CN1387923A - Heterogeneous cell-free hypodermal framework and its prepn - Google Patents
Heterogeneous cell-free hypodermal framework and its prepn Download PDFInfo
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- CN1387923A CN1387923A CN 02112478 CN02112478A CN1387923A CN 1387923 A CN1387923 A CN 1387923A CN 02112478 CN02112478 CN 02112478 CN 02112478 A CN02112478 A CN 02112478A CN 1387923 A CN1387923 A CN 1387923A
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Abstract
The present invention relates to the skin tissue engineering technology in medicine and is heterogeneous cell-free hypodermal cradle and its preparation. It is essential to close wound fast and effectively in treating serious burns, trauma and ulcer. In the present invention, sucking pig skin is used in making hypodermal cradle, and cell-free hypodermal cradle is modified in spatial structure and surface physical and chemical characteristics. That is, the cell-free hypodermal cradle is treated with trypsin to increase collagenous fibre interval to favor cell amplification in wound and speed vascularization and is soaked in glutaric dialdehyde solution to alter the surface physical and chemical characteristics of hypodermal cradle and to reduce immunologic rejection and occupancy.
Description
Technical field: the present invention relates to medical science skin tissue engineering technical field, is a kind of novel heterogeneous cell-free hypodermal framework and preparation method thereof.
Background technology: serious degree of depth burning, wound and difficult more ulcer etc. all need to solve fast the effectively problem of wound closure.Mainly utilize clinically at present, not only can increase new wound surface, and usually exist from the insufficient problem in body skin source for the large skin defect patient from body skin wound closure.For this reason, at the external structure Graftskin, this artificial skin is made up of dermis scaffold and seed cell, but does not still have ideal dermis scaffold so far by tissue engineering technique in hope.The dermis scaffold product A lloderm of at present external clinical practice
Be that allograft skin is obtained by the cell component of removing wherein, though have clinical effectiveness preferably, exist the source limited, cost an arm and a leg, problem such as viral infection.Though xenogenesis skin source is abundant, especially Corii Sus domestica, its dermis and people are comparatively approaching, but at present domesticly also do not have a matured product, zoopery and clinical test results show, its exist vascularization speed slow, have immunogenicity and occupancy problems such as (can not in time be replaced) by cambium.
Summary of the invention: it is the skin source that the present invention adopts the piglets skin with abundant source, by its space structure of appropriate change and Surface Physical Chemistry character, accelerates vascularization speed, reduction immunological rejection and space-occupying purpose thereby reach.
Preparation method is as follows:
Reagent and material: Dispase II enzyme Sigma company, trypsin Difco import packing, NaCl builds in Shanghai prosperous chemical reagent factory, SDS Shanghai China Shun biological engineering company limited, glutaraldehyde China Medicine (Group) Shanghai Chemical Reagent Co.,
Remove epidermis liquid: the aseptic aqueous solution of 0.025%Dispase II, 1mol/L NaCl
1. bark fetching: consider the color and luster and the collagen composition of skin, selecting white piglets skin for use is the skin source, and body weight is no more than 40kg, gets its veutro, back skin, prunes away except that subcutaneous tissue.
The preparation no basement membrane dermis scaffold so that with combine wound closure from body sword pachydermia:
(1) separate skin corium: utilization is split the skin machine and is further removed subcutaneous tissue and part skin corium, and reuse is split the skin machine and separated skin corium, and thickness is no more than 0.3mm, with 0.05% liquor hibitane soaking disinfection, and the abundant rinsing of physiological saline solution;
(2) remove cell component: the skin corium that is separated to was at room temperature used 0.5% SDS solution or 0.5%Triton X-100 solution effects 24～48 hours, vibration or stir, acellular dermis;
(3) modify space structure: with acellular dermis with the abundant rinsing of physiological saline solution, then prepare with 0.25% trypsin PBS solution, pH 7.0～7.4) soak, 4 ℃, act on 2～4 hours, appropriateness vibration or stirring, thereby the dermal collagen fibre gap increases, so that for wound surface cell proliferation and new collagenogenic secretion provide suitable space, accelerate vascularization speed, reduce space-occupying generation;
(4) modification of surfaces physical and chemical character: with above-mentioned acellular dermis with the abundant rinsing of physiological saline solution, then soak with 0.02% freshly prepared glutaraldehyde solution, act on 10～30min under the room temperature, so that, reduce immunogenicity with the amino acid residue pendant hydroxyl group sealing that exposes;
(5) sterilization: after the abundant rinsing of physiological saline solution, encapsulation,
60Co 100 Gy sterilization, no basement membrane dermis scaffold product.
3. preparation has the dermis scaffold of basement membrane:
(1) preparation cortex: utilize and to split subcutaneous tissue and the part skin corium that the skin machine is removed the piglets skin earlier, contain skin corium and epidermal area in the remaining cortex, thickness is no more than 0.3mm, with it with 0.05% liquor hibitane soaking disinfection, the abundant rinsing of physiological saline solution;
(2) remove epidermal area: the gained cortex is spent epidermis liquid soaked 24～48 hours for 4 ℃, throw off epidermal area, get skin corium.It is identical that all the other each steps and above-mentioned preparation do not have (2) of basement membrane dermis scaffold～(5), makes the dermis scaffold with basement membrane.
The specific embodiment:
Embodiment 1: the dermis scaffold of the no basement membrane of preparation
1. bark fetching: consider the color and luster and the collagen composition of skin, selecting white piglets skin for use is the skin source, and body weight is no more than 40kg, gets its veutro, back skin, prunes away except that subcutaneous tissue.
2. separation skin corium: utilize and split the skin machine and remove subcutaneous tissue and part skin corium earlier, reuse is split the skin machine and is separated required skin corium, thickness 0.3mm; With 0.05% liquor hibitane soaking disinfection, physiological saline solution rinsing 3 times, each 10 minutes;
3. remove cell component: the skin corium that is separated to 0.5% SDS solution, is at room temperature acted on 36 hours, vibration, acellular dermis.
4. modification space structure: with the abundant rinsing of physiological saline solution (3 times, each 10 minutes) acellular dermis, then with the preparation of 0.25% trypsin PBS solution, pH 7.0) soak, 4 ℃, act on 2 hours, the appropriateness vibration makes the increase of dermal collagen fibre gap;
5. modification of surfaces physical and chemical character: with the above-mentioned acellular dermis of the abundant rinsing of physiological saline solution (3 times, each 10 minutes), then soak with 0.02% freshly prepared glutaraldehyde solution, effect is 20 minutes under the room temperature;
6. sterilization: with the abundant rinsing of physiological saline solution (3 times, each 10 minutes), encapsulation,
60Co 100 Gy sterilization must not have the basement membrane dermis scaffold.
During use, dermis scaffold is placed on the degree of depth skin injury wound surface, covers from body sword pachydermia (0.3mm) above.Advantage: get the sword pachydermia can reach in conjunction with the dermis scaffold wound closure in the effect of pachydermia or full pachydermia, reduce the probability of skin donor site scar hyperplasia simultaneously; But the skin donor site quickly-healing can repeatedly supply skin in case of necessity.
Embodiment 2: preparation has the dermis scaffold of basement membrane
1. preparation cortex: utilize behind the bark fetching and split subcutaneous tissue and the part skin corium that the skin machine is removed the piglets skin earlier, contain skin corium and epidermal area in the remaining cortex, thickness is no more than 0.3mm, with it with 0.05% liquor hibitane soaking disinfection, physiological saline solution rinsing 3 times, each 10 minutes;
2. remove epidermal area: above-mentioned gained cortex spends epidermis liquid (aseptic aqueous solution that contains 0.025%Dispase II, 1mol/L NaCl) and soaked 36 hours for 4 ℃, throws off epidermal area, gets skin corium.All the other each steps must have the dermis scaffold of basement membrane with the 3-6 of embodiment 1.
During use, cultivate altogether with human fibroblasts, epidermis cell earlier, corium face plantation fibroblast, the substrate face is cultivated epidermis cell, obtains engineered Graftskin.Advantage: with the xenogenesis acellular dermis be support in the external structure Composite Skin, be used to seal degree of depth skin injury, its character and function approach from the body skin.Dependency to skin donor site is little.
Claims (7)
1. the preparation method of a heterogeneous cell-free hypodermal framework, comprise bark fetching, separate skin corium, remove cell component, sterilization, encapsulation, it is characterized in that also comprising to the space structure of acellular dermis layer and the modification of Surface Physical Chemistry character, also be about to acellular dermis layer trypsin immersion treatment, the dermal collagen fibre gap is increased; The reuse glutaraldehyde solution soaks, to change the materialization property on dermis scaffold surface.
2. by the preparation method of the described heterogeneous cell-free hypodermal framework of claim 1, it is characterized in that the xenogenesis skin is the piglets skin, isolating skin corium does not have basement membrane.
3. by the preparation method of the described heterogeneous cell-free hypodermal framework of claim 1, it is characterized in that the xenogenesis skin is the piglets skin, isolating skin corium has basement membrane.
4. the prepared dermis scaffold of the described heterogeneous cell-free hypodermal framework preparation method of claim 1.
5. the dermis scaffold of the prepared no basement membrane of the described heterogeneous cell-free hypodermal framework preparation method of claim 2.
6. the prepared dermis scaffold of the described heterogeneous cell-free hypodermal framework preparation method of claim 3 with basement membrane.
7. claim 4,5, the 6 described dermis scaffolds purposes that is used to seal burning, wound and wound surface.
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| Application Number | Priority Date | Filing Date | Title |
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| CN 02112478 CN1387923A (en) | 2002-07-11 | 2002-07-11 | Heterogeneous cell-free hypodermal framework and its prepn |
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| CN 02112478 CN1387923A (en) | 2002-07-11 | 2002-07-11 | Heterogeneous cell-free hypodermal framework and its prepn |
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100372511C (en) * | 2005-11-30 | 2008-03-05 | 烟台正海生物技术有限公司 | Acellular dermal matrix |
| CN100408107C (en) * | 2003-10-17 | 2008-08-06 | 胡杰 | Biological wound dressing |
| CN100415309C (en) * | 2003-07-03 | 2008-09-03 | 中国人民解放军第三○四医院 | Application of selective cell-removed pork skin as skin substitute of human body and its preparation method |
| CN1737129B (en) * | 2004-08-19 | 2010-10-06 | 丘纪屏 | Artificial skin transplant and its preparation method |
| CN101954116A (en) * | 2010-08-31 | 2011-01-26 | 长沙达瑞奇实业有限公司 | Manufacturing method of xenoskin used for burns and scalds |
| CN109529123A (en) * | 2018-11-08 | 2019-03-29 | 中国人民解放军第四军医大学 | The vascularization holostrome organization engineering skin and preparation method thereof that hydrogel, nano fiber scaffold and Skin Cell assemble layer by layer |
| CN110732042A (en) * | 2019-11-18 | 2020-01-31 | 上海白衣缘生物工程有限公司 | skin biological materials |
| CN110801533A (en) * | 2019-11-18 | 2020-02-18 | 上海白衣缘生物工程有限公司 | Biological material for otology |
-
2002
- 2002-07-11 CN CN 02112478 patent/CN1387923A/en active Pending
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100415309C (en) * | 2003-07-03 | 2008-09-03 | 中国人民解放军第三○四医院 | Application of selective cell-removed pork skin as skin substitute of human body and its preparation method |
| CN100408107C (en) * | 2003-10-17 | 2008-08-06 | 胡杰 | Biological wound dressing |
| CN1737129B (en) * | 2004-08-19 | 2010-10-06 | 丘纪屏 | Artificial skin transplant and its preparation method |
| CN100372511C (en) * | 2005-11-30 | 2008-03-05 | 烟台正海生物技术有限公司 | Acellular dermal matrix |
| CN101954116A (en) * | 2010-08-31 | 2011-01-26 | 长沙达瑞奇实业有限公司 | Manufacturing method of xenoskin used for burns and scalds |
| CN109529123A (en) * | 2018-11-08 | 2019-03-29 | 中国人民解放军第四军医大学 | The vascularization holostrome organization engineering skin and preparation method thereof that hydrogel, nano fiber scaffold and Skin Cell assemble layer by layer |
| CN109529123B (en) * | 2018-11-08 | 2021-02-19 | 中国人民解放军第四军医大学 | Vascularized full-layer tissue engineering skin formed by assembling hydrogel, nanofiber scaffold and skin cells layer by layer and preparation method thereof |
| CN110732042A (en) * | 2019-11-18 | 2020-01-31 | 上海白衣缘生物工程有限公司 | skin biological materials |
| CN110801533A (en) * | 2019-11-18 | 2020-02-18 | 上海白衣缘生物工程有限公司 | Biological material for otology |
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