CN113577381A - 基于微凝胶支架材料构建的可注射软骨及其应用 - Google Patents
基于微凝胶支架材料构建的可注射软骨及其应用 Download PDFInfo
- Publication number
- CN113577381A CN113577381A CN202110907991.1A CN202110907991A CN113577381A CN 113577381 A CN113577381 A CN 113577381A CN 202110907991 A CN202110907991 A CN 202110907991A CN 113577381 A CN113577381 A CN 113577381A
- Authority
- CN
- China
- Prior art keywords
- microgel
- cartilage
- scaffold material
- injectable
- culture
- 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
- 210000000845 cartilage Anatomy 0.000 title claims abstract description 112
- 239000000463 material Substances 0.000 title claims abstract description 84
- 238000004132 cross linking Methods 0.000 claims abstract description 24
- 210000001612 chondrocyte Anatomy 0.000 claims abstract description 23
- 210000004027 cell Anatomy 0.000 claims abstract description 15
- 238000004108 freeze drying Methods 0.000 claims abstract description 14
- 230000007547 defect Effects 0.000 claims abstract description 13
- 210000000130 stem cell Anatomy 0.000 claims abstract description 13
- 238000000338 in vitro Methods 0.000 claims abstract description 9
- 229920002521 macromolecule Polymers 0.000 claims abstract description 8
- 230000008439 repair process Effects 0.000 claims abstract description 8
- 230000008929 regeneration Effects 0.000 claims abstract description 3
- 238000011069 regeneration method Methods 0.000 claims abstract description 3
- 239000000017 hydrogel Substances 0.000 claims description 43
- 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 34
- 229920002674 hyaluronan Polymers 0.000 claims description 34
- 229960003160 hyaluronic acid Drugs 0.000 claims description 34
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 25
- 108010010803 Gelatin Proteins 0.000 claims description 22
- 239000008273 gelatin Substances 0.000 claims description 22
- 229920000159 gelatin Polymers 0.000 claims description 22
- 235000019322 gelatine Nutrition 0.000 claims description 22
- 235000011852 gelatine desserts Nutrition 0.000 claims description 22
- 229920000615 alginic acid Polymers 0.000 claims description 21
- 235000010443 alginic acid Nutrition 0.000 claims description 21
- 238000000034 method Methods 0.000 claims description 20
- 239000000783 alginic acid Substances 0.000 claims description 19
- 229960001126 alginic acid Drugs 0.000 claims description 19
- 150000004781 alginic acids Chemical class 0.000 claims description 19
- 238000006243 chemical reaction Methods 0.000 claims description 19
- 238000012258 culturing Methods 0.000 claims description 15
- 239000002243 precursor Substances 0.000 claims description 15
- 238000003756 stirring Methods 0.000 claims description 15
- 238000005516 engineering process Methods 0.000 claims description 12
- 125000003172 aldehyde group Chemical group 0.000 claims description 10
- 239000008367 deionised water Substances 0.000 claims description 10
- 229910021641 deionized water Inorganic materials 0.000 claims description 10
- 230000003068 static effect Effects 0.000 claims description 10
- 238000000502 dialysis Methods 0.000 claims description 8
- 239000011859 microparticle Substances 0.000 claims description 8
- 238000012216 screening Methods 0.000 claims description 8
- 239000000839 emulsion Substances 0.000 claims description 7
- 239000000499 gel Substances 0.000 claims description 7
- 238000000227 grinding Methods 0.000 claims description 7
- BHPQYMZQTOCNFJ-UHFFFAOYSA-N Calcium cation Chemical compound [Ca+2] BHPQYMZQTOCNFJ-UHFFFAOYSA-N 0.000 claims description 6
- 230000000735 allogeneic effect Effects 0.000 claims description 6
- 125000003277 amino group Chemical group 0.000 claims description 6
- 210000001185 bone marrow Anatomy 0.000 claims description 6
- 229910001424 calcium ion Inorganic materials 0.000 claims description 6
- 239000003431 cross linking reagent Substances 0.000 claims description 6
- SHKUUQIDMUMQQK-UHFFFAOYSA-N 2-[4-(oxiran-2-ylmethoxy)butoxymethyl]oxirane Chemical compound C1OC1COCCCCOCC1CO1 SHKUUQIDMUMQQK-UHFFFAOYSA-N 0.000 claims description 5
- 238000010382 chemical cross-linking Methods 0.000 claims description 5
- 125000000524 functional group Chemical group 0.000 claims description 5
- 239000011148 porous material Substances 0.000 claims description 5
- 239000004005 microsphere Substances 0.000 claims description 4
- 238000001338 self-assembly Methods 0.000 claims description 3
- 210000001188 articular cartilage Anatomy 0.000 claims description 2
- QTCANKDTWWSCMR-UHFFFAOYSA-N costic aldehyde Natural products C1CCC(=C)C2CC(C(=C)C=O)CCC21C QTCANKDTWWSCMR-UHFFFAOYSA-N 0.000 claims description 2
- 210000004728 ear cartilage Anatomy 0.000 claims description 2
- 210000001671 embryonic stem cell Anatomy 0.000 claims description 2
- 238000005286 illumination Methods 0.000 claims description 2
- ISTFUJWTQAMRGA-UHFFFAOYSA-N iso-beta-costal Natural products C1C(C(=C)C=O)CCC2(C)CCCC(C)=C21 ISTFUJWTQAMRGA-UHFFFAOYSA-N 0.000 claims description 2
- 210000002901 mesenchymal stem cell Anatomy 0.000 claims description 2
- 238000005507 spraying Methods 0.000 claims description 2
- 239000002537 cosmetic Substances 0.000 claims 2
- 239000006185 dispersion Substances 0.000 claims 1
- 238000007720 emulsion polymerization reaction Methods 0.000 claims 1
- 238000003701 mechanical milling Methods 0.000 claims 1
- 230000004069 differentiation Effects 0.000 abstract description 2
- 229920003023 plastic Polymers 0.000 abstract description 2
- 239000004033 plastic Substances 0.000 abstract description 2
- 210000003321 cartilage cell Anatomy 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 27
- 239000012071 phase Substances 0.000 description 24
- 229940014259 gelatin Drugs 0.000 description 20
- 229920001661 Chitosan Polymers 0.000 description 16
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 16
- 229920000642 polymer Polymers 0.000 description 14
- 238000010276 construction Methods 0.000 description 12
- 238000002360 preparation method Methods 0.000 description 11
- 239000002262 Schiff base Substances 0.000 description 9
- 150000004753 Schiff bases Chemical class 0.000 description 9
- 239000002609 medium Substances 0.000 description 9
- JQWHASGSAFIOCM-UHFFFAOYSA-M sodium periodate Chemical compound [Na+].[O-]I(=O)(=O)=O JQWHASGSAFIOCM-UHFFFAOYSA-M 0.000 description 8
- SQDAZGGFXASXDW-UHFFFAOYSA-N 5-bromo-2-(trifluoromethoxy)pyridine Chemical compound FC(F)(F)OC1=CC=C(Br)C=N1 SQDAZGGFXASXDW-UHFFFAOYSA-N 0.000 description 7
- 229920001287 Chondroitin sulfate Polymers 0.000 description 7
- 229940059329 chondroitin sulfate Drugs 0.000 description 7
- 239000007924 injection Substances 0.000 description 7
- 238000002347 injection Methods 0.000 description 7
- 239000002245 particle Substances 0.000 description 7
- 229920003169 water-soluble polymer Polymers 0.000 description 7
- DNIAPMSPPWPWGF-UHFFFAOYSA-N Propylene glycol Chemical compound CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- 229920001282 polysaccharide Polymers 0.000 description 6
- 239000005017 polysaccharide Substances 0.000 description 6
- 150000004804 polysaccharides Chemical class 0.000 description 6
- 108090000623 proteins and genes Proteins 0.000 description 6
- 210000001519 tissue Anatomy 0.000 description 6
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 5
- 239000001768 carboxy methyl cellulose Substances 0.000 description 5
- 235000010948 carboxy methyl cellulose Nutrition 0.000 description 5
- 125000002057 carboxymethyl group Chemical group [H]OC(=O)C([H])([H])[*] 0.000 description 5
- 239000008112 carboxymethyl-cellulose Substances 0.000 description 5
- 229940105329 carboxymethylcellulose Drugs 0.000 description 5
- 239000012153 distilled water Substances 0.000 description 5
- CERQOIWHTDAKMF-UHFFFAOYSA-M methacrylate group Chemical group C(C(=C)C)(=O)[O-] CERQOIWHTDAKMF-UHFFFAOYSA-M 0.000 description 5
- 235000018102 proteins Nutrition 0.000 description 5
- 102000004169 proteins and genes Human genes 0.000 description 5
- 229920002307 Dextran Polymers 0.000 description 4
- 241000283973 Oryctolagus cuniculus Species 0.000 description 4
- 239000006285 cell suspension Substances 0.000 description 4
- 238000001727 in vivo Methods 0.000 description 4
- 230000006698 induction Effects 0.000 description 4
- 239000000203 mixture Substances 0.000 description 4
- 238000012986 modification Methods 0.000 description 4
- 230000004048 modification Effects 0.000 description 4
- 239000002861 polymer material Substances 0.000 description 4
- 229920001184 polypeptide Polymers 0.000 description 4
- 108090000765 processed proteins & peptides Proteins 0.000 description 4
- 102000004196 processed proteins & peptides Human genes 0.000 description 4
- 102000008186 Collagen Human genes 0.000 description 3
- 108010035532 Collagen Proteins 0.000 description 3
- NWGKJDSIEKMTRX-AAZCQSIUSA-N Sorbitan monooleate Chemical compound CCCCCCCC\C=C/CCCCCCCC(=O)OC[C@@H](O)[C@H]1OC[C@H](O)[C@H]1O NWGKJDSIEKMTRX-AAZCQSIUSA-N 0.000 description 3
- 239000008346 aqueous phase Substances 0.000 description 3
- 229920001436 collagen Polymers 0.000 description 3
- 239000007857 degradation product Substances 0.000 description 3
- 229960002086 dextran Drugs 0.000 description 3
- AFOSIXZFDONLBT-UHFFFAOYSA-N divinyl sulfone Chemical compound C=CS(=O)(=O)C=C AFOSIXZFDONLBT-UHFFFAOYSA-N 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 238000011049 filling Methods 0.000 description 3
- 239000001963 growth medium Substances 0.000 description 3
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 3
- 230000003993 interaction Effects 0.000 description 3
- 229940057995 liquid paraffin Drugs 0.000 description 3
- 239000012528 membrane Substances 0.000 description 3
- 239000011259 mixed solution Substances 0.000 description 3
- 238000011580 nude mouse model Methods 0.000 description 3
- 238000006116 polymerization reaction Methods 0.000 description 3
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 3
- 238000003753 real-time PCR Methods 0.000 description 3
- 230000001105 regulatory effect Effects 0.000 description 3
- 238000010008 shearing Methods 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- OQILSTRGJVCFAG-UHFFFAOYSA-N 1-(oxiran-2-ylmethoxy)butan-1-ol Chemical compound CCCC(O)OCC1CO1 OQILSTRGJVCFAG-UHFFFAOYSA-N 0.000 description 2
- GJKGAPPUXSSCFI-UHFFFAOYSA-N 2-Hydroxy-4'-(2-hydroxyethoxy)-2-methylpropiophenone Chemical compound CC(C)(O)C(=O)C1=CC=C(OCCO)C=C1 GJKGAPPUXSSCFI-UHFFFAOYSA-N 0.000 description 2
- FHVDTGUDJYJELY-UHFFFAOYSA-N 6-{[2-carboxy-4,5-dihydroxy-6-(phosphanyloxy)oxan-3-yl]oxy}-4,5-dihydroxy-3-phosphanyloxane-2-carboxylic acid Chemical compound O1C(C(O)=O)C(P)C(O)C(O)C1OC1C(C(O)=O)OC(OP)C(O)C1O FHVDTGUDJYJELY-UHFFFAOYSA-N 0.000 description 2
- 102000016284 Aggrecans Human genes 0.000 description 2
- 108010067219 Aggrecans Proteins 0.000 description 2
- 102000004506 Blood Proteins Human genes 0.000 description 2
- 108010017384 Blood Proteins Proteins 0.000 description 2
- 101100328886 Caenorhabditis elegans col-2 gene Proteins 0.000 description 2
- 102000000503 Collagen Type II Human genes 0.000 description 2
- 108010041390 Collagen Type II Proteins 0.000 description 2
- 102000016942 Elastin Human genes 0.000 description 2
- 108010014258 Elastin Proteins 0.000 description 2
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 2
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 2
- 108010022355 Fibroins Proteins 0.000 description 2
- 101000711846 Homo sapiens Transcription factor SOX-9 Proteins 0.000 description 2
- WTDHULULXKLSOZ-UHFFFAOYSA-N Hydroxylamine hydrochloride Chemical compound Cl.ON WTDHULULXKLSOZ-UHFFFAOYSA-N 0.000 description 2
- JVTAAEKCZFNVCJ-UHFFFAOYSA-M Lactate Chemical compound CC(O)C([O-])=O JVTAAEKCZFNVCJ-UHFFFAOYSA-M 0.000 description 2
- 240000007711 Peperomia pellucida Species 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- 102100034204 Transcription factor SOX-9 Human genes 0.000 description 2
- 229940072056 alginate Drugs 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 2
- 230000003848 cartilage regeneration Effects 0.000 description 2
- 239000006143 cell culture medium Substances 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 150000002009 diols Chemical group 0.000 description 2
- 229920002549 elastin Polymers 0.000 description 2
- SWRGUMCEJHQWEE-UHFFFAOYSA-N ethanedihydrazide Chemical compound NNC(=O)C(=O)NN SWRGUMCEJHQWEE-UHFFFAOYSA-N 0.000 description 2
- 229940093476 ethylene glycol Drugs 0.000 description 2
- 210000002744 extracellular matrix Anatomy 0.000 description 2
- VOZRXNHHFUQHIL-UHFFFAOYSA-N glycidyl methacrylate Chemical compound CC(=C)C(=O)OCC1CO1 VOZRXNHHFUQHIL-UHFFFAOYSA-N 0.000 description 2
- 229910052739 hydrogen Inorganic materials 0.000 description 2
- 239000001257 hydrogen Substances 0.000 description 2
- WGCNASOHLSPBMP-UHFFFAOYSA-N hydroxyacetaldehyde Natural products OCC=O WGCNASOHLSPBMP-UHFFFAOYSA-N 0.000 description 2
- 238000011534 incubation Methods 0.000 description 2
- 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 description 2
- DCUFMVPCXCSVNP-UHFFFAOYSA-N methacrylic anhydride Chemical compound CC(=C)C(=O)OC(=O)C(C)=C DCUFMVPCXCSVNP-UHFFFAOYSA-N 0.000 description 2
- 229920005615 natural polymer Polymers 0.000 description 2
- 235000015097 nutrients Nutrition 0.000 description 2
- 239000002504 physiological saline solution Substances 0.000 description 2
- 238000011002 quantification Methods 0.000 description 2
- 150000003254 radicals Chemical class 0.000 description 2
- OARRHUQTFTUEOS-UHFFFAOYSA-N safranin Chemical compound [Cl-].C=12C=C(N)C(C)=CC2=NC2=CC(C)=C(N)C=C2[N+]=1C1=CC=CC=C1 OARRHUQTFTUEOS-UHFFFAOYSA-N 0.000 description 2
- 238000010186 staining Methods 0.000 description 2
- 238000007920 subcutaneous administration Methods 0.000 description 2
- 238000001356 surgical procedure Methods 0.000 description 2
- 238000003786 synthesis reaction Methods 0.000 description 2
- 229920001059 synthetic polymer Polymers 0.000 description 2
- 229920000936 Agarose Polymers 0.000 description 1
- 239000004971 Cross linker Substances 0.000 description 1
- 238000005698 Diels-Alder reaction Methods 0.000 description 1
- 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 description 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- 229920000663 Hydroxyethyl cellulose Polymers 0.000 description 1
- 239000004354 Hydroxyethyl cellulose Substances 0.000 description 1
- 229920002153 Hydroxypropyl cellulose Polymers 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 238000006845 Michael addition reaction Methods 0.000 description 1
- 241000872931 Myoporum sandwicense Species 0.000 description 1
- 108010064851 Plant Proteins Proteins 0.000 description 1
- 229920000463 Poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol) Polymers 0.000 description 1
- 229920000436 Poly(lactide-co-glycolide)-block-poly(ethylene glycol)-block-poly(lactide-co-glycolide) Polymers 0.000 description 1
- 229920002845 Poly(methacrylic acid) Polymers 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- 229920002873 Polyethylenimine Polymers 0.000 description 1
- 108010020346 Polyglutamic Acid Proteins 0.000 description 1
- 108010039918 Polylysine Proteins 0.000 description 1
- 239000004372 Polyvinyl alcohol Substances 0.000 description 1
- 229920002125 Sokalan® Polymers 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- 150000001408 amides Chemical class 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 235000021120 animal protein Nutrition 0.000 description 1
- 238000012925 biological evaluation Methods 0.000 description 1
- 229920001400 block copolymer Polymers 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 210000000988 bone and bone Anatomy 0.000 description 1
- 150000001718 carbodiimides Chemical class 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- RQFQJYYMBWVMQG-IXDPLRRUSA-N chitotriose Chemical compound O[C@@H]1[C@@H](N)[C@H](O)O[C@H](CO)[C@H]1O[C@H]1[C@H](N)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O2)N)[C@@H](CO)O1 RQFQJYYMBWVMQG-IXDPLRRUSA-N 0.000 description 1
- 238000012650 click reaction Methods 0.000 description 1
- 238000010668 complexation reaction Methods 0.000 description 1
- 238000009833 condensation Methods 0.000 description 1
- 230000005494 condensation Effects 0.000 description 1
- 238000006482 condensation reaction Methods 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 239000008380 degradant Substances 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 239000000412 dendrimer Substances 0.000 description 1
- 229920000736 dendritic polymer Polymers 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- OQORATGQIWQZBQ-UHFFFAOYSA-L dilithium phosphonato-(2,4,6-trimethyl-3-phenylphenyl)methanone Chemical compound 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 description 1
- 238000007865 diluting Methods 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000009881 electrostatic interaction Effects 0.000 description 1
- 238000010556 emulsion polymerization method Methods 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 229920000669 heparin Polymers 0.000 description 1
- 229960002897 heparin Drugs 0.000 description 1
- 235000019447 hydroxyethyl cellulose Nutrition 0.000 description 1
- 229940071826 hydroxyethyl cellulose Drugs 0.000 description 1
- 239000001863 hydroxypropyl cellulose Substances 0.000 description 1
- 235000010977 hydroxypropyl cellulose Nutrition 0.000 description 1
- 229940071676 hydroxypropylcellulose Drugs 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 238000001990 intravenous administration Methods 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 230000001678 irradiating effect Effects 0.000 description 1
- 229920000609 methyl cellulose Polymers 0.000 description 1
- 239000001923 methylcellulose Substances 0.000 description 1
- 235000010981 methylcellulose Nutrition 0.000 description 1
- 229960002900 methylcellulose Drugs 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 238000001000 micrograph Methods 0.000 description 1
- 238000002324 minimally invasive surgery Methods 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 239000003607 modifier Substances 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 230000001590 oxidative effect Effects 0.000 description 1
- 235000021118 plant-derived protein Nutrition 0.000 description 1
- 229920000762 poly(caprolactone)-poly(ethylene glycol)-poly(caprolactone) Polymers 0.000 description 1
- 229920001424 poly(ethylene gycol)-block-poly(l-lactic acid)-block-poly(ethylene glycol) Polymers 0.000 description 1
- 229920002401 polyacrylamide Polymers 0.000 description 1
- 229920000058 polyacrylate Polymers 0.000 description 1
- 239000004584 polyacrylic acid Substances 0.000 description 1
- 229920001223 polyethylene glycol Polymers 0.000 description 1
- 229920002643 polyglutamic acid Polymers 0.000 description 1
- 229920000656 polylysine Polymers 0.000 description 1
- -1 polymethacrylamide Polymers 0.000 description 1
- 229920000193 polymethacrylate Polymers 0.000 description 1
- 229920002451 polyvinyl alcohol Polymers 0.000 description 1
- 229920000036 polyvinylpyrrolidone Polymers 0.000 description 1
- 239000001267 polyvinylpyrrolidone Substances 0.000 description 1
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 description 1
- 230000017854 proteolysis Effects 0.000 description 1
- 150000003242 quaternary ammonium salts Chemical class 0.000 description 1
- 238000010791 quenching Methods 0.000 description 1
- 230000001172 regenerating effect Effects 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 230000007480 spreading Effects 0.000 description 1
- 238000003892 spreading Methods 0.000 description 1
- 230000017423 tissue regeneration Effects 0.000 description 1
- 238000004448 titration Methods 0.000 description 1
- 210000003462 vein Anatomy 0.000 description 1
- 230000000007 visual effect Effects 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/14—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
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/38—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells
- A61L27/3804—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells characterised by specific cells or progenitors thereof, e.g. fibroblasts, connective tissue cells, kidney cells
- A61L27/3817—Cartilage-forming cells, e.g. pre-chondrocytes
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/38—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells
- A61L27/3804—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells characterised by specific cells or progenitors thereof, e.g. fibroblasts, connective tissue cells, kidney cells
- A61L27/3834—Cells able to produce different cell types, e.g. hematopoietic stem cells, mesenchymal stem cells, marrow stromal cells, embryonic stem cells
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/38—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells
- A61L27/3839—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells characterised by the site of application in the body
- A61L27/3843—Connective tissue
- A61L27/3852—Cartilage, e.g. meniscus
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/38—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells
- A61L27/3895—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells using specific culture conditions, e.g. stimulating differentiation of stem cells, pulsatile flow conditions
-
- 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
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08H—DERIVATIVES OF NATURAL MACROMOLECULAR COMPOUNDS
- C08H1/00—Macromolecular products derived from proteins
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J3/00—Processes of treating or compounding macromolecular substances
- C08J3/02—Making solutions, dispersions, lattices or gels by other methods than by solution, emulsion or suspension polymerisation techniques
- C08J3/03—Making solutions, dispersions, lattices or gels by other methods than by solution, emulsion or suspension polymerisation techniques in aqueous media
- C08J3/075—Macromolecular gels
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J3/00—Processes of treating or compounding macromolecular substances
- C08J3/24—Crosslinking, e.g. vulcanising, of macromolecules
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J3/00—Processes of treating or compounding macromolecular substances
- C08J3/24—Crosslinking, e.g. vulcanising, of macromolecules
- C08J3/246—Intercrosslinking of at least two polymers
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J9/00—Working-up of macromolecular substances to porous or cellular articles or materials; After-treatment thereof
- C08J9/28—Working-up of macromolecular substances to porous or cellular articles or materials; After-treatment thereof by elimination of a liquid phase from a macromolecular composition or article, e.g. drying of coagulum
-
- 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
- A61L2400/00—Materials characterised by their function or physical properties
- A61L2400/06—Flowable or injectable implant compositions
-
- 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
-
- 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
- A61L2430/40—Preparation and treatment of biological tissue for implantation, e.g. decellularisation, cross-linking
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2201/00—Foams characterised by the foaming process
- C08J2201/04—Foams characterised by the foaming process characterised by the elimination of a liquid or solid component, e.g. precipitation, leaching out, evaporation
- C08J2201/048—Elimination of a frozen liquid phase
- C08J2201/0484—Elimination of a frozen liquid phase the liquid phase being aqueous
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2207/00—Foams characterised by their intended use
- C08J2207/10—Medical applications, e.g. biocompatible scaffolds
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2305/00—Characterised by the use of polysaccharides or of their derivatives not provided for in groups C08J2301/00 or C08J2303/00
- C08J2305/04—Alginic acid; Derivatives thereof
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2305/00—Characterised by the use of polysaccharides or of their derivatives not provided for in groups C08J2301/00 or C08J2303/00
- C08J2305/08—Chitin; Chondroitin sulfate; Hyaluronic acid; Derivatives thereof
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2389/00—Characterised by the use of proteins; Derivatives thereof
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2405/00—Characterised by the use of polysaccharides or of their derivatives not provided for in groups C08J2401/00 or C08J2403/00
- C08J2405/08—Chitin; Chondroitin sulfate; Hyaluronic acid; Derivatives thereof
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08K—Use of inorganic or non-macromolecular organic substances as compounding ingredients
- C08K3/00—Use of inorganic substances as compounding ingredients
- C08K3/16—Halogen-containing compounds
- C08K2003/162—Calcium, strontium or barium halides, e.g. calcium, strontium or barium chloride
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08K—Use of inorganic or non-macromolecular organic substances as compounding ingredients
- C08K5/00—Use of organic ingredients
- C08K5/04—Oxygen-containing compounds
- C08K5/15—Heterocyclic compounds having oxygen in the ring
- C08K5/151—Heterocyclic compounds having oxygen in the ring having one oxygen atom in the ring
- C08K5/1515—Three-membered rings
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Medicinal Chemistry (AREA)
- Biomedical Technology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Epidemiology (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Dermatology (AREA)
- General Health & Medical Sciences (AREA)
- Oral & Maxillofacial Surgery (AREA)
- Transplantation (AREA)
- Animal Behavior & Ethology (AREA)
- Cell Biology (AREA)
- Polymers & Plastics (AREA)
- Organic Chemistry (AREA)
- Botany (AREA)
- Zoology (AREA)
- Dispersion Chemistry (AREA)
- Materials Engineering (AREA)
- Developmental Biology & Embryology (AREA)
- Urology & Nephrology (AREA)
- Vascular Medicine (AREA)
- Hematology (AREA)
- Biochemistry (AREA)
- Rheumatology (AREA)
- Materials For Medical Uses (AREA)
Abstract
本发明涉及基于微凝胶支架材料构建的可注射软骨及其应用。首先将水溶性高分子交联形成微米尺寸的凝胶，冷冻干燥得到微凝胶支架材料。然后将软骨细胞或干细胞接种于微凝胶支架，经体外诱导分化、培养后，即可得到基于微凝胶支架材料构建的可注射软骨。本发明还提供基于微凝胶支架材料构建的可注射软骨在整形美容、软骨缺损修复与再生等领域的应用。本发明提供的基于微凝胶支架材料构建的可注射软骨采用负载软骨细胞的微凝胶进行体外培养获得，大大降低了制备可注射软骨所需的细胞量，从而大幅度降低传统可注射软骨的使用成本。
Description
技术领域
本发明涉及生物医学组织工程领域，尤其涉及一种基于微凝胶支架材料构建的可注射软骨及其应用。
背景技术
可注射软骨在软骨缺损填充与组织再生领域有着重要的应用前景。当前，可注射软骨可以通过取自体软骨制备软骨颗粒用于直接注射；也可以通过膜片技术，将自体软骨种子细胞在体外扩增，制成足够剂量且可用于注射的自体新生软骨微粒，从而实现自体软骨的注射回植。
但是，现在基于软骨膜片技术制备的可注射软骨也面临着一些问题，如力学强度不够、颗粒均质度不够、成本昂贵等。
组织工程理念的兴起为可注射软骨的构建提供了全新的思路，结合可注射型支架材料负载软骨细胞或干细胞有望解决现在可注射软骨技术的一些缺陷。在众多支架材料中，水凝胶材料以其高度含水、合适的力学强度，是组织工程与再生医学领域最为理想的支架材料。但是如何将水凝胶材料应用于可注射软骨制备上仍然存在一定的技术难度。
发明内容
本发明的目的在于提供一种基于微凝胶支架材料构建的可注射软骨及其应用。
本发明的目的可以通过以下技术方案来实现：
本发明的第一个目的是提供微凝胶支架材料的制备方法。
本发明中，微凝胶支架材料的制备方法为微凝胶经冷冻干燥后，即得到微凝胶支架材料；所述微凝胶是指将水凝胶制备成微米尺寸范围内的凝胶；所述水凝胶材料是由水溶性高分子通过物理交联或化学交联或光交联中的一种或多种交联方式获得。即，微凝胶支架材料的制备方法为：水溶性高分子经一定制备方法交联形成微米尺寸的凝胶，经冷冻干燥，即可得到微凝胶支架材料。
由于制备的微凝胶尺寸较小，往往小于注射器的内径，所以易于实现水凝胶的可注射。
本发明中，所述微凝胶的尺寸可以为1μm-1mm，优选为100μm-500μm。
在本发明的一个实施方式中，所述水凝胶材料是由水溶性高分子通过物理交联或化学交联或光交联中的一种或多种交联方式，或单一交联方式的多种材料组合交联形成。
在本发明的一个实施方式中，所述水溶性高分子选自天然高分子材料或合成高分子材料。
在本发明的一个实施方式中，所述天然高分子材料包括天然多糖类物质及其修饰物或降解物，蛋白及其修饰物或降解物。
在本发明的一个实施方式中，所述天然多糖类物质包括透明质酸、羧甲基纤维素、甲基纤维素、羟乙基纤维素、羟丙基纤维素、海藻酸、葡聚糖、琼脂糖、肝素、硫酸软骨素、乙二醇壳聚糖、丙二醇壳聚糖、壳聚糖乳酸盐、羧甲基壳聚糖或壳聚糖季铵盐。
在本发明的一个实施方式中，所述蛋白包括各种亲水或水溶性动植物蛋白、胶原蛋白、血清蛋白、丝素蛋白、弹性蛋白。
在本发明的一个实施方式中，所述蛋白降解物包括明胶或多肽。
在本发明的一个实施方式中，所述合成高分子材料包括两臂或多臂聚乙二醇、聚乙烯亚胺、树枝体、合成多肽、聚赖氨酸、聚谷氨酸、聚丙烯酸、聚甲基丙烯酸、聚丙烯酸酯、聚甲基丙烯酸酯、聚丙烯酰胺、聚甲基丙烯酰胺、聚乙烯醇、聚乙烯吡咯烷酮。
在本发明的一个实施方式中，所述水凝胶材料优选为天然多糖类或蛋白类高分子，进一步优选为透明质酸、明胶。
本发明中，将水凝胶制备成微米尺寸范围内的凝胶的方法包括机械研磨法、乳液聚合法、微流控技术、自组装、喷雾法等。优选为机械研磨法、微流控技术。
在本发明的一个实施方式中，微凝胶制备可实现的实施方式：将透明质酸溶于去离子水，加入交联剂搅拌反应，然后将交联透明质酸倒入透析袋中，用去离子水透析2-3d，除去多余的交联剂，通过机械研磨的方式制备交联透明质酸的微凝胶，然后冷冻干燥，并筛选合适尺寸的微颗粒，即可得到所述的交联透明质酸微凝胶。
上述交联剂可以选用BDDE(丁二醇缩水甘油醚)，DVS(二乙烯基砜)，ADH(乙二酸二酰肼)，EDC(碳二亚胺)，GMA(甲基丙烯酸缩水甘油酯)。进一步优选为BDDE或DVS交联剂。
在本发明的一个实施方式中，微凝胶制备可实现的实施方式：将水凝胶前体溶液作为连续相(水相)，以质量比为8:2的液体石蜡和司盘80的混合溶液作为分散相(油相)。利用注射泵将水相和油相从不同的通道注射到微流控芯片中，在两相交汇处，水相被油相剪切得到单乳液液滴，然后在管道中继续被向前推进，通过交联反应形成微凝胶，并收集至烧杯中。然后冷冻干燥，并筛选合适尺寸的微颗粒，即可得到所述的微凝胶。
上述水凝胶可以选用透明质酸、羧甲基纤维素、海藻酸、葡聚糖、硫酸软骨素、乙二醇壳聚糖、羧甲基壳聚糖、胶原蛋白、丝素蛋白、弹性蛋白、明胶、多肽。优选为透明质酸、海藻酸、硫酸软骨素、乙二醇壳聚糖、明胶。
上述交联反应可以选用物理交联或化学交联或光交联，所述物理交联包括热缩合(温敏性)：聚异丙基丙烯酰胺(PNIPAAm)、嵌段共聚物(PEO-PPO-PEO、PLGA-PEG-PLGA、PEG-PLLA-PEG、PCL-PEG-PCL等)；自组装作用：亲疏水作用、氢键作用、主客体相互作用；离子交联：海藻酸与钙离子；静电相互作用：壳聚糖与磷酸类物质。所述化学交联包括叠氮-炔点击反应、巯基-迈克尔加成反应、酰胺缩合反应、席夫碱反应、狄尔斯-阿尔德反应等。所述光交联是指在光源照射下生成的自由基，引发含甲基丙烯酸酯基的高分子衍生物上双键官能团的聚合反应。
在本发明的一个实施方式中，物理交联水凝胶采用海藻酸与钙离子的交联，即海藻酸与钙离子通过络合作用交联制备水凝胶，即海藻酸水凝胶。所述海藻酸水凝胶可实现的实施方式：将海藻酸高分子溶于生物相容性介质，配成一定浓度的水凝胶前体溶液，加入一定量的钙离子溶液搅拌均匀后，即可获得物理交联的海藻酸水凝胶。
在本发明的一个实施方式中，化学交联水凝胶采用席夫碱反应制备，即含醛基的高分子衍生物与含胺基的高分子衍生物通过席夫碱反应交联制备水凝胶。所述席夫碱水凝胶可实现的实施方式：将含醛基的高分子衍生物和含氨基的高分子衍生物分别溶于生物相容性介质，配成一定浓度的水凝胶前体溶液，混合均匀后，即可获得化学交联的席夫碱水凝胶。
所述含醛基的高分子衍生物的制备方法为邻二醇氧化法，即利用高碘酸钠氧化含邻二醇结构的高分子衍生物得到醛基官能团(参考文献Brendan P.Purcell,David Lobb,Jason A.Burdick,et al.Nat.Mater.2014,13,653.)。所述含醛基的高分子衍生物可实现的实施方式：将含有邻二醇结构的水溶性高分子衍生物于蒸馏水中溶解，加入一定量的高碘酸钠，室温下搅拌反应5-12h，加入乙二醇淬灭反应。然后将反应液倒入透析袋中透析2-3d，然后冷冻干燥，即可得到所述的含醛基的高分子衍生物。反应中，水溶性高分子中的邻二醇结构与高碘酸钠的摩尔比优选为1:0.1-2；高分子溶液的质量浓度优选为1.0％-10％w/v。
所述含醛基的高分子衍生物的制备方法中，含有邻二醇结构的水溶性高分子衍生物可以为多糖类(如葡聚糖、透明质酸、羧甲基纤维素、海藻酸、硫酸软骨素等)，优选为透明质酸、硫酸软骨素。
所述含胺基的高分子衍生物可以是天然含胺基多糖类亲水或水溶性高分子及其修饰物或降解物(如乙二醇壳聚糖、丙二醇壳聚糖、壳聚糖乳酸盐、羧甲基壳聚糖、壳寡糖等)；也可以是生物或经微生物表达后提取的蛋白及其改性物或降解物(如胶原，血清蛋白及明胶等)。优选为明胶、羧甲基壳聚糖。
在本发明的一个实施方式中，光交联构建的水凝胶材料是通过光引发聚合交联反应制备，即光引发剂在光源照射下生成的自由基，引发含甲基丙烯酸酯基的高分子衍生物上双键官能团的聚合反应，从而制备光交联水凝胶。
在本发明的一个实施方式中，光交联水凝胶可实现的实施方式：将含甲基丙烯酸酯基的高分子衍生物和光引发剂溶于生物相容性介质，配成一定浓度的水凝胶前体溶液，在254nm-450nm(优选为365nm或405nm)波长的光源照射下，即可获得光交联水凝胶。
所述含甲基丙烯酸酯基的高分子衍生物的制备方法：将含羟基或胺基的水溶性高分子溶于去离子水，冷却至0-4℃，加入甲基丙烯酸酐，再缓慢滴加5M NaOH，反应24h，然后将反应液倒入透析袋中，用去离子水透析2-3d，然后冷冻干燥，即可得到所述的含甲基丙烯酸酯基的高分子衍生物。
上述含羟基或胺基的多糖类(如：透明质酸、海藻酸、羧甲基纤维素、羧甲基壳聚糖、葡聚糖、硫酸软骨素等)、含羟基或胺基的蛋白或多肽类(如：明胶等)，优选为透明质酸、明胶、海藻酸、羧甲基纤维素、硫酸软骨素，进一步优选为透明质酸、明胶。
上述光交联水凝胶可实现的实施方式中，用于构建光交联水凝胶材料的光引发剂可以选用I 2959(2-羟基-4'-(2-羟乙氧基)-2-甲基苯丙酮)或LAP(苯基-2,4,6-三甲基苯甲酰基膦酸锂)。
在本发明的一个实施方式中，所述生物相容性介质选自蒸馏水、生理盐水、缓冲液或细胞培养基溶液。根据不同的应用，可选取不同的生物相容性介质。
在本发明的一个实施方式中，所述一定浓度的水凝胶前体溶液可以为0.1％w/v-60％w/v，优选为1％w/v-20％w/v。
本发明还提供基于上述方法获得的微凝胶支架材料。
本发明的第二个目的是提供基于微凝胶支架材料构建可注射软骨的方法。
本发明中，基于微凝胶支架材料构建可注射软骨的方法：提取自体或异体的软骨细胞或干细胞，在体外诱导分化、扩增并传代，然后接种于微凝胶支架材料，孵育，经一定的培养方式培养一段时间即可获得可注射软骨。
本发明中，所述软骨细胞来源于自体或异体的软骨组织，优选为耳软骨、关节软骨、半月板软骨、肋软骨等。所述干细胞来源于自体或异体的干细胞，优选为骨髓间充质干细胞、脂肪干细胞、胚胎干细胞等。
在本发明的一个实施方式中，孵育的时间为24小时。
本发明中，所述培养方式为静态培养或动态培养，静态培养方式即将负载细胞的凝胶微球放置于孔板中培养，动态培养方式即将负载细胞的微球放置于生物反应器中搅拌或加压培养。
本发明中，所述培养一段时间为1天-12月，优选为2周-12周。
在本发明的一个实施方式中，所述细胞优选自体耳软骨细胞，培养方式优选为搅拌型生物反应器培养，培养时间优选为4周。
在本发明的一个实施方式中，所述基于微凝胶支架材料构建可注射软骨的可实现的实施方式：通过机械研磨法制备BDDE交联的透明质酸微凝胶，并包裹软骨细胞，通过静态培养方式置于孔板中培养4周，即可获得可注射软骨。
在本发明的一个实施方式中，所述基于微凝胶支架材料构建可注射软骨的可实现的实施方式：将海藻酸高分子溶于生物相容性介质，配成一定浓度的水凝胶前体溶液，通过微流控技术制备一定尺寸的微凝胶，并加入一定量的钙离子溶液进行交联，并包裹软骨细胞，通过静态培养方式置于孔板中培养4周，即可获得可注射软骨。
在本发明的一个实施方式中，所述基于微凝胶支架材料构建可注射软骨的可实现的实施方式：将含醛基的高分子衍生物和含氨基的高分子衍生物分别溶于生物相容性介质，配成一定浓度的水凝胶前体溶液，通过微流控技术制备一定尺寸的微凝胶，并包裹软骨细胞，通过静态培养方式置于孔板中培养4周，即可获得可注射软骨。
在本发明的一个实施方式中，所述基于微凝胶支架材料构建可注射软骨的可实现的实施方式：将标记双键官能团的明胶溶于生物相容性介质，配置一定浓度的水凝胶前体溶液，通过微流控技术在光照下(365nm)制备明胶微凝胶，并包裹骨髓干细胞，通过生物反应器搅拌培养4周，即可获得可注射软骨。
在本发明的一个实施方式中，所述生物相容性介质选自蒸馏水、生理盐水、缓冲液或细胞培养基溶液。根据不同的应用，可选取不同的生物相容性介质。
在本发明的一个实施方式中，所述一定浓度的水凝胶前体溶液可以为0.1％w/v-60％w/v，优选为1％w/v-20％w/v。
本发明基于微凝胶支架材料构建可注射软骨的原理：水凝胶材料给细胞提供三维的培养环境，结合体外生物反应器培养，能够有效改善水凝胶内的营养物质交换，为细胞构建合适的三维培养体系。经过一段时间培养，负载的软骨细胞逐渐成熟，并且分泌出丰富的细胞外基质，最终形成成熟的软骨组织。另外，选用微凝胶支架材料能够实现组织工程软骨的可注射使用，从而对于软骨缺损实施微创填充。因此，运用本发明提供的方法基本可以通过微凝胶支架材料包裹软骨细胞构建可注射软骨。
本发明的第三个目的是提供基于微凝胶支架材料构建可注射软骨的方法获得的基于微凝胶材料构建的可注射软骨。
本发明的第四个目的是提供基于微凝胶支架材料构建的可注射软骨的应用。
本发明提供了基于微凝胶支架材料构建的可注射软骨在整形美容、软骨缺损修复与再生等方面的应用。
与现有技术相比，本发明具有如下优点及有益效果：
(1)与整块水凝胶相比，微凝胶作为软骨培养的支架材料，具有更大的比表面积，更利于细胞接种及营养物质交换。同时，微凝胶支架材料的制备更能控制粒径的均质度，从而保证可注射软骨的均质度；
(2)本发明提供的基于微凝胶支架材料构建的可注射软骨，可以通过注射方式进行软骨缺损填充，能够与当前主流的微创手术配合使用；
(3)传统的可注射软骨采用自体软骨颗粒或软骨膜片技术，而本发明提供的基于微凝胶支架材料构建的可注射软骨采用负载软骨细胞的微凝胶进行体外培养获得，大大降低了制备可注射软骨所需的细胞量，从而大幅度降低传统可注射软骨的使用成本。
附图说明
图1为明胶微凝胶材料的显微镜直观图和粒径分布图。
图2为明胶微凝胶材料负载细胞的扫描电镜图。
图3为基于微凝胶支架材料构建的可注射软骨的效果直观图。
图4为可注射软骨体内培养8周后的组织学图。
图5为可注射软骨体内培养8周后的PCR图。
具体实施方式
下面结合附图和具体实施例对本发明进行详细说明。
实施例一：基于交联透明质酸微凝胶支架材料构建可注射软骨
交联透明质酸微凝胶构建：将透明质酸(2g，340kDa)溶于100mL去离子水，加入0.1g BDDE(丁二醇缩水甘油醚)，并缓慢滴加0.1mL 1M NaOH溶液，继续搅拌反应5h，然后将交联透明质酸倒入透析袋中，用去离子水透析2-3d，除去多余的交联剂，通过机械研磨的方式制备交联透明质酸的微凝胶，然后冷冻干燥，并筛选100μm微颗粒，即可得到交联透明质酸微凝胶支架材料。
可注射软骨构建：从兔子耳朵分离软骨细胞，常规分离培养扩增，传代培养至第二代或第三代，收集并调节细胞悬液终浓度为10×106/mL，接种于上述交联透明质酸微凝胶支架材料，于离心管中孵育24小时后，软骨细胞可以铺展于微凝胶表面。然后，转移至搅拌型生物反应器中，放入培养箱中，于软骨诱导培养基中，继续搅拌培养4周，即可得到可注射软骨。
实施例二：基于海藻酸微凝胶支架材料构建可注射软骨
海藻酸微凝胶构建：配置10％w/v的海藻酸溶液，利用注射泵将其逐渐滴加于0.1MCaCl2溶液，交联形成微凝胶，并收集至烧杯中。然后冷冻干燥，并筛选100μm微颗粒，即可得到海藻酸微凝胶支架材料。
可注射软骨构建：从兔子耳朵分离软骨细胞，常规分离培养扩增，传代培养至第二代或第三代，收集并调节细胞悬液终浓度为10×106/mL，包裹于上述海藻酸微凝胶支架材料，于离心管中孵育24小时后，软骨细胞可以在微凝胶内生长。然后，转移至搅拌型生物反应器中，放入培养箱中，于软骨诱导培养基中，继续搅拌培养4周，即可得到可注射软骨。
实施例三：基于席夫碱微凝胶支架材料构建可注射软骨
氧化透明质酸(HAO)的合成：将透明质酸(2g，340kDa)溶于100mL蒸馏水中至完全溶解，将高碘酸钠(NaIO4，1g)溶于5mL蒸馏水中，然后缓慢滴加上述溶液，于室温下搅拌反应12h。反应结束后，滴加1mL乙二醇继续搅拌30min，然后将反应液倒入透析袋(MWCO 7000)中，用去离子水透析2-3d，冷冻干燥即可得到HAO(1.82g)。根据盐酸羟胺滴定法，可以计算出醛基的含量大约为35％。
席夫碱微凝胶构建：分别配置10％w/v的HAO溶液和10％w/v的壳聚糖溶液作为连续相(水相)，以质量比为8:2的液体石蜡和司盘80的混合溶液作为分散相(油相)。利用注射泵将水相和油相从不同的通道注射到微流控芯片中，在两相交汇处，水相被油相剪切得到单乳液液滴，然后在管道中继续被向前推进，交联形成微凝胶，并收集至烧杯中。然后冷冻干燥，并筛选100μm微颗粒，即可得到席夫碱微凝胶支架材料。
可注射软骨构建：从兔子耳朵分离软骨细胞，常规分离培养扩增，传代培养至第二代或第三代，收集并调节细胞悬液终浓度为10×106/mL，接种于上述席夫碱微凝胶支架材料，于离心管中孵育24小时后，软骨细胞可以铺展于微凝胶表面。然后，转移至搅拌型生物反应器中，放入培养箱中，于软骨诱导培养基中，继续搅拌培养4周，即可得到可注射软骨。
实施例四：基于光敏明胶微凝胶支架材料构建可注射软骨
甲基丙烯酸酯化明胶(GelMA)的合成：将明胶(1g)溶于10mL PBS(pH＝7.4)中，加热至50℃搅拌至完全溶解，加入0.5mL甲基丙烯酸酐，反应2-3h，反应后用40mL PBS稀释反应液，然后倒入透析袋(MWCO 7000)中，用去离子水透析2-3d，冷冻干燥即可得到甲基丙烯酸酯化明胶(0.9g)。根据核磁氢谱图，可计算出双键的含量大约为75％。
光敏明胶微凝胶构建：配置10％w/v的GelMA溶液(含0.2％LAP w/v光引发剂)作为连续相(水相)，以质量比为8:2的液体石蜡和司盘80的混合溶液作为分散相(油相)。利用注射泵将水相和油相从不同的通道注射到微流控芯片中，在两相交汇处，水相被油相剪切得到单乳液液滴，然后在管道中继续被向前推进，通过光照(365nm)，交联形成微凝胶，并收集至烧杯中。然后冷冻干燥，并筛选100μm微颗粒，即可得到明胶微凝胶支架材料(图1所示)。
可注射软骨构建：从兔子骨髓分离骨髓干细胞，常规分离培养扩增，传代培养至第二代或第三代，收集并调节细胞悬液终浓度为10×106/mL，接种于上述光敏明胶微凝胶支架材料，于离心管中孵育24小时后，软骨细胞可以铺展于微凝胶表面(图2所示)。然后，转移至搅拌型生物反应器中，放入培养箱中，于软骨诱导培养基中，继续搅拌培养4周，即可得到可注射软骨(图3所示)。
实施例五：可注射软骨的生物学评价
体外培养4周后，取材，进行大体观、组织学、q-PCR定量等软骨再生指标体外检测。实验结果表明，可注射软骨经培养后呈现白色的外观，逐渐表现为软骨样组织。组织学上可以观察到软骨细胞在支架材料上聚集，并分泌出细胞外基质，番红和二型胶原组织学呈现特异性染色。q-PCR的基因定量数据表明Col2，Aggrecan，SOX9的表达达到正常软骨组织的60％。
实施例六：可注射软骨应用于皮下软骨再生
采用裸鼠，按实施例四构建成熟的可注射软骨，将其注射至裸鼠背部皮下，在体内培养4周，通过静脉注射空气的方法处死实验中的裸鼠，并取样对实验修复效果进行评价。实验结果表明，经体内培养后，再生软骨组织呈现白色的外观，逐渐表现为成熟的软骨样组织。组织学上可以观察到支架材料逐渐降解，并逐渐被再生软骨组织取代，番红和二型胶原组织学呈现特异性染色(图4所示)，q-PCR的基因定量数据表明Col2，Aggrecan，SOX9的表达达到正常软骨组织的90％(图5所示)。
实施例七：可注射软骨应用于软骨缺损修复
采用新西兰雄性大白兔，每只兔子均在关节处制造直径为4mm的软骨缺损。实验前按体重随机分组(每组4只)：1.可注射软骨修复组；2.不做处理的空白组。手术过程中，首先按实施例四构建成熟的可注射软骨，然后将其注射至缺损部位。在手术3月后，通过静脉注射空气的方法处死实验中的兔子，并取样对实验修复效果进行评价。实验结果表明，使用可注射软骨处理的缺损部位取得了完全的软骨修复，而对照组中骨缺损部位没有得到任何修复。因此，该类可注射软骨对软骨缺损有很好的修复效果。
上述的对实施例的描述是为便于该技术领域的普通技术人员能理解和使用发明。熟悉本领域技术的人员显然可以容易地对这些实施例做出各种修改，并把在此说明的一般原理应用到其他实施例中而不必经过创造性的劳动。因此，本发明不限于上述实施例，本领域技术人员根据本发明的揭示，不脱离本发明范畴所做出的改进和修改都应该在本发明的保护范围之内。
Claims (10)
1.一种微凝胶支架材料，其特征在于，微凝胶经冷冻干燥后即得到微凝胶支架材料；
所述微凝胶是指将水凝胶制备成微米尺寸范围内的凝胶；
所述水凝胶材料是由水溶性高分子通过物理交联或化学交联或光交联中的一种或多种交联方式获得；
所述微凝胶的尺寸可以为1μm-1mm，优选为100μm-500μm。
2.根据权利要求1所述的一种微凝胶支架材料，其特征在于，将水凝胶制备成微米尺寸范围内的凝胶的方法包括机械研磨法、乳液聚合法、微流控技术、自组装、喷雾法。
3.根据权利要求1所述的一种微凝胶支架材料，其特征在于，所述微凝胶支架材料选自以下的一种：
将透明质酸溶于去离子水，加入交联剂搅拌反应，然后将交联透明质酸倒入透析袋中，用去离子水透析2-3d，除去多余的交联剂，通过机械研磨的方式制备交联透明质酸的微凝胶，然后冷冻干燥，并筛选合适尺寸的微颗粒，即得到交联透明质酸微凝胶；或，
将水凝胶前体溶液作为水相，选择分散相，将水相和油相从不同的通道注射到微流控芯片中，在两相交汇处，水相被油相剪切得到单乳液液滴，然后在管道中继续被向前推进，通过交联反应形成微凝胶，并收集，然后冷冻干燥，筛选合适尺寸的微颗粒，即得到微凝胶。
4.基于微凝胶支架材料构建可注射软骨的方法，其特征在于，提取自体或异体的软骨细胞或干细胞，在体外诱导分化、扩增并传代，然后接种于权利要求1所述微凝胶支架材料，孵育，经一定的培养方式培养一段时间即获得可注射软骨。
5.根据权利要求4所述基于微凝胶支架材料构建可注射软骨的方法，其特征在于，所述软骨细胞来源于自体或异体的软骨组织，优选为耳软骨、关节软骨、半月板软骨、肋软骨；
所述干细胞来源于自体或异体的干细胞，优选为骨髓间充质干细胞、脂肪干细胞、胚胎干细胞。
6.根据权利要求4所述基于微凝胶支架材料构建可注射软骨的方法，其特征在于，所述培养方式为静态培养或动态培养，静态培养方式即将负载细胞的凝胶微球放置于孔板中培养，动态培养方式即将负载细胞的微球放置于生物反应器中搅拌或加压培养。
7.根据权利要求4所述基于微凝胶支架材料构建可注射软骨的方法，其特征在于，所述培养一段时间为1天-12月，优选为2周-12周。
8.根据权利要求4所述基于微凝胶支架材料构建可注射软骨的方法，其特征在于，选择以下方法中的一种：
通过机械研磨法制备BDDE交联的透明质酸微凝胶，并包裹软骨细胞，通过静态培养方式置于孔板中培养，即获得可注射软骨；或，
将海藻酸高分子溶于生物相容性介质，配成水凝胶前体溶液，通过微流控技术制备一定尺寸的微凝胶，并加入钙离子溶液进行交联，并包裹软骨细胞，通过静态培养方式置于孔板中培养，即获得可注射软骨；或，
将含醛基的高分子衍生物和含氨基的高分子衍生物分别溶于生物相容性介质，配成水凝胶前体溶液，通过微流控技术制备一定尺寸的微凝胶，并包裹软骨细胞，通过静态培养方式置于孔板中培养，即获得可注射软骨；或，
将标记双键官能团的明胶溶于生物相容性介质，配置水凝胶前体溶液，通过微流控技术在光照下制备明胶微凝胶，并包裹骨髓干细胞，通过生物反应器搅拌培养，即获得可注射软骨。
9.基于微凝胶材料构建的可注射软骨，其特征在于，采用权利要求4-8中任一项所述方法制备得到。
10.权利要求9所述基于微凝胶材料构建的可注射软骨的应用，其特征在于，基于微凝胶支架材料构建的可注射软骨在制备整形美容、软骨缺损修复与再生材料上的应用。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110907991.1A CN113577381A (zh) | 2021-08-09 | 2021-08-09 | 基于微凝胶支架材料构建的可注射软骨及其应用 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110907991.1A CN113577381A (zh) | 2021-08-09 | 2021-08-09 | 基于微凝胶支架材料构建的可注射软骨及其应用 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN113577381A true CN113577381A (zh) | 2021-11-02 |
Family
ID=78256361
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110907991.1A Pending CN113577381A (zh) | 2021-08-09 | 2021-08-09 | 基于微凝胶支架材料构建的可注射软骨及其应用 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113577381A (zh) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114870079A (zh) * | 2022-04-21 | 2022-08-09 | 苏州大学 | 一种基于碳量子点的抗菌和促骨修复水凝胶及其制备方法与应用 |
| CN114984299A (zh) * | 2022-05-30 | 2022-09-02 | 浙江大学 | 一种用于治疗糖尿病创面的抗菌抗氧化水凝胶敷料及其制备方法 |
| CN116077736A (zh) * | 2022-09-26 | 2023-05-09 | 四川大学 | 基于多孔明胶复合微球的可注射骨修复支架及其制备方法 |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109837235A (zh) * | 2019-01-15 | 2019-06-04 | 浙江大学 | 水凝胶微载体在细胞的黏附、扩增、冻存和消化中的应用 |
-
2021
- 2021-08-09 CN CN202110907991.1A patent/CN113577381A/zh active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109837235A (zh) * | 2019-01-15 | 2019-06-04 | 浙江大学 | 水凝胶微载体在细胞的黏附、扩增、冻存和消化中的应用 |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114870079A (zh) * | 2022-04-21 | 2022-08-09 | 苏州大学 | 一种基于碳量子点的抗菌和促骨修复水凝胶及其制备方法与应用 |
| CN114984299A (zh) * | 2022-05-30 | 2022-09-02 | 浙江大学 | 一种用于治疗糖尿病创面的抗菌抗氧化水凝胶敷料及其制备方法 |
| CN114984299B (zh) * | 2022-05-30 | 2023-01-06 | 浙江大学 | 一种用于治疗糖尿病创面的抗菌抗氧化水凝胶敷料及其制备方法 |
| CN116077736A (zh) * | 2022-09-26 | 2023-05-09 | 四川大学 | 基于多孔明胶复合微球的可注射骨修复支架及其制备方法 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Feng et al. | Microgel assembly: Fabrication, characteristics and application in tissue engineering and regenerative medicine | |
| Zhu et al. | 3D bioprinting mesenchymal stem cell-laden construct with core–shell nanospheres for cartilage tissue engineering | |
| CN113577381A (zh) | 基于微凝胶支架材料构建的可注射软骨及其应用 | |
| Peng et al. | Characterization of a novel polyvinyl alcohol/chitosan porous hydrogel combined with bone marrow mesenchymal stem cells and its application in articular cartilage repair | |
| EP2882465B1 (en) | Methods of manufacturing hydrogel microparticles having living cells, and compositions for manufacturing a scaffold for tissue engineering | |
| Liu et al. | Microcryogels as injectable 3-D cellular microniches for site-directed and augmented cell delivery | |
| US8137696B2 (en) | Biomimetic composition reinforced by a polyelectrolytic complex of hyaluronic acid and chitosan | |
| US20140010790A1 (en) | Cell culture material based on microbial cellulose | |
| Bae et al. | Fabrication of hyaluronic acid hydrogel beads for cell encapsulation | |
| Zhang et al. | “All-in-one” zwitterionic granular hydrogel bioink for stem cell spheroids production and 3D bioprinting | |
| Yadav et al. | In vitro chondrogenesis with lysozyme susceptible bacterial cellulose as a scaffold | |
| WO2011059112A1 (ja) | 粒子含有細胞集合体 | |
| CN112245658A (zh) | 一种可注射晶胶微球细胞扩增载体及其制备方法 | |
| CN104587531B (zh) | 一种修复关节软骨损伤的凝胶支架的制备方法 | |
| Li et al. | Alginate/PEG based microcarriers with cleavable crosslinkage for expansion and non-invasive harvest of human umbilical cord blood mesenchymal stem cells | |
| Deptuła et al. | Application of 3D-printed hydrogels in wound healing and regenerative medicine | |
| CN115475281B (zh) | 组织工程软骨-骨复合体及其构建方法与应用 | |
| CN115475283B (zh) | 基于水凝胶材料构建的组织工程骨及其制备方法与应用 | |
| CN115804758A (zh) | 制备多孔干细胞微载体的方法、由该方法制备得到的多孔干细胞微载体及应用 | |
| CN116251237A (zh) | 一种负载单个软骨细胞的仿生微型培养支架及制备与应用 | |
| Zhou et al. | Microspheres for cell culture | |
| CN115702951B (zh) | 基于多孔水凝胶材料构建的血管化组织工程骨及其应用 | |
| Kulkarni et al. | Cell immobilization strategies for tissue engineering: Recent trends and future perspectives | |
| CN115702952A (zh) | 基于可注射型水凝胶支架材料构建的可注射骨及其应用 | |
| CN114849601B (zh) | 蛋白修饰微球及其应用 |
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 | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20211102 |
|
| RJ01 | Rejection of invention patent application after publication |