CN103143058B - Preparation of composite hydrogel tissue engineer cartilage repairing scaffold with biological activity - Google Patents
Preparation of composite hydrogel tissue engineer cartilage repairing scaffold with biological activity Download PDFInfo
- Publication number
- CN103143058B CN103143058B CN201310110593.2A CN201310110593A CN103143058B CN 103143058 B CN103143058 B CN 103143058B CN 201310110593 A CN201310110593 A CN 201310110593A CN 103143058 B CN103143058 B CN 103143058B
- Authority
- CN
- China
- Prior art keywords
- cartilage
- aqueous solution
- preparation
- tissue engineering
- paraffin
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Landscapes
- Materials For Medical Uses (AREA)
Abstract
The invention discloses a preparation method of a composite hydrogel tissue engineer cartilage repairing scaffold with biological activity. The preparation method comprises the steps of: preparing a paraffin model of the cartilage repairing scaffold; by taking the paraffin model as a template, selecting and fermenting-cultivating a bacterial strain capable of secreting bacterial cellulose; removing paraffin from a fermentation product, and purifying, dewatering and immersing the fermentation product, thus obtaining a bacterial cellulose hydrogel cartilage repairing scaffold containing 30-50% of CaC12 aqueous solution by weight; dissolving polyvinyl alcohol in an Na2HPO4 aqueous solution, thus obtaining a mixed solution A; enabling the mixed solution A to enter the bacterial cellulose hydrogel cartilage repairing scaffold by virtue of an immersion method to obtain a product; freezing-unfreezing the obtained product for multiple times, and irradiating the product by gamma rays, thus obtaining the composite hydrogel tissue engineer cartilage repairing scaffold with biological activity. Composite hydrogel prepared by the method has an appropriate appearance, a porous structure, an excellent mechanical property and excellent biological activity and can be used as a tissue engineer scaffold material for repairing cartilage tissue.
Description
Technical field
The present invention relates to a kind of biomaterial compound preparation field, particularly a kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support.
Background technology
Cartilage defect is difficult to self-regeneration, causes very large misery to patient.The reparation of cartilage defect is the thorny problem of puzzlement clinical treatment, and worldwide this kind of patient numbers is numerous, and along with social senilization's aggravation, cartilage defect sickness rate presents growing trend.Autologous, homogenous cartilage is mainly adopted to transplant and Endoprostheses to cartilage defect repair clinically.Cartilage transplantation, due to the limited source of cartilaginous tissue, significantly limit the application clinically of this method.Although it is sufficient that allogeneic cartilage transplants cartilaginous tissue source, there is certain immune rejection problems in this method; In addition, strong bonded can be reached between transplanting cartilage and adjacent cartilage and remain the difficult problem do not solved completely.Traditional prosthetic replacement's service life is short, and along with the growth of service life, easily produces material failure, the problem such as aging; In recent years, along with the development of tissue engineering technique, organization engineered cartilage is considered to the repair of cartilage method having application prospect most.Tissue engineering bone/cartilage reparation is with cartilaginous tissue recovery support for carrier, in conjunction with cartilage seed cell, somatomedin, by body or In vitro culture build cartilaginous tissue.Cartilaginous tissue recovery support has important function in this course, and it is cell, the reconstruction of tissue provides necessary three dimensions and mechanics support, plays the effect of analog cell epimatrix; There is good histocompatibility, applicable aperture and porosity, be beneficial to the propagation of chondrocyte and stick, and the infiltration of nutrient substance and the discharge of products of cellular metabolism; There is good biological activity, the cartilaginous tissue of reconstruction and the autologous bone strong bonded of bottom can be made.Therefore need cartilaginous tissue recovery support to have good biological activity, applicable mechanical property, desirable three-dimensional microstructures, and there is the macro morphology conformed to normal articular cartilage.
Polyvinyl alcohol (PVA) hydrogel is the water-soluble swollen body of the network-like structure that hydrophilic polyethene alcohol macromole is formed after crosslinked.PVA hydrogel has stable chemical property, good physical and mechanical properties, is easy to molding, with the good biocompatibility of tissue, especially the physical property that PVA hydrogel is similar to tissue performance, to excellent permeability, suitable autgmentability, the good pliability etc. such as hydrone, these characteristics make PVA hydrogel be used as the biomedical sectors such as wound dressing, drug release carrier, artificial tissue, organizational project.Recent study confirms that PVA hydrogel has the bio-mechanical property similar with human articular cartilage and good biocompatibility, the cartilage surface of energy reconstruct smooth after implant into body, alleviate wearing and tearing, Some substitute articular cartilage, delaying or stop the generation of osteoarthritis after traumatic, is a kind of desirable articular cartilage substitution material.But PVA hydrogel does not have biological activity compared with human articular cartilage, affect the fixing of cartilage and repair function, and the mechanical strength of PVA hydrogel is more weak, is difficult to when using as tissue engineering bracket control its exterior appearance and microstructure.Therefore for improving the performance of PVA hydrogel, the present invention by the bacteria cellulose aquagel compound of PVA hydrogel with the excipient ability and mechanical property with excellence, and adds the hydroxyapatite with good biological activity by reaction.
Bacterial cellulose (BC) is as a kind of excellent biomaterial, and the bacteria cellulose aquagel formed under moisture state has physics, the chemical property of its uniqueness: it has natural three-dimensional manometer network structure; Good mechanical property; High-hydrophilic, good ventilative, water suction, water permeability: excellent excipient ability, can accurately control its exterior appearance and microstructure when Bacterial cellulose is cultivated.Meanwhile, Bacterial cellulose also have in good body, biocompatibility in vitro and good biodegradability.Research shows, the mechanical property (stretch modulus and elastic modelling quantity) of bacteria cellulose aquagel excellence can reach the level similar to normal cartilage; Chondrocyte shows high-caliber growth on Bacterial cellulose simultaneously, and can maintain the migration propagation of cartilage differentiation, support chondrocyte.Hydroxyapatite is a kind of calcium phosphate biological ceramic, has good biocompatibility, interface biological activity and bone conductibility, synosteosis ability.The cartilaginous tissue of reconstruction and the autologous bone strong bonded of bottom can be made in tissue engineering bone/cartilage reparation.
The present invention adopts computed tomography, three-dimensional fast shaping technology, bacterial strain/paraffin mixed culture fermentation technology and the preparation of hydroxyapatite is combined with freeze-thaw method, prepares one and has bioactive composite aquogel tissue engineering bone/cartilage recovery support.This tissue engineering bone/cartilage recovery support has the inierpeneirating network structure that PVA hydrogel network shape structure and BC hydrogel network structure are formed, and hydroapatite particles is evenly embedded in inierpeneirating network structure inside simultaneously; The support obtained has the macro morphology and mechanical property that conform to normal articular cartilage, good biological activity, and desirable three-dimensional porous structure.Present invention process is simple, cost is low, pollution-free, the composite aquogel tissue engineering bone/cartilage recovery support of preparation can be used as tissue engineering bracket material and is applied to during the cartilaginous tissues such as articular cartilage reparation, meniscal repairs repair.
Summary of the invention
The object of this invention is to provide a kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support.Relate to a kind of biomaterial compound preparation field.Present invention process is simple, cost is low, pollution-free, and the composite aquogel tissue engineering bone/cartilage recovery support of preparation has applicable exterior appearance and loose structure, good mechanical property and biological activity, can be used as tissue engineering bracket material and be applied in the cartilaginous tissue reparations such as articular cartilage reparation, meniscal repairs.
A kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support that the present invention relates to.Comprise:
(1) adopt computed tomography to scan the cartilaginous tissue that expection is repaired, obtain the mathematical model of cartilaginous tissue three dimensional structure, utilize mathematical model to design the three-dimensional porous structure of timbering material inside, obtain its minus by Boolean calculation.Utilizing three-dimensional fast shaping technology, adopt three-dimensional rapid printer, take paraffin powder as moulding material, the paraffin minus of obtained repair of cartilage support.
(2) Bacterial cellulose fermentation: choose and the cellulosic bacterial strain activation of secreting bacteria can be prepared into seed mash, then seed mash is evenly dripped on the paraffin minus through sterilization treatment, add fermentation medium and cultivate 6 ~ 14d.
(3) tunning removes through paraffin, purification process and dehydration impregnation process obtain containing CaCl
2aqueous solution percentage by weight is the bacteria cellulose aquagel repair of cartilage support of 30 ~ 50%.By polyvinyl alcohol dissolution in Na
2hPO
4in aqueous solution, obtain mixed solution A.Adopt infusion process, make mixed solution A enter bacteria cellulose aquagel repair of cartilage internal stent, through freeze-thaw for several times, gamma-radiation radiation treatment obtains one and has bioactive composite aquogel tissue engineering bone/cartilage recovery support the product obtained.
As preferred technical scheme:
Wherein, a kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support as above, it is characterized in that: the three-dimensional porous structure of described design timbering material inside be every 5mm × 5mm processing plane on micropore in n × n array, micro-pore diameter is 200 ~ 500 μm, and micropore spacing is not less than 200 μm.The three-dimensional porous structure of computer modeling technique design, will finally affect the three-dimensional porous structure of tissue engineering bone/cartilage recovery support inside by paraffin minus, fermentation culture.In organizational project incubation, the aperture be applicable to and porosity, be beneficial to the propagation of chondrocyte and stick, and the infiltration of nutrient substance and the discharge of products of cellular metabolism.Therefore by regulating the aperture of three-dimensional porous quantity, position and micropore, aperture and the porosity of three-dimensional porous rack can be controlled when timbering material loose structure designs.
There is a preparation method for bioactive composite aquogel tissue engineering bone/cartilage recovery support as above, it is characterized in that: the described cellulosic bacterial strain of energy secreting bacteria is one or more in acetobacter xylinum, rhizobium, Sarcina, Rhodopseudomonas, achromobacter, Alcaligenes, Aerobacter or azotobacter.
A kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support as above, it is characterized in that: described bacterial strain/paraffin mixed culture fermentation method is: by bacterial strain concentration be 50 ~ 95% seed mash evenly drip on paraffin minus with dot matrix way, fermentation medium is added after 10 ~ 20min, fermentation culture temperature is 20 ~ 30 DEG C, and the time is 6 ~ 14d.
A kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support as above, it is characterized in that: described paraffin removes, purification treating method is: at the temperature of 50 ~ 70 DEG C, tunning soaks 5 ~ 10min respectively in distilled water, isopropyl alcohol and dehydrated alcohol, then at the temperature of 70 ~ 100 DEG C, the NaOH aqueous wash medium being 4 ~ 8% at percentage by weight washs 4 ~ 6h, more repeatedly rinses to neutrality with distilled water.With the tropina removed on deparaffnize, Bacterial cellulose and the residual media sticked on cellulose membrane.
There is a preparation method for bioactive composite aquogel tissue engineering bone/cartilage recovery support as above, it is characterized in that: described dehydration impregnation process method is that Bacterial cellulose is immersed in CaCl after high speed centrifugation, vacuum dehydration or press dewatering
2in aqueous solution, obtain through repeatedly dewatering, soaking containing CaCl
2aqueous solution percentage by weight is the bacteria cellulose aquagel of 30 ~ 50%.Described CaCl
2concentration of aqueous solution is 50 ~ 100mmol/L.Dehydration, immersion treatment can make CaCl on the basis not destroying bacteria cellulose aquagel three-dimensional net structure repeatedly
2aqueous solution enters Bacterial cellulose inside, and removes unnecessary hydrone.
There is a preparation method for bioactive composite aquogel tissue engineering bone/cartilage recovery support as above, it is characterized in that: described mixed solution A is that polyvinyl alcohol is dissolved in Na at 60 ~ 100 DEG C
2hPO
4the mixed solution obtained in aqueous solution, wherein polyvinyl alcohol molecule amount is 2 ~ 200,000, and alcoholysis degree is 87 ~ 99%, and the percentage by weight containing polyvinyl alcohol in mixed solution A is 10 ~ 25%; Na
2hPO
4the PO of aqueous solution
4ion concentration is CaCl
2in aqueous solution, the 0.5-0.7 of Ca ion concentration doubly, preferably 0.6 times.CaCl
2aqueous solution and Na
2hPO
4aqueous solution reacts to each other and can obtain hydroapatite particles, in this course Ca ion and PO
4the molar ratio of ion obtains hydroxyapatite [Ca
10(PO
4)
6(OH)
2] key.Polyvinyl alcohol-Na
2hPO
4aqueous solution enters containing CaCl
2the bacteria cellulose aquagel repair of cartilage internal stent of aqueous solution, due to Bacterial cellulose and polyvinyl alcohol molecule chain all having the deposition that a large amount of oh groups can promote Ca ion, cellulose nano-fibrous for template with Bacterial cellulose, CaCl
2with Na
2hPO
4solution reacts to each other formation hydroapatite particles.Simultaneously because the interaction of oh group and Ca ion on polyvinyl alcohol molecule chain facilitates the interaction between PVA strand, define the structure of similar " physical crosslinking point ", be conducive to the formation of PVA hydrogel; And hinder the increase of hydroapatite particles particle diameter, be conducive to the mechanical strength improving composite aquogel repair of cartilage support.
There is a preparation method for bioactive composite aquogel tissue engineering bone/cartilage recovery support as above, it is characterized in that: described freeze-thaw method is at-40 ~-10 DEG C, and freezing 6 ~ 24h, then at room temperature thaw 1 ~ 4h.The number of occurrence is 1 ~ 4 time.The freezing kinestate of the strand of PVA in Bacterial cellulose that makes " is freezed " to get off, and Interchain interaction and chain entanglement occur PVA strand around hydroapatite particles, are formed with sequence structure.After thawing, these compact ordered domains no longer separate, and become " entanglement point ".Have again new ordered domains to be formed when again freezing, these microcells are called " physical crosslinking point ".PVA molecular motion can be promoted, rearrange that be there is by the folding acquisition of strand the PVA hydrogel of crystallization/semicrystalline structure by freeze-thaw method.And the inierpeneirating network structure that PVA hydrogel network shape structure and BC hydrogel network structure are formed, hydroapatite particles is evenly embedded in inierpeneirating network structure inside, thus makes the physical property of composite aquogel and mechanical performance be greatly improved simultaneously.
Have a preparation method for bioactive composite aquogel tissue engineering bone/cartilage recovery support as above, it is characterized in that: described gamma-radiation radiation treatment, its irradiation dose is 15 ~ 60kGy.Under the effect of gamma-radiation radiation treatment, polyvinyl alcohol molecule interchain is crosslinked together by the free radical produced, the degree of cross linking increases with the increase of radiation dose, the crosslink density of hydrogel increases, in network structure, the size of micropore will reduce, thus make the reduction such as swelling ratio, water content of hydrogel, therefore the regulation and control to crosslink density can be realized by control radiation dose.Therefore the gamma-radiation irradiation energy adopted acts on PVA equably, and the crosslinking points of PVA is evenly distributed, and the degree of cross linking is easy to control, and can improve the crosslink density of PVA hydrogel further, with the mechanical property requirements making product reach human body cartilaginous tissue; And the sterilization that also can synchronously realize in radiation treatment process repair of cartilage support.
Compared with prior art, the invention has the beneficial effects as follows:
The present invention adopts computed tomography, three-dimensional fast shaping technology, bacterial strain/paraffin mixed culture fermentation technology and the preparation of hydroxyapatite is combined with freeze-thaw method, and preparation one has bioactive composite aquogel tissue engineering bone/cartilage recovery support.This tissue engineering bone/cartilage recovery support has the inierpeneirating network structure that PVA hydrogel network shape structure and BC hydrogel network structure are formed, and hydroapatite particles is evenly embedded in inierpeneirating network structure inside simultaneously; The repair of cartilage support obtained has the macro morphology and mechanical property that conform to normal articular cartilage, good biological activity, and desirable three-dimensional porous structure.Present invention process is simple, cost is low, pollution-free, the composite aquogel tissue engineering bone/cartilage recovery support of preparation can be used as tissue engineering bracket material and is applied to during the cartilaginous tissues such as articular cartilage reparation, meniscal repairs repair.
Detailed description of the invention
Below in conjunction with specific embodiment, set forth the present invention further.Should be understood that these embodiments are only not used in for illustration of the present invention to limit the scope of the invention.In addition should be understood that those skilled in the art can make various changes or modifications the present invention, and these equivalent form of values fall within the application's appended claims limited range equally after the content of having read the present invention's instruction.
Embodiment 1:
Adopt computed tomography to scan the cartilaginous tissue that expection is repaired, obtain the mathematical model of cartilaginous tissue three dimensional structure.Utilize mathematical model to design the three-dimensional porous structure of timbering material inside, the micropore in 9 × 9 arrays on the processing plane of every 5mm × 5mm, micro-pore diameter is 500 μm, and micropore spacing is not less than 200 μm.The repair of cartilage support designed obtains its minus by Boolean calculation, utilizes three-dimensional fast shaping technology, adopts three-dimensional rapid printer, take paraffin powder as moulding material, the paraffin minus of obtained repair of cartilage support.
Choose the cellulosic acetobacter xylinum of energy secreting bacteria, bacterial strain activation is prepared into seed mash, is that the seed mash of 50% evenly drips on paraffin minus with dot matrix way by bacterial strain concentration, after 10min, adds fermentation medium, fermentation culture temperature is 30 DEG C, and the time is 6d.Tunning, at 50 DEG C, soaks 10min respectively in distilled water, isopropyl alcohol and dehydrated alcohol.Then be in the NaOH aqueous solution of 4% at percentage by weight, at the temperature of 100 DEG C, heat 4h, more repeatedly rinse to neutrality with distilled water.By CaCl
2be dissolved in distilled water and be mixed with the CaCl that concentration is 50mmol/L
2aqueous solution.Adopt supercentrifugal process to carry out partial dehydration to Bacterial cellulose, and be immersed in CaCl
2in aqueous solution.Repeatedly dewater, soak after obtain containing CaCl
2aqueous solution percentage by weight is the bacteria cellulose aquagel repair of cartilage support of 50%.
At 60 DEG C, be 200,000 by molecular weight, alcoholysis degree be 87% polyvinyl alcohol be dissolved in the Na of 30mmol/L
2hPO
4in aqueous solution, obtaining containing polyvinyl alcohol weight percentage ratio is the mixed solution A of 10%, is then immersed in mixed solution A by bacteria cellulose aquagel.Complex after impregnation process is at-10 DEG C, and freezing 6h, at room temperature thaw 1h, and the number of occurrence is 4 times.Then adopt gamma-radiation radiation treatment, irradiation dose is 15kGy, obtains one and has bioactive composite aquogel tissue engineering bone/cartilage recovery support.
Embodiment 2:
Adopt computed tomography to scan the cartilaginous tissue that expection is repaired, obtain the mathematical model of cartilaginous tissue three dimensional structure.Utilize mathematical model to design the three-dimensional porous structure of timbering material inside, the micropore in 6 × 6 arrays on the processing plane of every 5mm × 5mm, micro-pore diameter is 200 μm, and micropore spacing is not less than 200 μm.The repair of cartilage support designed obtains its minus by Boolean calculation, utilizes three-dimensional fast shaping technology, adopts three-dimensional rapid printer, take paraffin powder as moulding material, the paraffin minus of obtained repair of cartilage support.
Choose the cellulosic rhizobium of energy secreting bacteria, bacterial strain activation is prepared into seed mash, is that the seed mash of 95% evenly drips on paraffin minus with dot matrix way by bacterial strain concentration, after 20min, adds fermentation medium, fermentation culture temperature is 20 DEG C, and the time is 14d.Tunning, at 70 DEG C, soaks 5min respectively in distilled water, isopropyl alcohol and dehydrated alcohol.Then be in the NaOH aqueous solution of 5% at percentage by weight, at the temperature of 90 DEG C, heat 5h, more repeatedly rinse to neutrality with distilled water.By CaCl
2be dissolved in distilled water and be mixed with the CaCl that concentration is 50mmol/L
2aqueous solution.Employing high speed centrifugation, method are carried out partial dehydration to Bacterial cellulose, and are immersed in CaCl
2in aqueous solution.Repeatedly dewater, soak after obtain containing CaCl
2aqueous solution percentage by weight is the bacteria cellulose aquagel repair of cartilage support of 50%.
At 60 DEG C, be 150,000 by molecular weight, alcoholysis degree be 88% polyvinyl alcohol be dissolved in the Na of 30mmol/L
2hPO
4in aqueous solution, obtaining containing polyvinyl alcohol weight percentage ratio is the mixed solution A of 25%, is then immersed in mixed solution A by bacteria cellulose aquagel.Complex after impregnation process is at-20 DEG C, and freezing 8h, at room temperature thaw 2h, and the number of occurrence is 4 times.Then adopt gamma-radiation radiation treatment, irradiation dose is 25kGy, obtains one and has bioactive composite aquogel tissue engineering bone/cartilage recovery support.
Embodiment 3:
Adopt computed tomography to scan the cartilaginous tissue that expection is repaired, obtain the mathematical model of cartilaginous tissue three dimensional structure.Utilize mathematical model to design the three-dimensional porous structure of timbering material inside, the micropore in 7 × 7 arrays on the processing plane of every 5mm × 5mm, micro-pore diameter is 250 μm, and micropore spacing is not less than 200 μm.The repair of cartilage support designed obtains its minus by Boolean calculation, utilizes three-dimensional fast shaping technology, adopts three-dimensional rapid printer, take paraffin powder as moulding material, the paraffin minus of obtained repair of cartilage support.
Choose the cellulosic Rhodopseudomonas of energy secreting bacteria, bacterial strain activation is prepared into seed mash, is that the seed mash of 65% evenly drips on paraffin minus with dot matrix way by bacterial strain concentration, after 10min, adds fermentation medium, fermentation culture temperature is 20 DEG C, and the time is 12d.Tunning, at 60 DEG C, soaks 10min respectively in distilled water, isopropyl alcohol and dehydrated alcohol.Then be in the NaOH aqueous solution of 6% at percentage by weight, at the temperature of 80 DEG C, heat 6h, more repeatedly rinse to neutrality with distilled water.By CaCl
2be dissolved in distilled water and be mixed with the CaCl that concentration is 100mmol/L
2aqueous solution.Adopt vacuum dehydration method to carry out partial dehydration to Bacterial cellulose, and be immersed in CaCl
2in aqueous solution.Repeatedly dewater, soak after obtain containing CaCl
2aqueous solution percentage by weight is the bacteria cellulose aquagel repair of cartilage support of 40%.
At 70 DEG C, be 120,000 by molecular weight, alcoholysis degree be 89% polyvinyl alcohol be dissolved in the Na of 60mmol/L
2hPO
4in aqueous solution, obtaining containing polyvinyl alcohol weight percentage ratio is the mixed solution A of 15%, is then immersed in mixed solution A by bacteria cellulose aquagel.Complex after impregnation process is at-30 DEG C, and freezing 10h, at room temperature thaw 3h, and the number of occurrence is 3 times.Then adopt gamma-radiation radiation treatment, irradiation dose is 35kGy, obtains one and has bioactive composite aquogel tissue engineering bone/cartilage recovery support.
Embodiment 4:
Adopt computed tomography to scan the cartilaginous tissue that expection is repaired, obtain the mathematical model of cartilaginous tissue three dimensional structure.Utilize mathematical model to design the three-dimensional porous structure of timbering material inside, the micropore in 8 × 8 arrays on the processing plane of every 5mm × 5mm, micro-pore diameter is 300 μm, and micropore spacing is not less than 200 μm.The repair of cartilage support designed obtains its minus by Boolean calculation, utilizes three-dimensional fast shaping technology, adopts three-dimensional rapid printer, take paraffin powder as moulding material, the paraffin minus of obtained repair of cartilage support.
Choose the cellulosic achromobacter of energy secreting bacteria, bacterial strain activation is prepared into seed mash, is that the seed mash of 75% evenly drips on paraffin minus with dot matrix way by bacterial strain concentration, after 20min, adds fermentation medium, fermentation culture temperature is 30 DEG C, and the time is 10d.Tunning, at 70 DEG C, soaks 5min respectively in distilled water, isopropyl alcohol and dehydrated alcohol.Then be in the NaOH aqueous solution of 7% at percentage by weight, at the temperature of 70 DEG C, heat 6h, more repeatedly rinse to neutrality with distilled water.By CaCl
2be dissolved in distilled water and be mixed with the CaCl that concentration is 100mmol/L
2aqueous solution.Adopt vacuum dehydration method to carry out partial dehydration to Bacterial cellulose, and be immersed in CaCl
2in aqueous solution.Repeatedly dewater, soak after obtain containing CaCl
2aqueous solution percentage by weight is the bacteria cellulose aquagel repair of cartilage support of 40%.
At 80 DEG C, be 100,000 by molecular weight, alcoholysis degree be 90% polyvinyl alcohol be dissolved in the Na of 60mmol/L
2hPO
4in aqueous solution, obtaining containing polyvinyl alcohol weight percentage ratio is the mixed solution A of 10%, is then immersed in mixed solution A by bacteria cellulose aquagel.Complex after impregnation process is at-40 DEG C, and freezing 12h, at room temperature thaw 4h, and the number of occurrence is 3 times.Then adopt gamma-radiation radiation treatment, irradiation dose is 45kGy, obtains one and has bioactive composite aquogel tissue engineering bone/cartilage recovery support.
Embodiment 5:
Adopt computed tomography to scan the cartilaginous tissue that expection is repaired, obtain the mathematical model of cartilaginous tissue three dimensional structure.Utilize mathematical model to design the three-dimensional porous structure of timbering material inside, the micropore in 8 × 8 arrays on the processing plane of every 5mm × 5mm, micro-pore diameter is 400 μm, and micropore spacing is not less than 200 μm.The repair of cartilage support designed obtains its minus by Boolean calculation, utilizes three-dimensional fast shaping technology, adopts three-dimensional rapid printer, take paraffin powder as moulding material, the paraffin minus of obtained repair of cartilage support.
Choose the cellulosic Alcaligenes of energy secreting bacteria, bacterial strain activation is prepared into seed mash, is that the seed mash of 85% evenly drips on paraffin minus with dot matrix way by bacterial strain concentration, after 15min, adds fermentation medium, fermentation culture temperature is 25 DEG C, and the time is 8d.Tunning, at 60 DEG C, soaks 10min respectively in distilled water, isopropyl alcohol and dehydrated alcohol.Then be in the NaOH aqueous solution of 8% at percentage by weight, at the temperature of 80 DEG C, heat 5h, more repeatedly rinse to neutrality with distilled water.By CaCl
2be dissolved in distilled water and be mixed with the CaCl that concentration is 75mmol/L
2aqueous solution.Adopt press dewatering method to carry out partial dehydration to Bacterial cellulose, and be immersed in CaCl
2in aqueous solution.Repeatedly dewater, soak after obtain containing CaCl
2aqueous solution percentage by weight is the bacteria cellulose aquagel repair of cartilage support of 30%.
At 100 DEG C, be 20,000 by molecular weight, alcoholysis degree be 99% polyvinyl alcohol be dissolved in the Na of 45mmol/L
2hPO
4in aqueous solution, obtaining containing polyvinyl alcohol weight percentage ratio is the mixed solution A of 25%, is then immersed in mixed solution A by bacteria cellulose aquagel.Complex after impregnation process is at-40 DEG C, and freezing 16h, at room temperature thaw 4h, and the number of occurrence is 2 times.Then adopt gamma-radiation radiation treatment, irradiation dose is 55kGy, obtains one and has bioactive composite aquogel tissue engineering bone/cartilage recovery support.
Embodiment 6
Adopt computed tomography to scan the cartilaginous tissue that expection is repaired, obtain the mathematical model of cartilaginous tissue three dimensional structure.Utilize mathematical model to design the three-dimensional porous structure of timbering material inside, the micropore in 7 × 7 arrays on the processing plane of every 5mm × 5mm, micro-pore diameter is 500 μm, and micropore spacing is not less than 200 μm.The repair of cartilage support designed obtains its minus by Boolean calculation, utilizes three-dimensional fast shaping technology, adopts three-dimensional rapid printer, take paraffin powder as moulding material, the paraffin minus of obtained repair of cartilage support.
Choose the cellulosic azotobacter of energy secreting bacteria, bacterial strain activation is prepared into seed mash, is that the seed mash of 95% evenly drips on paraffin minus with dot matrix way by bacterial strain concentration, after 20min, adds fermentation medium, fermentation culture temperature is 30 DEG C, and the time is 14d.Tunning, at 50 DEG C, soaks 10min respectively in distilled water, isopropyl alcohol and dehydrated alcohol.Then be in the NaOH aqueous solution of 6% at percentage by weight, at the temperature of 100 DEG C, heat 4h, more repeatedly rinse to neutrality with distilled water.By CaCl
2be dissolved in distilled water and be mixed with the CaCl that concentration is 75mmol/L
2aqueous solution.Adopt press dewatering method to carry out partial dehydration to Bacterial cellulose, and be immersed in CaCl
2in aqueous solution.Repeatedly dewater, soak after obtain containing CaCl
2aqueous solution percentage by weight is the bacteria cellulose aquagel repair of cartilage support of 30%.
At 100 DEG C, be 80,000 by molecular weight, alcoholysis degree be 95% polyvinyl alcohol be dissolved in the Na of 45mmol/L
2hPO
4in aqueous solution, obtaining containing polyvinyl alcohol weight percentage ratio is the mixed solution A of 15%, is then immersed in mixed solution A by bacteria cellulose aquagel.Complex after impregnation process is at-40 DEG C, and freezing 24h, at room temperature thaw 4h.Then adopt gamma-radiation radiation treatment, irradiation dose is 60kGy, obtains one and has bioactive composite aquogel tissue engineering bone/cartilage recovery support.
Claims (7)
1. there is a preparation method for bioactive composite aquogel tissue engineering bone/cartilage recovery support, comprising:
A () adopts computed tomography to scan the cartilaginous tissue that expection is repaired, obtain the mathematical model of cartilaginous tissue three dimensional structure, mathematical model is utilized to design the three-dimensional porous structure of timbering material inside, its minus is obtained by Boolean calculation, utilize three-dimensional fast shaping technology, adopting three-dimensional rapid printer, take paraffin powder as moulding material, the paraffin minus of obtained repair of cartilage support;
B () Bacterial cellulose ferments: choose and the cellulosic bacterial strain activation of secreting bacteria can be prepared into seed mash, then evenly dripped on the paraffin minus through sterilization treatment by seed mash, add fermentation medium and cultivate 6 ~ 14d;
C () tunning removes through paraffin, purification process and dehydration impregnation process obtain containing CaCl
2aqueous solution percentage by weight is the bacteria cellulose aquagel repair of cartilage support of 30 ~ 50%, by polyvinyl alcohol dissolution in Na
2hPO
4in aqueous solution, obtain mixed solution A, adopt infusion process, make mixed solution A enter bacteria cellulose aquagel repair of cartilage internal stent, through freeze-thaw for several times, gamma-radiation radiation treatment obtains one and has bioactive composite aquogel tissue engineering bone/cartilage recovery support the product obtained; Wherein said mixed solution A is that polyvinyl alcohol is dissolved in Na at 60 ~ 100 DEG C
2hPO
4the mixed solution obtained in aqueous solution, wherein polyvinyl alcohol molecule amount is 2 ~ 200,000, and alcoholysis degree is 87 ~ 99%, and the percentage by weight containing polyvinyl alcohol in mixed solution A is 10 ~ 25%; Wherein Na
2hPO
4the PO of aqueous solution
4ion concentration is CaCl
2in aqueous solution, the 0.5-0.7 of Ca ion concentration doubly.
2. a kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support as claimed in claim 1, is characterized in that: described can the cellulosic bacterial strain of secreting bacteria be one or more in acetobacter xylinum, rhizobium, Sarcina, Rhodopseudomonas, achromobacter, Alcaligenes, Aerobacter or azotobacter.
3. a kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support as claimed in claim 1, it is characterized in that: described paraffin removes, purification treating method is: at the temperature of 50 ~ 70 DEG C, tunning soaks 5 ~ 10min respectively in distilled water, isopropyl alcohol and dehydrated alcohol, then at the temperature of 70 ~ 100 DEG C, the NaOH aqueous wash medium being 4 ~ 8% at percentage by weight washs 4 ~ 6h, more repeatedly rinses to neutrality with distilled water.
4. a kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support as claimed in claim 1, it is characterized in that: described dehydration impregnation process method is that Bacterial cellulose is immersed in CaCl after high speed centrifugation, vacuum dehydration or press dewatering
2in aqueous solution, obtain through repeatedly dewatering, soaking containing CaCl
2aqueous solution percentage by weight is the bacteria cellulose aquagel of 30 ~ 50%, described CaCl
2concentration of aqueous solution is 50 ~ 100mmol/L.
5. a kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support as claimed in claim 1, is characterized in that:
Described mixed solution A is that polyvinyl alcohol is dissolved in Na at 60 ~ 100 DEG C
2hPO
4the mixed solution obtained in aqueous solution, wherein polyvinyl alcohol molecule amount is 2 ~ 200,000, and alcoholysis degree is 87 ~ 99%, and the percentage by weight containing polyvinyl alcohol in mixed solution A is 10 ~ 25%; Na
2hPO
4the PO of aqueous solution
4ion concentration is CaCl
20.6 times of Ca ion concentration in aqueous solution.
6. a kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support as claimed in claim 1, it is characterized in that: described freeze-thaw method is at-40 ~-10 DEG C, freezing 6 ~ 24h, then at room temperature thaw 1 ~ 4h, and the number of occurrence is 1 ~ 4 time.
7. a kind of preparation method with bioactive composite aquogel tissue engineering bone/cartilage recovery support as claimed in claim 1, it is characterized in that: described gamma-radiation radiation treatment, its irradiation dose is 15 ~ 60kGy.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310110593.2A CN103143058B (en) | 2013-04-01 | 2013-04-01 | Preparation of composite hydrogel tissue engineer cartilage repairing scaffold with biological activity |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310110593.2A CN103143058B (en) | 2013-04-01 | 2013-04-01 | Preparation of composite hydrogel tissue engineer cartilage repairing scaffold with biological activity |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103143058A CN103143058A (en) | 2013-06-12 |
| CN103143058B true CN103143058B (en) | 2015-03-25 |
Family
ID=48541556
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310110593.2A Active CN103143058B (en) | 2013-04-01 | 2013-04-01 | Preparation of composite hydrogel tissue engineer cartilage repairing scaffold with biological activity |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103143058B (en) |
Families Citing this family (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103893818B (en) * | 2014-03-14 | 2015-07-01 | 华南理工大学 | Osteochondral three-dimensional stent with regular interpenetrating network structure and preparation method thereof |
| CN106492286B (en) * | 2016-09-19 | 2019-04-16 | 盐城工业职业技术学院 | A kind of fibroin/bacteria cellulose composite hydrogel and its preparation method and application |
| FI128467B (en) * | 2016-12-30 | 2020-05-29 | Teknologian Tutkimuskeskus Vtt Oy | Three dimensional printing with biomaterial |
| CN107670114A (en) * | 2017-10-07 | 2018-02-09 | 北京林业大学 | A kind of medical bio composite based on the enhancing of Eucalyptus nano micro crystal cellulose and preparation method thereof |
| CN113321821A (en) * | 2021-05-31 | 2021-08-31 | 南方医科大学 | Transparent ion-conductive hydrogel and preparation method and application thereof |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US8110222B2 (en) * | 2002-11-15 | 2012-02-07 | Ut-Battelle, Llc. | Composite material |
| CN102525688A (en) * | 2011-12-08 | 2012-07-04 | 浙江工业大学 | Manufacture method of tissue engineering bracket with both internal microstructure and individualized appearance |
| CN102961784B (en) * | 2012-11-29 | 2014-09-10 | 华南理工大学 | BC (Bacterial Cellulose)/PVA (Polyvinyl Alcohol) composite material, as well as preparation method and application thereof |
-
2013
- 2013-04-01 CN CN201310110593.2A patent/CN103143058B/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN103143058A (en) | 2013-06-12 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN101461963B (en) | Multiplex composite bone tissue engineering bracket material capable of degrading gradiently and preparation method thereof | |
| CN103877617B (en) | Two cross-linked hydrogel of injectable fibroin protein-alginate and preparation method thereof and using method | |
| CN101302486B (en) | Acetobacter xylinum and method for preparing nano-cellulose skin tissue repair material by using the same | |
| CN102430155B (en) | Cellular silk fibroin porous scaffold, and preparation method thereof | |
| CN103143058B (en) | Preparation of composite hydrogel tissue engineer cartilage repairing scaffold with biological activity | |
| CN102058902B (en) | Method for preparing mesh-shaped bionic bone porous stent material | |
| CN103044693A (en) | Preparation method for bacterial cellulose/polyvinyl alcohol composite hydrogel | |
| CN107213529B (en) | Preparation method of degradable medical high-molecular three-dimensional material for improving adhesion and osteogenic performance of osteoblasts | |
| CN101274108B (en) | Compound porous bracket and method of producing the same | |
| CN107349475B (en) | The artificial organ engineering skin and preparation method thereof that nano fibrous membrane is layering with stem cell | |
| CN104117098A (en) | Magnetic hydroxyl apatite/polymer tridimensional porous stent material with oriented magnetic field and preparation method thereof | |
| CN101264341A (en) | Three-dimensional porous tissue engineering bracket material, preparation and application thereof | |
| CN105664260A (en) | Method for preparing bone tissue engineering three-dimensional porous support based on graphene/silk fibroin | |
| CN103893825A (en) | Method for preparing bacterial cellulose compounded amnion extracellular matrix material containing collagen | |
| CN107137763A (en) | A kind of study of vascularized tissue engineering bone and preparation method thereof | |
| CN103159961B (en) | Preparation method of mineralized bacterial cellulose/polyvinyl alcohol composite aquagel cartilage repairing material | |
| CN101176798B (en) | Complex stephanoporate bracket of calcium sulphate and freeze drying bone as well as preparation method thereof | |
| CN103272283B (en) | Mineralized bacterial cellulose three-dimensional porous bone tissue restoration scaffold preparation method | |
| Nemati et al. | Nano bacterial cellulose for biomedical applications: A mini review focus on tissue engineering | |
| Salihu et al. | Overview of inexpensive production routes of bacterial cellulose and its applications in biomedical engineering | |
| CN101856516B (en) | Preparation of collagen-chitosan-laser micropore dermal matrix composite membranes | |
| CN101496915B (en) | Heterogeneous dermis reticular layer stent without basement membrane and cell as well as preparation method thereof | |
| CN111249523B (en) | Bone-like composite material support and preparation method thereof | |
| CN103691002A (en) | Bacterial cellulose/collagen/hydroxyapatite composite material, and preparation and application thereof | |
| CN111467570A (en) | MicroRNA-loaded tissue engineering scaffold and preparation method thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant |