CN109701077A - A kind of micropore regenerating tissues matrix and its preparation and application - Google Patents
A kind of micropore regenerating tissues matrix and its preparation and application Download PDFInfo
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Abstract
The present invention provides a kind of micropore regenerating tissues matrix and its preparation and application.The micropore regenerating tissues matrix is the flaky material of porous structure, there is equally distributed micropore on material, and the aperture of micropore is 0.15~0.20mm, and pitch of holes is 0.8~1.2mm, the collagenous fibres protruded into the oriented hole of micropore inner surface.It is complete that micropore regenerating tissues matrix proposed by the present invention takes off cell, non-immunogenicity, the presence of acellular poison sexual factor, and the collagenous fibres of regenerating tissues matrix, reticular fibre, elastic fibers and its composition three-dimensional structure destroy it is smaller, it is preferable with surface of a wound compactness, there are many collagenous fibres protruded into hole for its micropore inner surface, it is possible to provide more moves into cell attachment site.
Description
Technical field
The invention belongs to biomedical materials fields, and in particular to a kind of micropore regenerating tissues matrix, and preparation method thereof
And application.
Background technique
Regenerating tissues matrix (Regenerative Tissue Matrix, RTM) is by human skin through serial physical, change
Method removes the cell component in epidermis and skin corium, removes allosome bring immunogenicity, remains the collagen of corium
Fiber, reticular fibre, elastomer and its composition three-D space structure.Nineteen ninety-five, Lifecell Corp., the U.S. first reported
The preparation method of acellular dermal matrix, while having carried out facing in its substitution reparation of Dermal defect caused by burn, wound
Bed application study.Later research finds acellular dermal matrix, except can be used for burning, in addition to the wound repairs such as wound, but also as
The substitute of mucous membrane and soft tissue is widely used in the subjects such as plastic surgery, oral cavity, neurosurgery, ophthalmology, ear-nose-throat department neck
Domain.
A kind of ideal regenerating tissues matrix (RTM) should be eliminated as much as having antigenic cell component, retain more
The basic structure of complete collagenous fibres ingredient and tissue, makes dermal matrix have preferable histocompatbility, the stability of height
The features such as with appropriate flexibility.And these characteristics of regenerating tissues matrix (RTM) depend mainly on its constituent and structure,
This is directly related with preparation method.Although the preparation of existing Xenogenic acellular dermal matrix comparative maturity,
It prepares and there is also some problems in applying, such as: trypsin process and NaCl-SDS method, and disadvantage is that de- cell degree is not thorough,
There is immune response in various degree, tissue compatible difference causes transplanting survival rate lower；Dispase II-Triton and NaOH disappears
Erosion method, disadvantage are that preparation process is complicated, and method used is larger to natural leather ultra microstructure brokenization, extracellular matrix at
Code insurance stays poor, there is certain damaging action to corium basilar memebrane.It and is to prevent collagen collagenous fibres mistake in preparation method
It spends after degrading and slowing down transplanting by the speed of proteases for decomposing, is crosslinked using glutaraldehyde, because glutaraldehyde has centainly thin
Cellular toxicity can cause body wound repair slow, extend repair time.
During dental implant and huge mole removal skin repair, having sizeable micropore on dermal matrix be can promote
Blood plasma, free cell, fibroblast isoreactivity ingredient penetrate into cell-less corium ground substance, maintain the battalion of dermal matrix early stage
Support supply.During dental implant, root of the tooth is wrapped up using micropore decellularized vascular matrix, micropore can promote teeth roots into fibre
Dimension cell is moved into, adheres to and is expanded, and acceleration is merged with body tissue, so that the survival rate of dental implant and shortening be promoted to repair
The multiple time.After the removal of huge mole, during using micropore decellularized vascular matrix to carry out skin repair as sticking patch, corium base
Micropore in matter can prevent the formation of hypohydrops, pneumatosis and hemotoncus, and blood plasma and free cell is promoted to penetrate into dermal matrix table
Face promotes the survival rate of skin-grafting.
Unified understanding there is no to punching and interporal lacuna size on dermal matrix at present, but when aperture is larger will form it is dotted or
Netted scar, and when aperture is smaller, histocyte is difficult to move into, and vascularization speed is slow.Therefore suitable micropore size is selected
And to provide more cell attachment sites around aperture particularly important to the vascularization of dermal matrix.
Laser boring is to reach practical laser processing technology earliest, is widely used in metal, leather, glass, timber
The processing of equal materials.Laser drilling has the advantages that trepanning velocity is fast, high-efficient, and aperture is small, micropore is evenly distributed.It will swash
Light cheesing techniques introduce the processing of Allodermis Matrix matrix micropores, and uniformly densely covered micropore can be produced on dermal matrix, reduce
The appearance for transplanting skin surface scar, may advantageously facilitate being uniformly distributed for capilary, improve dermal matrix transplanting survival to reach
Rate and the purpose for improving healing quality.
In the preparation method and cheesing techniques of existing Allodermis Matrix, for the presence for avoiding immunogenic substance, majority is adopted
With more violent preparation condition, result causes dermal matrix reticular fibre and elastic fibers largely to degrade, so that collagen is fine
Three-dimensional structure brokenization of dimension greatly, reduces the transplanting success of dermal matrix；In dermal matrix cheesing techniques, relatively mostly use
Dermal matrix or wet dermal matrix after rehydration are punched, and drilling technology majority is prepared in acellular dermal matrix and completed
It carries out afterwards, aperture majority is controlled in 80-140um, but aperture inner surface cell attachment site is less.Chinese patent
CN105688286A discloses a kind of laser micropore acellular dermal matrix and preparation method thereof, and pulse laser is used in patent
Device punches dermal matrix, and raw material is using the wet acellular dermal matrix prepared, although its pore-size distribution is equal
It is even, but cell attachment site is less in its aperture.Chinese patent CN200510126108.6 discloses a kind of acellular dermal base
Matter and preparation method thereof cleans mammal corium using high-concentration sodium hydroxide in patent repeatedly and obtains acellular dermal base
Matter, alkali process condition acutely cause part collagenous degeneration, protein cleavage to affect original protein structure in dermal matrix.
Summary of the invention
To solve above-mentioned problem of the prior art, the present invention provides a kind of micropore regenerating tissues matrix.Micropore regeneration
It is complete that periplast takes off cell, non-immunogenicity, the presence of acellular poison sexual factor, and the collagenous fibres of periplast, netted
The three-dimensional structure destruction of fiber, elastic fibers and its composition is smaller, preferable with surface of a wound compactness, and there are many stretches micropore inner surface
Enter the collagenous fibres in hole, it is possible to provide more move into cell attachment site.
Second object of the present invention is to propose the preparation method of the micropore regenerating tissues matrix.By to freeze-drying
Skin corium matrix afterwards carries out laser boring, then expands de- cell processing, oscillation cleaning and amino acid solution using rehydration, lye
Immersion treatment obtains micropore Allodermis Matrix.
Third object of the present invention is to propose the application of the micropore regenerating tissues matrix.
Realize the technical solution of above-mentioned purpose of the present invention are as follows:
A kind of micropore regenerating tissues matrix, the micropore regenerating tissues matrix are the flaky material of porous structure, material
On have an equally distributed micropore, the aperture of micropore is 0.15~0.20mm, and pitch of holes is 0.8~1.2mm, and micropore inner surface is oriented
The collagenous fibres protruded into hole.
A kind of preparation method of micropore regenerating tissues matrix, comprising the following steps:
1) heterogenous skin pre-processes: the heterogenous skin for scraping off subcutaneous tissue being cleaned in physiological saline, then in chlorination
It is impregnated in sodium solution, removes skin epidermis, then clean in physiological saline, obtain Allodermis Matrix；
2) freeze-drying process: Allodermis Matrix being put into freeze drying protectant and is impregnated, be put into after shaping it is quick-frozen in liquid nitrogen, so
Vacuum freeze drying afterwards；
3) punching is handled: being punched using laser-processing system to the Allodermis Matrix after freeze-drying；
4) lye expands de- cell processing: the Allodermis Matrix after punching is put into physiological saline and/or freeze drying protectant
Then middle rehydration removes skin corium inner cell using lye expansion process, then is with oscillation cleaning in physiological saline to efflux
It is neutral.
Further, in step 1), the concentration of the sodium chloride solution is 0.8~1.2mol/L, unit area skin graft
Solid-liquid ratio with liquid is 1cm2: (0.5~2) mL, soaking time are 18~30h, and soaking temperature is 24~32 DEG C.
Wherein, freeze drying protectant described in step 2) is trehalose weak brine solution, the concentration of trehalose is 200~
600mmol/L, weak brine solution therein are made of 1 part of physiological saline and two parts of waters for injection, unit area skin graft and liquid
The solid-liquid ratio of body is 1cm2: (0.5~2) ml, soaking time are 4~6h, and soaking temperature is 30~40 DEG C.
Preferably, after step 2) freeze drying protectant impregnates, by the Allodermis Matrix hard frame (material of the hard frame
Matter can mainly bear the low temperature of liquid nitrogen just, such as polyimides, polytrifluorochloroethylene) shaping is fixed, it is kept fixed state
Carry out the freeze-drying process and punching processing.Frame can be removed by having beaten hole.
A preferred technical solution of the present invention is the operation of liquid nitrogen flash freezer described in step 2) are as follows: -6~-2 DEG C of pre-coolings,
Then quick-frozen 20~60s in liquid nitrogen is hung in cotton rope；
The operation of the vacuum freeze drying are as follows: the precooling temperature that vacuum freeze drier is arranged is -30~-40 DEG C, is put
It carries out vacuumizing frozen dried after entering Allodermis Matrix.
Further, the frozen dried that vacuumizes is divided into two stages, the stage 1: temperature lower than -30 DEG C, it is true
Reciprocal of duty cycle maintains 14h under conditions of being less than 3Pa；Stage 2: being warming up to 20 DEG C, maintains 4h under conditions of vacuum degree is less than 3Pa；It is whole
During a vacuum freeze drying, vacuum degree is not higher than 30Pa.
Wherein, laser-processing system described in step 3) is multi-functional ultraviolet system of processing, uses diode pumping solid
Volumetric laser, wavelength 355nm；
The laser boring parameter are as follows: aperture is 0.15~0.20mm, and pitch of holes is 0.8~1.2mm.
Wherein, lye described in the step 4) takes off cell expansion processing, processing mode are as follows: using containing 0.1~
The Na of 0.5mol/L2CO3It is handled with mass fraction for the solution of 0.1~0.4%SDS；
Or, first using the Na of 0.1~0.5mol/L2CO3Then base extraction uses mass fraction for 0.1~0.4% again
SDS processing；
Preferably, the solid-liquid ratio of skin graft surface area and the solution of processing is (1~3) cm2: 1mL vibrates 1~3h, stands leaching
2~6h is steeped, oscillation and immersion treatment 2-5 times are repeated；
It is highly preferred that the material after lye expansion process is put into the solution of amino acid or surfactant at immersion
Reason, then handled through physiological saline oscillation cleaning, radiation sterilizing, it dispenses；
The amino acid solution of the immersion treatment are as follows: in every liter of solution contain 2g threonine, 2g serine, 1g glutamic acid and
1g asparatate.
Application of the micropore regenerating tissues matrix of the present invention as dental implant packing material and huge mole sticking patch.
The beneficial effects of the present invention are:
The de- cell of micropore regenerating tissues matrix proposed by the present invention is complete, and non-immunogenicity, acellular poison sexual factor is deposited
, and collagenous fibres in matrix, reticular fibre, elastic fibers and its composition three-dimensional structure destroy smaller, be bonded with the surface of a wound
Preferably, there are many collagenous fibres protruded into hole for micropore inner surface, it is possible to provide more moves into cell attachment site for degree.
The preparation method of micropore regenerating tissues matrix of the present invention carries out laser boring, micropore using dry Allodermis Matrix
After rehydration, lye expansion process, protrudes into the collagenous fibres in hole in the presence of many in micropore, be conducive to the attachment for moving into cell and
Proliferation improves the survival rate of regenerating tissues matrix transplanting.
This method, using freezing rehydration technical process, can help to cell detachment before lye expands removal cell technique,
Shorten base extraction intensity, is conducive to the three-dimensional knot for mitigating regenerating tissues Collagen fiber, reticular fibre, elastic fibers composition
The destruction of structure.
This method uses laser boring mode, and precision is high, speed is fast, and micropore is uniformly distributed, and can reduce transplanting epidermis face
The appearance of scar improves wound healing quality.
Compared with existing preparation method, The present invention reduces glutaraldehydes to the cross-linking process of regenerating tissues matrix, avoids
The introducing of cytotoxicity factor.
Detailed description of the invention
Fig. 1 is the photo (light aobvious × 10) under 1 micropore regenerating tissues matrix optical microscope of embodiment.
Fig. 2 is the electromicroscopic photograph of 1 micropore regenerating tissues matrix micropores inner surface of embodiment.
Fig. 3 is the photo (light aobvious × 10) under 2 micropore regenerating tissues matrix optical microscope of embodiment.
Fig. 4 is the electromicroscopic photograph of 2 micropore regenerating tissues matrix micropores inner surface of embodiment.
Specific embodiment
The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention..
In embodiment, unless otherwise noted, used method is the method for this field routine.The present invention relates to
Raw material or the commercially available acquisition of reagent.
Illustrate technical solution of the present invention below by way of specific embodiment.
Embodiment 1
Micropore regenerating tissues Matrix formulation procedure of the present invention the following steps are included:
1. washing: heterogenous skin being taken out from storage protection liquid, after scraping off subcutaneous tissue, is put into physiological saline and vibrates
Cleaning, the area of skin graft and the solid-liquid ratio (cm of physiological saline2: mL) it is 1:1, feed liquid total volume is no more than oscillation container capacity
50%, frequency of oscillation 30rpm, oscillation cleaning time are 1.5h, every oscillation 0.5h, replace a physiological saline, resonance is swung clear
It washes 3 times.
2. removing epidermis: the heterogenous skin that oscillation cleaning is crossed is put into fill and impregnate in 1mol/L NaCl solution, material
Liquor ratio (cm2: mL) it is 1:1, under the conditions of 28 DEG C, impregnate for 24 hours.After the completion of immersion, heterogenous skin is taken out, tears epidermis off.It will
The skin for eliminating epidermis obtains Allodermis Matrix layer according still further to step 1 oscillation cleaning.
3. liquid nitrogen frozen: Allodermis Matrix layer is put into the low concentration of trehalose containing 400mmol/L salt water (by 1 part of physiological saline
Formed with 2 parts of waters for injection) in, under the conditions of 37 DEG C, it is incubated for 5h.By the skin graft shaping after incubation, clamped with the frame of hard
Four sides, make skin graft keep it is smooth, prevent during the freeze-drying process dermal matrix roll up convenient for post laser punching.By shaping
After skin graft afterwards is pre-chilled under the conditions of -4 DEG C, quick-frozen 30s in liquid nitrogen is hung in.
4. freeze-drying: the Allodermis Matrix after will be quick-frozen is put into is pre-chilled to -30~-40 DEG C of vacuum freeze drying rapidly
In machine, opens vacuum pump and carry out vacuum drying processing, drying box vacuum degree is extracted into 10Pa or less.Temperature lower than -30 DEG C,
Vacuum degree maintains 14h under conditions of being less than 3Pa；Then 20 DEG C are warming up to, maintains 4h under conditions of vacuum degree is less than 3Pa, it is whole
In a process of vacuum drying, vacuum degree is not higher than 30Pa.
5. laser punching: setting the parameter of multi-functional ultraviolet laser machining system (German-Chinese DL500U), select the purple of 355nm
The Allodermis Matrix of freeze-drying is vacantly placed in frame by outer light, and skin graft is made, there are 5mm distance is greater than, to prevent bottom apart from bottom plate
Portion's waste heat causes skin graft to lose；Laser boring parameter are as follows: punching spacing 1.0mm, aperture 0.2mm carry out matrix form punching, obtain
Dry micropore Allodermis Matrix.
6. rehydration: dry micropore Allodermis Matrix being put into physiological saline, solid-liquid ratio (cm2: mL) it is 1:1, oscillation
20min is impregnated, micropore Allodermis Matrix is obtained.
7. lye expands de- cell processing: micropore Allodermis Matrix is added to the Na containing 0.2mol/L2CO3And 0.2%SDS
Solution in carry out immersion treatment, solid-liquid ratio (cm2: mL) 1:1,50% of feed liquid total volume no more than oscillator volume, oscillation
Frequency is 30rpm, vibrates 2h, is standing and soaking 4h, repeats oscillation cleaning and immersion treatment 4 times.Again by dermal matrix according to step 1
The cleaning solution of oscillation cleaning to outflow is neutral (pH6-8), obtains micropore acellular dermal matrix.
8. amino acid solution immersion treatment: decellularized vascular matrix matrix being added to amino acid solution and (is contained in every L solution
Have 2g threonine, 2g serine, 1g glutamic acid and 1g asparatate) in, immersion treatment 1h is vibrated, immersion is then allowed to stand
0.5h.Dermal matrix is obtained into micropore regenerating tissues matrix according to step 1 oscillation cleaning again.
9. sterilization: using Co60 radiation sterilizing, irradiation dose 25KGy.
10. encapsulation: sterilized micropore regenerating tissues matrix being immersed in PBS buffer solution, and with plastic bag sealing packet
Dress.
Obtained product photo is shown in Fig. 1, and micropore regenerating tissues matrix made from the present embodiment is the sheet material of porous structure
Expect there is equally distributed micropore on material, aperture is 0.15~0.20mm, and pitch of holes is 0.8~1.2mm, and micropore inner surface has
The collagenous fibres protruded into hole, electromicroscopic photograph referring to fig. 2.
Embodiment 2
The micropore regenerating tissues Matrix formulation procedure of the present embodiment the following steps are included:
Step 1-5 is same as Example 1；
6. rehydration is handled: being successively decreased rehydration mode using gradient, i.e., the micropore Allodermis Matrix of freeze-drying is placed in 400mmol/L's
In trehalose low concentration saline solutions, solid-liquid ratio (cm2: ml) it is 1:1,15min is impregnated in oscillation；It then takes out and is added to
In 200mmol/L trehalose low concentration salt water, solid-liquid ratio (cm2: ml) are as follows: 15min is impregnated in 1:1, oscillation；Then it is placed in no sea again
In the physiological saline of algae sugar, solid-liquid ratio (cm2: ml) are as follows: 15min is impregnated in 1:1, oscillation, i.e. acquisition micropore Allodermis Matrix.
7. lye expands de- cell processing: micropore Allodermis Matrix is added to the Na of 0.2mol/L2CO3In carry out at immersion
Reason, solid-liquid ratio (cm2: ml) 1:1, feed liquid total volume 50%, frequency of oscillation 30rpm no more than oscillator volume, oscillation 2h,
It is standing and soaking 4h, is repeated oscillation cleaning immersion treatment 4 times.It is different that dermal matrix is obtained into de- cell according to step 1 oscillation cleaning again
Body dermal matrix.
8. surfactant is handled: by decellularized vascular matrix matrix, being added to oscillation treatment in 0.2%SDS solution, expect
Liquor ratio (cm2: ml) 1:1, the processing time is 1h, for sloughing the cell fragment of dermal matrix remnants, reduces immunogenicity.By table
Activating agent cell free dermal matrix in face obtains decellularized vascular matrix matrix according to step 1 oscillation cleaning.
9. amino acid solution immersion treatment: decellularized vascular matrix matrix being added to amino acid solution and (is contained in every L solution
Have 2g threonine, 2g serine, 1g glutamic acid and 1g asparatate) in, immersion treatment 1h is vibrated, immersion is then allowed to stand
0.5h.Dermal matrix is obtained into micropore regenerating tissues matrix according to step 1 oscillation cleaning again.
10. sterilization: using Co60 radiation sterilizing, irradiation dose 25KGy.
11. encapsulation: sterilized micropore regenerating tissues matrix being immersed in PBS buffer solution, and with plastic bag sealing packet
Dress.
Obtained product photo is shown in Fig. 3, and micropore regenerating tissues matrix made from the present embodiment is the sheet material of porous structure
Expect there is equally distributed micropore on material, aperture is 0.15~0.20mm, and pitch of holes is 0.8~1.2mm, and micropore inner surface has
The collagenous fibres protruded into hole, referring to fig. 4.
Comparative example 1
The micropore regenerating tissues Matrix formulation procedure of this comparative example the following steps are included:
Steps 1 and 2 are same as Example 1.
3. base extraction: Allodermis Matrix is added in 0.6% NaOH solution and is impregnated, solid-liquid ratio (cm2:ml) 1:1, material
Liquid total volume is no more than 50%, frequency of oscillation 30rpm of oscillator volume, vibrates 1h, is standing and soaking 5h, coprocessing 18h.Again
Dermal matrix is handled according to step 1 oscillation cleaning.
4. surfactant is handled: Allodermis Matrix matrix is added in 0.2%SDS solution and carries out immersion treatment, feed liquid
Than (cm2: mL) 1:1,50% of feed liquid total volume no more than oscillator volume, frequency of oscillation 30rpm, oscillation treatment 0.5h.
Dermal matrix is handled according to step 1 oscillation cleaning again.
5. glutaraldehyde cross-linking is handled: Allodermis Matrix matrix is added to 0.2% glutaraldehyde solution immersion treatment, solid-liquid ratio
(cm2: mL) 1:1, immersion treatment 20min.Dermal matrix is handled according to step 1 oscillation cleaning again.
6. laser punching: setting the parameter of multi-functional ultraviolet laser machining system, select the ultraviolet light of 355nm, by allosome
Dermal matrix is vacantly placed in frame, and for dermal matrix apart from bottom plate there are certain distance, laser boring parameter is to punch spacing
1.0mm, aperture 0.15mm carry out matrix form punching.Micropore Allodermis Matrix matrix is handled according to step 1 oscillation cleaning again.
7. sterilization: using Co60 radiation sterilizing, irradiation dose 25KGy.
8. encapsulation: by sterilized micropore regenerating tissues matrix, being impregnated with PBS buffer solution, also packed with plastic bag sealing.
Experimental example 1: the performance detection of regenerating tissues matrix
The biology performances such as the mechanical property of regenerating tissues matrix prepared to embodiment 1,2 and comparative example 1 carry out
Test, performance indicator includes tensile strength, suture strength, extension elongation rate.Specific testing result is as shown in table 1.The results show that
Embodiment 1,2 products and the better compactness of skin are preferable, and toughness and elasticity are good, and are able to satisfy sticking patch after huge mole is removed and stitch
Close the requirement of intensity.
1 micropore regenerating tissues matrix testing result of table
| Project | Tensile strength (MPa) | Suture strength (N) | Extension elongation rate (%) |
| Embodiment 1 | 6.2 | 19.5 | 14.0 |
| Embodiment 2 | 6.4 | 19.8 | 14.1 |
| Comparative example 1 | 4.7 | 18.2 | 11.2 |
2 animal model test of experimental example
Experimental subjects: taking healthy SD rat 48, and male and female are unlimited, and rat body weight is 250 ± 10g.
Experimental procedure:
1. the yellow Jackets (50mg/kg) using 3% carry out intraperitoneal anesthesia to rat.
2. SD rat is fixed on operating table, back unhairing, iodine disinfection, aseptic towel is spread.
3. being removed the peel using drum dermatome, adjustment drum dermatome takes skin thickness to suitable position, and pulling handle is complete
Full layers skin takes skin, obtains self razor graft, and skin graft size is 2.5cm × 2.5cm.
4. rat is then randomly divided into 3 groups, every group 16.Example 1 group: surface of a wound transplanting uses the preparation of embodiment 1
The micropore regenerating tissues matrix that method obtains；2 groups of embodiment: the micropore that surface of a wound transplanting is obtained using the preparation method of embodiment 2
Regenerating tissues matrix；Control group: autologous transplanting razor graft.
5. after micropore regenerating tissues matrix and grafts, being covered with petrolatum gauze and aseptic dressing, and fixed, operation
Single cage is raised afterwards, and fixed, dressing after 2 weeks.
Experimental result:
Two weeks after skin graft and micropore regenerating tissues matrix transfer operation, 2 groups of regenerating tissues matrix of example 1 group and embodiment
Major part survives, and two groups of phenomenons have fragmentary spot to be dispersed in the surface of a wound, the micropore regenerating tissues matrix of deep layer is in skin without significant difference
Undertissue is completely embedded, and control group surface of a wound major part razor graft survives well.4 weeks after transfer operation, example 1 group and implementation
2 groups of example have been approached healing, and blood fortune is good, and transplantation site has furfur, the close healing of the control group surface of a wound.6 weeks after transfer operation, implement
1 group of example and 2 groups of embodiment, the surface of a wound heals substantially, and for two groups of appearances without significant difference, the surface of a wound is more smooth；Control group surface of a wound base
This healing.
Each group surface of a wound skin grafts survival rate is as shown in table 2.
The postoperative each group skin grafts survival rate of table 2 compares (%, ± s)
Note: compared with the control group,*P<0.05
Postoperative 2 weeks and 4 weeks, 2 groups of transplanting successes of example 1 group and embodiment were lower than control group (P < 0.05)；Postoperative 6
Week, example 1 group and 2 groups of embodiment not statistically significant compared with the control group (P > 0.05).
The test of 3 dental implant of experimental example
Case selection: section receives in 3 crescent moons the patient for needing dental implant 23, male 13, female 10, patient oral cavity
It is hygienic good.
Material: 1 micropore regenerating tissues matrix of embodiment.
Operation method: preoperative planning carries out examination of mouth to all plantation cases, uses the width of calliper to measure alveolar ridge
Degree, the height of slot ridge.Planting body implantation and the implantation of micropore regenerating tissues matrix, the micropore regenerating tissues base of the suitable size of clip
Periplast is folded and is implanted into according to the degree that lip side is recessed by matter, and connective tissue faces outwardly.It is anti-that postoperative routine gives antibiotic
Inflammation treatment.
Observation method: tracing study patient is 1 week postoperative, 3 weeks, 3 months, transplanting regenerating tissues matrix wound healing in 6 months
Situation.And using taking x-rays assessment plantation sport adjacent teeth and the relationship of surrounding tissue.
As a result: no obvious tumefaction is removed in observation operation, does not occur the symptoms such as fash, fever, mono-anesthesia, pain, bleeding.Art
3 months X-rays show the fitting of peri-implant fine jade tissue tight without transmission gaps afterwards, and no obvious row bone resorption occurs.Postoperative 6
Moon observation planting body survival rate 100%.
Although above by embodiment, the present invention is described, however, it will be appreciated by those skilled in the art that without departing from
Under the premise of spirit of that invention and essence, to the improvement and modification that the present invention is done, it is within the scope of protection of the invention interior.
Claims (10)
1. a kind of micropore regenerating tissues matrix, which is characterized in that the micropore regenerating tissues matrix is the sheet of porous structure
Material has equally distributed micropore on material, and the aperture of micropore is 0.15~0.20mm, and pitch of holes is 0.8~1.2mm, micropore
The collagenous fibres protruded into the oriented hole of inner surface.
2. a kind of preparation method of micropore regenerating tissues matrix, which comprises the following steps:
1) heterogenous skin pre-processes: the heterogenous skin for scraping off subcutaneous tissue is cleaned in physiological saline, it is then molten in sodium chloride
It is impregnated in liquid, removes skin epidermis, then clean in physiological saline, obtain Allodermis Matrix；
2) freeze-drying process: Allodermis Matrix being put into freeze drying protectant and is impregnated, be put into after shaping it is quick-frozen in liquid nitrogen, then very
Vacuum freecing-dry；
3) punching is handled: being punched using laser-processing system to the Allodermis Matrix after freeze-drying；
4) lye expands de- cell processing: the Allodermis Matrix after punching being put into physiological saline and/or freeze drying protectant multiple
Then water removes skin corium inner cell using lye expansion process, then with oscillation cleaning in physiological saline to efflux be neutral.
3. preparation method according to claim 2, which is characterized in that in step 1), the concentration of the sodium chloride solution
For 0.8~1.2mol/L, the solid-liquid ratio of unit area skin graft and liquid is 1cm2: (0.5~2) mL, soaking time be 18~
30h, soaking temperature are 24~32 DEG C.
4. preparation method according to claim 2, which is characterized in that freeze drying protectant described in step 2) is seaweed malt sugar
Saline solution, wherein the concentration of trehalose is 200~600mmol/L, and weak brine solution is injected by 1 part of physiological saline and two parts
It is formed with water, the solid-liquid ratio of unit area skin graft and liquid is 1cm2: (0.5~2) ml, soaking time are 4~6h, soaking temperature
It is 30~40 DEG C.
5. preparation method according to claim 2, which is characterized in that, will be described different after step 2) freeze drying protectant impregnates
Body corium is fixed with hard frame, and the state that is kept fixed carries out the freeze-drying process and punching processing.
6. preparation method according to claim 2, which is characterized in that the operation of liquid nitrogen flash freezer described in step 2) are as follows: -6
~-2 DEG C of pre-coolings, then hang in quick-frozen 20~60s in liquid nitrogen with cotton rope；
The operation of the vacuum freeze drying are as follows: the precooling temperature that vacuum freeze drier is arranged is -30~-40 DEG C, is put into different
It carries out vacuumizing frozen dried after body corium.
7. preparation method according to claim 6, which is characterized in that the frozen dried that vacuumizes is divided into two stages,
Stage 1: 14h is maintained under conditions of temperature is less than 3Pa lower than -30 DEG C, vacuum degree；Stage 2: 20 DEG C are warming up to, in vacuum degree
Less than maintaining 4h under conditions of 3Pa；In entire process of vacuum drying, vacuum degree is not higher than 30Pa.
8. preparation method according to claim 2, which is characterized in that laser-processing system described in step 3) is multi-functional
Ultraviolet system of processing uses diode-pumped solid laser, wavelength 355nm；
The laser boring parameter are as follows: aperture is 0.15~0.20mm, and pitch of holes is 0.8~1.2mm.
9. preparation method according to claim 3, which is characterized in that lye described in step 4) takes off at cell expansion
Reason, processing mode are as follows: use the Na containing 0.1~0.5mol/L2CO3With mass fraction be 0.1~0.4%SDS solution into
Row processing；
Or, first using the Na of 0.1~0.5mol/L2CO3Base extraction, then use again mass fraction for 0.1~0.4%SDS at
Reason；
Preferably, the solid-liquid ratio of skin graft surface area and the solution of processing is (1~3) cm2: 1mL vibrates 1~3h, it is standing and soaking 2~
6h repeats oscillation and immersion treatment 2~5 times；
It is highly preferred that the material after lye expansion process is put into immersion treatment in the solution of amino acid, then shake through physiological saline
Swing cleaning, radiation sterilizing, packing；
The amino acid solution of the immersion treatment are as follows: contain 2g threonine, 2g serine, 1g glutamic acid and 1g days in every liter of solution
L-aminobutanedioic acid.
10. application of the micropore regenerating tissues matrix described in claim 1 as dental implant packing material and huge mole sticking patch.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114980938A (en) * | 2020-12-21 | 2022-08-30 | 爱恩斯生物科技(昆山)有限公司 | Acellular nerve graft material and method for producing same |
| CN116212094A (en) * | 2023-05-08 | 2023-06-06 | 天津锦安旭瑞生物科技有限公司 | Active cell xenogeneic dermis repairing biological auxiliary material and preparation method thereof |
Citations (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1343523A (en) * | 2000-09-19 | 2002-04-10 | 中国人民解放军第二军医大学 | Millipore non-cell xanoepidermis substitute |
| CN102188751A (en) * | 2010-03-19 | 2011-09-21 | 温州医学院附属第一医院 | Laser micropore porcine acellular dermal matrix and preparation method thereof |
| CN102293690A (en) * | 2011-06-07 | 2011-12-28 | 天津市托福医用原子能科技有限公司 | Preparation method of freeze-thawing xenogenic laser microporous irradiated acellular dermal matrix and product thereof |
| WO2012142419A1 (en) * | 2011-04-14 | 2012-10-18 | Lifecell Corporation | Regenerative materials |
| CN102869392A (en) * | 2010-02-26 | 2013-01-09 | Cg生物技术有限公司 | Method for producing acellular dermal matrix, and acellular dermal matrix produced by same |
| CN105126170A (en) * | 2015-08-18 | 2015-12-09 | 深圳兰度生物材料有限公司 | Acellular dermal matrix and preparing method of acellular dermal matrix |
| CN107007886A (en) * | 2017-03-03 | 2017-08-04 | 北京博辉瑞进生物科技有限公司 | A kind of biological tissue's host material, preparation method and its usage |
| CN107412867A (en) * | 2017-05-10 | 2017-12-01 | 广州润虹医药科技股份有限公司 | A kind of preparation method of Xenogenic acellular dermal matrix |
| US20170368231A1 (en) * | 2016-06-23 | 2017-12-28 | Dermagenesis, Llc | Bioengineered Regenerative Graft Matrix, and Methods for Making Thereof |
-
2019
- 2019-01-29 CN CN201910084392.7A patent/CN109701077B/en active Active
Patent Citations (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1343523A (en) * | 2000-09-19 | 2002-04-10 | 中国人民解放军第二军医大学 | Millipore non-cell xanoepidermis substitute |
| CN102869392A (en) * | 2010-02-26 | 2013-01-09 | Cg生物技术有限公司 | Method for producing acellular dermal matrix, and acellular dermal matrix produced by same |
| CN102188751A (en) * | 2010-03-19 | 2011-09-21 | 温州医学院附属第一医院 | Laser micropore porcine acellular dermal matrix and preparation method thereof |
| WO2012142419A1 (en) * | 2011-04-14 | 2012-10-18 | Lifecell Corporation | Regenerative materials |
| CN102293690A (en) * | 2011-06-07 | 2011-12-28 | 天津市托福医用原子能科技有限公司 | Preparation method of freeze-thawing xenogenic laser microporous irradiated acellular dermal matrix and product thereof |
| CN105126170A (en) * | 2015-08-18 | 2015-12-09 | 深圳兰度生物材料有限公司 | Acellular dermal matrix and preparing method of acellular dermal matrix |
| US20170368231A1 (en) * | 2016-06-23 | 2017-12-28 | Dermagenesis, Llc | Bioengineered Regenerative Graft Matrix, and Methods for Making Thereof |
| CN107007886A (en) * | 2017-03-03 | 2017-08-04 | 北京博辉瑞进生物科技有限公司 | A kind of biological tissue's host material, preparation method and its usage |
| CN107412867A (en) * | 2017-05-10 | 2017-12-01 | 广州润虹医药科技股份有限公司 | A kind of preparation method of Xenogenic acellular dermal matrix |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114980938A (en) * | 2020-12-21 | 2022-08-30 | 爱恩斯生物科技(昆山)有限公司 | Acellular nerve graft material and method for producing same |
| CN116212094A (en) * | 2023-05-08 | 2023-06-06 | 天津锦安旭瑞生物科技有限公司 | Active cell xenogeneic dermis repairing biological auxiliary material and preparation method thereof |
| CN116212094B (en) * | 2023-05-08 | 2023-08-04 | 天津锦安旭瑞生物科技有限公司 | Active cell xenogeneic dermis repairing biological auxiliary material and preparation method thereof |
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