CN104288129A - Stem cell micro-needle patch for resisting wrinkles and removing freckles and preparation method thereof - Google Patents
Stem cell micro-needle patch for resisting wrinkles and removing freckles and preparation method thereof Download PDFInfo
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- CN104288129A CN104288129A CN201410519509.7A CN201410519509A CN104288129A CN 104288129 A CN104288129 A CN 104288129A CN 201410519509 A CN201410519509 A CN 201410519509A CN 104288129 A CN104288129 A CN 104288129A
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Abstract
The invention discloses a stem cell micro-needle patch for resisting wrinkles and removing freckles and a preparation method thereof, and particularly relates to a micro-needle transdermal patch which is prepared of a anti-wrinkle and freckle removing essence and is injected into subcutaneous tissues so as to achieve cosmetic results, wherein the anti-wrinkle and freckle removing essence is prepared from stem cells, hyaluronic acid, epidermal growth factors, vitamin E and the like. The micro-needle patch has the advantages that the anti-wrinkle and freckle removing essence is introduced to a corium layer of skin by a microtubule, active ingredients permeate in the skin effectively and efficiently so that skin cells directly absorb all of the active ingredients in the anti-wrinkle and freckle removing essence, good effects are achieved, and finally the skin is vitalized from inside to outside so as to thoroughly relieve wrinkles and color spots from the skin. The stem cell micro-needle patch is used for resisting wrinkles and removing freckles and is safe, effective and fundamentally free of wounds.
Description
Technical field
The present invention relates to a kind of stem cell microneedle patch for dewrinkling spot-eliminating and preparation method thereof, particularly, relate to and a kind ofly utilize the dewrinkling spot-eliminating essence of the preparation such as stem cell, hyaluronic acid, epidermal growth factor and vitamin E to be injected in subcutaneous tissue by micropin percutaneous plaster to reach cosmetic result, belong to regeneration medicine technology and anti-aging beauty-care service field.
Background technology
The body cell moment is in regenerative cell and substitutes in the process of old and feeble degenerated cell, when cell turnover speed is less than cell ageing speed, produce and upgrade deficiency, just start to occur old and feeble sign, the wrinkle of skin of face and the formation of mottle are a kind of key character and index.The cell that can play regeneration function in body is exactly stem cell, and that is the source of skin aging is exactly the deficiency of stem cell updating ability.Clinical research in recent years shows, stem cell is widely used in facial plasty, and as embryonic stem cell and fat-derived stem cells etc., stem cell can play reduce wrinkle, speckle dispelling to face and cure the effect of wound.By carrying out stem-cell therapy to ageing skin, skin thickness the is had increase of significance, the collagen content in corium also presents significance to be increased, thus the skin reached human body is old and feeble produces good reparation and beautification function.
Stem cell is the pluripotent cell that a class has self renewal, infinite copy and Multidirectional Differentiation ability, and medical circle is called " general-purpose cell ".Fat stem cell has extremely strong in-vitro multiplication ability, can to cell direction differentiation such as fat, bone, muscle, endothelium, islets of langerhans, nerves.Compared with other stem cell, the advantages such as fat stem cell has that biological characteristics is stable, source is sufficient, easily obtain and condition of in vitro culture is simple are the study hotspots in organizational project and regenerative medicine field.The mechanism of, speckle dispelling crease-resistant about fat stem cell, mainly contain following some: 1, fat stem cell can secrete cytokines and somatomedin, plays the function of senile cell of nutrition, reparation and replacement body injury; 2, fat stem cell can provide antioxidant, scavenging activated oxygen, and heat shock protein, to local, makes local toxicity material be eliminated, and promotes biological cells and tissues reparation; 3, fat stem cell can increase collagen secretion, engulfs pigment removal mottle, improves skin elasticity.
At present, drug cosmetics and surgery beauty treatment are all a kind of interim measures of curing the symptoms, not the disease, and there is excessive risk, complication, side effect with pain with become silted up blue or green.And Wicresoft's beauty treatment relies on efficiently, safely, be easy to operation and recover the advantage ever more popular such as fast.Wherein micropin beauty treatment is asked beautiful person to favor by great majority.Skin particularly horny layer has barrier action to Drug Percutaneous Absorption, hinder the absorption of medicine, micropin can pierce through the stratum corneum barrier of skin, produce the infiltration that microchannel increases medicine, medicine infiltrates the capillary network of dermal layer of the skin by micropin hole, with blood circulation through treatment target position, enough, administration object fast can be reached, greatly reduce the dosage of medicine and improve the curative effect of medicine.Sharp because micropin is thin, general paracentesis depth, only at horny layer and dermis, does not touch teleneuron, decreases the degree of injury to body tissue, stimulates skin corium simultaneously, percutaneous self-healing ability, promotes ossein hypertrophy.Existing research has confirmed safety and the nonirritant of micropin.Therefore, micropin transdermal imports the best mode that medicine mode is safe, efficient and painless administration.
The present invention fully utilizes stem cell transplantation technology and micropin percutaneous plaster medicine-feeding technology, provides a kind of stem cell transplantation dewrinkling spot-eliminating beauty treatment new method.Utilize micropin percutaneous plaster technology can increase stem cell transfer efficiency, direct arrival skin needs the position of beauty treatment, can fix a point, given layer, quantitative effective ingredient is delivered directly to required skin histology, containing fat stem cell, hyaluronic acid, epidermal growth factor, vitamin E in dewrinkling spot-eliminating essence prepared by the present invention, source pure natural.
The hyaluronic acid that dewrinkling spot-eliminating essence in the present invention contains has moisturizing, maintenance and Nutrition to skin, it can pin large quantity of moisture, skin is made to keep full high resilience, reduce wrinkle, also can mild dilation blood capillary, increase blood circulation, improve intermediate supersession, promote the absorption of skin-nourishing and the discharge of refuse, hyaluronic acid is again good Percutaneous absorption enhancer while moisturizing.
The epidermal growth factor that dewrinkling spot-eliminating essence in the present invention contains has and smoothes away wrinkles and prevent the effect of mottle: 1, epidermal growth factor can promote that cytotrophy material is initiatively transported in cell from extracellular, increases intracellular nutrition.Promote skin corium emiocytosis rubber polymer fibril, make dermal tissue full, thus reduce and smooth away wrinkles.2, the propagation of epidermal growth factor to epidermis cell has stronger facilitation, and the accumulation such as residual of less pigment and dead cell, eliminates mottle and pigmentation, cutaneous manifestations is gone out and tenderly to have no time in vain.
The vitamin E contained in dewrinkling spot-eliminating essence in the present invention is topmost antioxidant, can delay skin aging, and its main mechanism is: 1, can directly remove peroxide and free radical, play antioxidation.2, LPO inhibitor, eliminating chloasma, the activity of restraint of tyrosinase, thus reduce melanin.3, protect skin from the injury of ultraviolet and pollution, reduce the deposition of cicatrix and pigment.
Summary of the invention
The object of the present invention is to provide a kind of stem cell microneedle patch for dewrinkling spot-eliminating and preparation method thereof, stem cell, hyaluronic acid, epidermal growth factor and vitamin E etc. can be injected in subcutaneous tissue the cosmetic result reaching dewrinkling spot-eliminating by microneedle patch of the present invention.
For realizing above-mentioned technical purpose, reach above-mentioned technique effect, the present invention is achieved through the following technical solutions:
A kind of stem cell microneedle patch for dewrinkling spot-eliminating, the microneedle array be made up of silicon or metal material and forming containing the liquid storage cylinder of stem cell essence and adhesive-layer with dewrinkling spot-eliminating function, described reserve liquid layer contains liquid injection port, and the main component of described stem cell essence and concentration of volume percent are stem cell suspension 80%, hyaluronic acid solution 5%, epidermal growth factor solution 5%, vitamin E solution 5% and normal saline 5% respectively.
Further, described microneedle array is a kind of empty micropin array, and described hollow array micropin diameter is 40 ~ 80 μm.
Further, described liquid storage cylinder and microneedle array are connected as a single entity.
Further, described adhesive-layer is medical grade, is made up of adherent layer and adhered layer.
Further, described stem cell suspension is made up of fat stem cell and normal saline, and described hyaluronic acid solution concentration is 0.5%, and described epidermal growth factor solution concentration is 50-100 μ g/ml, and described vitamin E solution concentration is 10mg/ml.
Further, described fat stem cell concentration is 1 × 10
6~ 1 × 10
7individual/mL, described fat stem cell is prepared from by separation and Culture in human fatty tissue.
For a preparation method for the stem cell microneedle patch of dewrinkling spot-eliminating, comprise the following steps:
1, the microneedle array that silicon or metal material are made is made by microelectronic manufacturing technology or micro-molded technology, and sterilizing is for subsequent use;
2, preparing respectively and form stem cell suspension by fat stem cell and normal saline, is wherein 1 × 10 containing fat stem cell concentration
6~ 1 × 10
7individual/mL, concentration is 0.5% hyaluronic acid solution, and concentration is 50-100 μ g/ml epidermal growth factor solution, and concentration is 10mg/ml vitamin E solution and normal saline;
3, stem cell essence is prepared: solution above-mentioned steps prepared is prepared into stem cell essence according to formula mixing, concentration of volume percent is stem cell suspension 80%, hyaluronic acid solution 5%, epidermal growth factor solution 5%, vitamin E solution 5% and normal saline 5% respectively, then uses 500 object cell screen filtrations;
4, the stem cell essence after filtration is injected the liquid storage cylinder of microneedle array one with syringe; Then enclose medical grade adhesive-layer and be namely prepared into stem cell microneedle patch.
Advantageous Effects of the present invention is:
1, the invention discloses a kind of stem cell microneedle patch for dewrinkling spot-eliminating, by the mode of microneedle array intramuscular injection, stem cell essence can be made to infiltrate human body skin smoothly and be absorbed by vasoganglion, and required active component is delivered directly to optimal absorption position and carries out dewrinkling spot-eliminating; The small wound that micropin causes can't cause cicatrix, on the contrary can the regeneration of chafe, activation human epidermal cell, thus repairs the wrinkle of human body skin; The pain that microneedle patch effectively can be avoided beauty treatment local injection and cause and infection, safe and reliable.
2, microneedle patch of the present invention containing stem cell essence, the active component contained source pure natural also can reach the effect of dewrinkling spot-eliminating, is in particular in:
A, stem cell used are fat stem cell, fat stem cell has biological characteristics and stablizes, source is sufficient, the advantage such as to draw materials convenient, fat stem cell injects skin through micropin, can rejuvenation skin, improve the circulation of skin emblem and lymph circulation, increase skin elasticity can be reached, improve wrinkle of skin, the effect of dispelling speckles and whitening face skin, the dewrinkling spot-eliminating essence utilizing fat stem cell to prepare really accomplishes the biological beauty realizing skin from cellular level, safer in dewrinkling spot-eliminating application, more effectively, stability is stronger.
B, hyaluronic acid have moisturizing, maintenance and Nutrition to skin, it can pin large quantity of moisture, skin is made to keep full high resilience, reduce wrinkle, also can mild dilation blood capillary, increase blood circulation, improve intermediate supersession, promote the absorption of skin-nourishing and the discharge of refuse, hyaluronic acid is again good Percutaneous absorption enhancer while moisturizing.
C, epidermal growth factor have and smooth away wrinkles and prevent the effect of mottle: epidermal growth factor can promote that cytotrophy material is initiatively transported in cell from extracellular, increase intracellular nutrition, promote skin corium emiocytosis rubber polymer fibril, make dermal tissue full, thus reduce and smooth away wrinkles; The propagation of epidermal growth factor to epidermis cell has stronger facilitation, and the accumulation such as residual of less pigment and dead cell, eliminates mottle and pigmentation, cutaneous manifestations is gone out and tenderly to have no time in vain.
D, vitamin E are topmost antioxidants, can delay skin aging, and its main mechanism is: can directly remove peroxide and free radical, play antioxidation; LPO inhibitor, eliminating chloasma, the activity of restraint of tyrosinase, thus reduce melanin; Protection skin, from the injury of ultraviolet and pollution, reduces the deposition of cicatrix and pigment.
Above-mentioned explanation is only the general introduction of technical solution of the present invention, in order to better understand technological means of the present invention, and can be implemented according to the content of description, coordinates accompanying drawing to be described in detail as follows below with preferred embodiment of the present invention.The specific embodiment of the present invention is provided in detail by following examples and accompanying drawing thereof.
accompanying drawing explanation
Accompanying drawing described herein is used to provide a further understanding of the present invention, and form a application's part, schematic description and description of the present invention, for explaining the present invention, does not form inappropriate limitation of the present invention.In the accompanying drawings:
Fig. 1 is dewrinkling spot-eliminating stem cell microneedle patch administration schematic diagram, empty micropin skin puncture releasing liquid and and diffusion;
Fig. 2 is fat stem cell growth curve;
Fig. 3 A is fat stem cell adipogenic induction cultivation figure, inverted microscope 40 × take pictures; Fig. 3 B is oil red O stain figure after adipogenic induction, inverted microscope 100 × take pictures.
Detailed description of the invention
Below with reference to the accompanying drawings and in conjunction with the embodiments, describe the present invention in detail.
The preparation of embodiment one stem cell microneedle patch
1, the microneedle array that silicon or metal material are made is made by microelectronic manufacturing technology or micro-molded technology, and sterilizing is for subsequent use.
2, prepare stem cell suspension, the stem cell that the present invention contains is fat stem cell, fat stem cell to prepare extraction step as follows:
The first step, obtains human fatty tissue under aseptic condition, shreds the cleaning of rear PBS solution and removes hemocyte;
Second step, adopts Digestive system (containing 0.1%
collagenase Type and 2.5% pancreatin, form with 1:1 ratio mixed preparing) digestion fatty tissue, repeatedly shred 170r/min to pasty state and shake digestion 30 minutes;
3rd step, draws the liquid of lower floor's individual cells, and move in complete medium (DMEM in high glucose containing 10% hyclone) and stop digestion, then centrifugal 10 minutes of 1500r/min, is resuspended in after removing supernatant in complete medium, puts into 37 DEG C, 5% CO
2cultivate in incubator;
4th step, first time changed liquid in cultivation after 12 ~ 24 hours, first time can be carried out after observation of cell is adherent and change liquid, after this liquid was changed every 2 ~ 3 days, go down to posterity after merging until Growth of Cells, after 0.25% pancreatin (containing 0.2%EDTA) conventional digestion, go down to posterity at 1: 3 is inoculated into lasting amplification in vitro cultivation in new culture dish;
5th step, get the fat stem cell in growth conditions good 3rd generation to the 9th generation respectively, form under sight glass, when cell fusion reaches 80% ~ 90%, collect the stem cell suspension be prepared into containing fat stem cell and normal saline, and make the concentration of fat stem cell be 1 × 10
6~ 1 × 10
7individual/mL.
3, preparing concentration is respectively 0.5% hyaluronic acid solution, and concentration is 50 ~ 100 μ g/ml epidermal growth factor solution, and concentration is 10mg/ml vitamin E solution and normal saline.
4, stem cell essence is prepared: solution above-mentioned steps prepared is prepared into stem cell essence according to formula mixing, concentration of volume percent is stem cell suspension 80%, hyaluronic acid solution 5%, epidermal growth factor solution 5%, vitamin E solution 5% and normal saline 5% respectively, obtains the stem cell essence of dewrinkling spot-eliminating after then using 500 object cell screen filtrations.
5, the stem cell essence syringe after above-mentioned filtration is injected the liquid storage cylinder that microneedle array is connected as one; Then enclose medical grade adhesive-layer and be namely prepared into stem cell microneedle patch.
The essence of dewrinkling spot-eliminating imports in the skin corium of skin by microtubule by microneedle patch, as shown in Figure 1, active component infiltrates in skin effectively efficiently, Skin Cell is allowed directly to absorb whole active component in dewrinkling spot-eliminating essence, assimilation effect is close to a hundred per cent, finally reach and make skin live again from inside to outside, thoroughly improve wrinkle and the mottle of skin quality.
The qualification of embodiment two fat stem cell
1, the drafting of growth curve
Get well-grown P3 for cell, after 0.25% pancreatin (containing 0.2%EDTA) digestion, with 2 × 10
4the concentration of/ml is inoculated in 24 orifice plates, every hole 1 mL.Plantation 0,24,48,72,96,120,144,168h detects cell OD value.Method: every hole adds MTT 20 μ L, be placed in 37 DEG C, 5% CO2 incubator continues cultivation 4 hours, collected by the cell centrifugation in hole, every hole adds 150 μ L DMSO, shake after 10 minutes, detection OD value automatically under microplate reader 490 nm wavelength.Experiment in triplicate.Be transverse axis with incubation time after statistical analysis, OD value is longitudinal axis drafting growth curve, as shown in Fig. 2 fat stem cell growth curve: cell proliferation is in good condition, without exception.
2, to differentiation and the qualification of adipose cell
Get well-grown P4 fat subsitutes stem cell, after 0.25% pancreatin (containing 0.2%EDTA) digestion, with 2 × 10
4the concentration of individual/ml is inoculated in 6 orifice plates of preset poly-D-lysine pretreatment coverslip.Cell reaches more than 80% when merging, add adipogenic induction liquid (containing 1 μm of ol/L dexamethasone, 0.5m mol/L IBMX, 10mg/L insulin, 100mg/L indomethacin and containing volume fraction 10% hyclone), fat maintenance medium (complete medium containing 10mg/L insulin) is changed into after 3d, 24h changes fat induced liquid again into and again induces about 18d altogether, carries out oil red O stain qualification.After adipogenic induction, a large amount of cell of Microscopic observation becomes similar round and polygon by spindle shape, and have bright, that refractivity is good fat in a large number to drip in cytoplasm, oil red O stain becomes strong positive reaction, presents redness, proves that it becomes fat ability, as shown in Figure 3.
Embodiment three facial VISIA skin test effect, take following scheme:
Experimental subject: volunteer 20 is from Wuxi in experiment, and skin of face produces wrinkle and mottle due to scytitis, aging or cicatrix.Volunteer Clinical observation forbid edible pungent, stimulate food, smoking cessation and alleviating alcohol addiction.Volunteer, from clinical first 2 weeks, does not use hormone medicine and Claritin, does not use adjunct cosmetic.Volunteer's aspiration is participated in the experiment, and signs Informed Consent Form.
Test method: VISIA skinanalysis apparatus is made up of natural light, polarized light and ultraviolet light 3 kinds of light sources, use Mirror medical image system, the facial camera work that circulates and loupe instrument, the skin aging indexs such as quantitative analysis texture, pore, speckle and wrinkle, accurately by the progress that numerical value display improves.Before treatment, after treatment, patient's face carries out VISIA skin test, carries out percentile scoring to the texture of patient skin, pore, speckle and wrinkle, record data.Scoring × 100% before treated effect=(before treatment scoring after scoring-treatment)/patient treatment.
The use of stem cell microneedle patch: the stem cell microneedle patch prepared is pasted and is injected in nasolabial fold, the position that there is pitting wrinkle at wrinkles on one's forehead, glabella stricture of vagina, crows-feet place, and (or) there is the position of speckle in buccal, temples.Total cellular score>=1 × 10 that every patient obtains
7, once, continuous be for 7 times a course for the treatment of to every two days pasters.
Experimental result: treat the forward and backward VISIA professional skin image analysis system treatment front and back appraisal result that compares and show, treat forward and backward measured value and improve in wrinkle and mottle obviously.Mottle improvement group amplitude is 19.30 ± 5.58%; It is 18.90 ± 7.26% that wrinkle improves amplitude,
p< 0.05.All there were significant differences for the improvement amplitude illustrating in wrinkle and mottle texture and wrinkle.
The foregoing is only the preferred embodiments of the present invention, be not limited to the present invention, for a person skilled in the art, the present invention can have various modifications and variations.Within the spirit and principles in the present invention all, any amendment done, equivalent replacement, improvement etc., all should be included within protection scope of the present invention.
Claims (7)
1. the stem cell microneedle patch for dewrinkling spot-eliminating, it is characterized in that, the microneedle array be made up of silicon or metal material and forming containing the liquid storage cylinder of stem cell essence and adhesive-layer with dewrinkling spot-eliminating function, described reserve liquid layer contains liquid injection port, and the main component of described stem cell essence and concentration of volume percent are stem cell suspension 80%, hyaluronic acid solution 5%, epidermal growth factor solution 5%, vitamin E solution 5% and normal saline 5% respectively.
2. a kind of stem cell microneedle patch for dewrinkling spot-eliminating according to claim 1, is characterized in that: described microneedle array is a kind of empty micropin array, and described hollow array micropin diameter is 40 ~ 80 μm.
3. a kind of stem cell microneedle patch for dewrinkling spot-eliminating according to claim 1, is characterized in that: described liquid storage cylinder and microneedle array are connected as a single entity.
4. a kind of stem cell microneedle patch for dewrinkling spot-eliminating according to claim 1, is characterized in that: described adhesive-layer is medical grade, is made up of adherent layer and adhered layer.
5. a kind of stem cell microneedle patch for dewrinkling spot-eliminating according to claim 1, it is characterized in that: described stem cell suspension is made up of fat stem cell and normal saline, described hyaluronic acid solution concentration is 0.5%, described epidermal growth factor solution concentration is 50-100 μ g/ml, and described vitamin E solution concentration is 10mg/ml.
6. a kind of stem cell microneedle patch for dewrinkling spot-eliminating according to claim 1, is characterized in that: described fat stem cell concentration is 1 × 10
6~ 1 × 10
7individual/mL, described fat stem cell is prepared from by separation and Culture in human fatty tissue.
7., for a preparation method for the stem cell microneedle patch of dewrinkling spot-eliminating, it is characterized in that: comprise the following steps:
Step 1) makes by microelectronic manufacturing technology or micro-molded technology the microneedle array that silicon or metal material make, and sterilizing is for subsequent use;
Step 2) prepare respectively and form stem cell suspension by fat stem cell and normal saline, be wherein 1 × 10 containing fat stem cell concentration
6~ 1 × 10
7individual/mL, concentration is 0.5% hyaluronic acid solution, and concentration is 50-100 μ g/ml epidermal growth factor solution, and concentration is 10mg/ml vitamin E solution and normal saline;
Step 3) prepares stem cell essence: solution above-mentioned steps prepared is prepared into stem cell essence according to formula mixing, concentration of volume percent is stem cell suspension 80%, hyaluronic acid solution 5%, epidermal growth factor solution 5%, vitamin E solution 5% and normal saline 5% respectively, then uses 500 object cell screen filtrations;
Stem cell essence after filtration is injected the liquid storage cylinder of microneedle array one by step 4) with syringe; Then enclose medical grade adhesive-layer and be namely prepared into stem cell microneedle patch.
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