CN102718928A - Hydrogel for intelligent desorption of cell sheet layer and application of hydrogel - Google Patents
Hydrogel for intelligent desorption of cell sheet layer and application of hydrogel Download PDFInfo
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- CN102718928A CN102718928A CN2012102282709A CN201210228270A CN102718928A CN 102718928 A CN102718928 A CN 102718928A CN 2012102282709 A CN2012102282709 A CN 2012102282709A CN 201210228270 A CN201210228270 A CN 201210228270A CN 102718928 A CN102718928 A CN 102718928A
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Abstract
The invention relates to hydrogel for intelligent desorption of a cell sheet layer and an application of the hydrogel. The hydrogel is formed by dissolving functional monomers M1, chemical or physical crosslinking agents M2 and water-solubility biomacromolecules M3 in solvents 4 and adopting an oxidation reduction initiating system for free radical polymerization, and the mass percentages of M1, M2, M3 and M4 are respectively 5 to 10 percent, 2 to 10 percent, 1 to 15 percent and 75 to 92 percent. The hydrogel has excellent temperature or pH value response characteristics and biocompatibility, and the fast multiplication and desorption of cells can be effectively promoted. The cell sheet layer formed after the desorption maintains the complete cell epimatrix, and the biological function damage of the cells caused by an enzyme hydrolysis method and the cell damage caused by degradation ingredients of support frame materials are avoided. The cell sheet layer after the intelligent desorption by the hydrogel can be used for treating and repairing relevant tissues and organs of skin, corneal epithelium and parodontium ligament.
Description
Technical field
The present invention relates to biomedical engineering and intelligent macromolecule material field, be specifically related to a kind of cell sheets intelligence desorption hydrogel and application thereof.
Background technology
For a long time, donor in short supply with graft-rejection be two big obstacles of organ transplantation development.Problem such as low and degraded generation inflammation causes its clinical application curative effect not good enough because of intensity for the natural materials that uses at present and synthetic timbering material.The cell sheets technology is a new technology of results seed cell in the organizational project, and need not support can successfully transplant, and the immunogenicity of having avoided causing after the conventional stent materials graft, brings out problems such as inflammatory reaction, optical interference.
Cell sheets (Cell Sheet) medical treatment is that patient's self cell taking-up back is cultivated in petridish, then the lamella that is proliferated into is transplanted in affected part, impels the technology of affected part neomorph.When cell sheets is attached to when causing on the organ that function can't recover because of grave illness or severely injured, the material that is equivalent to tethelin that is called as cytokine just can be secreted from cell sheets, impels trichangiectasia, and damaged cells will be brought back to life reproduction.Because it is patient's self a cell, therefore do not produce rejection.The possibility that can make the patient of diseases such as mellitus, periodontopathy, lung qi leakage, hepatic insufficiency and joint function disturbance obtain curing through the cell sheets technology improves greatly.Chinese patent CN101896600A discloses a kind of cell sheets and its working method of patterning.Said method comprises step: provide to be arranged in suprabasil successive cell sheets, said substrate comprises transfer of shapes character and/or changeable surface property; Make said successive cell sheets be exposed to releasing agent with the patterning mode; After being exposed to said releasing agent, wash said cell sheets, to remove by the cell of said releasing agent influence and to use second cell type to refill the breach that after being removed, has stayed by the cell of said releasing agent influence.
(polyN-isopropylacrylamide PNIPAAm) has the temperature sensitive performance to PNIPAM, and its volume phase transition temperature is～32 ℃.Yamato etc. (Tissue Eng., 2001,7:473.) PNIPAAm is grafted to the Tissue Culture Plate surface, cell is gone up in inoculation under 37 ℃ of conditions, because PNIPAAm has hydrophobicity, thereby cell can stick, stretches and breed; When reducing below the temperature to 32 ℃, the PNIPAAm possess hydrophilic property, volumetric expansion, cell and culture plate surface isolation, whole cell crowd is then keeping under the iuntercellular connection state with the lamellar morphology desorption, thereby obtains the cell sheets of reservation cytoactive.Through changing the biological function damage that temperature harvested cell lamella has avoided the enzyme digestion pair cell to cause, and can directly be attached to tissue and need not stitching, avoid the degrade injury of composition pair cell of timbering material simultaneously.
The multicomponent system that hydrogel is made up of the superpolymer with three-dimensional crosslinked network structure of hydrophilic polymer chain structure and medium carries etc. at medicine controlled releasing, soft tissue support structure and cell bag and to have a good application prospect aspect the bio-medical material.Hydrogel material just develops towards intelligent direction as the transfer system of filling deficient support, bioactive molecules sustained release carrier and tissue/cell.Temperature-sensitive hydrogel is the macromolecule hydrogel that its volume changes with variation of ambient temperature; To be its volume change the macromolecule hydrogel that changes with environment pH value to the pH sensitive aquagel, has in this gellike macromolecular network to be dissociated into ionic group (carboxyl, sulfonic group or amino), can capture or discharge proton according to the variation of environment pH value.Utilize the soft moisture performance and the sensitivity characteristic of temperature or pH response water gel, then can be through changing the lamella desorption that temperature or pH value realize cell.Pure synthetic water gel cell compatibility is relatively poor, in the synthetic water gel, introduces natural biological consistency monomer such as collagen, chitosan and silk gum etc., then can promote the propagation and the desorption of cell.
Can find out the report that the present temperature sensitive property of no-trump still or pH sensitive aqueous gel, biocompatibility macromolecule and cell sheets desorption technology combine by last analysis.The present invention has taken all factors into consideration temperature or the pH value sensitivity characteristic and the soft moisture performance of hydrogel; And biocompatibility macromolecule good cell consistency; With deionized water as solvent; Adopt radical polymerization, making can be through the hydrogel of attemperation or the automatic desorption of pH value realization cell sheets, and can be applicable to the treatment and the reparation of tissue such as skin, corneal epithelium and periodontium ligament and organ.
Summary of the invention
The technical problem that quasi-solution of the present invention is determined is: for effectively promoting cell adhesion; Avoid the active damage that causes with function etc. of traditional enzymolysis process pair cell, the reduction complication relevant with traditional organizational project simultaneously is like inflammatory reaction etc.; A kind of cell sheets intelligence desorption hydrogel that can be applicable to biomedical engineering is provided; This hydrogel had both had excellent biological compatibility, can promote the cell fast breeding, and having excellent temperature or pH value response again can; Only just can between wetting ability and hydrophobicity, change fast through changing temperature or pH value gel, thus the intelligent control of realization cell proliferation and desorption.
A kind of cell sheets intelligence desorption hydrogel of the present invention; Be dissolved among the solvent M4 by function monomer M1, chemistry or physical crosslinking agent M2 and water-soluble biological polymer M3 and adopt redox initiation system to carry out radical polymerization to form, the mass percent of described M1, M2, M3 and M4 is M1:8-15%, M2:1-10%; M3:1-15%; M4:60-90% changes temperature or pH value back cell from the hydrogel surface desorption, forms laminated structure.
Described M1 has temperature sensitivity or pH value susceptibility; Be N-NSC 11448 or N; N '-two ethyl acrylamides or two ethoxy propylene acid amides or methylacrylic acid N, a kind of in N-dimethylaminoethyl or acrylic or methacrylic acid or the methylene-succinic acid, with and blend.
Described M2 is chemical cross-linking agent or physical crosslinking agent, is N, and N '-methylene-bisacrylamide or TGM 1 or piperazine are attained a kind of in acrylic amide or LUTARALDEHYDE or hectorite or magnesium soap stone or sauconite or the lithium magnesium silicate, with and composition thereof.
Described M3 is the water-soluble biological polymer, a kind of in starch or Xylo-Mucine or sodium-alginate or chitosan or Sodium salts humic acids or gelatin or melon glue or Vinylpyrrolidone polymer or beta-cyclodextrin or the sericin, with and composition thereof.
Described M4 is a deionized water.
Described hydrogel desorption cell is a kind of in mouse fibroblast or corneal cell or Skeletal Muscle Cell or mesenchymal stem cells MSCs or skin fiber archeocyte or Human umbilical vein endothelial cells or Islet cells or the periodontal ligament stem cell.
Described hydrogel temperature be 3-15 ℃ or pH value during for 2-5 proliferating cells form cell sheets from the rapid desorption of hydrogel surface.
Cell sheets behind the described hydrogel intelligence desorption is implanted the tissue that receives the position point, makes up three-dimensional structure through the stack of homotype or heterocyst lamella.
Cell sheets behind the described hydrogel intelligence desorption is in the treatment of skin, corneal epithelium and periodontium ligament tissue or organ and the application in the reparation.
Principle of the present invention is: temperature sensitive or pH value response water gel has good temperature and pH value susceptibility, and when envrionment temperature was higher or lower than phase transition temperature or critical pH respectively, the variation of close and distant water can take place gel, and its volume shrinks or the swelling phenomenon.When high temperature or low pH value; Because hydrophobic grouping plays a leading role or acid ion is protonated, the gel macro manifestations is a volumetric shrinkage, simultaneously because hydrogel contains biocompatibility macromolecule; Cell sticks in gel surface fast, sprawl, breed the quantity that reaches certain after; Drop to below the phase transition temperature envrionment temperature or environment pH value is raised to more than the critical pH, at this moment gel volume expands, and is attached to the automatic desorption of cell of gel surface; Thereby obtain having highly active cell sheets, but reprint to tissue defect place fast breedings such as skins.
Compared with prior art, the present invention has following beneficial effect:
(1) to can be used for the hydrogel component of cell sheets intelligence desorption simple, easy to operate in the present invention, has good temperature or pH value response characteristic, and biocompatibility, can effectively promote the fast breeding and the desorption of cell.
(2) cell compatibility of hydrogel and cell proliferation or desorption rate can wait through monomeric charge ratio, degree of crosslinking and temperature of reaction.
(3) the cell sheets desorption is simple and easy to do, only just can realize the automatic desorption of cell through controlled temperature or pH value, thus the damage of having avoided traditional enzymolysis process pair cell activity, function etc. to cause.
(4) cell sheets that obtains has kept the intact cell epimatrix; Do not need timbering material; Through the mutual range upon range of three-dimensional tissue that obtains full cell of identical or different cell sheets; Overcome the drawback of traditional tissue engineering bracket degraded, had a good application prospect in fields such as periodontal and skin tissue regenerations.
Embodiment
Below in conjunction with specific embodiment the present invention is done further to set forth in detail.Should understand; These embodiment only be used to the present invention is described and be not used in the restriction scope of the present invention; Other any do not run counter to change, the modification done under spirit of the present invention and the principle, substitutes, combination, simplify; All should be the substitute mode of equivalence, be included within protection scope of the present invention.Should be understood that in addition those skilled in the art can do various changes or modification to the present invention after the content of having read the present invention's instruction, these equivalent form of values fall within the application's appended claims institute restricted portion equally.
Embodiment 1
A kind of cell sheets intelligence desorption hydrogel, by N-NSC 11448, N, N '-methylene-bisacrylamide and Xylo-Mucine are dissolved in and adopt ammonium persulfate/Tetramethyl Ethylene Diamine initiator system to carry out radical polymerization in the deionized water to form; Described NSC 11448, N; The mass percent of N '-methylene-bisacrylamide, Xylo-Mucine and deionized water is 10%, 2%, 8%; 80%; Reducing the hydrogel envrionment temperature is 5 ℃, and the skin fiber archeocyte of breeding 7 days forms laminated structure from the hydrogel surface desorption.Skin fiber archeocyte lamella behind the desorption can be used for the treatment and the reparation of burn and scald epiderm skin.
Embodiment 2
A kind of cell sheets intelligence desorption hydrogel is dissolved in by N-NSC 11448, hectorite and sodium-alginate and adopts ammonium persulfate/Tetramethyl Ethylene Diamine initiator system to carry out radical polymerization in the deionized water to form, and the mass percent of described NSC 11448, hectorite, sodium-alginate and deionized water is 11%; 1%; 3%, 85%, reducing the hydrogel envrionment temperature is 4 ℃; The corneal cell of breeding 7 days forms laminated structure from the hydrogel surface desorption.Corneal cell lamella behind the desorption can be used for the treatment and the reparation of cornea.
Embodiment 3
A kind of cell sheets intelligence desorption hydrogel, by methylacrylic acid N, N-dimethylaminoethyl, TGM 1 and Sodium salts humic acids are dissolved in and adopt ammonium persulfate/Tetramethyl Ethylene Diamine initiator system to carry out radical polymerization in the deionized water to form; Described methylacrylic acid N; The mass percent of N-dimethylaminoethyl, TGM 1, Sodium salts humic acids and deionized water is 8%, 3%, 4%; 85%; Reducing the hydrogel envrionment temperature is 3 ℃, and the mesenchymal stem cells MSCs of breeding 7 days forms laminated structure from the hydrogel surface desorption.Mesenchymal stem cells MSCs lamella behind the desorption can be used for sclerodermatous treatment and reparation.
Embodiment 4
A kind of cell sheets intelligence desorption hydrogel, by N, N '-two ethyl acrylamides, TGM 1 and Vinylpyrrolidone polymer are dissolved in and adopt ammonium persulfate/Tetramethyl Ethylene Diamine initiator system to carry out radical polymerization in the deionized water to form; Described N; The mass percent of N '-two ethyl acrylamides, TGM 1, Vinylpyrrolidone polymer and deionized water is 10%, 4%, 6%; 80%; Reducing the hydrogel envrionment temperature is 4 ℃, and the Human umbilical vein endothelial cells of breeding 7 days forms laminated structure from the hydrogel surface desorption.Human umbilical vein endothelial cells lamella behind the desorption can be used for the treatment and the reparation of cardiac muscular tissue.
Embodiment 5
A kind of cell sheets intelligence desorption hydrogel, by blend, the N of NSC 11448 and vinylformic acid (mass ratio is 3: 1), the mixture of N '-methylene-bisacrylamide/hectorite and sericin are dissolved in and adopt ammonium persulfate/Tetramethyl Ethylene Diamine initiator system to carry out radical polymerization in the deionized water to form; Described NSC 11448 and acrylic acid blend, N; The mass percent of N '-methylene-bisacrylamide, sericin and deionized water is 12%, 6%, 12%; 80%; Reducing the hydrogel envrionment temperature is 5 ℃, and the periodontal ligament stem cell of breeding 7 days forms laminated structure from the hydrogel surface desorption.Periodontal ligament stem cell lamella behind the desorption can be used for the treatment and the reparation of periodontitis.
Embodiment 6
A kind of cell sheets intelligence desorption hydrogel, by methylacrylic acid N, the blend of N-dimethylaminoethyl and methylacrylic acid (mass ratio is 4: 1), N; N '-methylene-bisacrylamide and chitosan are dissolved in and adopt ammonium persulfate/Tetramethyl Ethylene Diamine initiator system to carry out radical polymerization in the deionized water to form described methylacrylic acid N, the blend of N-dimethylaminoethyl and methylacrylic acid, N; The mass percent of N '-methylene-bisacrylamide, chitosan and deionized water is 14%, 8%, 3%; 75%; Reducing the hydrogel envrionment temperature is 3 ℃, and the skin fiber archeocyte of breeding 7 days forms laminated structure from the hydrogel surface desorption.Skin fiber archeocyte lamella behind the desorption can be used for the treatment and the reparation of burn and scald epiderm skin.
Claims (9)
1. cell sheets intelligence desorption hydrogel and application thereof; It is characterized in that being dissolved among the solvent M4 by function monomer M1, chemistry or physical crosslinking agent M2 and water-soluble biological polymer M3 and adopt redox initiation system to carry out radical polymerization to form, the mass percent of described M1, M2, M3 and M4 is M1:8-15%, M2:1-10%; M3:1-15%; M4:60-90% changes temperature or pH value back cell from the hydrogel surface desorption, forms laminated structure.
2. a kind of cell sheets intelligence desorption hydrogel according to claim 1; It is characterized in that described M1 has temperature sensitivity or pH value susceptibility; Be N-NSC 11448 or N; N '-two ethyl acrylamides or two ethoxy propylene acid amides or methylacrylic acid N, a kind of in N-dimethylaminoethyl or acrylic or methacrylic acid or the methylene-succinic acid, with and blend.
3. a kind of cell sheets intelligence desorption hydrogel according to claim 1; It is characterized in that described M2 is chemical cross-linking agent or physical crosslinking agent; Be N; N '-methylene-bisacrylamide or TGM 1 or piperazine are attained a kind of in acrylic amide or LUTARALDEHYDE or hectorite or magnesium soap stone or sauconite or the lithium magnesium silicate, with and composition thereof.
4. a kind of cell sheets intelligence desorption hydrogel according to claim 1; It is characterized in that described M3 is the water-soluble biological polymer; A kind of in starch or Xylo-Mucine or sodium-alginate or chitosan or Sodium salts humic acids or gelatin or melon glue or Vinylpyrrolidone polymer or beta-cyclodextrin or the sericin, with and composition thereof.
5. a kind of cell sheets intelligence desorption hydrogel according to claim 1 is characterized in that described M4 is a deionized water.
6. a kind of cell sheets intelligence desorption hydrogel according to claim 1 is characterized in that described hydrogel desorption cell is a kind of in mouse fibroblast or corneal cell or Skeletal Muscle Cell or mesenchymal stem cells MSCs or skin fiber archeocyte or Human umbilical vein endothelial cells or Islet cells or the periodontal ligament stem cell.
7. a kind of cell sheets intelligence desorption hydrogel according to claim 1, it is characterized in that described hydrogel temperature be 3-15 ℃ or pH value during for 2-5 proliferating cells from the rapid desorption of hydrogel surface, the formation cell sheets.
8. a kind of cell sheets intelligence desorption hydrogel according to claim 1 is characterized in that the cell sheets behind the described hydrogel intelligence desorption is implanted the tissue that receives the position point, makes up three-dimensional structure through the stack of homotype or heterocyst lamella.
9. a kind of cell sheets intelligence desorption hydrogel according to claim 1 and application thereof, it is characterized in that behind the described hydrogel intelligence desorption cell sheets the treatment of skin, corneal epithelium and periodontium ligament tissue or organ with repair in application.
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Cited By (18)
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| CN103159896A (en) * | 2013-04-15 | 2013-06-19 | 西北师范大学 | Preparation of cellulose grafted acrylate copolymer and application of copolymer as adsorbent |
| CN103520777A (en) * | 2013-10-08 | 2014-01-22 | 天津工业大学 | Highly-tough hole-adjustable gel artificial blood vessel and making method thereof |
| CN103554369A (en) * | 2013-11-11 | 2014-02-05 | 江苏大学 | Double-stimulus response hydrogel with cyclodextrin and preparation method thereof |
| CN103603079A (en) * | 2013-11-15 | 2014-02-26 | 无锡中科光远生物材料有限公司 | Preparation method of responsive nanofiber for delivering anti-cancer medicaments |
| CN103966158A (en) * | 2014-05-28 | 2014-08-06 | 中国人民解放军第四军医大学 | Preparation method of specific extracellular matrix ECM of periodontium and application thereof |
| CN105218835A (en) * | 2015-09-25 | 2016-01-06 | 西北师范大学 | A kind of preparation of keratin based sensitive high-molecular gel and the application as pharmaceutical carrier |
| CN105254917A (en) * | 2015-11-03 | 2016-01-20 | 中山大学 | Method for preparing cell sheet from sodium alginate hydrogel |
| CN105879046A (en) * | 2016-04-18 | 2016-08-24 | 天津工业大学 | Preparation method of beta-cyclodextrin modified organic hectorite based antibacterial agent |
| CN106421925A (en) * | 2016-09-08 | 2017-02-22 | 中国医学科学院生物医学工程研究所 | Preparation method of pancreas islet culture scaffold based on glycosaminoglycan bionic extracellular matrix hydrogel |
| CN106880869A (en) * | 2015-12-16 | 2017-06-23 | 西安组织工程与再生医学研究所 | A kind of preparation method of pure titanium implant and periodontium complex |
| CN107141389A (en) * | 2017-06-15 | 2017-09-08 | 吉林大学 | A kind of preparation method of high intensity, the light-operated desorption hydrogel of biological stickiness |
| CN107619452A (en) * | 2016-07-14 | 2018-01-23 | 山东大学（威海） | A kind of preparation method of the chitosan-based hydrogel of physical crosslinking with efficient pH sensitiveness and high mechanical properties |
| CN109929072A (en) * | 2019-03-22 | 2019-06-25 | 华东师范大学 | A kind of freeze proof polysaccharide composite starch hydrogel and preparation method and application |
| CN110101903A (en) * | 2019-04-09 | 2019-08-09 | 温州医科大学 | A kind of BG compound rest of inducible hypoxemia and its application |
| CN110665063A (en) * | 2019-10-28 | 2020-01-10 | 中国人民解放军第四军医大学 | 3D biological printing ink and preparation method thereof, tissue engineering scaffold and preparation method thereof |
| CN112250798A (en) * | 2020-09-18 | 2021-01-22 | 生物岛实验室 | Preparation method of hydrogel and mesenchymal stem cells |
| CN114272427A (en) * | 2021-12-31 | 2022-04-05 | 山东大学 | Preparation method of multifunctional colloid patch capable of being pasted with wet surface |
| CN115466708A (en) * | 2022-09-16 | 2022-12-13 | 四川大学 | Method for controllably adhering and releasing cells |
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| CN103159896A (en) * | 2013-04-15 | 2013-06-19 | 西北师范大学 | Preparation of cellulose grafted acrylate copolymer and application of copolymer as adsorbent |
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| CN103554369A (en) * | 2013-11-11 | 2014-02-05 | 江苏大学 | Double-stimulus response hydrogel with cyclodextrin and preparation method thereof |
| CN103554369B (en) * | 2013-11-11 | 2016-07-06 | 江苏大学 | A kind of preparation method of the double stimuli responsive hydrogel with cyclodextrin |
| CN103603079A (en) * | 2013-11-15 | 2014-02-26 | 无锡中科光远生物材料有限公司 | Preparation method of responsive nanofiber for delivering anti-cancer medicaments |
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| CN103966158A (en) * | 2014-05-28 | 2014-08-06 | 中国人民解放军第四军医大学 | Preparation method of specific extracellular matrix ECM of periodontium and application thereof |
| CN103966158B (en) * | 2014-05-28 | 2016-09-21 | 中国人民解放军第四军医大学 | A kind of preparation method and applications of periodontal tissue specific cell epimatrix ECM |
| CN105218835A (en) * | 2015-09-25 | 2016-01-06 | 西北师范大学 | A kind of preparation of keratin based sensitive high-molecular gel and the application as pharmaceutical carrier |
| CN105218835B (en) * | 2015-09-25 | 2017-12-12 | 西北师范大学 | A kind of preparation of keratin base sensitive high-molecular gel and the application as pharmaceutical carrier |
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| CN105879046B (en) * | 2016-04-18 | 2019-09-20 | 天津工业大学 | The preparation method of the organic lithium soapstone based antimicrobials of beta-cyclodextrin modified |
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| CN107141389A (en) * | 2017-06-15 | 2017-09-08 | 吉林大学 | A kind of preparation method of high intensity, the light-operated desorption hydrogel of biological stickiness |
| CN109929072B (en) * | 2019-03-22 | 2021-03-26 | 华东师范大学 | Antifreeze polysaccharide composite starch hydrogel and preparation method and application thereof |
| CN109929072A (en) * | 2019-03-22 | 2019-06-25 | 华东师范大学 | A kind of freeze proof polysaccharide composite starch hydrogel and preparation method and application |
| CN110101903A (en) * | 2019-04-09 | 2019-08-09 | 温州医科大学 | A kind of BG compound rest of inducible hypoxemia and its application |
| CN110101903B (en) * | 2019-04-09 | 2021-11-02 | 温州医科大学 | BG composite scaffold capable of inducing hypoxia and application thereof |
| CN110665063A (en) * | 2019-10-28 | 2020-01-10 | 中国人民解放军第四军医大学 | 3D biological printing ink and preparation method thereof, tissue engineering scaffold and preparation method thereof |
| CN112250798A (en) * | 2020-09-18 | 2021-01-22 | 生物岛实验室 | Preparation method of hydrogel and mesenchymal stem cells |
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| CN115466708A (en) * | 2022-09-16 | 2022-12-13 | 四川大学 | Method for controllably adhering and releasing cells |
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