | 2019-05-07 | 华子昂 | Hair growth method and application using stem cell reconstructed hair follicles |
| CN110551692A (en) * | 2019-07-09 | 2019-12-10 | 吕贯廷 | hFGF9 gene modified mesenchymal stem cell and preparation method and application thereof |
| CN110917334A (en) * | 2019-11-27 | 2020-03-27 | 浙江卫未生物医药科技有限公司 | Hair regenerating agent based on hair follicle cell dryness maintenance |
| CN110878287A (en) * | 2019-12-05 | 2020-03-13 | 伯仕利生物科技发展（盐城）有限公司 | Preparation method and application of stem cell supernatant over-expressing TAT-KGF |
| CN114748507A (en) * | 2022-02-17 | 2022-07-15 | 浙江卫未生物医药科技有限公司 | Human hair follicle mesenchymal stem cell and CGF composition and preparation method and application thereof |
| CN115918610A (en) * | 2023-02-24 | 2023-04-07 | 中日友好医院(中日友好临床医学研究所) | Preparation method and application of stress liver depression and spleen deficiency type alopecia areata-like mouse model |
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