CN105326862A - Stem cell preparation for treating burned skin and preparation method thereof - Google Patents
Stem cell preparation for treating burned skin and preparation method thereof Download PDFInfo
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- CN105326862A CN105326862A CN201510779644.XA CN201510779644A CN105326862A CN 105326862 A CN105326862 A CN 105326862A CN 201510779644 A CN201510779644 A CN 201510779644A CN 105326862 A CN105326862 A CN 105326862A
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Abstract
The invention provides a stem cell preparation for treating burned skin and a preparation method thereof. The stem cell preparation contains epidermal stem cell suspension, wherein silk fibroin, chitosan and VC are added to the epidermal stem cell suspension. The stem cell preparation for treating the burned skin is the epidermal stem cell suspension and can promote epidermal cell growth for a long time and shorten the wound healing time. In addition, the high polymer materials including the silk fibroin, the chitosan and the VC are added into the epidermal stem cell suspension so that the cell viability can be improved, and the cell survival time can be prolonged.
Description
Technical field
The present invention relates to stem cell medicine and preparation method thereof, particularly relate to a kind of stem cell medicine being used for the treatment of burned skin and preparation method thereof.
Background technology
Studies have reported that, wound surface covering after severe trauma, large-area burns, anaplasty has become a very important problem, except autologous patient skin-grafting, people adopted allograft of skin, and other associated restoration materials carry out flap coverage, but no matter autologous or heterogenous skin transplants the restriction being all subject to donor source and immunological rejection.Urgently find a kind of desirable natural drug without rejection, the reparation of promotion skin injury, promotion skin regeneration.
Mescenchymal stem cell (mesenchymalstemcells, MSC) is one group of foreign cell group coming from substrate, can obtain from human body great majority tissue.Lot of experiments shows, and mescenchymal stem cell has epidermis cell differentiation potential, the healing of the skin that can promote to be wound.Owing to there is the hidden danger of virus contamination in bone marrow derived mescenchymal stem cell, and along with the growth of donor age, obvious downward trend appears in its cell quantity and amplification, differentiation capability, is not suitable for batch preparation.And epidermal stem cells is present in a kind of skin progenitor cell in the gap between stratum basale and hair follicle root, as the specific stem cells of skin histology, the chief functional cells of skin metabolism can not only be maintained, also be closely related with the reparation of wound surface, keratinocyte is divided in wound surface environment, be transferred to skin wound and repairing skin wound surfaces, therefore, epidermal stem cells is one of desirable seed cell of skin tissue engineering.
Summary of the invention
The technical problem that the present invention need solve is to provide a kind of promotion epidermal growth, shortens the stem cell medicine of the treatment burned skin of wound healing time.
For solving above-mentioned technical problem, the present invention devises a kind of stem cell medicine for the treatment of burned skin, and it comprises epidermal stem cells suspension, is also added with fibroin albumen, chitosan and VC in described epidermal stem cells suspension.
Improve further as the present invention, the concentration of described epidermal stem cells suspension epidermal stem cells is 1 × 10
6~ 1 × 10
8individual/ml.
Present invention also offers a kind of preparation method of stem cell medicine of above-mentioned treatment burned skin, it comprises the following steps:
Get the epidermal tissue of people, adopt trypsin and EDTA digest mixed liquor and digest, serum termination, blow and beat gently to epidermal graft with suction pipe and be translucent, after mistake 200 order screen filtration, more centrifugal, abandon supernatant and obtain Cell sap;
Above-mentioned Cell sap is seeded in the culture bottle of IV collagen type finishing, after sticking 10 ~ 20min fast, collects adherent cell and cultivate, add serum-free DMEM, put 37 DEG C, CO
2concentration is cultivate in the incubator of 5%, within every 3 days, changes liquid 1 time, when cell grows up to 70% ~ 80% warm state, adds fibroin albumen, chitosan and VC, make epidermal stem cells suspension formulation.
Improve further as the present invention, adding fibroin albumen, before chitosan and VC, also comprise at high temperature under high pressure, to fibroin albumen, chitosan and VC carry out sterilizing.
Stem cell medicine of the present invention is epidermal stem cells suspension formulation, can promote epidermal growth for a long time, shortens the wound healing time.And the present invention is by fibroin albumen, and chitosan and VC macromolecular material add in stem cell living cells suspension, can improve Cell viability, extend the cell survival time.
Detailed description of the invention
Technical scheme of the present invention is understood better in order to make relevant technical staff in the field, below in conjunction with embodiment of the present invention, technical scheme in embodiment of the present invention is clearly and completely described, obviously, described embodiment is only the present invention's part embodiment, instead of whole embodiments.
The invention provides a kind of stem cell medicine for the treatment of burned skin, be used for the treatment of the wound of burnt degree skin, promote skin healing.
Stem cell medicine of the present invention is epidermal stem cells suspension formulation, and wherein, the concentration of epidermal stem cells is 1 × 10
6~ 1 × 10
8individual/ml, is also added with fibroin albumen in this epidermal stem cells suspension, chitosan and VC.
The preparation method of stem cell medicine of the present invention comprises:
First, get the epidermal tissue of people, adopt trypsin and EDTA digest mixed liquor and digest, serum termination, blow and beat gently to epidermal graft with suction pipe and be translucent, after mistake 200 order screen filtration, more centrifugal, abandon supernatant and obtain Cell sap;
Again above-mentioned Cell sap is seeded in the culture bottle of IV collagen type finishing, after sticking 10 ~ 20min fast, collects adherent cell and cultivate, add serum-free DMEM, put 37 DEG C, CO
2concentration is cultivate in the incubator of 5%, within every 3 days, changes liquid 1 time, when cell grows up to 70% ~ 80% warm state, then adds sterilized fibroin albumen at high temperature under high pressure, chitosan and VC, makes epidermal stem cells suspension formulation.
Below, the present invention is further described in conjunction with specific embodiments.
Embodiment one
The separation and Culture of epidermal stem cells: through Ethics Committee's approval and the informed consent of the family members that are correlated with, obtain less than 12 years old child's foreskin from minor operation room of Affiliated Hospital of Nantong University the 3rd, child is all without diseases of urinary system.Foreskin after ring-type excision is got under aseptic condition, after 0.5g/L chlorhexidine acetate solution soaking sterilization 10min, normal saline flushing repeatedly, carefully rejects subcutaneous tissue with cutting, and is then cut into the skin bit of 5mm × 5mm size, obtain the epidermal tissue of people, shred epidermis, then add 0.25WT% trypsin and 0.02WT%EDTA (tetraacethyl diaminoethanes) digests mixed liquor, volume ratio is 1: 1,15ml, at 37 DEG C, CO
2digest 5min in the incubator of concentration 5%, serum stops, and blows and beats gently to epidermal graft be translucent with suction pipe, centrifugal with 1200r/min again after crossing 200 order screen filtration, abandons supernatant and obtains Cell sap.Cell sap is seeded in the culture bottle of IV collagen type finishing, after sticking 10 ~ 20min fast, collects adherent cell and cultivate, add serum-free DMEM15ml, put 37 DEG C, CO
2concentration is cultivate in the incubator of 5%, within every 3 days, changes liquid 1 time, and when cell grows up to 70% ~ 80% warm state (be substantially paved with bottle at the bottom of), adjustment cell concentration is 1 × 10
6/ cm
2, obtain epidermal stem cells suspension.
Preparation: take fibroin albumen 50-200mg, 1-2g chitosan, 50-200mgV
cin 121 DEG C of autoclavings after 15 minutes, join above-mentioned epidermal stem cells suspension, make homogeneous epidermis single cell suspension.
For verifying the remarkable result of stem cell medicine provided by the invention further, be specifically described by following experiment and experimental data.
Confirmatory experiment:
Laboratory animal grouping and model construction: take consubstantiality counter point, make two III degree of sheet cutting mechanism at 20 Sprague-Dawley rats (SD rat) back.Left side wound surface adopts stem cell medicine of the present invention, is loaded in syringe by stem cell medicine of the present invention, and be applied in left side, give 200 μ L, concentration is 1 × 10
6individual/ml, right side wound surface adopts the living cells suspension of common stem cell (not adding fibroin albumen, chitosan and VC) as a control group, and rear gauze covers wrapping.Under the observation of continuous many days, the wound area of both sides wound surface, relative to the ratio of the wound area of first day, sees the following form 1:
| Preparation group | Matched group | |
| 1st day | 1 | 1 |
| 2nd day | 0.94±0.01 | 0.99±0.02 |
| 3rd day | 0.82±0.02 | 0.98±0.02 |
| 4th day | 0.75±0.01 | 0.95±0.01 |
| 5th day | 0.61±0.03 | 0.94±0.01 |
| 8th day | 0.35±0.01 | 0.92±0.03 |
| 10th day | 0.17±0.03 | 0.86±0.02 |
The ratio value of table 1 rat both sides wound area
Result shows: along with natural law increases, the wound area of left side wound surface is more and more less, and it is effective gradually react stem cell medicine of the present invention, and speed of wound healing is faster than matched group, and result points out stem cell medicine of the present invention to promote wound healing, the shortening course of disease.
Below only have expressed one embodiment of the present invention, it describes comparatively concrete and detailed, but therefore can not be interpreted as the restriction to the scope of the claims of the present invention.It should be pointed out that for the person of ordinary skill of the art, without departing from the inventive concept of the premise, can also make some distortion and improvement, these all belong to protection scope of the present invention.Therefore, the protection domain of patent of the present invention should be as the criterion with claims.
Claims (4)
1. treat a stem cell medicine for burned skin, it is characterized in that, comprise epidermal stem cells suspension, be also added with fibroin albumen in described epidermal stem cells suspension, chitosan and VC.
2. a kind of stem cell medicine for the treatment of burned skin according to claims 1, is characterized in that, the concentration of described epidermal stem cells suspension epidermal stem cells is 1 × 10
6~ 1 × 10
8individual/ml.
3. treat a preparation method for the stem cell medicine of burned skin, it is characterized in that, comprise the following steps:
Get the epidermal tissue of people, adopt trypsin and EDTA digest mixed liquor and digest, serum termination, blow and beat gently to epidermal graft with suction pipe and be translucent, after mistake 200 order screen filtration, more centrifugal, abandon supernatant and obtain Cell sap;
Above-mentioned Cell sap is seeded in the culture bottle of IV collagen type finishing, after sticking 10 ~ 20min fast, collects adherent cell and cultivate, add serum-free DMEM, put 37 DEG C, CO
2concentration is cultivate in the incubator of 5%, within every 3 days, changes liquid 1 time, when cell grows up to 70% ~ 80% warm state, adds fibroin albumen, chitosan and VC, make epidermal stem cells suspension formulation.
4. a kind of preparation method for the treatment of the stem cell medicine of burned skin according to claim 3, is characterized in that,
Adding fibroin albumen, before chitosan and VC, also comprise at high temperature under high pressure, to fibroin albumen, chitosan and VC carry out sterilizing.
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| CN201510779644.XA CN105326862A (en) | 2015-11-16 | 2015-11-16 | Stem cell preparation for treating burned skin and preparation method thereof |
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| CN201510779644.XA CN105326862A (en) | 2015-11-16 | 2015-11-16 | Stem cell preparation for treating burned skin and preparation method thereof |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108048393A (en) * | 2017-12-12 | 2018-05-18 | 谢举临 | A kind of method of new rapid amplifying skin epidermis substrate stem cell |
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| CN101914495A (en) * | 2010-07-22 | 2010-12-15 | 吉林大学 | Culture method for largely amplifying hair follicle stem cells in vitro |
| CN102552323A (en) * | 2012-01-17 | 2012-07-11 | 浙江大学 | Medicine for accelerating skin repair and regeneration, preparation method thereof and application thereof |
| US20140271584A1 (en) * | 2013-03-15 | 2014-09-18 | The Research Foundation For The State University Of New York | Methods and Compositions for Direct Reprogramming of Somatic Cells to Stem Cells, and Uses of these Cells |
| CN104877954A (en) * | 2015-05-19 | 2015-09-02 | 暨南大学 | Stem cell niche as well as preparation method and application thereof |
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Patent Citations (4)
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| CN101914495A (en) * | 2010-07-22 | 2010-12-15 | 吉林大学 | Culture method for largely amplifying hair follicle stem cells in vitro |
| CN102552323A (en) * | 2012-01-17 | 2012-07-11 | 浙江大学 | Medicine for accelerating skin repair and regeneration, preparation method thereof and application thereof |
| US20140271584A1 (en) * | 2013-03-15 | 2014-09-18 | The Research Foundation For The State University Of New York | Methods and Compositions for Direct Reprogramming of Somatic Cells to Stem Cells, and Uses of these Cells |
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| 吕国忠等: "体外培养表皮干细胞复合高分子支架原位修复深度烧伤创面的研究", 《中华损伤与修复杂志》 * |
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| CN108048393A (en) * | 2017-12-12 | 2018-05-18 | 谢举临 | A kind of method of new rapid amplifying skin epidermis substrate stem cell |
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Address after: 518057, room 2, building 10, Shenzhen biological incubation center, No. 302, Nanshan District hi tech, Shenzhen, Guangdong Applicant after: ISTEM REGENERATIVE MEDICINE SCI-TECH CO., LTD. Address before: 518057, Guangdong Province, Nanshan District high tech Zone, South Ring Road, No. 29, No. 02 building, international students, building No. 15 Applicant before: ISTEM REGENERATIVE MEDICINE SCI-TECH CO., LTD. |
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