LT6309B - Porous three dimensional cellulose based scaffold and method - Google Patents
Porous three dimensional cellulose based scaffold and method Download PDFInfo
- Publication number
- LT6309B LT6309B LT2014117A LT2014117A LT6309B LT 6309 B LT6309 B LT 6309B LT 2014117 A LT2014117 A LT 2014117A LT 2014117 A LT2014117 A LT 2014117A LT 6309 B LT6309 B LT 6309B
- Authority
- LT
- Lithuania
- Prior art keywords
- bone
- cellulose
- particles
- dimensional
- gel
- Prior art date
Links
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/40—Composite materials, i.e. containing one material dispersed in a matrix of the same or different material
- A61L27/44—Composite materials, i.e. containing one material dispersed in a matrix of the same or different material having a macromolecular matrix
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/3604—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix characterised by the human or animal origin of the biological material, e.g. hair, fascia, fish scales, silk, shellac, pericardium, pleura, renal tissue, amniotic membrane, parenchymal tissue, fetal tissue, muscle tissue, fat tissue, enamel
- A61L27/3608—Bone, e.g. demineralised bone matrix [DBM], bone powder
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
- A61L27/56—Porous materials, e.g. foams or sponges
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2430/00—Materials or treatment for tissue regeneration
- A61L2430/02—Materials or treatment for tissue regeneration for reconstruction of bones; weight-bearing implants
Abstract
Description
Išradimas priklauso medžiagų inžinerijos sričiai ir gali būti panaudotas odontologijoje ar kitos srities chirurgijoje kaulo audinio regeneracijai defekto vietoje.The invention relates to the field of material engineering and can be used in dentistry or other surgery for bone tissue regeneration at the site of the defect.
Trimatis porėtas celiuliozės karkasas, pasižymintis biosuderinamumu, geromis osteokondukcinėmis ir mechaninėmis savybėmis, gaunamas sudarant kompo zitą iš gamtinio polimero - celiuliozės ir biogeninių kalcio šaltinių, būtent, autogeninio (nuosavo) kaulo, alogeninio (kito tos pačios rūšies, t.y. žmogaus, kito individo) kaulo, ksenogeninio (kitos rūšies organizmo) kaulo dalelių. Kompozito struktūra, tinkama kaulo audinio regeneracijai, formuojama liofilizacijos būdu ar veikiant superkriziniu skysčiu, prieš tai inkliudavus vandeniniais alkoholio tirpalais. Gali būti blokelio ar granulių pavidalo, gali būti pagaminta pasta.Three-dimensional porous cellulose backbone with biocompatibility, good osteoconductive and mechanical properties, obtained by composite of natural polymer - cellulose and biogenic sources of calcium, namely, autogenous (native) bone, allogeneic (other human, other species) bone , xenogeneic (other species of bone) bone particles. The structure of the composite, suitable for bone tissue regeneration, is formed by lyophilization or by the action of a supercritical fluid, preincubated with aqueous alcoholic solutions. May be in the form of blocks or granules, or a paste may be produced.
Kaulo audinio atstatymui defekto srityje odontologai naudoja kaulo pakaitalus. Tai gali būti:Bone substitutes are used by dentists to repair bone tissue in the defect area. That might be:
- autogeniniai pakaitalai - nuosavas kaulas iš kitos to paties žmogaus vietos. Privalumai - neiššaukia imuninės sistemos atsako. Trūkumai - reikalinga papildoma chirurginė operacija kaulo išėmimui, neretai trūksta kaulo, be to sunku jam suteikti reikiamą formą;- autogenous substitutes - own bone from another site of the same person. Benefits - Does not trigger the immune system to respond. Disadvantages - extra surgery is needed to remove the bone, often bone is missing and it is difficult to give it the required shape;
- alogeniniai kaulo pakaitalai - kito tos pačios rūšies (t.y. žmogaus) individo kaulas. Pagal fizikines savybes tinkamas, bet galima pernešti kito žmogaus ligų užkratus;- allogeneic bone substitutes - bone of another individual of the same species (i.e. human). Physically suitable but capable of transmitting infectious diseases to another person;
- ksenogeniniai kaulo pakaitalai - tai gyvūno, dažniausiai jauno jautuko kaulas, kuriame išdeginti baltymai ir likusi tik mineralinė dalis. Jo morfologija ne visai atitinka žmogaus kaulo struktūrai. Jis pernelyg greitai rezorbuojasi organizme;- Xenogeneic bone substitutes are the bone of an animal, usually a young bull, in which the protein and only the mineral remains. Its morphology does not quite match the structure of human bone. It is absorbed too quickly by the body;
- sintetiniai kaulo pakaitalai. Tarpe jų, populiariausi - trikalcio fosfatas (TCP) ir hidroksiapatitas (HA).- Synthetic bone substitutes. Among them, tricalcium phosphate (TCP) and hydroxyapatite (HA) are the most popular.
Dauguma kaulo pakaitalų yra miltelių ar granulių (0,5-2 mm) pavidalo.Most bone substitutes are in the form of powder or granules (0.5 to 2 mm).
Pagrindinis jų trūkumas - jie dažnai sušoka į monolitinį gabalą, kuriame neįauga kraujagyslės. Susidaro nepakankamo stiprumo darinys, kuris trupa įsukant metalinį implantą,Their main disadvantage is that they often jump into a monolithic, non-vascularized piece. This results in an inadequate structure that crumbles when the metal implant is screwed in,
Siekiant gauti porėtą trimatę struktūrą, įvairios formos kaulo pakaitalų dalelės sumaišomos ir iš jų gaminami agregatai, įvairiais būdais suklijuojant, supresuojant ar sukepant daleles. Tam naudojamos įvairios kaulinės dalelės, TCP, HA (VVinterbottomIn order to obtain a porous three-dimensional structure, bone substitute particles of various shapes are mixed and assembled from these by various means of adhesion, compression or sintering. Various bony particles, TCP, HA (Winterbottom
J.M. et ai. Implant, method of making šame and ūse of the implant for the treatment of bone defects. Patent No. US 6478825 B 1, 2002-11-02) arba šios mineralinės medžiagos kartu su polimero dalelėmis (Giomo T. PLGA/HA hydroxyapatite composite bone grafts and method of making. Pub. No. US 2013/02 18291 Al, 201308-22). Trimatės struktūros kompozitai taip pat sudaromi elektrinio verpimo metodu (Mei Wei, Fei Peng, Zhi-kang Xu. Electrospun apatite/polymer nano-composite scaffolds. Patent No. US7879093 B2, 2011-02-01) arba naudojant sparčios prototipų gamybos technologijas (angį. Rapid Prototyping) (Sawkins, M.J., et ai. 3D Cell and scaffold patteming strategies in tissue engineering. Recent Patente on Biomedical Engineering, 2013, vol. 6, no. 1, p. 3-21; Teoh, S.H., et ai. Three-dimensional bioresorbable scaffolds for tissue engineering applications. Patent No. US8071007 B 1, 2011-12-06). Naudojami sintetiniai polimerai: poli( s-kaprolaktonas), poli(L-pieno rūgštis), poli(glikolio rūgštis), poli(D,L-pieno-ko-glikolio rūgštis), polifosfazenai, polipropilenfumaratas (Muhammad, I.S.; Xiaoxue, X.; Li, L. A review on biodegradable polymeric materials for bone tissue engineering applications. Journal of Materials Science, 2009, p. 5713-5724). Iš sintetinių polimerų plačiausiai naudojamas polifs-kaprolaktonas) (Dhandayuthapani, B., et ai. Polymeric scaffolds in tissue engineering application: A review. International Journal of Polymer Science, 2011,vol.2011,p.l-19).J.M. et al. Implant, a method of making this and a mustache for the treatment of bone defects. Patent No. U.S. Pat. No. 6,478,825 B 1, 02-11-2002) or these minerals together with polymer particles (Giomo T. PLGA / HA hydroxyapatite composite bone grafts and method of making. . Three-dimensional composites are also made by electric spinning (Mei Wei, Fei Peng, Zhi-kang Xu. Electrospun apatite / polymer nano-composite scaffolds. Patent No. US7879093 B2, 2011-02-01) or by rapid prototype production technology (carbon). (Rapid Prototyping) (Sawkins, MJ, et al., 3D Cell and Scaffold Patching Strategies in Tissue Engineering. Recent Patent on Biomedical Engineering, 2013, Vol. 6, No. 1, pp. 3-21; Teoh, SH, et al. . Three-Dimensional Bioresorbable Scaffolds for Tissue Engineering Applications. Patent No. US8071007 B 1, 2011-12-06). Synthetic polymers used: poly (s-caprolactone), poly (L-lactic acid), poly (glycolic acid), poly (D, L-lactic-co-glycolic acid), polyphosphazenes, polypropylene fumarate (Muhammad, IS; Xiaoxue, X) .; Li, L. A Review of Biodegradable Polymeric Materials for Bone Tissue Engineering Applications (Journal of Materials Science, 2009, pp. 5713-5724). Of the synthetic polymers, the most widely used is polyphaprolactone) (Dhandayuthapani, B., et al., Polymeric scaffolds in tissue engineering application: A review. International Journal of Polymer Science, 2011, pp.19-19).
Tačiau sintetiniai polimerai pasižymi mažesniu biosuderinamumu nei natūralūs, jie neretai sukelia audinių nekrozę, jų skilimo produktai, kaip kad glikolio rūgštis ir kiti rūgštiniai junginiai gali padidinti vietinį rūgštingumą, kas gali sukelti audinių pažeidimus. Be to, polimerams yrant organizme gali susidaryti ir toksiški metabolitai. Aprašyti gavimo būdai sudėtingi ir neužtikrina reikiamo porėtumo, nes dalelės yra sulipdomos arba sukepinamos. Be to, svarbus trūkumas yra tai, kad kalcio šaltiniais naudojami sintetinės kilmės mineraliniai junginiai - TCP ir HA.However, synthetic polymers have less biocompatibility than natural polymers, often causing tissue necrosis, and their degradation products, such as glycolic acid and other acidic compounds, can increase local acidity, which can cause tissue damage. In addition, toxic metabolites may also be formed in the body through degradation of polymers. The described methods of preparation are complex and do not provide the required porosity because the particles are adherent or sintered. In addition, an important disadvantage is that calcium sources use synthetic mineral compounds TCP and HA.
Artimiausias prototipas yra celiuliozės pagrindu gaunamas trimatis karkasas, kurio paviršius mineralizuojamas modeliniame kūno skystyje (SBF -angį. simulated body jluid) (Petrauskaite, o. et.al. Biomimetic mineralization on a macroporous cellulose-based matrix for bone regeneration. BioMed Research International. ISSN 2314-6133. 2013, vol. 2013, p. 1-9). Šiuo atveju porėtas karkasas yra gaunamas iš regeneruotos celiuliozės gelio liofilizacijos būdu. Gaunamas tolygiai porėtas blokas, pasižymintis morfologija, atitinkančia natūralaus kaulo struktūrą.The closest prototype is a cellulose-based three-dimensional scaffold whose surface is mineralized in a simulated body fluid (SBF) (Petrauskaite, et al.). Biomimetic mineralization on a macroporous cellulose-based matrix for bone regeneration. BioMed Research International. ISSN 2314-6133. 2013, vol. 2013, pp. 1-9). In this case, the porous backbone is derived from regenerated cellulose gel by lyophilization. A uniformly porous block with a morphology corresponding to the structure of the natural bone is obtained.
Prie esminių metodo trūkumų galima priskirti tai, kad celiuliozė nepasižymi bioaktyvumu, nesuauga su kauliniais audiniais, pasižymi prastomis osteokondukcinėmis savybėmis. Nors šių savybių pagerinimui karkaso paviršius mineralizuojamas SBF tirpale, tačiau gaunamas padengimas sintetiniais mineralais, kurie visada pasižymi mažesniu biosuderinamumu nei natūralūs. Be to, mineralizuojant SBF-e, padengiamas tik paviršius, ir mineralų kiekis kompozituose sudaro tik iki 12 procentų nuo celiuliozės svorio, todėl tokio karkaso osteokondukcinės savybės pagerėja nežymiai. Be to, gautas karkasas pasižymi prastomis mechaninėmis savybėmis (Jungo modulis 4 MPa).One of the main drawbacks of the method is that cellulose is not bioactivated, does not grow with bone tissue, and has poor osteoconductive properties. Although the surface of the skeleton is mineralized in SBF solution to improve these properties, it is coated with synthetic minerals, which always have lower biocompatibility than natural ones. In addition, SBF-e mineralization only covers the surface, and the content of minerals in the composites is up to 12 percent by weight of the cellulose, resulting in a slight improvement in the osteoconductive properties of such a scaffold. In addition, the resulting skeleton exhibits poor mechanical properties (Jungian modulus 4 MPa).
Šio išradimo tikslas - gauti trimatį porėtą celiuliozės karkasą, kuris pasi žymėtų biosuderinamumu ir geromis osteokondukcinėmis bei mechaninėmis savybėmis ir struktūra, atitinkančią natūralaus kaulo morfologiją.It is an object of the present invention to provide a three-dimensional porous cellulose backbone which is characterized by biocompatibility and good osteoconductive and mechanical properties and structure consistent with natural bone morphology.
Šis tikslas pasiekiamas sudarant celiuliozės kompozitą su biogeninių kaulų 0,01-2000 mikronų dydžio dalelėmis, esant masių santykiui 1: 0,12-6,0. Atskiru atveju celiuliozės kompo zitas sudaromas su sepijos kaulo, kuriame yra mineralinių junginių ir polisacharidas chitinas, 0,01-2000 mikronų dydžio dalelėmis. Atskiru atveju karkaso porų paviršius padengiamas kolagenu, trombocitais praturtintu fibrinu, įvairiais augimo faktoriais, terapeutiniais priedais, kamieninėmis ląstelėmis. Trimatės porėtos struktūros, atitinkančios kaulo morfologiją, karkasas gaunamas celiuliozės geli formuojant iš acetilceliuliozės tirpalo su įterptomis biogeninio kaulo dalelėmis masių santykiu 1: 0,12-6,0, po to jį inkliuduojant 10-30 % vandeniniu alkoholio tirpalu ir liofilizuojant arba veikiant gelį superkriziniu anglies dioksidu 24-70 MPa slėgyje 3580°C temperatūroje. Gautas blokelio formos karkasas gali būti sumalamas iki norimo dydžio granulių ir/ar iš jų pagaminama pasta.This object is achieved by forming a cellulose composite with a particle size of 0.01 to 2000 microns in biogenic bone at a weight ratio of 1: 0.12 to 6.0. In an individual case, the cellulose composite is formed from 0.01 to 2000 micron particles of sepia bone containing mineral compounds and polysaccharide chitin. On a case-by-case basis, the surface of skeletal pores is coated with collagen, platelet-enriched fibrin, various growth factors, therapeutic additives, and stem cells. The three-dimensional porous structure, corresponding to bone morphology, is obtained by forming a cellulose gel with a 1: 0.12-6.0 weight ratio of acetylcellulose solution with biogenic bone particles embedded in it, and then lyophilizing or gel supercritical lyophilization. carbon dioxide at a pressure of 24-70 MPa at 3580 ° C. The resulting block-shaped frame can be ground to a desired size of pellets and / or a paste made from them.
Gaunamo karkaso privalumai - naudojami ne sintetiniai, o natūralūs komponentai: gamtinis polimeras - celiuliozė ir smulkinti biogeniniai kaulai. Celiuliozė yra necitotoksiška, pasižymi biosuderinamumu. Jos irimo produktai nėra toksiški. Biogeniniais kaulais gali būti naudojami autogeninis, alogeninis ar ksenogeninis (bet kurio gyvūno) kaulas. Atskiru atveju gali būti naudojamas smulkintas sepijos kaulas, kuriame be mineralinių junginių yra ir polisacharidas chitinas, pasižymintis priešuždegiminėmis, antimikrobinėmis ir koaguliacinėmis savybėmis, kas suteikia papildomus privalumus. Celiuliozės ir smulkintų kaulų dalelių masių santykis kompozitel: 0,12-6,0.The benefits of the resulting scaffold are the use of natural, non-synthetic components: natural polymer - cellulose and comminuted biogenic bones. Cellulose is non-cytotoxic and has biocompatibility. Its degradation products are non-toxic. Autogenous, allogeneic, or xenogeneic (of any animal) bone can be used as biogenic bone. On a case-by-case basis, shredded sepia bone can be used, which, in addition to mineral compounds, also contains polysaccharide chitin, which has anti-inflammatory, antimicrobial and coagulant properties, which provides additional benefits. Cellulose to crushed bone particle weight ratio in composite: 0.12-6.0.
Išradimo rezultatus iliustruoja brėžiniai. Fig. 1 - 1-ame pavyzdyje pagaminto karkaso skerspjūvio vaizdas, gautas mikrokompiuterinės tomografijos metodu. Fig. 2 - 1-ame pavyzdyje pagaminto karkaso trimatis vaizdas, gautas mikrokompiuterinės tomografijos metodu. Fig. 3 - ll-ame pavyzdyje pagaminto karkaso skerspjūvio vaizdas, gautas mikrokompiuterinės tomografijos metodu. Fig. 4 - ll-ame pavyzdyje pagaminto karkaso trimatis vaizdas, gautas mikrokompiuterinės tomografijos metodu. Fig. 5 - ląstelių proliferacija karkasuose.The results of the invention are illustrated in the drawings. FIG. 1 - 1 is a cross-sectional view of a scaffold fabricated by microcomputer tomography. FIG. 2 is a 3D view of a scaffold fabricated in Example 1 obtained by microcomputer tomography. FIG. Fig. 3 is a cross-sectional view of a skeleton fabricated in Example III obtained by microcomputer tomography. FIG. - A three-dimensional image of the skeleton produced in Example 4L obtained by microcomputer tomography. FIG. 5 - Cell proliferation in scaffolds.
Būdo atlikimas.Performance of the way.
Smulkinti kaulai įvedami į acetilceliuliozės tirpalą, iš kurio gaunamas regeneruotos celiuliozės gelis su įterptomis kaulų dalelėmis. Smulkintų kaulų dalelių dydis 0,01-2000 mikronų, optimalus dydis - iki 200 mikronų. Porėta struktūra, atitinkanti kaulo morfologiją, suformuojama celiuliozės gelį su įterptomis biogeninių kaulo dalelėmis liofiiizuojant arba veikiant gelį superkriziniu anglies dioksidu 24-70 MPa slėgyje 35-80 - °C temperatūroje, prieš tai jį inkliudavus 10-30 % vandeniniais alkoholio tirpalais. Galima naudoti įvairius vandenyje tirpius alkoholius, tačiau tinkamiausias yra etanolis. Kompozitas pasižymi geromis mechaninėmis savybėmis Jungo modulis sudaro ne mažiau 8 MPa. Kompozito morfologija (porėta struktūra) pilnai atitinka žmogaus kaulo morfologijai (lentelė). Poros savo dydžiu tinkamos vaskuliarizacijai (kraujagyslių tinklui susidaryti) ir ląstelių proliferacijai. Bandymais su pelėmis nustatyta, kad kompozite po 2 savaičių buvo išplitęs kraujagyslių tinklas. Poros tarpusavyje susisiekiančios, todėl užtikrina maistinių medžiagų ir metabolitų transportą.The broken bones are injected into a solution of acetylcellulose, which produces a regenerated cellulose gel with embedded bone particles. The particle size of the crushed bone is 0.01-2000 microns, the optimum size is up to 200 microns. The porous structure, corresponding to bone morphology, is formed by lyophilizing a cellulose gel with embedded biogenic bone particles or by exposure to supercritical carbon dioxide at a pressure of 24-70 MPa at 35-80 ° C prior to incubation with 10-30% aqueous alcoholic solutions. A variety of water-soluble alcohols can be used, but ethanol is preferred. The composite has good mechanical properties. The Jungian modulus is at least 8 MPa. The composite morphology (porous structure) is fully consistent with human bone morphology (Table). The pores are suitable in size for vascularization (formation of the vascular network) and cell proliferation. In mice, the composite was found to have a dilated vascular network after 2 weeks. Couples communicate with each other and thus transport nutrients and metabolites.
Lentelė. Pagamintų kompozitų ir natūralaus kaulo struktūriniai parametraiTable. Structural parameters of manufactured composites and natural bone
Tiriant žmogaus osteoblastų linijos ląstelių MG-63 proliferaciją, nustatyta, kad pagaminti karkasai pasižymi geresnėmis osteokondukcinėmis savybėmis nei prototipe gautas karkasas (Fig. 5).When investigating the proliferation of human osteoblast lineage cell MG-63, it was found that the produced scaffolds have better osteoconductive properties than the prototype-derived scaffold (Fig. (Fig.5). 5).
Osteokondukcinių savybių ivertinimo metodas. Tyrimui naudojamos žmogaus osteoblastų linijos ląstelės MG-63 (ATCCVV CRL_1427™) (American Type Collection Culture, JAV). Išaugintos ląstelės sėjamos ant mėginių prieš tai juos sterilizavus UV jonizuojančia spinduliuote, švitinant 24 vai. Mėginiai, mirkomi 1 vai. mitybinėje terpėje, patalpinami j 24 duobučių plokšteles (vienas mėginys vienoje plokštelėje) ir supilstoma ant jų ląstelių suspensija. Karkasai su pasėtomis ląstelėmis laikomi termostate 37 C temperatūroje esant 5 % CO2. Tiriamos medžiagos poveikis ląstelių augimui vertinamas, nustačius DNR kiekį po 1, 3 ir 7 dienų. DNR kiekis nustatomas, naudojant fluorochromą Quant-iTP PicoGreen® (Life Technologies, JAV) pagal gamintojo nurodymus.Method for evaluation of osteoconductive properties. Human osteoblast line MG-63 cells (ATCCVV CRL_1427 ™) (American Type Collection Culture, USA) were used for the assay. The cultured cells are seeded on samples after having been sterilized by UV irradiation for 24 hours. Samples soaked for 1 hour. in culture medium, place them in 24-well plates (one sample per plate) and place the cell suspension thereon. Frames with seeded cells are stored in a thermostat at 37 ° C and 5% CO 2 . The effect of the test substance on cell growth is assessed by measuring the amount of DNA at 1, 3 and 7 days. DNA content was determined using the fluorochrome Quant-iTP PicoGreen® (Life Technologies, USA) according to the manufacturer's instructions.
Karkaso porų paviršius gali būti padengtas kolagenu, trombocitais praturtintu fibrinu, įvairiais augimo faktoriais, terapeutiniais priedais, kamieninėmis ląstelėmis. Karkasas gaunamas blokelio formos, iš kurio galima lengvai išpjauti norimo pavidalo implantus, gali būti sum altas iki norimo dydžio granulių, gali būti suformuota pasta.The surface of skeletal pores can be coated with collagen, platelet-enriched fibrin, various growth factors, therapeutic additives, stem cells. The frame is obtained in the form of a block from which implants of the desired shape can be easily cut, granules of the desired size can be milled, and a paste can be formed.
Pasta gaminama sumaišant sumaltą kompozitą su gliceroliu, polietilenglikoliu (molekulinė masė 400-600) ar kitu hidrogeliu.The paste is made by mixing the ground composite with glycerol, polyethylene glycol (molecular weight 400-600) or other hydrogel.
pavyzdys g acetilceliuliozės ištirpina 261 ml acetono-amoniako tirpalo (tūrių santykis lygus 1 :0,45), prideda 15 galogeninio kaulo granulių (granulių dydis neviršija 200 mikronų), gerai išmaišo. Gautą dispersiją išpūsto į norimo tūrio ir formos indus ir išlaiko, kol susiformuoja kietas gelis. Gautą gelį kruopščiai išplauna distiliuotu vandeniu. Po to, 24 valandas palaiko 25 % etanolio tirpale ir liofilizuoja. Kompozito Jungo modulis sudaro 8 MPa. Mėginio skerspjūvio vaizdas, gautas mikrokompiuterinės tomografijos metodu, pateiktas Fig. 1, o trimatis vaizdas - Fig. 2. Ląstelių proliferacijos karkase duomenys pateikti Fig. 5.sample g dissolved in 261 ml of acetone-ammonia solution (volume ratio: 1: 0.45), add 15 gallogenic bone beads (granule size not exceeding 200 microns), mix well. The resulting dispersion is blown into vessels of desired volume and shape and retained until a solid gel is formed. The resulting gel is thoroughly washed with distilled water. Subsequently, it is kept in 25% ethanol solution for 24 hours and lyophilized. The Jungian modulus of the composite is 8 MPa. A cross-sectional view of the sample obtained by microcomputer tomography is shown in Figs. 1, and the three-dimensional view of Figs. 2. Cell proliferation scaffold data are presented in Figs. 5.
pavyzdys g acetilceliuliozės ištirpina 267 ml acetono-amoniako tirpalo (tūrių santykis lygus 1 :0,5), prideda 12 g susmulkinto (dalelių dydis neviršija 100 mikronų) sepijos kaulo, gerai išmaišo. Gautą dispersiją išpūsto į norimo tūrio ir formos indus ir išlaiko, kol susiformuoja kietas gelis. Gautą geli kruopščiai išplauna distiliuotu vandeniu. Po to, 24 valandas palaiko 20 % etanolio tirpale ir liofilizuoja.Sample 1 g of acetylcellulose dissolves in 267 ml of acetone-ammonia solution (volume ratio: 1: 0.5), adds 12 g of crushed (particle size up to 100 microns) sepia bone, mix well. The resulting dispersion is blown into vessels of desired volume and shape and retained until a solid gel is formed. The resulting gel is thoroughly washed with distilled water. Subsequently, it is kept in 20% ethanol solution for 24 hours and lyophilized.
Iš susmulkinto sepijos kaulo, prieš jį naudojant, išplauna baltymus. Tam sepijos kaulo miltelius užpila 0,5 M natrio šarmo tirpalu ir maišo 5 vai. 80 °C temperatūroje. Po to kruopščiai išplauna distiliuotu vandeniu, išdžiovina ir persijoja.Protein is washed from the crushed bone of cuttlefish before being used. For this purpose, the sepia bone powder is filled with 0.5 M sodium hydroxide solution and stirred for 5 hours. At 80 ° C. It is then thoroughly washed with distilled water, dried and sieved.
Kompozito Jungo modulis sudaro 10 MPa. Mėginio skerspjūvio vaizdas, gautas mikrokompiuterinės tomografijos metodu, pateiktas Fig. 3, o trimatis vaizdas Fig. 4. Ląstelių proliferacijos karkase duomenys pateikti Fig. 5.The Jungian modulus of the composite is 10 MPa. A cross-sectional view of the sample obtained by microcomputer tomography is shown in Figs. 3, and the three-dimensional view of FIG. 4. Cell proliferation scaffold data are presented in Figs. 5.
pavyzdys g acetilceliuliozės ištirpina 10 1,5 ml acetono-amoniako tirpalo (tūrių santykis lygus 1 :0,45), prideda 12 g smulkinto ksenogeninio kaulo (jautuko) granulių (granulių dydis neviršija 200 mikronų), gerai išmaišo. Gautą dispersiją išpūsto į norimo tūrio ir formos indus ir išlaiko, kol susiformuoja kietas gelis. Gautą geli kruopščiai išplauna distiliuotu vandeniu ir veikia superkriziniu anglies dioksidu esant 80 °C temperatūrai, 30 MPa slėgyje.Sample g dissolves 10 g of acetylcellulose in 1.5 ml of acetone-ammonia solution (volume ratio: 1: 0.45), adds 12 g of crushed xenogeneic bone (bull) (granules not exceeding 200 microns), mixes well. The resulting dispersion is blown into vessels of desired volume and shape and retained until a solid gel is formed. The resulting gel is thoroughly washed with distilled water and exposed to supercritical carbon dioxide at 80 ° C under a pressure of 30 MPa.
Kompozito Jungo modulis sudaro 20 MPa. Ląstelių proliferacijos karkase duomenys pateikti Fig. 5.The Jungian modulus of the composite is 20 MPa. Cell proliferation scaffold data are presented in Figs. 5.
pavyzdysexample
Pagaminto karkaso pavirstų padengia kolageno sluoksniu. Tam įvairios kilmės (žmogaus, kiaulės, žiurkės, karvės, rekombinantinio) I tipo kolageną ištirpina 0,1 M acto rūgštyje pagal Sigma Aldrich pateikiamą metodiką, po to tirpalą praskiedžia 10 kartų fosfatiniu buferiu ir titruoja su 0,1 M natrio šarmu iki pH 7. Norimos formos karkasą patalpina į centrifūgos mėgintuvėlį, užpila kolageno tirpalu ir centrifuguoja 10 min 4000-6000 aps/min greičiu. Po to karkasą liofilizuoja.The surface of the fabricated frame is coated with a layer of collagen. For this, type I collagen of various origins (human, porcine, rat, cow, recombinant) is dissolved in 0.1 M acetic acid according to the procedure provided by Sigma Aldrich, then diluted 10 times with phosphate buffer and titrated with 0.1 M sodium hydroxide to pH 7 Place the frame of the desired shape in a centrifuge tube, fill with collagen solution and centrifuge for 10 min at 4000-6000 rpm. The frame is then lyophilized.
Claims (7)
Priority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| LT2014117A LT6309B (en) | 2014-10-13 | 2014-10-13 | Porous three dimensional cellulose based scaffold and method |
| PCT/IB2015/057597 WO2016059508A1 (en) | 2014-10-13 | 2015-10-05 | A 3d porous cellulose scaffold for bone tissue engineering and the method for its preparation |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| LT2014117A LT6309B (en) | 2014-10-13 | 2014-10-13 | Porous three dimensional cellulose based scaffold and method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| LT2014117A LT2014117A (en) | 2016-05-10 |
| LT6309B true LT6309B (en) | 2016-08-25 |
Family
ID=54345553
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| LT2014117A LT6309B (en) | 2014-10-13 | 2014-10-13 | Porous three dimensional cellulose based scaffold and method |
Country Status (2)
| Country | Link |
|---|---|
| LT (1) | LT6309B (en) |
| WO (1) | WO2016059508A1 (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| RU2660588C1 (en) * | 2017-07-18 | 2018-07-06 | Федеральное государственное учреждение "Федеральный научно-исследовательский центр "Кристаллография и фотоника" Российской академии наук" | Method of hydrogel hardening |
| CN114904052B (en) * | 2022-05-06 | 2024-02-02 | 南京友德邦医疗科技有限公司 | Novel degradable magnesium alloy vascular stent and preparation method thereof |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6478825B1 (en) | 2001-11-28 | 2002-11-12 | Osteotech, Inc. | Implant, method of making same and use of the implant for the treatment of bone defects |
| US7879093B2 (en) | 2007-03-26 | 2011-02-01 | University Of Connecticut | Electrospun apatite/polymer nano-composite scaffolds |
| US8071007B1 (en) | 2000-09-20 | 2011-12-06 | Osteopore International Pte. Ltd. | Three-dimensional bioresorbable scaffolds for tissue engineering applications |
| US20130218291A1 (en) | 2012-02-21 | 2013-08-22 | Thierry Giorno | Plga/ha hydroxyapatite composite bone grafts and method of making |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6340477B1 (en) * | 2000-04-27 | 2002-01-22 | Lifenet | Bone matrix composition and methods for making and using same |
-
2014
- 2014-10-13 LT LT2014117A patent/LT6309B/en not_active IP Right Cessation
-
2015
- 2015-10-05 WO PCT/IB2015/057597 patent/WO2016059508A1/en active Application Filing
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US8071007B1 (en) | 2000-09-20 | 2011-12-06 | Osteopore International Pte. Ltd. | Three-dimensional bioresorbable scaffolds for tissue engineering applications |
| US6478825B1 (en) | 2001-11-28 | 2002-11-12 | Osteotech, Inc. | Implant, method of making same and use of the implant for the treatment of bone defects |
| US7879093B2 (en) | 2007-03-26 | 2011-02-01 | University Of Connecticut | Electrospun apatite/polymer nano-composite scaffolds |
| US20130218291A1 (en) | 2012-02-21 | 2013-08-22 | Thierry Giorno | Plga/ha hydroxyapatite composite bone grafts and method of making |
Non-Patent Citations (2)
| Title |
|---|
| DHANDAYUTHAPANI, B., ET AL: "Polymeric scaffolds in tissue engineering application:", INTERNATIONAL JOURNAL OF POLYMER SCIENCE, 2011, pages 19 |
| MUHAMMAD, I.S.; XIAOXUE, X.; LI, L.: "review on biodegradable polymeric materials for bone tissue engineering applications", JOURNAL OF MATERIALS SCIENCE, 2009, pages 5713 - 5724, XP019750027, DOI: doi:10.1007/s10853-009-3770-7 |
Also Published As
| Publication number | Publication date |
|---|---|
| WO2016059508A1 (en) | 2016-04-21 |
| LT2014117A (en) | 2016-05-10 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Levengood et al. | Chitosan-based scaffolds for bone tissue engineering | |
| CA2677992C (en) | A collagen/hydroxyapatite composite scaffold, and process for the production thereof | |
| JP4873555B2 (en) | Method for producing porous body containing apatite / collagen composite fiber | |
| KR101027630B1 (en) | Preparation method of porous scaffold by hyaluronic acid - collagen natural polymer for cartilage resuscitation | |
| JP2010273847A (en) | High-density porous composite | |
| KR20130037324A (en) | Composition and method for 3d printing stacking of scaffold adequate to multi-material for bone regenerating | |
| JP2009132601A (en) | Porous body comprising apatite/collagen composite fiber and its production method | |
| Liao et al. | Size matters: Effects of PLGA‐microsphere size in injectable CPC/PLGA on bone formation | |
| US10517993B2 (en) | Porous composite, bone regeneration material, and method for producing porous composite | |
| US8465582B2 (en) | Process for producing inorganic interconnected 3D open cell bone substitutes | |
| KR102014248B1 (en) | A preparation method of injectable extracellular matrix based hydrogel derived from decellularized porcine skin loaded with bi-phasic calcium phosphate | |
| KR101379894B1 (en) | Bone graft materials using transgenic pig bone, method of preparing the same, and ceramic materials for biomedical application comprising the same, and | |
| LT6309B (en) | Porous three dimensional cellulose based scaffold and method | |
| JP5881206B2 (en) | Bone regeneration material | |
| RU2554804C1 (en) | Method for preparing calcium phosphate composite | |
| Yu et al. | Antler collagen/chitosan scaffolds improve critical calvarial defect healing in rats | |
| JP6018477B2 (en) | Artificial bone-cartilage complex and method for producing the same | |
| KR102494197B1 (en) | Cartilage component based bio ink for manufacturing microtia therapeutic construct and a method of making the same | |
| EP3697461A1 (en) | Porous material | |
| JP2021115165A (en) | Porous complex | |
| Pan et al. | Assessment of the suitability of a new composite as a bone defect filler in a rabbit model | |
| JP5589188B2 (en) | Bone prosthetic material and method for producing bone prosthetic material | |
| JP2006239093A (en) | Keratin porous material | |
| Srimora et al. | Evaluation of physical properties of bone scaffolds prepared from polycaprolactone microspheres | |
| Wirjosentono et al. | Biocompatibility of Chitosan/Collagen/PVA Nanocomposite Containing Calcium Apatite |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| BB1A | Patent application published |
Effective date: 20160510 |
|
| FG9A | Patent granted |
Effective date: 20160825 |
|
| MM9A | Lapsed patents |
Effective date: 20171013 |