CN113456303A - Osteochondral scaffold and preparation method and application thereof - Google Patents
Osteochondral scaffold and preparation method and application thereof Download PDFInfo
- Publication number
- CN113456303A CN113456303A CN202110903051.5A CN202110903051A CN113456303A CN 113456303 A CN113456303 A CN 113456303A CN 202110903051 A CN202110903051 A CN 202110903051A CN 113456303 A CN113456303 A CN 113456303A
- Authority
- CN
- China
- Prior art keywords
- layer
- cartilage
- bone
- parts
- hydrogel
- 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.)
- Pending
Links
- 238000002360 preparation method Methods 0.000 title claims abstract description 30
- 239000010410 layer Substances 0.000 claims abstract description 262
- 210000000845 cartilage Anatomy 0.000 claims abstract description 171
- 210000000988 bone and bone Anatomy 0.000 claims abstract description 125
- 239000012790 adhesive layer Substances 0.000 claims abstract description 32
- 201000009859 Osteochondrosis Diseases 0.000 claims abstract description 5
- 239000000017 hydrogel Substances 0.000 claims description 116
- 239000000463 material Substances 0.000 claims description 74
- 239000000203 mixture Substances 0.000 claims description 66
- 108010010803 Gelatin Proteins 0.000 claims description 58
- 229920000159 gelatin Polymers 0.000 claims description 58
- 239000008273 gelatin Substances 0.000 claims description 58
- 235000019322 gelatine Nutrition 0.000 claims description 58
- 235000011852 gelatine desserts Nutrition 0.000 claims description 58
- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 claims description 48
- 229920002674 hyaluronan Polymers 0.000 claims description 48
- 229960003160 hyaluronic acid Drugs 0.000 claims description 48
- 238000004132 cross linking Methods 0.000 claims description 44
- 238000000034 method Methods 0.000 claims description 40
- 229920001287 Chondroitin sulfate Polymers 0.000 claims description 37
- 229940059329 chondroitin sulfate Drugs 0.000 claims description 37
- SQDAZGGFXASXDW-UHFFFAOYSA-N 5-bromo-2-(trifluoromethoxy)pyridine Chemical compound FC(F)(F)OC1=CC=C(Br)C=N1 SQDAZGGFXASXDW-UHFFFAOYSA-N 0.000 claims description 36
- 238000010146 3D printing Methods 0.000 claims description 35
- 238000007639 printing Methods 0.000 claims description 33
- 229920001610 polycaprolactone Polymers 0.000 claims description 26
- 239000004626 polylactic acid Substances 0.000 claims description 26
- 229920000747 poly(lactic acid) Polymers 0.000 claims description 24
- 239000004632 polycaprolactone Substances 0.000 claims description 24
- 239000000499 gel Substances 0.000 claims description 22
- 238000001723 curing Methods 0.000 claims description 21
- 239000002861 polymer material Substances 0.000 claims description 21
- 230000000975 bioactive effect Effects 0.000 claims description 20
- 210000004027 cell Anatomy 0.000 claims description 20
- 239000001506 calcium phosphate Substances 0.000 claims description 18
- 238000004108 freeze drying Methods 0.000 claims description 18
- 239000011148 porous material Substances 0.000 claims description 18
- 230000001737 promoting effect Effects 0.000 claims description 18
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 claims description 18
- 239000002609 medium Substances 0.000 claims description 17
- 229910052588 hydroxylapatite Inorganic materials 0.000 claims description 16
- XYJRXVWERLGGKC-UHFFFAOYSA-D pentacalcium;hydroxide;triphosphate Chemical compound [OH-].[Ca+2].[Ca+2].[Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O XYJRXVWERLGGKC-UHFFFAOYSA-D 0.000 claims description 16
- 239000002904 solvent Substances 0.000 claims description 16
- 229940078499 tricalcium phosphate Drugs 0.000 claims description 16
- 229910000391 tricalcium phosphate Inorganic materials 0.000 claims description 16
- 235000019731 tricalcium phosphate Nutrition 0.000 claims description 16
- 238000011068 loading method Methods 0.000 claims description 15
- 239000007857 degradation product Substances 0.000 claims description 14
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 13
- 238000011049 filling Methods 0.000 claims description 12
- 239000003999 initiator Substances 0.000 claims description 12
- 210000002901 mesenchymal stem cell Anatomy 0.000 claims description 12
- 229920001282 polysaccharide Polymers 0.000 claims description 12
- 239000005017 polysaccharide Substances 0.000 claims description 12
- 102000009618 Transforming Growth Factors Human genes 0.000 claims description 11
- 108010009583 Transforming Growth Factors Proteins 0.000 claims description 11
- 239000002562 thickening agent Substances 0.000 claims description 11
- 229920001661 Chitosan Polymers 0.000 claims description 10
- 239000005313 bioactive glass Substances 0.000 claims description 10
- 238000007710 freezing Methods 0.000 claims description 10
- 230000008014 freezing Effects 0.000 claims description 10
- 239000003292 glue Substances 0.000 claims description 10
- 230000017854 proteolysis Effects 0.000 claims description 10
- ZRKFYGHZFMAOKI-QMGMOQQFSA-N tgfbeta Chemical compound C([C@H](NC(=O)[C@H](C(C)C)NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CC(C)C)NC(=O)CNC(=O)[C@H](C)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](NC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)CCSC)C(C)C)[C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](C)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(=O)N1[C@@H](CCC1)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(C)C)C(O)=O)C1=CC=C(O)C=C1 ZRKFYGHZFMAOKI-QMGMOQQFSA-N 0.000 claims description 10
- 230000004048 modification Effects 0.000 claims description 9
- 238000012986 modification Methods 0.000 claims description 9
- 238000001816 cooling Methods 0.000 claims description 8
- 230000008021 deposition Effects 0.000 claims description 8
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 claims description 6
- 239000002202 Polyethylene glycol Substances 0.000 claims description 6
- 239000004372 Polyvinyl alcohol Substances 0.000 claims description 6
- DNIAPMSPPWPWGF-UHFFFAOYSA-N Propylene glycol Chemical compound CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 claims description 6
- 238000001125 extrusion Methods 0.000 claims description 6
- 229910010272 inorganic material Inorganic materials 0.000 claims description 6
- 239000011147 inorganic material Substances 0.000 claims description 6
- 229920001223 polyethylene glycol Polymers 0.000 claims description 6
- 229920002451 polyvinyl alcohol Polymers 0.000 claims description 6
- 239000002994 raw material Substances 0.000 claims description 6
- 238000010382 chemical cross-linking Methods 0.000 claims description 5
- 229920000058 polyacrylate Polymers 0.000 claims description 5
- -1 polymethacrylamide Polymers 0.000 claims description 5
- 229920000036 polyvinylpyrrolidone Polymers 0.000 claims description 5
- 239000001267 polyvinylpyrrolidone Substances 0.000 claims description 5
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 claims description 5
- 239000000843 powder Substances 0.000 claims description 5
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 claims description 4
- 241001465754 Metazoa Species 0.000 claims description 4
- 108010064851 Plant Proteins Proteins 0.000 claims description 4
- AUNGANRZJHBGPY-SCRDCRAPSA-N Riboflavin Chemical compound OC[C@@H](O)[C@@H](O)[C@@H](O)CN1C=2C=C(C)C(C)=CC=2N=C2C1=NC(=O)NC2=O AUNGANRZJHBGPY-SCRDCRAPSA-N 0.000 claims description 4
- 229920002125 Sokalan® Polymers 0.000 claims description 4
- 230000000735 allogeneic effect Effects 0.000 claims description 4
- 235000021120 animal protein Nutrition 0.000 claims description 4
- 239000006143 cell culture medium Substances 0.000 claims description 4
- 238000001035 drying Methods 0.000 claims description 4
- 210000001671 embryonic stem cell Anatomy 0.000 claims description 4
- 238000005286 illumination Methods 0.000 claims description 4
- 210000004263 induced pluripotent stem cell Anatomy 0.000 claims description 4
- 235000021118 plant-derived protein Nutrition 0.000 claims description 4
- 229920001184 polypeptide Polymers 0.000 claims description 4
- 238000004321 preservation Methods 0.000 claims description 4
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 4
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 4
- 235000018102 proteins Nutrition 0.000 claims description 4
- 102000004169 proteins and genes Human genes 0.000 claims description 4
- 108090000623 proteins and genes Proteins 0.000 claims description 4
- 239000008213 purified water Substances 0.000 claims description 4
- GJKGAPPUXSSCFI-UHFFFAOYSA-N 2-Hydroxy-4'-(2-hydroxyethoxy)-2-methylpropiophenone Chemical group CC(C)(O)C(=O)C1=CC=C(OCCO)C=C1 GJKGAPPUXSSCFI-UHFFFAOYSA-N 0.000 claims description 3
- 229920002845 Poly(methacrylic acid) Polymers 0.000 claims description 3
- 159000000007 calcium salts Chemical class 0.000 claims description 3
- 210000001612 chondrocyte Anatomy 0.000 claims description 3
- 230000002255 enzymatic effect Effects 0.000 claims description 3
- 230000010412 perfusion Effects 0.000 claims description 3
- 238000000016 photochemical curing Methods 0.000 claims description 3
- 239000002504 physiological saline solution Substances 0.000 claims description 3
- 229920002401 polyacrylamide Polymers 0.000 claims description 3
- 239000004584 polyacrylic acid Substances 0.000 claims description 3
- 229920000193 polymethacrylate Polymers 0.000 claims description 3
- 239000012266 salt solution Substances 0.000 claims description 3
- JFJNVIPVOCESGZ-UHFFFAOYSA-N 2,3-dipyridin-2-ylpyridine Chemical compound N1=CC=CC=C1C1=CC=CN=C1C1=CC=CC=N1 JFJNVIPVOCESGZ-UHFFFAOYSA-N 0.000 claims description 2
- LCPVQAHEFVXVKT-UHFFFAOYSA-N 2-(2,4-difluorophenoxy)pyridin-3-amine Chemical compound NC1=CC=CN=C1OC1=CC=C(F)C=C1F LCPVQAHEFVXVKT-UHFFFAOYSA-N 0.000 claims description 2
- 229920000936 Agarose Polymers 0.000 claims description 2
- 102000004506 Blood Proteins Human genes 0.000 claims description 2
- 108010017384 Blood Proteins Proteins 0.000 claims description 2
- 108010049931 Bone Morphogenetic Protein 2 Proteins 0.000 claims description 2
- 108010049951 Bone Morphogenetic Protein 3 Proteins 0.000 claims description 2
- 108010049955 Bone Morphogenetic Protein 4 Proteins 0.000 claims description 2
- 108010049976 Bone Morphogenetic Protein 5 Proteins 0.000 claims description 2
- 108010049974 Bone Morphogenetic Protein 6 Proteins 0.000 claims description 2
- 108010049870 Bone Morphogenetic Protein 7 Proteins 0.000 claims description 2
- 102100024506 Bone morphogenetic protein 2 Human genes 0.000 claims description 2
- 102100024504 Bone morphogenetic protein 3 Human genes 0.000 claims description 2
- 102100024505 Bone morphogenetic protein 4 Human genes 0.000 claims description 2
- 102100022526 Bone morphogenetic protein 5 Human genes 0.000 claims description 2
- 102100022525 Bone morphogenetic protein 6 Human genes 0.000 claims description 2
- 102100022544 Bone morphogenetic protein 7 Human genes 0.000 claims description 2
- 102100022545 Bone morphogenetic protein 8B Human genes 0.000 claims description 2
- 229920002134 Carboxymethyl cellulose Polymers 0.000 claims description 2
- 102000012422 Collagen Type I Human genes 0.000 claims description 2
- 108010022452 Collagen Type I Proteins 0.000 claims description 2
- 102000000503 Collagen Type II Human genes 0.000 claims description 2
- 108010041390 Collagen Type II Proteins 0.000 claims description 2
- AUNGANRZJHBGPY-UHFFFAOYSA-N D-Lyxoflavin Natural products OCC(O)C(O)C(O)CN1C=2C=C(C)C(C)=CC=2N=C2C1=NC(=O)NC2=O AUNGANRZJHBGPY-UHFFFAOYSA-N 0.000 claims description 2
- 229920002307 Dextran Polymers 0.000 claims description 2
- 102000016942 Elastin Human genes 0.000 claims description 2
- 108010014258 Elastin Proteins 0.000 claims description 2
- 108010022355 Fibroins Proteins 0.000 claims description 2
- 108010090290 Growth Differentiation Factor 2 Proteins 0.000 claims description 2
- 102100040892 Growth/differentiation factor 2 Human genes 0.000 claims description 2
- HTTJABKRGRZYRN-UHFFFAOYSA-N Heparin Chemical compound OC1C(NC(=O)C)C(O)OC(COS(O)(=O)=O)C1OC1C(OS(O)(=O)=O)C(O)C(OC2C(C(OS(O)(=O)=O)C(OC3C(C(O)C(O)C(O3)C(O)=O)OS(O)(=O)=O)C(CO)O2)NS(O)(=O)=O)C(C(O)=O)O1 HTTJABKRGRZYRN-UHFFFAOYSA-N 0.000 claims description 2
- 101000899368 Homo sapiens Bone morphogenetic protein 8B Proteins 0.000 claims description 2
- 239000004354 Hydroxyethyl cellulose Substances 0.000 claims description 2
- 229920000663 Hydroxyethyl cellulose Polymers 0.000 claims description 2
- 229920002153 Hydroxypropyl cellulose Polymers 0.000 claims description 2
- JVTAAEKCZFNVCJ-UHFFFAOYSA-M Lactate Chemical compound CC(O)C([O-])=O JVTAAEKCZFNVCJ-UHFFFAOYSA-M 0.000 claims description 2
- 229920002873 Polyethylenimine Polymers 0.000 claims description 2
- 210000002449 bone cell Anatomy 0.000 claims description 2
- 229910000019 calcium carbonate Inorganic materials 0.000 claims description 2
- 229910000389 calcium phosphate Inorganic materials 0.000 claims description 2
- 235000011010 calcium phosphates Nutrition 0.000 claims description 2
- 239000001768 carboxy methyl cellulose Substances 0.000 claims description 2
- 235000010948 carboxy methyl cellulose Nutrition 0.000 claims description 2
- 125000002057 carboxymethyl group Chemical group [H]OC(=O)C([H])([H])[*] 0.000 claims description 2
- 239000008112 carboxymethyl-cellulose Substances 0.000 claims description 2
- 125000004386 diacrylate group Chemical group 0.000 claims description 2
- 229920002549 elastin Polymers 0.000 claims description 2
- SEACYXSIPDVVMV-UHFFFAOYSA-L eosin Y Chemical compound [Na+].[Na+].[O-]C(=O)C1=CC=CC=C1C1=C2C=C(Br)C(=O)C(Br)=C2OC2=C(Br)C([O-])=C(Br)C=C21 SEACYXSIPDVVMV-UHFFFAOYSA-L 0.000 claims description 2
- FVTCRASFADXXNN-SCRDCRAPSA-N flavin mononucleotide Chemical compound OP(=O)(O)OC[C@@H](O)[C@@H](O)[C@@H](O)CN1C=2C=C(C)C(C)=CC=2N=C2C1=NC(=O)NC2=O FVTCRASFADXXNN-SCRDCRAPSA-N 0.000 claims description 2
- 239000011768 flavin mononucleotide Substances 0.000 claims description 2
- 229940013640 flavin mononucleotide Drugs 0.000 claims description 2
- FVTCRASFADXXNN-UHFFFAOYSA-N flavin mononucleotide Natural products OP(=O)(O)OCC(O)C(O)C(O)CN1C=2C=C(C)C(C)=CC=2N=C2C1=NC(=O)NC2=O FVTCRASFADXXNN-UHFFFAOYSA-N 0.000 claims description 2
- 238000000227 grinding Methods 0.000 claims description 2
- 229920000669 heparin Polymers 0.000 claims description 2
- 229960002897 heparin Drugs 0.000 claims description 2
- 235000019447 hydroxyethyl cellulose Nutrition 0.000 claims description 2
- 229940071826 hydroxyethyl cellulose Drugs 0.000 claims description 2
- 239000001863 hydroxypropyl cellulose Substances 0.000 claims description 2
- 235000010977 hydroxypropyl cellulose Nutrition 0.000 claims description 2
- 239000004615 ingredient Substances 0.000 claims description 2
- 229920000609 methyl cellulose Polymers 0.000 claims description 2
- 239000001923 methylcellulose Substances 0.000 claims description 2
- 235000010981 methylcellulose Nutrition 0.000 claims description 2
- 230000000921 morphogenic effect Effects 0.000 claims description 2
- 239000008055 phosphate buffer solution Substances 0.000 claims description 2
- 230000009145 protein modification Effects 0.000 claims description 2
- 150000003242 quaternary ammonium salts Chemical class 0.000 claims description 2
- 235000019192 riboflavin Nutrition 0.000 claims description 2
- 239000002151 riboflavin Substances 0.000 claims description 2
- 229960002477 riboflavin Drugs 0.000 claims description 2
- 235000019231 riboflavin-5'-phosphate Nutrition 0.000 claims description 2
- CHQMHPLRPQMAMX-UHFFFAOYSA-L sodium persulfate Substances [Na+].[Na+].[O-]S(=O)(=O)OOS([O-])(=O)=O CHQMHPLRPQMAMX-UHFFFAOYSA-L 0.000 claims description 2
- 150000004804 polysaccharides Chemical class 0.000 claims 6
- 229920001577 copolymer Polymers 0.000 claims 2
- 239000003607 modifier Substances 0.000 claims 2
- LNAZSHAWQACDHT-XIYTZBAFSA-N (2r,3r,4s,5r,6s)-4,5-dimethoxy-2-(methoxymethyl)-3-[(2s,3r,4s,5r,6r)-3,4,5-trimethoxy-6-(methoxymethyl)oxan-2-yl]oxy-6-[(2r,3r,4s,5r,6r)-4,5,6-trimethoxy-2-(methoxymethyl)oxan-3-yl]oxyoxane Chemical compound CO[C@@H]1[C@@H](OC)[C@H](OC)[C@@H](COC)O[C@H]1O[C@H]1[C@H](OC)[C@@H](OC)[C@H](O[C@H]2[C@@H]([C@@H](OC)[C@H](OC)O[C@@H]2COC)OC)O[C@@H]1COC LNAZSHAWQACDHT-XIYTZBAFSA-N 0.000 claims 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-M Chloride anion Chemical compound [Cl-] VEXZGXHMUGYJMC-UHFFFAOYSA-M 0.000 claims 1
- KJTLSVCANCCWHF-UHFFFAOYSA-N Ruthenium Chemical compound [Ru] KJTLSVCANCCWHF-UHFFFAOYSA-N 0.000 claims 1
- 239000000783 alginic acid Substances 0.000 claims 1
- 229920000615 alginic acid Polymers 0.000 claims 1
- 235000010443 alginic acid Nutrition 0.000 claims 1
- 229960001126 alginic acid Drugs 0.000 claims 1
- 150000004781 alginic acids Chemical class 0.000 claims 1
- 229940105329 carboxymethylcellulose Drugs 0.000 claims 1
- 229960002086 dextran Drugs 0.000 claims 1
- OQORATGQIWQZBQ-UHFFFAOYSA-L dilithium phosphonato-(2,4,6-trimethyl-3-phenylphenyl)methanone Chemical group C1(=CC=CC=C1)C=1C(=C(C(=O)P([O-])([O-])=O)C(=CC=1C)C)C.[Li+].[Li+] OQORATGQIWQZBQ-UHFFFAOYSA-L 0.000 claims 1
- 229940093476 ethylene glycol Drugs 0.000 claims 1
- 229940071676 hydroxypropylcellulose Drugs 0.000 claims 1
- FZWBNHMXJMCXLU-BLAUPYHCSA-N isomaltotriose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1OC[C@@H]1[C@@H](O)[C@H](O)[C@@H](O)[C@@H](OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C=O)O1 FZWBNHMXJMCXLU-BLAUPYHCSA-N 0.000 claims 1
- 229960002900 methylcellulose Drugs 0.000 claims 1
- 229910052707 ruthenium Inorganic materials 0.000 claims 1
- 208000027418 Wounds and injury Diseases 0.000 abstract description 4
- 230000006378 damage Effects 0.000 abstract description 3
- 208000014674 injury Diseases 0.000 abstract description 2
- OOIBFPKQHULHSQ-UHFFFAOYSA-N (3-hydroxy-1-adamantyl) 2-methylprop-2-enoate Chemical compound C1C(C2)CC3CC2(O)CC1(OC(=O)C(=C)C)C3 OOIBFPKQHULHSQ-UHFFFAOYSA-N 0.000 description 36
- 230000007547 defect Effects 0.000 description 21
- 239000000243 solution Substances 0.000 description 20
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical group ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 18
- 101100172132 Mus musculus Eif3a gene Proteins 0.000 description 16
- 239000002131 composite material Substances 0.000 description 15
- IXPNQXFRVYWDDI-UHFFFAOYSA-N 1-methyl-2,4-dioxo-1,3-diazinane-5-carboximidamide Chemical compound CN1CC(C(N)=N)C(=O)NC1=O IXPNQXFRVYWDDI-UHFFFAOYSA-N 0.000 description 13
- 230000008439 repair process Effects 0.000 description 13
- 235000010413 sodium alginate Nutrition 0.000 description 12
- 239000000661 sodium alginate Substances 0.000 description 12
- 229940005550 sodium alginate Drugs 0.000 description 12
- 230000000694 effects Effects 0.000 description 11
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 10
- 239000002953 phosphate buffered saline Substances 0.000 description 10
- 210000001519 tissue Anatomy 0.000 description 10
- 230000015572 biosynthetic process Effects 0.000 description 9
- 238000003786 synthesis reaction Methods 0.000 description 9
- 239000008367 deionised water Substances 0.000 description 8
- 229910021641 deionized water Inorganic materials 0.000 description 8
- 238000005160 1H NMR spectroscopy Methods 0.000 description 6
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 6
- 238000005481 NMR spectroscopy Methods 0.000 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- 238000007796 conventional method Methods 0.000 description 6
- 238000000151 deposition Methods 0.000 description 6
- 150000004676 glycans Chemical class 0.000 description 6
- 229910052739 hydrogen Inorganic materials 0.000 description 6
- 239000001257 hydrogen Substances 0.000 description 6
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 description 5
- 239000004472 Lysine Substances 0.000 description 5
- 241000283973 Oryctolagus cuniculus Species 0.000 description 5
- 238000006243 chemical reaction Methods 0.000 description 5
- 125000003588 lysine group Chemical group [H]N([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])(N([H])[H])C(*)=O 0.000 description 5
- 238000002791 soaking Methods 0.000 description 5
- 238000005303 weighing Methods 0.000 description 5
- 210000001185 bone marrow Anatomy 0.000 description 4
- 238000012258 culturing Methods 0.000 description 4
- 238000010586 diagram Methods 0.000 description 4
- 238000005516 engineering process Methods 0.000 description 4
- 238000000465 moulding Methods 0.000 description 4
- 239000011347 resin Substances 0.000 description 4
- 229920005989 resin Polymers 0.000 description 4
- 238000001228 spectrum Methods 0.000 description 4
- 230000007704 transition Effects 0.000 description 4
- 229920002148 Gellan gum Polymers 0.000 description 3
- 229920000084 Gum arabic Polymers 0.000 description 3
- 241000406668 Loxodonta cyclotis Species 0.000 description 3
- 241000978776 Senegalia senegal Species 0.000 description 3
- 235000010489 acacia gum Nutrition 0.000 description 3
- 239000000205 acacia gum Substances 0.000 description 3
- 239000007864 aqueous solution Substances 0.000 description 3
- 210000001188 articular cartilage Anatomy 0.000 description 3
- 238000004113 cell culture Methods 0.000 description 3
- 238000000502 dialysis Methods 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 230000002349 favourable effect Effects 0.000 description 3
- 235000010492 gellan gum Nutrition 0.000 description 3
- 239000000216 gellan gum Substances 0.000 description 3
- 239000001963 growth medium Substances 0.000 description 3
- DCUFMVPCXCSVNP-UHFFFAOYSA-N methacrylic anhydride Chemical compound CC(=C)C(=O)OC(=O)C(C)=C DCUFMVPCXCSVNP-UHFFFAOYSA-N 0.000 description 3
- 238000003756 stirring Methods 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 239000000230 xanthan gum Substances 0.000 description 3
- 235000010493 xanthan gum Nutrition 0.000 description 3
- 229920001285 xanthan gum Polymers 0.000 description 3
- 229940082509 xanthan gum Drugs 0.000 description 3
- NIXOWILDQLNWCW-UHFFFAOYSA-M Acrylate Chemical compound [O-]C(=O)C=C NIXOWILDQLNWCW-UHFFFAOYSA-M 0.000 description 2
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 2
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 2
- COLNVLDHVKWLRT-QMMMGPOBSA-N L-phenylalanine Chemical compound OC(=O)[C@@H](N)CC1=CC=CC=C1 COLNVLDHVKWLRT-QMMMGPOBSA-N 0.000 description 2
- 108010019160 Pancreatin Proteins 0.000 description 2
- 229920003171 Poly (ethylene oxide) Polymers 0.000 description 2
- 239000004696 Poly ether ether ketone Substances 0.000 description 2
- 239000004697 Polyetherimide Substances 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 description 2
- ROOXNKNUYICQNP-UHFFFAOYSA-N ammonium persulfate Chemical compound [NH4+].[NH4+].[O-]S(=O)(=O)OOS([O-])(=O)=O ROOXNKNUYICQNP-UHFFFAOYSA-N 0.000 description 2
- DEGAKNSWVGKMLS-UHFFFAOYSA-N calcein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC(CN(CC(O)=O)CC(O)=O)=C(O)C=C1OC1=C2C=C(CN(CC(O)=O)CC(=O)O)C(O)=C1 DEGAKNSWVGKMLS-UHFFFAOYSA-N 0.000 description 2
- 210000003321 cartilage cell Anatomy 0.000 description 2
- 239000006285 cell suspension Substances 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 238000009472 formulation Methods 0.000 description 2
- 239000003517 fume Substances 0.000 description 2
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 2
- 238000011065 in-situ storage Methods 0.000 description 2
- 239000013642 negative control Substances 0.000 description 2
- 235000015097 nutrients Nutrition 0.000 description 2
- 229960002378 oftasceine Drugs 0.000 description 2
- 230000011164 ossification Effects 0.000 description 2
- 229940055695 pancreatin Drugs 0.000 description 2
- COLNVLDHVKWLRT-UHFFFAOYSA-N phenylalanine Natural products OC(=O)C(N)CC1=CC=CC=C1 COLNVLDHVKWLRT-UHFFFAOYSA-N 0.000 description 2
- 229920002530 polyetherether ketone Polymers 0.000 description 2
- 229920001601 polyetherimide Polymers 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 238000004611 spectroscopical analysis Methods 0.000 description 2
- 210000000130 stem cell Anatomy 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000001291 vacuum drying Methods 0.000 description 2
- 229920002554 vinyl polymer Polymers 0.000 description 2
- RYHBNJHYFVUHQT-UHFFFAOYSA-N 1,4-Dioxane Chemical compound C1COCCO1 RYHBNJHYFVUHQT-UHFFFAOYSA-N 0.000 description 1
- NIXOWILDQLNWCW-UHFFFAOYSA-N Acrylic acid Chemical compound OC(=O)C=C NIXOWILDQLNWCW-UHFFFAOYSA-N 0.000 description 1
- 206010007710 Cartilage injury Diseases 0.000 description 1
- 102000008186 Collagen Human genes 0.000 description 1
- 108010035532 Collagen Proteins 0.000 description 1
- QTANTQQOYSUMLC-UHFFFAOYSA-O Ethidium cation Chemical class C12=CC(N)=CC=C2C2=CC=C(N)C=C2[N+](CC)=C1C1=CC=CC=C1 QTANTQQOYSUMLC-UHFFFAOYSA-O 0.000 description 1
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 1
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 1
- 108010049003 Fibrinogen Proteins 0.000 description 1
- 102000008946 Fibrinogen Human genes 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 208000012659 Joint disease Diseases 0.000 description 1
- LBPJJBJKHHBVBI-LQDZTQBFSA-N Mollin Chemical compound CC(=O)O[C@H]1[C@H](Oc2cc3oc(C)cc(=O)c3c(O)c2C)O[C@H](CO)[C@@H](O)[C@@H]1O LBPJJBJKHHBVBI-LQDZTQBFSA-N 0.000 description 1
- LBPJJBJKHHBVBI-UHFFFAOYSA-N Mollin Natural products CC(=O)OC1C(O)C(O)C(CO)OC1Oc2cc3OC(=CC(=O)c3c(O)c2C)C LBPJJBJKHHBVBI-UHFFFAOYSA-N 0.000 description 1
- 125000003047 N-acetyl group Chemical group 0.000 description 1
- 208000037273 Pathologic Processes Diseases 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 239000004698 Polyethylene Substances 0.000 description 1
- 229920000491 Polyphenylsulfone Polymers 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 1
- 208000013201 Stress fracture Diseases 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 229910001870 ammonium persulfate Inorganic materials 0.000 description 1
- 230000002051 biphasic effect Effects 0.000 description 1
- 210000004204 blood vessel Anatomy 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 229960001631 carbomer Drugs 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 238000006555 catalytic reaction Methods 0.000 description 1
- 150000001768 cations Chemical class 0.000 description 1
- 230000024245 cell differentiation Effects 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 230000012292 cell migration Effects 0.000 description 1
- 230000003833 cell viability Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 229920001436 collagen Polymers 0.000 description 1
- 238000013329 compounding Methods 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 230000008034 disappearance Effects 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 238000012137 double-staining Methods 0.000 description 1
- 238000005553 drilling Methods 0.000 description 1
- 238000012407 engineering method Methods 0.000 description 1
- ZMMJGEGLRURXTF-UHFFFAOYSA-N ethidium bromide Chemical compound [Br-].C12=CC(N)=CC=C2C2=CC=C(N)C=C2[N+](CC)=C1C1=CC=CC=C1 ZMMJGEGLRURXTF-UHFFFAOYSA-N 0.000 description 1
- 229960005542 ethidium bromide Drugs 0.000 description 1
- 230000005284 excitation Effects 0.000 description 1
- 210000002744 extracellular matrix Anatomy 0.000 description 1
- 229940012952 fibrinogen Drugs 0.000 description 1
- 210000000968 fibrocartilage Anatomy 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- DDTGNKBZWQHIEH-UHFFFAOYSA-N heptalene Chemical compound C1=CC=CC=C2C=CC=CC=C21 DDTGNKBZWQHIEH-UHFFFAOYSA-N 0.000 description 1
- 210000004394 hip joint Anatomy 0.000 description 1
- 210000003035 hyaline cartilage Anatomy 0.000 description 1
- 238000002513 implantation Methods 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 238000001802 infusion Methods 0.000 description 1
- 230000010354 integration Effects 0.000 description 1
- 238000001990 intravenous administration Methods 0.000 description 1
- 210000000629 knee joint Anatomy 0.000 description 1
- 230000003902 lesion Effects 0.000 description 1
- 239000004973 liquid crystal related substance Substances 0.000 description 1
- 210000002751 lymph Anatomy 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- 238000001000 micrograph Methods 0.000 description 1
- 230000005012 migration Effects 0.000 description 1
- 238000013508 migration Methods 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 210000004409 osteocyte Anatomy 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 231100000915 pathological change Toxicity 0.000 description 1
- 230000036285 pathological change Effects 0.000 description 1
- 230000009054 pathological process Effects 0.000 description 1
- 230000035515 penetration Effects 0.000 description 1
- UEZVMMHDMIWARA-UHFFFAOYSA-M phosphonate Chemical compound [O-]P(=O)=O UEZVMMHDMIWARA-UHFFFAOYSA-M 0.000 description 1
- 239000000049 pigment Substances 0.000 description 1
- 229920003023 plastic Polymers 0.000 description 1
- 239000004033 plastic Substances 0.000 description 1
- 229920001983 poloxamer Polymers 0.000 description 1
- 229920001643 poly(ether ketone) Polymers 0.000 description 1
- 229920001652 poly(etherketoneketone) Polymers 0.000 description 1
- 229920000573 polyethylene Polymers 0.000 description 1
- 229920005644 polyethylene terephthalate glycol copolymer Polymers 0.000 description 1
- YBCAZPLXEGKKFM-UHFFFAOYSA-K ruthenium(iii) chloride Chemical compound [Cl-].[Cl-].[Cl-].[Ru+3] YBCAZPLXEGKKFM-UHFFFAOYSA-K 0.000 description 1
- 231100000241 scar Toxicity 0.000 description 1
- 238000001338 self-assembly Methods 0.000 description 1
- 210000000323 shoulder joint Anatomy 0.000 description 1
- 239000002356 single layer Substances 0.000 description 1
- 238000007711 solidification Methods 0.000 description 1
- 230000008023 solidification Effects 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
- 239000012128 staining reagent Substances 0.000 description 1
- 239000007858 starting material Substances 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
- 238000005491 wire drawing Methods 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
- A61F2/00—Filters implantable into blood vessels; Prostheses, i.e. artificial substitutes or replacements for parts of the body; Appliances for connecting them with the body; Devices providing patency to, or preventing collapsing of, tubular structures of the body, e.g. stents
- A61F2/02—Prostheses implantable into the body
- A61F2/28—Bones
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
- A61F2/00—Filters implantable into blood vessels; Prostheses, i.e. artificial substitutes or replacements for parts of the body; Appliances for connecting them with the body; Devices providing patency to, or preventing collapsing of, tubular structures of the body, e.g. stents
- A61F2/02—Prostheses implantable into the body
- A61F2/30—Joints
- A61F2/30756—Cartilage endoprostheses
-
- 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
- A61L27/02—Inorganic materials
- A61L27/12—Phosphorus-containing materials, e.g. apatite
-
- 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
- A61L27/14—Macromolecular materials
- A61L27/18—Macromolecular materials obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds
-
- 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
- A61L27/14—Macromolecular materials
- A61L27/20—Polysaccharides
-
- 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
- A61L27/14—Macromolecular materials
- A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
- A61L27/222—Gelatin
-
- 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
-
- 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
- A61L27/52—Hydrogels or hydrocolloids
-
- 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
- A61L27/54—Biologically active materials, e.g. therapeutic substances
-
- 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
- A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
- A61F2/00—Filters implantable into blood vessels; Prostheses, i.e. artificial substitutes or replacements for parts of the body; Appliances for connecting them with the body; Devices providing patency to, or preventing collapsing of, tubular structures of the body, e.g. stents
- A61F2/02—Prostheses implantable into the body
- A61F2/28—Bones
- A61F2002/2835—Bone graft implants for filling a bony defect or an endoprosthesis cavity, e.g. by synthetic material or biological material
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
- A61F2/00—Filters implantable into blood vessels; Prostheses, i.e. artificial substitutes or replacements for parts of the body; Appliances for connecting them with the body; Devices providing patency to, or preventing collapsing of, tubular structures of the body, e.g. stents
- A61F2/02—Prostheses implantable into the body
- A61F2/30—Joints
- A61F2002/30001—Additional features of subject-matter classified in A61F2/28, A61F2/30 and subgroups thereof
- A61F2002/30003—Material related properties of the prosthesis or of a coating on the prosthesis
- A61F2002/3006—Properties of materials and coating materials
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
- A61F2/00—Filters implantable into blood vessels; Prostheses, i.e. artificial substitutes or replacements for parts of the body; Appliances for connecting them with the body; Devices providing patency to, or preventing collapsing of, tubular structures of the body, e.g. stents
- A61F2/02—Prostheses implantable into the body
- A61F2/30—Joints
- A61F2/30767—Special external or bone-contacting surface, e.g. coating for improving bone ingrowth
- A61F2002/3093—Special external or bone-contacting surface, e.g. coating for improving bone ingrowth for promoting ingrowth of bone tissue
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
- A61F2/00—Filters implantable into blood vessels; Prostheses, i.e. artificial substitutes or replacements for parts of the body; Appliances for connecting them with the body; Devices providing patency to, or preventing collapsing of, tubular structures of the body, e.g. stents
- A61F2/02—Prostheses implantable into the body
- A61F2/30—Joints
- A61F2/3094—Designing or manufacturing processes
- A61F2002/30985—Designing or manufacturing processes using three dimensional printing [3DP]
-
- 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
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/40—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
- A61L2300/412—Tissue-regenerating or healing or proliferative agents
- A61L2300/414—Growth factors
-
- 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
-
- 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
- A61L2430/06—Materials or treatment for tissue regeneration for cartilage reconstruction, e.g. meniscus
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Veterinary Medicine (AREA)
- Oral & Maxillofacial Surgery (AREA)
- Transplantation (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Medicinal Chemistry (AREA)
- Epidemiology (AREA)
- Dermatology (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Dispersion Chemistry (AREA)
- Cardiology (AREA)
- Orthopedic Medicine & Surgery (AREA)
- Heart & Thoracic Surgery (AREA)
- Vascular Medicine (AREA)
- Rheumatology (AREA)
- Inorganic Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Molecular Biology (AREA)
- Materials For Medical Uses (AREA)
Abstract
The invention discloses an osteochondral scaffold and a preparation method and application thereof. The osteochondral scaffold comprises a cartilage layer, an adhesion layer and a bone layer, wherein two sides of the adhesion layer are respectively connected with the cartilage layer and the bone layer; one or more of the cartilage layer, the adhesive layer and the bone layer are porous structures. The osteochondral scaffold can be used for repairing osteochondral defects, can fully capture cells, has a strong osteochondral injury repairing function, and is simple and convenient to operate.
Description
Technical Field
The invention relates to a bone cartilage scaffold and a preparation method and application thereof.
Background
Articular cartilage lesions, as a fundamental pathological process, are involved in almost all clinically common early stages of pathological changes in joint diseases. Because the cartilage tissue lacks blood vessels and lymph distribution, has low content of cartilage cells, lacks progenitor cells necessary for cell differentiation, is embedded in thick extracellular matrix, has high migration difficulty, cannot effectively move to a damaged part to participate in repair, has very poor self-repair capability, and is difficult to naturally repair even tiny cartilage defects. The emergence of tissue engineering technology provides a brand new thought and method for the treatment of articular cartilage injury.
At present, the design of the tissue engineering bone cartilage scaffold is mainly divided into the following categories: 1) the bone adopts a bracket, the cartilage does not adopt the bracket, namely, the cartilage cells with high density are directly planted above the bone bracket; 2) two scaffold materials suitable for bone and cartilage construction are adopted, tissue engineering bone and cartilage are formed by in vitro culture respectively, and then the tissue engineering bone and cartilage are assembled into a tissue engineering bone-cartilage complex by methods such as adhesion, surgical suture or sequential implantation; 3) the bone and the cartilage are all integrated single-layer scaffolds made of the same scaffold material; 4) the osteochondral part is an integrated double-layer bracket constructed by two different bracket materials.
The double-layered osteochondral scaffold has more excellent characteristics because its layered structure is designed according to the growth requirements of bone and cartilage. However, the bone cartilage biphasic scaffold has the following problems: 1) the upper layer cartilage material has poor mechanical property, absorbs water and deforms after being implanted into a body, and the degradation rate is high; 2) the cartilage after repair is fibrocartilage rather than hyaline cartilage; 3) the newly formed cartilage is poorly integrated with the surrounding cartilage tissue. The existing clinical products repair osteochondral defects by perfusion, have no through hole structure, cells cannot enter the inside of the bracket and only stay on the surface, and the repair effect is not good; in addition, scar tissue or ossification may occur; 4) the mechanical strength of the connecting part is not enough, and the connecting part is easy to separate; it is easy to prevent the penetration of the cartilage layer of the bone layer, resulting in cell migration and nutrient transport failure.
Disclosure of Invention
The invention provides an osteochondral scaffold and a preparation method and application thereof, aiming at solving the defects of insufficient cell capture and poor damage repair function of the osteochondral scaffold in the prior art. The osteochondral scaffold can fully capture cells, has strong osteochondral injury repair function and is simple and convenient to operate.
In order to achieve the purpose, the invention adopts the following technical scheme:
the invention provides a bone cartilage support, which comprises a cartilage layer, a bonding layer and a bone layer, wherein two sides of the bonding layer are respectively connected with the cartilage layer and the bone layer; one or more of the cartilage layer, the adhesive layer and the bone layer are porous structures.
In the present invention, preferably, the cartilage layer, the adhesion layer and the bone layer are all porous structures; more preferably, the hole of the cartilage layer and the hole of the adhesion layer are communicated with the hole of the bone layer. The holes of the cartilage layer, the adhesive layer and the bone layer may or may not be fully aligned, preferably fully aligned.
In the present invention, the diameter of the pores of the cartilage layer and/or the bone layer is 50 to 350 μm, preferably 200 to 280 μm, for example 250 μm; preferably, the pore diameter of the pores of the bone layer is equal to the pore diameter of the pores of the cartilage layer. The pore size of the cartilage layer and the bone layer are both selected to be suitable for capturing cells and cell growth.
In the present invention, the holes of the cartilage layer and/or the bone layer are preferably arranged in a vertically crossing manner.
In the present invention, the porosity of the cartilage layer and/or the bone layer is 20% to 70%, preferably 40% to 60%, for example 50%.
In the present invention, the osteochondral scaffold may have a porosity of 20% to 70%, preferably 40% to 60%, for example 50%.
In the present invention, the adhesion layer is a transition layer between the cartilage layer and the bone layer, and the connection between the cartilage layer and the bone layer can be achieved without being necessarily by adhesion.
Preferably, the adhesive layer does not cover or partially cover the pores of the bone layer and/or the cartilage layer. That is, the adhesive layer covers only a portion or all of the non-porous areas of the bone layer and the cartilage layer to ensure that the adhesive layer does not block the pores of the bone layer and the cartilage layer. The holes of the adhesion layer can be consistent with the holes of the cartilage layer and the holes of the bone layer, so that the three layers are communicated.
In the present invention, the shape of the osteochondral scaffold is not particularly limited, and the osteochondral scaffold may be cut according to the size of a defect site during use.
For example, the osteochondral scaffold is a cylinder. The diameter of the cylinder can be 2-30 mm, preferably 2-20 mm, and more preferably 3-10 mm; the height of the cylinder can be 2-10 mm, preferably 3-6 mm.
For example, the osteochondral scaffold is a cuboid. The bottom surface of the cuboid can be a square, and the side length of the square can be 2-30 mm, preferably 2-20 mm, and more preferably 3-10 mm; the height of the cuboid is preferably 2-10 mm, more preferably 3-6 mm.
In the present invention, the height ratio of the bone layer and the cartilage layer may be 1: (0.1 to 1), preferably 1: (0.2-0.5).
In the present invention, the height of the adhesion layer may be 5 μm to 2mm, preferably 0.1 to 2mm, and more preferably 0.5 to 1 mm.
In the present invention, the material of the cartilage layer may be a cartilage layer material conventional in the art, preferably a hydrogel material. Wherein the hydrogel material may be one or more of a single-network hydrogel material, an interpenetrating network hydrogel material, and a composite cross-linked hydrogel material. Hydrogel materials formed by a single cross-linking means are referred to as single network hydrogel materials. Hydrogel materials formed by two or more crosslinking modes are called interpenetrating network hydrogel materials, or double network hydrogel materials. The hydrogel material is prepared by compounding and crosslinking a plurality of gellable components in the same crosslinking mode, and is called a compound crosslinked hydrogel material. The hydrogel material is preferably a photo-crosslinked hydrogel material, and more preferably a composite photo-crosslinked hydrogel material.
In the present invention, the cartilage layer is preferably also loaded with a cartilage promoting component. Wherein the cartilage promoting component may comprise bioactive factors and/or cells. Wherein, the biological active factor preferably comprises transforming growth factor TGF alpha or TGF beta. The cells may include autologous or allogeneic chondrocytes, mesenchymal stem cells, embryonic stem cells or induced pluripotent stem cells.
In the present invention, the material of the bone layer may be medical polymer material conventional in the art, preferably polylactic acid (PLA), polylactic-co-glycolic acid (PLGA), or Polycaprolactone (PCL).
In the present invention, the material of the bone layer may also be a hydrogel material, which is as described above.
In the present invention, the bone layer is preferably also loaded with a bone promoting component. Wherein the bone promoting component may include one or more of a bioactive inorganic material, a bioactive factor, and a cell.
Wherein the bioactive inorganic material preferably comprises one or more of hydroxyapatite, calcium phosphate, calcium carbonate and bioactive glass. The mass percentage of the bioactive inorganic material in the bone layer can be 0.1 wt% to 70 wt%, preferably 1 wt% to 50 wt%, and more preferably 2.5 wt% to 30 wt%.
The bioactive factors preferably include one or more of the transforming growth factors TGF alpha, TGF beta, bone morphogenic proteins BMP-2, BMP-3, BMP-4, BMP-5, BMP-6, BMP-7, BMP-8, and BMP-9, and cartilage inducing compounds (e.g., KGN, etc.).
The cells may include autologous or allogeneic bone cells, mesenchymal stem cells, embryonic stem cells, or induced pluripotent stem cells.
In certain embodiments, the material of the adhesive layer may be a hydrogel material, as previously described.
In certain embodiments, the adhesive layer may be formed from medical glues conventional in the art. The medical glue may be selected from, for example, conpide, Greensea (Greensea), elephant, seamano, morbix, mollin, 3M, heptalen, fuaile, Idabao (IDEALPLAST), kaiki or aunov.
In the present invention, preferably, the cartilage layer, the adhesive layer and the bone layer are all hydrogel materials. At this time, the gellable component in the cartilage layer, the adhesive layer, and the bone layer is graded in concentration.
The invention also provides a preparation method of the osteochondral scaffold, which comprises the following steps: connecting the bone layer and the cartilage layer to form an adhesive layer at the joint; one or more of the cartilage layer, the adhesive layer and the bone layer are porous structures.
In the present invention, the preparation method of the cartilage layer may be conventional in the art, and is generally prepared by using a hydrogel composition as a raw material and performing crosslinking and curing.
In the present invention, the hydrogel composition refers to a hydrogel-forming raw material composition, and includes at least a gellable component and a gelling medium. The gellable component may be a component that can be cured to form a gel, as is conventional in the art, and typically includes natural gellable components and/or synthetic gellable components.
The natural gellable component may be conventional in the art and preferably comprises one or more of a natural protein, a natural protein modification, a natural protein degradation product, a modification of a natural protein degradation product, a natural polysaccharide modification, a natural polysaccharide degradation product and a modification of a natural polysaccharide degradation product.
The natural protein comprises one or more of various hydrophilic animal and plant proteins, water-soluble animal and plant proteins, type I collagen, type II collagen, serum protein, silk fibroin and elastin. The natural protein degradation product preferably comprises gelatin (Gel) or a polypeptide. The modified substance of the natural protein degradation product is preferably a methacrylated natural protein degradation product, and more preferably is methacrylated gelatin (GelMA).
The natural polysaccharide comprises one or more of Hyaluronic Acid (HA), carboxymethyl cellulose, methyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, sodium alginate, dextran, agarose, heparin, Chondroitin Sulfate (CS), ethylene glycol chitosan, propylene glycol chitosan, chitosan lactate, carboxymethyl chitosan and chitosan quaternary ammonium salt, preferably Hyaluronic Acid (HA) and/or Chondroitin Sulfate (CS). The natural polysaccharide modification is preferably a methacrylated natural polysaccharide, such as methacrylated hyaluronic acid (HAMA) or methacrylated Chondroitin Sulfate (CSMA).
Wherein the synthetic gellable component may be conventional in the art, and preferably comprises one or more of two-or multi-arm polyethylene glycol diacrylate, polyethyleneimine, a synthetic polypeptide, polyacrylic acid, polymethacrylic acid, polyacrylate, polymethacrylate, polyacrylamide, polymethacrylamide, polyvinyl alcohol, and polyvinylpyrrolidone.
In the present invention, the gellable component preferably includes methacrylated gelatin (GelMA) and methacrylated hyaluronic acid (HAMA); more preferably, it comprises methacrylated gelatin (GelMA), methacrylated hyaluronic acid (HAMA) and methacrylated Chondroitin Sulfate (CSMA).
In the present invention, the gel medium may be conventional in the art, and preferably is one or more of purified water, physiological saline, cell culture medium, calcium salt solution, and phosphate buffered saline (PBS solution). Wherein the normal saline is 0.9% NaCl aqueous solution. The cell culture medium may be a cell culture medium conventional in the art, such as DMEM, DMEM/F12, RPMI 1640, and the like, commonly used media. The phosphate buffer solution may be conventional in the art, and preferably has a pH of 7.4.
In the present invention, the amount of the gel medium may be conventional in the art, preferably such that in the hydrogel composition: 5 to 30 percent of methacryloylated gelatin (GelMA), 0.5 to 2 percent of methacryloylated hyaluronic acid (HAMA), 0.1 to 5 percent of methacryloylated Chondroitin Sulfate (CSMA) and 0.01 to 1 percent of photoinitiator; wherein the percentage is the mass (g) of the component contained per 100mL of the gel medium.
In the present invention, the hydrogel composition may further include a photoinitiator. Wherein, the photoinitiator can be a photoinitiator which is conventional in the field, and preferably is a blue light initiator, a UV light initiator or a green light initiator; the blue light initiator is preferably phenyl-2, 4, 6-trimethylbenzoyllithium phosphonate (LAP), riboflavin, flavin mononucleotide, eosin Y or terpyridine ruthenium chloride/sodium persulfate (Ru/SPS (1/10)); the UV photoinitiator is preferably 2-hydroxy-2-methyl-1- [4- (2-hydroxyethoxy) phenyl ] -1-propanone (I2959).
In the present invention, the hydrogel composition may further include a thickener, and the amount of the thickener is preferably 0.1 to 25 parts. The thickener may be conventional in the art, and is preferably one or more of polyethylene oxide (PEO), polyethylene glycol (PEG), sodium alginate (Alg), hyaluronic acid, polyvinylpyrrolidone, gum arabic, gellan gum, and xanthan gum.
When the thickening agent comprises sodium alginate, the using amount of the sodium alginate is preferably 0.5-2 parts. When the thickener includes hyaluronic acid, the amount of hyaluronic acid is preferably 0.5 to 2 parts. When the thickener includes polyvinylpyrrolidone, the polyvinylpyrrolidone is preferably used in an amount of 2 to 10 parts. When the thickener includes gum arabic, the amount of the gum arabic is preferably 0.1 to 25 parts. When the thickener comprises gellan gum, the amount of the gellan gum is preferably 0.1 to 2 parts. When the thickener includes xanthan gum, the amount of the xanthan gum is preferably 0.1 to 5 parts.
In the present invention, the hydrogel composition may further include a synthetic type photosensitive material. The amount of the synthetic photosensitive material is preferably 5 to 30 parts. The synthetic type photosensitive material may be conventional in the art, and preferably includes one or more of polyethylene glycol acrylate (PEGDA), polyacrylic acid, polymethacrylic acid, polyacrylate, polymethacrylate, polyacrylamide, and polymethacrylamide. The synthetic photosensitive material is preferably polyethylene glycol acrylate.
In a preferred embodiment of the present invention, the hydrogel composition of the cartilage layer comprises the following components in parts by weight: 1-50 parts of methacryloylated gelatin, 0-30 parts of methacryloylated hyaluronic acid, 0-30 parts of methacryloylated chondroitin sulfate, 0.01-1 part of photoinitiator and a gel medium.
The amount of the methacrylated gelatin is preferably 1 to 30 parts, more preferably 1 to 20 parts, still more preferably 2 to 15 parts, still more preferably 5 to 15 parts, such as 8 parts, 10 parts or 12 parts.
The methacrylated gelatin may be conventional in the art, commercially available, or may be obtained by methacrylating gelatin (Gel) by methods conventional in the art.
The degree of methacrylation of the methacrylated gelatin is preferably 40% to 80%. Wherein, the degree of methacryloylation of the methacryloylated gelatin is calculated by using a nuclear magnetic resonance hydrogen spectrum (1H NMR), and specifically comprises the following components: selecting the integral area of a phenylalanine standard peak (7.1-7.4 ppm) as 1, and calculating the percentage of the peak area reduction of the lysine signal at the position of 2.8-2.95 ppm before and after gelatin modification, namely:
GelMA (peak area of lysine signal at 2.8-2.95 ppm in GelMA-peak area of lysine signal at 2.8-2.95 ppm in GelMA)/Gel peak area of lysine signal at 2.8-2.95 ppm in GelMA is 100%.
The amount of the methacryloylated hyaluronic acid is preferably 0.1 to 20 parts, more preferably 0.5 to 10 parts, still more preferably 1 to 3 parts, such as 1.5 parts or 2 parts.
The methacrylated hyaluronic acid may be conventional in the art, commercially available, or may be obtained by methacrylating Hyaluronic Acid (HA) by methods conventional in the art. The molecular weight of the methacryloylated hyaluronic acid can be 1-8000 kDa, preferably 100-1000 kDa, and more preferably 500-950 kDa.
The degree of methacryloylation of the methacryloylated hyaluronic acid is 20% to 60%, preferably 30% to 50%. Wherein the degree of methacryloylation of the methacryloylated hyaluronic acid is determined by nuclear magnetic resonance hydrogen spectroscopy (1H NMR), specifically:
the degree of methacrylation of HAMA ═ peak area at 5.6ppm for methacrylamide-vinyl group/peak area at 1.9ppm for N-acetylglucose, × 100%.
The amount of the methacryloylated chondroitin sulfate is preferably 0.1 to 20 parts, more preferably 0.5 to 20 parts, still more preferably 0.5 to 5 parts, and still more preferably 1 to 3 parts, such as 1 part, 2 parts, 2.5 parts, or 3 parts.
The methacrylated chondroitin sulfate can be obtained by conventional methods in the field, can be obtained commercially, and can also be obtained by performing methacrylation on Chondroitin Sulfate (CS) by the conventional methods in the field. The molecular weight of the methacryloylated chondroitin sulfate can be 5-50 kDa, and preferably 10-40 kDa.
The degree of methacrylation of the methacrylated chondroitin sulfate is preferably 30% to 50%. Wherein, the degree of methacryloylation of the methacryloylated chondroitin sulfate is calculated by using a nuclear magnetic resonance hydrogen spectrum (1H NMR), and specifically comprises the following components:
the degree of methacrylation of CSMA ═ peak area at 5.6ppm of methacrylamide-vinyl group/peak area at 1.9ppm of N-acetylglucose, × 100%.
Wherein the mass ratio of the methacrylated gelatin to the methacrylated hyaluronic acid may be (1-30): (0.5 to 10), preferably (2 to 15): (1-3), for example, 5: 2.
Wherein the mass ratio of the methacrylated gelatin to the methacrylated hyaluronic acid to the methacrylated chondroitin sulfate can be (1-30): (0.5-10): (0.5 to 20), preferably (2 to 15): 1: (1-3), for example, 10:1:3, 5:2:2, 15:1:1 or 8:1: 3.
In the present invention, preferably, the hydrogel composition of the cartilage layer comprises the following components in parts by weight:
5-15 parts of methacryloylated gelatin (GelMA),
0.5 to 2 parts of methacryloylated hyaluronic acid (HAMA),
and 0.1-0.5 part of photoinitiator.
In the present invention, preferably, the hydrogel composition of the cartilage layer comprises the following components in parts by weight:
5-15 parts of methacryloylated gelatin (GelMA),
0.5 to 2 parts of methacryloylated hyaluronic acid (HAMA),
0.5 to 3 parts of methacryloylated Chondroitin Sulfate (CSMA),
and 0.1-0.5 part of photoinitiator.
In the present invention, preferably, the hydrogel composition of the cartilage layer comprises the following components in parts by weight:
5-15 parts of methacryloylated gelatin (GelMA),
1-2 parts of sodium alginate (Alg),
and 0.1-0.5 part of photoinitiator.
In a specific embodiment of the invention, the hydrogel composition of the cartilage layer comprises the following components in parts by weight:
5 parts of methacrylated gelatin (GelMA),
2 portions of methacrylated hyaluronic acid (HAMA),
2 portions of methacryloylated Chondroitin Sulfate (CSMA),
and 0.25 part of a photoinitiator.
In a specific embodiment of the invention, the hydrogel composition of the cartilage layer comprises the following components in parts by weight:
10 parts of methacrylated gelatin (GelMA),
1 part of methacrylated hyaluronic acid (HAMA),
3 portions of methacryloylated Chondroitin Sulfate (CSMA),
and 0.25 part of a photoinitiator.
In a specific embodiment of the invention, the hydrogel composition of the cartilage layer comprises the following components in parts by weight:
15 parts of methacrylated gelatin (GelMA),
1 part of methacrylated hyaluronic acid (HAMA),
1 part of methacryloylated Chondroitin Sulfate (CSMA),
and 0.25 part of a photoinitiator.
In a specific embodiment of the invention, the hydrogel composition of the cartilage layer comprises the following components in parts by weight:
8 parts of methacrylated gelatin (GelMA),
1 part of methacrylated hyaluronic acid (HAMA),
3 portions of methacryloylated Chondroitin Sulfate (CSMA),
and 0.25 part of a photoinitiator.
In a specific embodiment of the invention, the hydrogel composition of the cartilage layer comprises the following components in parts by weight:
8 parts of methacrylated gelatin (GelMA),
1 part of methacrylated hyaluronic acid (HAMA),
3 portions of methacryloylated Chondroitin Sulfate (CSMA),
2 parts of sodium alginate, namely 2 parts of sodium alginate,
and 0.25 part of a photoinitiator.
In a specific embodiment of the invention, the hydrogel composition of the cartilage layer comprises the following components in parts by weight:
8 parts of methacrylated gelatin (GelMA),
1 part of methacrylated hyaluronic acid (HAMA),
3 portions of methacryloylated Chondroitin Sulfate (CSMA),
10 parts of PEGDA, namely 10 parts of PEGDA,
and 1 part of photoinitiator.
In a specific embodiment of the invention, the hydrogel composition of the cartilage layer comprises the following components in parts by weight:
5 parts of methacrylated gelatin (GelMA),
2 portions of methacrylated hyaluronic acid (HAMA),
and 0.5 part of photoinitiator.
In a specific embodiment of the invention, the hydrogel composition of the cartilage layer comprises the following components in parts by weight:
5 parts of methacrylated gelatin (GelMA),
2 parts of sodium alginate, namely 2 parts of sodium alginate,
and 0.5 part of photoinitiator.
In the present invention, the crosslinking curing means may include one or more of physical crosslinking, chemical crosslinking, enzymatic crosslinking, and photo-crosslinking; preferably including photo-crosslinking.
Wherein, the physical crosslinking can be carried out by the conventional method in the field, such as self-assembly solidification of collagen at about 37 ℃. Wherein the chemical crosslinking can be performed by a method conventional in the art, for example, a methacrylated material forms a gel under catalysis of ammonium persulfate, or sodium alginate forms a gel by crosslinking with a divalent metal cation.
Wherein the enzymatic crosslinking may be performed using methods conventional in the art, e.g., fibrinogen is catalyzed by a protease to form a gel.
Wherein the photo-crosslinking can be performed under light irradiation using a method conventional in the art; preferably, the photo-crosslinking has a wavelength of 365-405 nm and an intensity of 5-50 mW/cm2Under light irradiation; more preferably, the photocrosslinking is carried out at a wavelength of 405nm and an intensity of 10mW/cm2Is carried out under light irradiation.
In the invention, the preparation method of the cartilage layer can be a perfusion method, and comprises the following steps: pouring the hydrogel composition of the cartilage layer into a cartilage mould for photocrosslinking to obtain cured hydrogel; and (3) freeze-drying the solidified hydrogel to obtain the hydrogel.
Wherein the cartilage mold can be designed according to the shape and size of the required cartilage by adopting the conventional method in the field.
In the invention, the preparation method of the cartilage layer can be a direct 3D printing method, which comprises the following steps: 3D printing is carried out on the hydrogel composition of the cartilage layer, and meanwhile, photocrosslinking is carried out, so that cured hydrogel is obtained; and (3) freeze-drying the solidified hydrogel to obtain the hydrogel.
In a preferred embodiment, the method for preparing the cartilage layer comprises the following steps: printing the hydrogel composition by adopting an extrusion type 3D printer, and curing and forming by using blue light, wherein the heat preservation temperature is 30-37 ℃, the printing environment temperature is 22-25 ℃, the printing pressure is 20-40 PSI, the printing speed is 4-8mm/s, the filling rate is 40-60%, and the illumination intensity is 5-20 mW/cm2。
In the invention, the preparation method of the cartilage layer can be an indirect 3D printing method, also called a 3D printing demolding method or a 3D engineering method, and comprises the following steps:
s1, 3D printing is carried out on the sacrificial material to obtain a cartilage layer mold; wherein the sacrificial material is a hard polymer material which can be dissolved in a solvent;
s2, pouring the hydrogel composition of the cartilage layer into the cartilage layer mould for crosslinking to obtain a solidified hydrogel-cartilage layer mould complex;
s3, demolding: dissolving the cartilage layer mould by using a solvent to obtain cured hydrogel;
s4, freezing and drying the obtained solidified hydrogel to obtain the cartilage layer.
Wherein the hard polymer material is defined as: 3D printing is carried out on the high polymer material by taking the cubic support with the size of 10 x 10mm as a target, the size error of the actually formed support is within 10%, and the high polymer material can be called as a hard high polymer material. The cubic scaffold is a standard for determining whether the polymer material is a hard polymer material, and does not limit the shape that the material can be formed into. Adopt stereoplasm macromolecular material to carry out 3D and print, the form maintains well, can realize high accuracy and print.
Conventional sacrificial materials commonly used in the industry at present, such as pluronic, carbomer, gelatin particles, sucrose, etc., cannot maintain morphology during 3D printing, and cannot achieve fine printing. The hard polymer material which does not conform to the invention is PEEK (polyether ether ketone), PEKK (polyether ketone), PEI (polyetherimide) or PPSU (high performance medical grade plastic), for example, the materials can be printed in an FDM mode, the stability is very good, the materials are not suitable for being used as sacrificial materials, and the materials cannot be removed under the conventional conditions.
Wherein the sacrificial material is preferably biocompatible. The "biocompatible" criteria are: the cell viability was above 75% by biocompatibility testing using methods conventional in the art. The sacrificial material is preferably transparent or translucent.
Wherein, the sacrificial material is preferably polylactic acid (PLA), Polycaprolactone (PCL), polyethylene terephthalate-1, 4-cyclohexane dimethanol ester (PETG), polyvinyl alcohol (PVA) or synthetic photosensitive resin. The synthetic photosensitive resin is preferably a polyacrylate type photosensitive resin.
In step S1, preferably, the pigment is mixed in the sacrificial material, and then the colored sacrificial material is subjected to 3D printing to obtain a colored mold. This has the effect that, when the mold is removed in step S3, the disappearance of the color can be used as an indicator for monitoring the success of the mold removal.
In step S1, the 3D printing method may be a printing method that is conventional in the art and can realize a precise fine structure, and is preferably an extrusion method (i.e., a melt-deposition method) or a photo-curing method. The light curing mode can be a three-dimensional light curing molding technology (SLA), a digital light projection technology (DLP) or a liquid crystal display technology (LCD).
In step S1, the shape, size and structure of the cartilage layer mold may be designed according to the desired cartilage layer according to conventional methods in the art.
In step S2, a suitable crosslinking method may be selected according to the composition of the hydrogel composition, and the operation and conditions of crosslinking are as described above.
In step S3, the solvent may be selected according to the characteristics of the sacrificial material forming the mold, and may dissolve the mold. The solvent is preferably dichloromethane, chloroform, tetrahydrofuran, 1, 4-dioxane, purified water, physiological saline, calcium salt solution, Phosphate Buffered Saline (PBS), or a culture medium.
In step S4, the freeze-drying time is preferably 8-24 hours; preferably a pre-cooling step is performed prior to said freeze-drying; the pre-cooling temperature is preferably-20 ℃, and the pre-cooling time is preferably 1-3 h.
In the present invention, the method for preparing the cartilage layer preferably further comprises the step of loading a cartilage-promoting component. The method of loading the cartilage promoting component may be conventional in the art. The cartilage promoting component is as previously described. When the cartilage-promoting component is a bioactive factor, the bioactive factor is typically loaded by soaking the cartilage layer in a bioactive factor solution, wherein the concentration of the bioactive factor solution may be 1 μ g/mL to 200 μ g/mL, preferably 5 μ g/mL to 100 μ g/mL.
In the present invention, the preparation method of the bone layer may be conventional in the art, and preferably, the material of the bone layer is subjected to 3D printing. Wherein the material of the bone layer is as described above. The parameters of the 3D printing may be selected according to the structure and material of the bone layer using methods conventional in the art. The 3D printing is preferably performed using a fused deposition 3D printer.
In the present invention, preferably, the method for preparing the bone layer further comprises the step of loading a bone-promoting component. The methods of loading the bone promoting component may be conventional in the art, for example: grinding the material of the bone layer into powder and then adding the bone promoting component; alternatively, the material of the bone layer is dissolved in a solvent, and then the bone promoting ingredient is incorporated, and then the solvent is volatilized. The bone promoting component is as previously described. The solvent may be an organic solvent. When the bone-promoting component is a bioactive factor, the bioactive factor is typically loaded by soaking the bone layer in a bioactive factor solution, wherein the concentration of the bioactive factor solution may be 1 μ g/mL to 200 μ g/mL, preferably 5 μ g/mL to 100 μ g/mL.
In a preferred embodiment, the bone layer is provided by performing 3D printing on the hydroxyapatite-loaded PLA by using a fused deposition 3D printer, wherein the printing head temperature is 210 ℃, the platform temperature is 50-60 ℃, the printing speed is 50-60 mm/s, and the filling rate is 40% -60%.
In a preferred embodiment, the bone layer is provided by performing 3D printing on the PCL loaded with tricalcium phosphate by using a fused deposition type 3D printer, wherein the printing temperature is 140 to 150 ℃, the printing speed is 50 to 60mm/s, and the filling rate is 40 to 60%.
In a preferred embodiment, the bone layer is provided by 3D printing of GelMA loaded with tricalcium phosphate using an extrusion photocuring 3D printer, wherein the temperature is 30-35 ℃, the printing environment temperature is 22-25 ℃, the printing pressure is 20-40 PSI, the printing speed is 4-8mm/s, the filling rate is 40-60%, and the illumination intensity is 5-50 mW/cm2。
In the present invention, the method for preparing the bone layer may be a method in which the hydrogel composition is used as a raw material and is cross-linked and cured. When the bone layer is prepared by cross-linking and curing the hydrogel composition as a raw material, the hydrogel composition and the cross-linking and curing are as described above.
In a preferred embodiment, the hydrogel composition of the bone layer comprises the following components: 5 to 30 percent of methylacryloylated gelatin (GelMA), 2.5 to 50 percent of bioactive glass and 0.01 to 1 percent of photoinitiator; wherein the percentages are mass (g) of the components contained per 100mL of the gel medium.
In the present invention, the connection is generally made by connecting the bone layer and the cartilage with a medical glue. The operation condition of the connection can be determined according to the use method of the medical glue. The medical glue is as described above.
In some embodiments, the osteochondral scaffold is formed integrally by a 3D printing demolding method, which specifically includes the following steps:
s1, 3D printing is carried out on the sacrificial material to obtain the osteochondral scaffold mold; wherein the sacrificial material is a hard polymer material which can be dissolved in a solvent;
s2, sequentially pouring hydrogel compositions of a bone layer, a bonding layer and a cartilage layer into the osteochondral scaffold mold for crosslinking to obtain a cured hydrogel-osteochondral scaffold mold complex;
s3, demolding: dissolving the osteochondral scaffold mould by using a solvent to obtain cured hydrogel;
s4, freezing and drying the obtained solidified hydrogel to obtain the osteochondral scaffold.
In the above embodiment, preferably, the hydrogel composition of the bone layer comprises the following components: 5 to 30 percent of methylacryloylated gelatin (GelMA), 2.5 to 50 percent of bioactive glass and 0.01 to 1 percent of photoinitiator; the hydrogel composition of the adhesive layer comprises the following components: 5 to 30 percent of methylacryloylated gelatin (GelMA), 10 to 20 percent of bioactive glass and 0.01 to 1 percent of photoinitiator; the hydrogel composition of the cartilage layer comprises the following components: 5 to 30 percent of methacrylated gelatin (GelMA), 0.5 to 2 percent of methacrylated hyaluronic acid (HAMA), 0.5 to 5 percent of methacrylated Chondroitin Sulfate (CSMA) and 0.01 to 1 percent of photoinitiator; wherein the percentage is the mass (g) of the component contained per 100mL of the gel medium.
In the above embodiments, the sacrificial material, the hard polymer material, the 3D printing manner, the crosslinking operation and conditions, the solvent, and the freeze-drying are as described above.
The invention also provides the osteochondral scaffold prepared by the preparation method.
The invention also provides application of the osteochondral scaffold in repairing cartilage defects.
In the present invention, the cartilage defect may be a osteochondral complex defect or a simple cartilage defect. The osteochondral defect may be located in a knee joint, a hip joint, or a shoulder joint.
In repairing the osteochondral composite defect, the osteochondral scaffold may be used in the following manner: drilling a hole downwards to a bone layer at the osteochondral composite defect, removing redundant bone layer matrix after reaching a marrow cavity, and putting the osteochondral scaffold material at the osteochondral composite defect. At this time, bone marrow flowing out of the bone layer is rich in mesenchymal stem cells, and bone marrow-derived mesenchymal stem cells are captured in the scaffold while flowing through the osteochondral scaffold material. After the operation is finished, the wound is sutured, and the repair of the osteochondral complex defect is finished.
In repairing a simple cartilage defect, the osteochondral scaffold may be used in the following manner: and placing the cartilage layer of the osteochondral scaffold at the defect, and carrying out microfracture treatment at the cartilage defect until bone marrow flows out. At this time, bone marrow emerging from the bone layer is rich in mesenchymal stem cells, and the cells are captured in the scaffold while flowing through the osteochondral scaffold. After the operation is finished, the wound is sutured, and the repair of the simple cartilage defect is finished.
On the basis of the common knowledge in the field, the above preferred conditions can be combined randomly to obtain the preferred embodiments of the invention.
The reagents and starting materials used in the present invention are commercially available.
The positive progress effects of the invention are as follows:
1. the osteochondral scaffold provided by the invention has a refined through hole structure, ensures the vertical and horizontal through, is favorable for fully capturing cells when the scaffold is used for filling osteochondral defects, and is also favorable for transporting nutrient substances and metabolic wastes, thereby being favorable for defect repair. Furthermore, the stem cells can be induced to differentiate and grow into the chondrocytes and the osteocytes by selecting proper cartilage layer materials and bone layer materials.
2. The osteochondral scaffold has an excellent adhesive layer structure, and can firmly connect a cartilage layer and a bone layer to achieve the purpose of integrated integration; the adhesive layer is used as the transition layer of the bone layer to prevent ossification of the cartilage layer. The osteochondral scaffold can realize osteochondral full-layer repair.
3. The osteochondral scaffold has simple and convenient clinical operation mode and practicability, and provides a brand-new and effective solution for repairing cartilage defect and osteochondral composite defect clinically.
Drawings
FIG. 1 is a schematic structural view of an osteochondral scaffold according to embodiments 1 to 4 of the present invention.
Fig. 2 is a camera photograph of the osteochondral scaffold of example 1 of the present invention.
FIG. 3 is a photomicrograph of an osteochondral scaffold according to example 1 of the present invention.
FIG. 4 is a microphotograph of MSC cells cultured on osteochondral scaffolds in example 1, which is an effect of the present invention.
FIG. 5 is a photograph showing a general observation of a damaged joint in group (a) in example 2 of the effect of the present invention.
FIG. 6 is a photograph showing a general observation of a damaged joint in group (b) in example 2 in which the present invention is effective.
FIG. 7 is a photograph showing a general observation of a damaged joint in group (c) in example 2 in which the effect of the present invention is exhibited.
Detailed Description
The invention is further illustrated by the following examples, which are not intended to limit the scope of the invention. The experimental methods without specifying specific conditions in the following examples were selected according to the conventional methods and conditions, or according to the commercial instructions.
Example 1
1. Preparation of cartilage layer
(1) Synthesis of methacryloylated gelatin (GelMA): dissolving gelatin (1g) in 10mL of PBS (pH 7.4), heating to 50 ℃, stirring until the gelatin is completely dissolved, adding 0.5mL of methacrylic anhydride, reacting for 2-3 h, diluting the reaction solution with 40mL of PBS after the reaction, pouring the reaction solution into a dialysis bag (MWCO7000), dialyzing for 2-3d with deionized water, and freeze-drying to obtain methacrylated gelatin (0.9 g). According to nuclear magnetic resonance hydrogen spectrum (1H NMR), selecting the integral area of a phenylalanine standard peak (7.1-7.4 ppm) as 1, and calculating the percentage of the peak area reduction of the lysine signal at the position of 2.8-2.95 ppm before and after gelatin modification to obtain the methacryloylation degree of the methacryloylated gelatin of 65%.
(2) Methyl propylSynthesis of enoylated hyaluronic acid (HAMA): dissolving hyaluronic acid (1g, 900kDa) in 100mL deionized water, cooling to 0-4 ℃, adding 5mL methacrylic anhydride, slowly dropwise adding 5mL 5M NaOH aqueous solution, reacting for 24h, pouring the reaction solution into a dialysis bag (MWCO7000), dialyzing for 2-3d with deionized water, and freeze-drying to obtain methacrylated hyaluronic acid (0.9 g). According to nuclear magnetic resonance hydrogen spectrum (1H NMR), the degree of methacrylation of HAMA was calculated to be 40% (degree of methacrylation of HAMA ═ peak area of methacrylamide-vinyl at 5.6 ppm/peak area of N-acetylglucose at 1.9ppm ×. 100%).
(3) Formulation of hydrogel composition of 5% GelMA/2% HAMA/0.5% LAP: 0.05g of GelMA, 0.02g of HAMA and 5mg of LAP were weighed out and dissolved in 1mL of PBS solution (pH 7.4) to prepare 5% GelMA/2% HAMA/0.5% LAP at 37 ℃.
(4) Preparing a GelMA/HAMA cartilage layer by an infusion method:
the hydrogel composition was poured into a preformed cylindrical mold (diameter 5mm, height 3mm) at a wavelength of 405nm and an intensity of 10mW/cm2The light source is used for realizing photo-crosslinking, and then GelMA/HAMA photo-crosslinking curing hydrogel can be obtained; and (3) placing the prepared GelMA/HAMA photo-crosslinking curing hydrogel in a refrigerator at the temperature of-20 ℃ for freezing for 2h, and then freeze-drying by using a freeze dryer to obtain the GelMA/HAMA cartilage layer.
(5) Loading of TGF β: soaking the GelMA/HAMA cartilage layer in 10 microgram/mL TGF beta solution, after fully adsorbing for 12h, placing the obtained product in a refrigerator at the temperature of-20 ℃ for freezing for 2h, and then freeze-drying by using a freeze dryer to obtain the TGF beta-loaded cartilage layer which is marked as the TGF beta-GelMA/HAMA cartilage layer.
2. Preparation of bone layer
(1) Preparation of hydroxyapatite-loaded polylactic acid polymer material (HAP/PLA)
Weighing 5g of polylactic acid (PLA) to be dissolved in 20mL of dichloromethane until the PLA is completely dissolved, adding 1g of Hydroxyapatite (HAP), stirring to obtain a uniform solution, placing the solution in a fume hood to volatilize the dichloromethane, placing the solution in a vacuum drying oven to dry for 24h, crushing the solution into powder by a crusher, and preparing the powder into a wire rod with the diameter of 1.75mm by a wire drawing machine to obtain the HAP/PLA high polymer material.
(2)3D printing HAP/PLA high polymer material
The HAP/PLA high polymer material is printed and molded by a fused deposition type 3D printer (the printing temperature is 210 ℃, the platform temperature is 50 ℃, the printing speed is 60mm/s, the filling rate is 50%, and the layer height is 0.1mm) to obtain the HAP/PLA bone layer. The resulting HAP/PLA bone layer was cylindrical (5 mm diameter, 3mm height), had a porosity of 50% and a pore size of 250 μm.
3. Preparation of osteochondral scaffolds
Connecting the TGF beta-GelMA/HAMA cartilage layer with the HAP/PLA bone layer by medical glue colloidal gold elephant, forming a bonding layer at the joint, wherein the thickness of the bonding layer is about 100 mu m, thus obtaining the TGF beta-GelMA/HAMA-HAP/PLA osteochondral scaffold, the structural schematic diagram of which is shown in figure 1, and the TGF beta-GelMA/HAMA-HAP/PLA cartilage scaffold comprises a cartilage layer 1, a bonding layer 2 and a bone layer 3 from top to bottom; the picture of the object camera is shown in figure 2; the microstructure was observed by microscope as shown in FIG. 3.
In this example, if TGF β loading is not performed, the GelMA/HAMA-HAP/PLA osteochondral scaffold is obtained.
Example 2
1. Preparation of cartilage layer
(1) Synthesis of methacryloylated gelatin (GelMA): the same as in example 1.
(2) Synthesis of methacrylated hyaluronic acid (HAMA): the same as in example 1.
(3) Formulation of 5% GelMA/2% HAMA/0.5% LAP hydrogel composition: the same as in example 1.
(4) Preparing a cartilage layer by a direct 3D printing method:
printing the hydrogel composition by using an extrusion type 3D printer, and carrying out blue light curing molding (the heat preservation temperature is 37 ℃, the platform temperature is 22 ℃, the printing pressure is 20PSI, the printing speed is 5mm/s, and the filling rate is 50%); and (3) placing the prepared GelMA/HAMA photo-crosslinking curing hydrogel in a refrigerator at the temperature of-20 ℃ for freezing for 2h, and then freeze-drying by using a freeze dryer to obtain the GelMA/HAMA cartilage layer. The resulting GelMA/HAMA cartilage layer was cylindrical (diameter 5mm, height 1mm), had a porosity of 50% and a pore size of 300. mu.m.
(5) Loading of TGF β: the same as in example 1.
2. Preparing a bone layer: the same as in example 1.
3. Preparation of osteochondral scaffolds
The TGF beta-GelMA/HAMA cartilage layer and the HAP/PLA bone layer are connected by medical glue colloidal gold elephant, an adhesive layer is formed at the connecting part, the thickness of the adhesive layer is about 100 mu m, and the TGF beta-GelMA/HAMA-HAP/PLA osteochondral scaffold is obtained, and the structural schematic diagram of the TGF beta-GelMA/HAMA-HAP/PLA osteochondral scaffold is shown in figure 1.
In this example, if TGF β loading is not performed, the GelMA/HAMA-HAP/PLA osteochondral scaffold is obtained.
Example 3
1. Preparation of cartilage layer
(1) Synthesis of methacryloylated gelatin (GelMA): the same as in example 1.
(2) Preparing a sodium alginate/gelatin composite cross-linked hydrogel composition: 0.02g of sodium alginate (Alg), 0.05g of GelMA and 5mg of LAP were weighed out and dissolved in 1mL of PBS solution (pH 7.4) to prepare a hydrogel composition of 2% Alg/5% GelMA/0.5% LAP at 37 ℃.
(3) Pouring the hydrogel composition into a preformed mold at a wavelength of 405nm and an intensity of 10mW/cm2The light source realizes photocrosslinking; removing the shaped hydrogel from the mold in 0.1M CaCl2Soaking for 2h to realize chemical crosslinking, and obtaining the composite photo-crosslinking curing hydrogel; and then, placing the prepared composite photo-crosslinking curing hydrogel in a refrigerator at the temperature of-20 ℃ for freezing for 2h, and then freeze-drying by using a freeze dryer to obtain the Alg/GelMA cartilage layer.
(4) Loading of Mesenchymal Stem Cells (MSCs): digesting MSC with pancreatin, centrifuging to collect cells, dripping cell suspension on the above Alg/GelMA cartilage layer, incubating for 1h, adding culture medium, and culturing at 37 deg.C/5% CO2Culturing in a cell culture box under the conditions for 7d to obtain the MSC-Alg/GelMA cartilage layer.
2. Preparation of bone layer
(1) Preparation of tricalcium phosphate (TCP) -loaded polycaprolactone polymer material (TCP/PCL)
Weighing 5g of Polycaprolactone (PCL) to be dissolved in 20mL of dichloromethane until the Polycaprolactone (PCL) is completely dissolved, adding 1g of tricalcium phosphate (TCP), stirring to obtain a uniform solution, placing the solution in a fume hood to be dried for 24 hours in a vacuum drying oven after dichloromethane is volatilized, and crushing the solution by a crusher to obtain powder, thus preparing the TCP/PCL high polymer material.
(2)3D printing of a TCP/PCL high polymer material: the TCP/PCL high polymer material is printed and molded by a fused deposition type 3D printer (printing temperature: 140 ℃, printing pressure: 40PSI, printing speed: 50mm/s, filling rate: 50%) to obtain the TCP/PCL bone layer. The obtained TCP/PCL bone layer is a cylinder (diameter 5mm, height 3mm), porosity is 50%, and pore diameter is 300 μm.
3. Preparation of osteochondral scaffolds
Connecting the MSC-Alg/GelMA cartilage layer and the TCP/PCL cartilage layer by medical glue, and forming an adhesive layer at the joint, wherein the thickness of the adhesive layer is about 100 mu m, so that the MSC-Alg/GelMA-TCP/PCL cartilage scaffold is obtained, and the structural schematic diagram is shown in figure 1.
In this embodiment, if MSC loading is not performed, the Alg/GelMA-TCP/PCL osteochondral scaffold is obtained.
Example 4
1. Preparation of cartilage layer
(1) Synthesis of methacryloylated gelatin (GelMA): the same as in example 1.
(2) Preparing a sodium alginate/gelatin composite cross-linked hydrogel composition: the same as in example 3.
(3) Preparing an Alg/GelMA cartilage layer by a 3D printing demolding method:
s1, designing a proper cartilage layer mold according to the cartilage layer, and carrying out 3D printing molding on the polyacrylate photosensitive resin;
s2, pouring the hydrogel composition into the cartilage layer mould at the wavelength of 405nm and the intensity of 10mW/cm2Under the irradiation of the light source, the in-situ photocrosslinking is realized, and then 0.1M CaCl is added2Soaking for 2h to realize chemical crosslinking to obtain an Alg/GelMA composite photo-crosslinking cured hydrogel-cartilage layer mold complex;
s3, demolding: dissolving a cartilage layer mold in the Alg/GelMA composite photo-crosslinking curing hydrogel-cartilage layer mold complex by using dichloromethane to obtain Alg/GelMA composite photo-crosslinking curing hydrogel;
s4, placing the prepared Alg/GelMA composite photo-crosslinking curing hydrogel in a refrigerator at the temperature of-20 ℃ for freezing for 2h, and then freeze-drying by using a freeze dryer to obtain the Alg/GelMA cartilage layer. Wherein the obtained Alg/GelMA cartilage layer is cylindrical (diameter is 5mm, height is 1mm), porosity is 50%, and pore diameter is 250 μm.
(4) Loading of Mesenchymal Stem Cells (MSCs): the same as in example 3.
2. Preparing a bone layer: the same as in example 3.
3. Preparation of osteochondral scaffolds
Connecting the MSC-Alg/GelMA cartilage layer and the TCP/PCL cartilage layer by medical glue, and forming an adhesive layer at the joint, wherein the thickness of the adhesive layer is about 100 mu m, so that the MSC-Alg/GelMA-TCP/PCL cartilage scaffold is obtained, and the structural schematic diagram is shown in figure 1.
In this embodiment, if MSC loading is not performed, the Alg/GelMA-TCP/PCL osteochondral scaffold is obtained.
Example 5
1. Synthesis of gellable Components
(1) Synthesis of methacryloylated gelatin (GelMA): the same as in example 1.
(2) Synthesis of methacrylated hyaluronic acid (HAMA): the same as in example 1.
(3) Synthesis of methacryloylated Chondroitin Sulfate (CSMA): dissolving chondroitin sulfate (10g, 30kDa) in 100mL of deionized water, cooling to 0-4 ℃, adding 50mL of methacrylic anhydride, slowly dropwise adding 50mL of 5M NaOH aqueous solution, reacting for 24 hours, pouring the reaction solution into a dialysis bag (MWCO7000), dialyzing for 2-3 days with deionized water, and freeze-drying to obtain the methacryloyl chondroitin sulfate (9 g). From nuclear magnetic resonance hydrogen spectroscopy (1H NMR), the degree of methacryloylation of CSMA was calculated to be 40% (degree of methacryloylation of CSMA ═ peak area of methacrylamide-vinyl at 5.6 ppm/peak area of N-acetyl glucose at 1.9ppm ×. 100%).
2. Formulating hydrogel compositions
(1) Hydrogel composition of cartilage layer: weighing 20g of methacrylated gelatin (GelMA), 1g of methacrylated hyaluronic acid (HAMA) and 1g of methacrylated Chondroitin Sulfate (CSMA), dissolving in deionized water at 50 ℃, and adding 0.1g of initiator LAP to prepare hydrogel of the cartilage layer, wherein the percentage is the mass (g) of the components contained in each 100mL of gel medium;
(2) hydrogel composition of transition layer: weighing 20g of methacrylated gelatin (GelMA) and 10g of bioactive glass, dissolving in deionized water at 50 ℃, and adding 0.25g of initiator LAP to prepare hydrogel of the transition layer; wherein the percentage is the mass (g) of the component contained per 100mL of the gel medium;
(3) hydrogel composition of bone layer: weighing 20g of methacrylated gelatin (GelMA) and 50g of bioactive glass, dissolving in deionized water at 50 ℃, and adding 0.2g of initiator LAP to prepare hydrogel of a bone layer; wherein the percentages are mass (g) of the components contained per 100mL of the gel medium.
3. 3D printing demolding method integrated forming
S1, designing a proper osteochondral scaffold mold according to the osteochondral scaffold, and printing and molding polyvinyl alcohol (PVA) in a 3D mode;
s2, sequentially pouring the hydrogel composition of the bone layer, the adhesive layer and the cartilage layer into the osteochondral scaffold die, and controlling the wavelength at 405nm and the intensity at 10mW/cm2Realizing in-situ photocrosslinking under the irradiation of the light source to obtain a photocrosslinking cured hydrogel-osteochondral scaffold mold complex;
s3, demolding: dissolving the osteochondral scaffold mold in the photo-crosslinking cured hydrogel-osteochondral scaffold mold complex by purified water to obtain photo-crosslinking cured hydrogel;
s4, placing the prepared photo-crosslinking curing hydrogel in a refrigerator at the temperature of-20 ℃ for freezing for 2h, and freeze-drying by using a freeze dryer to obtain the integrally formed osteochondral scaffold.
The obtained osteochondral scaffold was a cuboid (bottom surface 30mm x 30mm, height 3mm), with a porosity of 50% and a pore size of 250 μm.
Effect example 1: cytocompatibility testing of osteochondral scaffolds
GelMA/HAMA-HAP/PLA osteochondral scaffolds prepared in example 1 and Alg/GelMA-TCP/PCL osteochondral scaffolds prepared in example 4 were used as examples.
Mesenchymal stem cells (MS)C) Digesting with pancreatin, centrifuging to collect cells, dripping the cell suspension onto the osteochondral scaffold, incubating for 1 hr, adding culture medium, and culturing at 37 deg.C/5% CO2Culturing for 24h in a cell culture box under the condition. Before testing, the cell culture was aspirated and washed several times with PBS, followed by addition of l mL of live/dead double staining reagent (10. mu.M calcein and 15. mu.M ethidium dimer in 5mL PBS) and incubation with cells for 30min at 37 ℃.
And observing the adhesion and survival condition of cells in the osteochondral scaffold by using a confocal fluorescence microscope. The living cells show the activity of calcein staining and emit green fluorescence at 433 nm; dead cells were stained with ethidium bromide and emitted red fluorescence under 543nm excitation. As shown in FIG. 4, the osteochondral scaffold of the present invention has good cell compatibility and can grow into the through hole structure of the scaffold material.
Effect example 2: application of osteochondral scaffold in rabbit osteochondral composite defect repair
GelMA/HAMA-HAP/PLA osteochondral scaffolds prepared in example 1 were used as an example.
New Zealand male white rabbits are adopted, and a osteochondral complex defect model is established for each rabbit. The groups were randomized by body weight before the experiment (3 per group): a: blank control group; b: bone layer scaffold (HAP/PLA) negative control group; c: osteochondral scaffold (GelMA/HAMA-HAP/PLA) group. In operation, the bracket is filled in the rabbit articular cartilage complex defect. After 12 weeks of operation, rabbits in the experiment were sacrificed by intravenous air, and the injured joint was extracted to evaluate the effect of the experimental repair. The general observation photographs of the damaged joint are shown in FIGS. 5 to 7. Fig. 5 is a blank control group, in which little new tissue was visible due to the absence of the stent. FIG. 6 is a negative control group with only a scaffold of bone layer, since cartilage layer is missing, no new cartilage is grown at all, and only the scaffold of bone layer which has not degraded is seen. Fig. 7 shows the osteochondral scaffold group, and it can be seen that new tissues are formed at the place where the osteochondral scaffold is implanted, and the new tissues have similar appearance to surrounding normal tissues, and have better repairing effect.
Claims (10)
1. The osteochondral scaffold comprises a cartilage layer, an adhesion layer and a bone layer, wherein two sides of the adhesion layer are respectively connected with the cartilage layer and the bone layer; one or more of the cartilage layer, the adhesive layer and the bone layer are porous structures.
2. The osteochondral scaffold of claim 1, wherein the cartilage layer, the adhesive layer and the bone layer are all porous structures; preferably, the hole of the cartilage layer and the hole of the adhesion layer are communicated with the hole of the bone layer; the holes of the cartilage layer, the adhesive layer and the bone layer may or may not be fully aligned, preferably fully aligned; and/or the adhesive layer does not cover or partially covers the hole of the cartilage layer and/or the bone layer;
and/or the diameter of the pores of the cartilage layer and/or the bone layer is 50 to 350 μm, preferably 200 to 280 μm, such as 250 μm; preferably, the pore diameter of the pores of the bone layer is equal to the pore diameter of the pores of the cartilage layer;
and/or the holes of the cartilage layer and/or the bone layer are preferably distributed in a vertically crossed arrangement;
and/or the porosity of the cartilage layer and/or the bone layer is between 20% and 70%, preferably between 40% and 60%, for example 50%;
and/or the osteochondral scaffold has a porosity of 20% to 70%, preferably 40% to 60%, for example 50%.
3. The osteochondral scaffold of claim 1, wherein the osteochondral scaffold is a cylinder; the diameter of the cylinder is preferably 2 to 20mm, more preferably 3 to 10 mm; the height of the cylinder is preferably 2 to 10mm, more preferably 3 to 6 mm;
or the osteochondral scaffold is a cuboid; the bottom surface of the cuboid can be a square, and the side length of the square can be 2-30 mm, preferably 2-20 mm, and more preferably 3-10 mm; the height of the cuboid is preferably 2-10 mm, more preferably 3-6 mm;
and/or the height ratio of the bone layer to the cartilage layer is 1: (0.1 to 1), preferably 1: (0.2 to 0.5);
and/or the height of the adhesive layer is 5 μm to 2mm, preferably 0.1 to 2mm, more preferably 0.5 to 1 mm;
and/or the material of the cartilage layer is hydrogel material; and/or, the cartilage layer is loaded with a cartilage promoting component; wherein the cartilage promoting component preferably comprises a bioactive factor, preferably transforming growth factor TGF α or TGF β, and/or a cell, preferably comprising autologous or allogeneic chondrocytes, mesenchymal stem cells, embryonic stem cells or induced pluripotent stem cells;
and/or the bone layer is made of polylactic acid, polylactic acid-glycolic acid copolymer or polycaprolactone;
and/or, the bone layer is loaded with a bone promoting component; wherein the bone promoting component preferably comprises one or more of a bioactive inorganic material, a bioactive factor, and a cell;
wherein the bioactive inorganic material preferably comprises one or more of hydroxyapatite, calcium phosphate, calcium carbonate and bioactive glass; the mass percentage of the bioactive inorganic material in the bone layer can be 0.1 wt% -70 wt%, preferably 1 wt% -50 wt%, more preferably 2.5 wt% -30 wt%;
the bioactive factor preferably comprises one or more of transforming growth factors TGF alpha, TGF beta, bone morphogenic proteins BMP-2, BMP-3, BMP-4, BMP-5, BMP-6, BMP-7, BMP-8, and BMP-9;
the cells preferably comprise autologous or allogeneic bone cells, mesenchymal stem cells, embryonic stem cells or induced pluripotent stem cells;
and/or the adhesive layer is formed by medical glue; alternatively, the material of the adhesive layer is a hydrogel material.
4. A method for preparing the osteochondral scaffold of any one of claims 1-3, comprising the steps of: connecting the bone layer and the cartilage layer to form an adhesive layer at the joint; one or more of the cartilage layer, the adhesive layer and the bone layer are porous structures.
5. The method for preparing an osteochondral scaffold according to claim 4, wherein the cartilage layer is prepared by using a hydrogel composition as a raw material and cross-linking and curing the hydrogel composition; wherein the hydrogel composition comprises at least a gellable component and a gelling medium;
the gellable component preferably comprises a natural gellable component and/or a synthetic gellable component; wherein the natural gellable component preferably comprises one or more of a natural protein, a natural protein modification, a natural protein degradation product, a modification of a natural protein degradation product, a natural polysaccharide modification, a natural polysaccharide degradation product, and a modification of a natural polysaccharide degradation product; the natural protein preferably comprises one or more of various hydrophilic animal and plant proteins, water-soluble animal and plant proteins, type I collagen, type II collagen, serum protein, silk fibroin and elastin; the natural protein degradation product preferably comprises gelatin or a polypeptide; the modifier of the natural protein degradation product is preferably a methacrylated natural protein degradation product, and is more preferably methacrylated gelatin; the natural polysaccharide preferably comprises one or more of hyaluronic acid, carboxymethyl cellulose, methyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, alginic acid, dextran, agarose, heparin, chondroitin sulfate, ethylene glycol chitosan, propylene glycol chitosan, chitosan lactate, carboxymethyl chitosan and chitosan quaternary ammonium salt, more preferably hyaluronic acid and/or chondroitin sulfate; the natural polysaccharide modifier is preferably a methacrylated natural polysaccharide, such as methacrylated hyaluronic acid or methacrylated chondroitin sulfate;
the synthetic gellable component preferably comprises one or more of two-or multi-arm polyethylene glycol diacrylate, polyethyleneimine, a synthetic polypeptide, polyacrylic acid, polymethacrylic acid, polyacrylate, polymethacrylate, polyacrylamide, polymethacrylamide, polyvinyl alcohol, and polyvinylpyrrolidone;
preferably, the gellable component comprises methacrylated gelatin and methacrylated hyaluronic acid; more preferably, the gelable components of the hydrogel include methacrylated gelatin, methacrylated hyaluronic acid, and methacrylated chondroitin sulfate;
preferably, the gel medium is one or more of purified water, physiological saline, cell culture medium, calcium salt solution and phosphate buffer solution;
preferably, the hydrogel composition of the cartilage layer comprises the following components in parts by weight: 1-50 parts of methacryloylated gelatin, 0-30 parts of methacryloylated hyaluronic acid, 0-30 parts of methacryloylated chondroitin sulfate, 0.01-1 part of photoinitiator and a gel medium;
wherein, the amount of the methacrylated gelatin is preferably 1 to 30 parts, more preferably 1 to 20 parts, still more preferably 2 to 15 parts, still more preferably 5 to 15 parts, such as 8 parts, 10 parts or 12 parts;
wherein the amount of the methacrylated hyaluronic acid is preferably 0.1 to 20 parts, more preferably 0.5 to 10 parts, still more preferably 1 to 3 parts, such as 1.5 parts or 2 parts;
wherein, the amount of the methacrylated chondroitin sulfate is preferably 0.1 to 20 parts, more preferably 0.5 to 20 parts, still more preferably 0.5 to 5 parts, and still more preferably 1 to 3 parts, such as 1 part, 2 parts, 2.5 parts, or 3 parts;
wherein the mass ratio of the methacrylated gelatin to the methacrylated hyaluronic acid may be (1-30): (0.5 to 10), preferably (2 to 15): (1-3), for example 5: 2;
wherein the mass ratio of the methacrylated gelatin to the methacrylated hyaluronic acid to the methacrylated chondroitin sulfate can be (1-30): (0.5-10): (0.5 to 20), preferably (2 to 15): 1: (1-3), for example 10:1:3, 5:2:2, 15:1:1 or 8:1: 3;
preferably, the amount of the gel medium is such that in the hydrogel composition: 5-30% of methacryloylated gelatin, 0.5-2% of methacryloylated hyaluronic acid, 0.1-5% of methacryloylated chondroitin sulfate and 0.01-1% of photoinitiator; wherein the percentage is the mass (g) of the component contained per 100mL of the gel medium;
preferably, the photoinitiator is a blue light initiator, an ultraviolet light initiator or a green light initiator; the blue light initiator is preferably lithium phenyl-2, 4, 6-trimethylbenzoylphosphonate, riboflavin, flavin mononucleotide, eosin Y or ruthenium terpyridine chloride/sodium persulfate; the UV initiator is preferably 2-hydroxy-2-methyl-1- [4- (2-hydroxyethoxy) phenyl ] -1-propanone;
optionally, the hydrogel composition of the cartilage layer further comprises a thickening agent, and the amount of the thickening agent is preferably 0.1-25 parts;
optionally, the hydrogel composition of the cartilage layer further comprises a synthetic photosensitive material, and the amount of the synthetic photosensitive material is preferably 5-30 parts;
wherein the crosslinking curing means preferably comprises one or more of physical crosslinking, chemical crosslinking, enzymatic crosslinking, and photo-crosslinking; wherein the photo-crosslinking preferably has a wavelength of 365 to 405nm and an intensity of 5 to 50mW/cm2Under light irradiation; the photocrosslinking is preferably carried out at a wavelength of 405nm and an intensity of 10mW/cm2Under light irradiation;
and/or, the preparation method of the cartilage layer further comprises the step of loading the cartilage promoting component.
6. The method for preparing an osteochondral scaffold according to claim 5, wherein the cartilage layer is prepared by a perfusion method comprising the steps of: pouring the hydrogel composition of the cartilage layer into a cartilage mould for photocrosslinking to obtain cured hydrogel; freezing and drying the solidified hydrogel to obtain the hydrogel;
or the preparation method of the cartilage layer is a direct 3D printing method, and comprises the following steps: 3D printing is carried out on the hydrogel composition of the cartilage layer, and meanwhile, photocrosslinking is carried out, so that cured hydrogel is obtained; freezing and drying the solidified hydrogel to obtain the hydrogel;
preferably, the preparation method of the cartilage layer comprises the following stepsThe following steps: printing the hydrogel composition of the cartilage layer by adopting an extrusion type 3D printer, and curing and forming by using blue light, wherein the heat preservation temperature is 30-37 ℃, the printing environment temperature is 22-25 ℃, the printing pressure is 20-40 PSI, the printing speed is 4-8mm/s, the filling rate is 40-60%, and the illumination intensity is 5-20 mW/cm2；
Or the preparation method of the cartilage layer is a 3D printing demolding method, and comprises the following steps:
s1, 3D printing is carried out on the sacrificial material to obtain a cartilage layer mold; wherein the sacrificial material is a hard polymer material which can be dissolved in a solvent;
s2, pouring the hydrogel composition of the cartilage layer into the cartilage layer mould for crosslinking to obtain a solidified hydrogel-cartilage layer mould complex;
s3, demolding: dissolving the cartilage layer mould by using a solvent to obtain cured hydrogel;
s4, freeze-drying the obtained solidified hydrogel to obtain the cartilage layer;
wherein the freeze drying time is preferably 8-24 h; preferably a pre-cooling step is performed prior to said freeze-drying;
the pre-cooling temperature is preferably-20 ℃, and the pre-cooling time is preferably 1-3 h.
7. The method for preparing an osteochondral scaffold according to claim 4, wherein the bone layer is prepared by 3D printing of the material of the bone layer; the 3D printing is preferably performed using a fused deposition 3D printer; the material of the bone layer is preferably polylactic acid, polylactic acid-glycolic acid copolymer or polycaprolactone;
preferably, the bone layer is provided by performing 3D printing on the PLA loaded with the hydroxyapatite by using a fused deposition type 3D printer, wherein the temperature of a printing head is 210 ℃, the temperature of a platform is 50-60 ℃, the printing speed is 50-60 mm/s, and the filling rate is 40-60%;
preferably, the bone layer is provided by performing 3D printing on the PCL loaded with the tricalcium phosphate by using a fused deposition type 3D printer, wherein the printing temperature is 140-150 ℃, the printing speed is 50-60 mm/s, and the filling rate is 40% -60%;
preferably, the bone layer is provided by performing 3D printing on GelMA loaded with tricalcium phosphate by adopting an extrusion type photocuring 3D printer, wherein the heat preservation temperature is 30-35 ℃, the printing environment temperature is 22-25 ℃, the printing pressure is 20-40 PSI, the printing speed is 4-8mm/s, the filling rate is 40-60%, and the illumination intensity is 5-50 mW/cm2；
And/or the preparation method of the bone layer is that the hydrogel composition is used as a raw material and is prepared by crosslinking and curing;
preferably, the hydrogel composition of the bone layer comprises the following components: 5 to 30 percent of methylacryloylated gelatin, 2.5 to 50 percent of bioactive glass and 0.01 to 1 percent of photoinitiator; wherein the percentage is the mass (g) of the component contained per 100mL of the gel medium;
and/or, the preparation method of the bone layer further comprises the step of loading a bone promoting component;
the preferred method of loading a bone-promoting component comprises: grinding the material of the bone layer into powder and then adding the bone promoting component; alternatively, the material of the bone layer is dissolved in a solvent, and then the bone promoting ingredient is incorporated, and then the solvent is volatilized.
8. The method for preparing an osteochondral scaffold according to claim 4, wherein the osteochondral scaffold is molded by 3D printing, comprising the steps of:
s1, 3D printing is carried out on the sacrificial material to obtain the osteochondral scaffold mold; wherein the sacrificial material is a hard polymer material which can be dissolved in a solvent;
s2, sequentially pouring hydrogel compositions of a bone layer, a bonding layer and a cartilage layer into the osteochondral scaffold mold for crosslinking to obtain a cured hydrogel-osteochondral scaffold mold complex;
s3, demolding: dissolving the osteochondral scaffold mould by using a solvent to obtain cured hydrogel;
s4, freeze-drying the obtained solidified hydrogel to obtain the osteochondral scaffold;
preferably, the hydrogel composition of the bone layer comprises the following components: 5 to 30 percent of methylacryloylated gelatin, 2.5 to 50 percent of bioactive glass and 0.01 to 1 percent of photoinitiator; the hydrogel composition of the adhesive layer comprises the following components: 5 to 30 percent of methylacryloylated gelatin, 10 to 20 percent of bioactive glass and 0.01 to 1 percent of photoinitiator; the hydrogel composition of the cartilage layer comprises the following components: 5 to 30 percent of methacrylated gelatin, 0.5 to 2 percent of methacrylated hyaluronic acid, 0.5 to 5 percent of methacrylated chondroitin sulfate and 0.01 to 1 percent of photoinitiator; wherein the percentage is the mass (g) of the component contained per 100mL of the gel medium.
9. An osteochondral scaffold prepared by the method of any one of claims 4 to 8.
10. Use of an osteochondral scaffold of any one of claims 1-3 and 9 in repairing an osteochondral defect.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2020107851611 | 2020-08-06 | ||
| CN202010785161 | 2020-08-06 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN113456303A true CN113456303A (en) | 2021-10-01 |
Family
ID=77867664
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110903051.5A Pending CN113456303A (en) | 2020-08-06 | 2021-08-06 | Osteochondral scaffold and preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113456303A (en) |
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114230678A (en) * | 2021-12-13 | 2022-03-25 | 珠海通桥医疗科技有限公司 | Photocrosslinking hydrogel embolization system for intravascular treatment and use method |
| CN114452443A (en) * | 2021-12-24 | 2022-05-10 | 佛山科学技术学院 | Composition for preparing articular cartilage scaffold and preparation method and application thereof |
| CN114870092A (en) * | 2022-05-20 | 2022-08-09 | 广州骊贝生物科技有限公司 | Bone substitute complex and preparation method and application thereof |
| CN115212002A (en) * | 2022-06-15 | 2022-10-21 | 苏州大学 | A 3D bioprinted scaffold for repairing cartilage defects and its preparation |
| CN115282339A (en) * | 2022-07-28 | 2022-11-04 | 四川大学 | A kind of cross-linked hyaluronic acid/hydroxyapatite injectable material, preparation method and application |
| CN115554475A (en) * | 2022-09-29 | 2023-01-03 | 中国人民解放军总医院第一医学中心 | Osteochondral scaffold and preparation method thereof |
| CN115581812A (en) * | 2022-09-29 | 2023-01-10 | 中国人民解放军总医院第一医学中心 | Cartilage scaffold and preparation method thereof |
| CN116688224A (en) * | 2022-02-28 | 2023-09-05 | 苏州大学 | Composite hydrogel scaffold for promoting osteochondral regeneration and its preparation method and application |
| CN116763995A (en) * | 2023-06-20 | 2023-09-19 | 中国矿业大学 | Bone cartilage integrated bracket with directional pore structure and preparation method thereof |
| CN118045235A (en) * | 2024-02-04 | 2024-05-17 | 深圳市第二人民医院(深圳市转化医学研究院) | Double-layer hydrogel bioactive scaffold and preparation method and application thereof |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20100233234A1 (en) * | 2009-03-12 | 2010-09-16 | New Jersey Institute Of Technology | System and method for a hydrogel and hydrogel composite for cartilage repair applications |
| US8475531B1 (en) * | 2009-04-21 | 2013-07-02 | Scott A. Maxson | Anchored multi-phasic osteochondral construct |
| CN107485731A (en) * | 2017-07-28 | 2017-12-19 | 上海理工大学 | Three layers of compound rest of integration and preparation method of a kind of articular cartilage reparation |
| CN108201632A (en) * | 2016-12-20 | 2018-06-26 | 重庆润泽医药有限公司 | A kind of articular cartilage repaiies scaffold |
| CN108525012A (en) * | 2018-03-13 | 2018-09-14 | 华南理工大学 | Gradient hybridization timbering material and preparation method thereof is repaired in a kind of bone cartilage integration |
| CN109364302A (en) * | 2018-09-21 | 2019-02-22 | 王翀 | A kind of preparation method of bone cartilage repair material and tissue engineering bracket |
-
2021
- 2021-08-06 CN CN202110903051.5A patent/CN113456303A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20100233234A1 (en) * | 2009-03-12 | 2010-09-16 | New Jersey Institute Of Technology | System and method for a hydrogel and hydrogel composite for cartilage repair applications |
| US8475531B1 (en) * | 2009-04-21 | 2013-07-02 | Scott A. Maxson | Anchored multi-phasic osteochondral construct |
| CN108201632A (en) * | 2016-12-20 | 2018-06-26 | 重庆润泽医药有限公司 | A kind of articular cartilage repaiies scaffold |
| CN107485731A (en) * | 2017-07-28 | 2017-12-19 | 上海理工大学 | Three layers of compound rest of integration and preparation method of a kind of articular cartilage reparation |
| CN108525012A (en) * | 2018-03-13 | 2018-09-14 | 华南理工大学 | Gradient hybridization timbering material and preparation method thereof is repaired in a kind of bone cartilage integration |
| CN109364302A (en) * | 2018-09-21 | 2019-02-22 | 王翀 | A kind of preparation method of bone cartilage repair material and tissue engineering bracket |
Non-Patent Citations (1)
| Title |
|---|
| MARCO COSTANTINI 等: "3D bioprinting of BM-MSCs-loaded ECM biomimetic hydrogels for in vitro neocartilage formation", 《BIOFABRICATION》 * |
Cited By (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114230678A (en) * | 2021-12-13 | 2022-03-25 | 珠海通桥医疗科技有限公司 | Photocrosslinking hydrogel embolization system for intravascular treatment and use method |
| CN114230678B (en) * | 2021-12-13 | 2023-09-01 | 珠海通桥医疗科技有限公司 | Photocrosslinked hydrogel embolism system for intravascular treatment and application method |
| CN114452443A (en) * | 2021-12-24 | 2022-05-10 | 佛山科学技术学院 | Composition for preparing articular cartilage scaffold and preparation method and application thereof |
| CN116688224A (en) * | 2022-02-28 | 2023-09-05 | 苏州大学 | Composite hydrogel scaffold for promoting osteochondral regeneration and its preparation method and application |
| CN114870092B (en) * | 2022-05-20 | 2023-08-22 | 广州骊贝生物科技有限公司 | Bone substitute complex, preparation method and application thereof |
| CN114870092A (en) * | 2022-05-20 | 2022-08-09 | 广州骊贝生物科技有限公司 | Bone substitute complex and preparation method and application thereof |
| CN115212002A (en) * | 2022-06-15 | 2022-10-21 | 苏州大学 | A 3D bioprinted scaffold for repairing cartilage defects and its preparation |
| CN115282339B (en) * | 2022-07-28 | 2023-02-28 | 四川大学 | A cross-linked hyaluronic acid/hydroxyapatite injectable material, preparation method and application |
| CN115282339A (en) * | 2022-07-28 | 2022-11-04 | 四川大学 | A kind of cross-linked hyaluronic acid/hydroxyapatite injectable material, preparation method and application |
| CN115581812A (en) * | 2022-09-29 | 2023-01-10 | 中国人民解放军总医院第一医学中心 | Cartilage scaffold and preparation method thereof |
| CN115554475A (en) * | 2022-09-29 | 2023-01-03 | 中国人民解放军总医院第一医学中心 | Osteochondral scaffold and preparation method thereof |
| CN115554475B (en) * | 2022-09-29 | 2023-09-05 | 中国人民解放军总医院第一医学中心 | Osteochondral scaffold and preparation method thereof |
| CN116763995A (en) * | 2023-06-20 | 2023-09-19 | 中国矿业大学 | Bone cartilage integrated bracket with directional pore structure and preparation method thereof |
| CN116763995B (en) * | 2023-06-20 | 2025-10-03 | 中国矿业大学 | Bone-cartilage integrated scaffold with directional pore structure and preparation method thereof |
| CN118045235A (en) * | 2024-02-04 | 2024-05-17 | 深圳市第二人民医院(深圳市转化医学研究院) | Double-layer hydrogel bioactive scaffold and preparation method and application thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN113456303A (en) | Osteochondral scaffold and preparation method and application thereof | |
| US20230293306A1 (en) | Bionic tissue stent, preparation method therefor and application thereof | |
| Chen et al. | 3D printing electrospinning fiber-reinforced decellularized extracellular matrix for cartilage regeneration | |
| CN213130115U (en) | Osteochondral scaffold | |
| CN109381749B (en) | Bone tissue repair ink, composition, scaffold, preparation method and kit | |
| Li et al. | Recent progress in tissue engineering and regenerative medicine | |
| Liu et al. | Development of biodegradable scaffolds for tissue engineering: a perspective on emerging technology | |
| Peng et al. | Emerging nanostructured materials for musculoskeletal tissue engineering | |
| CN109364302A (en) | A kind of preparation method of bone cartilage repair material and tissue engineering bracket | |
| Park et al. | Current advances in three-dimensional tissue/organ printing | |
| CN101147810B (en) | Cell-biodegradable material composite and its preparation method and application | |
| US20140012393A1 (en) | Complex support body for regenerating bone-cartilage, method for manufacturing thereof, and composition for treating bone and cartilage related diseases comprising same as active ingredient | |
| Yang et al. | In vitro and in vivo study on an injectable glycol chitosan/dibenzaldehyde-terminated polyethylene glycol hydrogel in repairing articular cartilage defects | |
| Piard et al. | Cell-laden 3D printed scaffolds for bone tissue engineering | |
| CN110302428A (en) | Cartilage-bone-bone marrow composite tissue structure and method based on living cell 3D printing | |
| Li et al. | Osteogenesis and chondrogenesis of biomimetic integrated porous PVA/gel/V‐n‐HA/pa6 scaffolds and BMSCs construct in repair of articular osteochondral defect | |
| Deepika et al. | Applications of nanoscaffolds in tissue engineering | |
| Jahani et al. | Additive manufacturing techniques for fabrication of bone scaffolds for tissue engineering applications | |
| Tamay et al. | Bioinks—materials used in printing cells in designed 3D forms | |
| Gao et al. | Application of photocrosslinkable hydrogels based on photolithography 3D bioprinting technology in bone tissue engineering | |
| Ahadian et al. | Biomaterials in tissue engineering | |
| Schagemann et al. | Morphology and function of ovine articular cartilage chondrocytes in 3-d hydrogel culture | |
| CN111282021A (en) | Meniscus composite scaffold and preparation method thereof | |
| CN115040694B (en) | Biocomposite scaffold for bone regeneration with dual functions of blood vessels and nerves and its preparation method and application | |
| CN106039420B (en) | Fibrin material and preparation method thereof for cartilage reparation integrated with subchondral bone |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |