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WO2024098285A1 - Matériau de réparation de tissu commandé par programme d'exosome et son procédé de préparation - Google Patents

Matériau de réparation de tissu commandé par programme d'exosome et son procédé de préparation Download PDF

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WO2024098285A1
WO2024098285A1 PCT/CN2022/130867 CN2022130867W WO2024098285A1 WO 2024098285 A1 WO2024098285 A1 WO 2024098285A1 CN 2022130867 W CN2022130867 W CN 2022130867W WO 2024098285 A1 WO2024098285 A1 WO 2024098285A1
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exosome
stem cells
flexible substrate
tissue repair
poly
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Chinese (zh)
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杜学敏
彭明星
赵启龙
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Shenzhen Institute of Advanced Technology of CAS
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Shenzhen Institute of Advanced Technology of CAS
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    • 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
    • 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
    • A61L27/54—Biologically active materials, e.g. therapeutic substances
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M1/00—Apparatus for enzymology or microbiology
    • C12M1/42—Apparatus for the treatment of microorganisms or enzymes with electrical or wave energy, e.g. magnetism, sonic waves
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor

Definitions

  • the present application relates to the fields of biomaterials and regenerative medicine, and in particular to an exosome-controlled tissue repair material and a preparation method thereof.
  • Tissue repair is of great significance in dealing with tissue and organ damage, aging, pathology and other aspects.
  • the behavioral functions of cells involved in tissue repair need to be regulated in an orderly manner. Therefore, it is crucial to create a cell microenvironment that can regulate cell behavioral functions in an orderly manner.
  • the natural cell microenvironment mainly achieves orderly regulation of cell behavioral functions through the interaction between the extracellular matrix and cells and the interaction between cells.
  • exosome vesicles are the main way to achieve intercellular communication: upstream cells release exosome vesicles containing bioactive molecules such as growth factors and nucleic acid fragments, which are received and activated by downstream cells to regulate the behavioral functions of downstream cells.
  • exosome vesicles Based on the powerful cell behavioral function regulation ability of exosome vesicles, it has been widely used in the field of tissue regeneration and repair in recent years.
  • exosome vesicles secreted by stem cells often contain more abundant cell-regulating bioactive molecules, which have a significant promoting effect on the regeneration and repair of various tissues and organs.
  • tissue repair materials and strategies based on exosomes mainly load the extracted exosomes into tissue repair materials, and then release them in a controlled release manner to achieve regulation of cell behavior and function and even promote tissue regeneration and repair.
  • tissue repair materials and strategies based on exosomes mainly load the extracted exosomes into tissue repair materials, and then release them in a controlled release manner to achieve regulation of cell behavior and function and even promote tissue regeneration and repair.
  • such strategies have achieved good results in different fields of tissue regeneration and repair, on the one hand, such strategies will lose the ability to continuously promote tissue repair as the loaded exosomes are exhausted; on the other hand, it is difficult to match the differentiated needs for bioactive molecules at different stages of tissue regeneration and repair, and to regulate the types of bioactive molecules in exosomes on demand.
  • One of the purposes of this application is to provide an exosome-controlled tissue repair material.
  • the thickness of the flexible substrate is 50-500 ⁇ m.
  • the surface morphology of the flexible substrate is a plane or a three-dimensional structure
  • the three-dimensional structure includes at least one of a microgroove array, a microcone array, and a microcolumn array.
  • the three-dimensional structure has a width of 50 nm-50 ⁇ m, a height of 50 nm-50 ⁇ m, and a pitch of 50 nm-50 ⁇ m.
  • the flexible substrate is at least one of piezoelectric material, photodeformable material composite piezoelectric material, photovoltaic material, upconversion material composite photovoltaic material, photothermal material composite pyroelectric material, magnetocaloric material composite pyroelectric material, and piezoelectric ion gel.
  • the piezoelectric material includes a piezoelectric crystal or a piezoelectric ceramic or a polyvinylidene fluoride ferroelectric polymer or a piezoelectric polymer
  • the piezoelectric crystal includes a quartz crystal or lithium gallate or lithium germanate or titanium germanate or lithium tantalate
  • the piezoelectric ceramic includes barium titanate or lead zirconate titanate or lead metaniobate or lithium barium lead niobate
  • the polyvinylidene fluoride ferroelectric polymer includes poly(vinylidene fluoride) or poly(vinylidene fluoride-trifluoroethylene) [P(VDF-TrFE) copolymer] or poly(vinylidene fluoride-chlorofluoroethylene) [P(VDF-CFE) copolymer] or poly(vinylidene fluoride-chlorotrifluoroethylene) [P(VDF-CTFE) copolymer] Polymer] or poly
  • the photodeformable material composite piezoelectric material is a combination of the following photodeformable materials and any one of the piezoelectric materials; wherein the photodeformable material includes at least one of a photoisomerization material or a ferroelectric inorganic photodeformable material, and the photoisomerization material includes at least one of azobenzene and its derivatives, spiropyran and its derivatives; the ferroelectric inorganic photodeformable material includes at least one of lead titanate, barium titanate, potassium niobate, lithium niobate, lithium tantalate, bismuth layered perovskite structure ferroelectric, tungsten bronze type ferroelectric, bismuth ferrite, potassium dihydrogen phosphate, triammonium sulfate, Roche salt, and perovskite type organic metal halide ferroelectric.
  • the photovoltaic material includes at least one of the following substances: organic photovoltaic materials of polyacetylene, polythiophene, polyaniline, polypyrrole, and derivatives and copolymers thereof.
  • the up-conversion material composite photovoltaic material is a combination of any one of the following up-conversion materials and the piezoelectric material; wherein the up-conversion material includes at least one of yttrium oxide, yttrium oxysulfide, lanthanum fluoride, sodium yttrium fluoride, and sodium gadolinium fluoride.
  • the photothermal material composite pyroelectric material includes any combination of photothermal material and pyroelectric material; wherein the photothermal material includes at least one of carbon black, carbon nanotubes, graphene, black phosphorus, polydopamine, gold nanorods, and gallium-indium alloy liquid metal; the pyroelectric material includes at least one of polyvinylidene fluoride ferroelectric polymers and perovskite ferroelectric ceramics, and the polyvinylidene fluoride ferroelectric polymers include poly(vinylidene fluoride), poly(vinylidene fluoride-trifluoroethylene) [P(VDF-TrFE) copolymer], poly(vinylidene fluoride-chlorofluoroethylene) [P(VDF-CFE) copolymer], poly(vinylidene fluoride At least one of poly(vinylidene fluoride-trifluoroethylene) [P(VDF-CTFE) copolymer
  • the magnetocaloric material composite pyroelectric material is a combination of any one of the following magnetocaloric materials and the pyroelectric material; wherein the magnetocaloric material includes at least one of Ru-Fe-B alloy, ferroferric oxide, iron, cobalt, nickel, and gadolinium.
  • the piezoelectric ion gel material includes at least one of a polyacrylic acid and polyacrylamide composite gel, a polyacrylic acid and chitosan composite gel, a sodium alginate and polyacrylamide composite gel, a polyacrylic acid and choline composite gel, a sodium alginate and choline composite gel, a methacrylated gelatin and choline composite gel, and a methacrylated hyaluronic acid and choline composite gel.
  • the stem cells include at least one of totipotent stem cells, pluripotent stem cells, induced pluripotent stem cells and adult unipotent stem cells
  • the totipotent stem cells include embryonic stem cells
  • the pluripotent stem cells include mesenchymal stem cells
  • the adult unipotent stem cells include at least one of corneal stem cells or neural stem cells or endothelial progenitor cells or endothelial stem cells.
  • the hydrogel is at least one of the following materials: alginate, chitosan, gelatin and its derivatives, collagen and its derivatives, hyaluronic acid and its derivatives, extracellular matrix proteins and their derivatives, silk fibroin and its derivatives, agarose, carrageenan, dextran, basement membrane matrix, polycaprolactone, polyethylene glycol and its derivatives, propylene glycol and ethylene oxide block polymers, polyvinyl pyrrolidone.
  • the hydrogel layer has a thickness of 50-500 ⁇ m.
  • the second purpose of the present application is to provide a method for preparing an exosome-controlled tissue repair material, comprising the following steps:
  • the hydrogel is pre-crosslinked on the surface of the flexible substrate so that the stem cells are encapsulated on the surface of the flexible substrate, thereby obtaining the exosome-controlled tissue repair material.
  • the step of preparing the flexible substrate specifically includes the following steps: preparing the flexible substrate by a casting method or a spin coating method.
  • a planar or three-dimensional structure is formed on the surface of the flexible substrate by photolithography, plasma dry etching or mechanical processing, and the three-dimensional structure includes at least one of a microgroove array, a microcone array and a micropillar array.
  • the seeding density of the stem cells is 10 3 -10 6 cells/cm 2 .
  • the cross-linking method is at least one of ionic cross-linking and ultraviolet light cross-linking.
  • the exosome-controlled tissue repair material includes: a flexible substrate, stem cells grown on the surface of the flexible substrate, and a hydrogel for encapsulating the stem cells on the surface of the flexible substrate.
  • the stem cells can continuously release exosome vesicles containing bioactive molecules.
  • the flexible substrate can convert external field energy into electricity, and stimulate the stem cells growing on its surface to guide differentiation, thereby regulating the proportion and content of different types of bioactive molecules in the released exosome vesicles, so as to meet the differentiated needs of tissues and organs for bioactive molecules at different stages of their regeneration and repair.
  • the exosome-controlled tissue repair material provided by the present application, through the continuous and controllable release of stem cell exosome vesicles in the external field-mediated material, orderly regulates the behavior and function of cells in damaged tissues and organs, thereby promoting the regeneration and repair of tissues and organs.
  • FIG1 is a schematic diagram of the structure of an exosome-controlled tissue repair material provided in an embodiment of the present application.
  • FIG. 2 is a flow chart showing the steps of applying for the exosome-controlled tissue repair material provided in this embodiment.
  • first and second are used for descriptive purposes only and should not be understood as indicating or implying relative importance or implicitly indicating the number of the indicated technical features. Therefore, a feature defined as “first” or “second” may explicitly or implicitly include one or more of the features. In the description of this application, the meaning of “plurality” is two or more, unless otherwise clearly and specifically defined.
  • Figure 1 is a schematic structural diagram of an exosome-controlled tissue repair material provided by an embodiment of the present application, including: a flexible substrate 110, stem cells 120 grown on the surface of the flexible substrate 110, and a hydrogel 130 for encapsulating the stem cells 120 on the surface of the flexible substrate 110.
  • the stem cells 120 can continuously release exosome vesicles containing bioactive molecules.
  • the flexible substrate 110 can convert external field energy into electricity and stimulate the stem cells 120 growing on its surface to guide differentiation in a directional manner, thereby regulating the proportion and content of different types of bioactive molecules in the exosome vesicles released by them.
  • the external field energy includes light or magnetic field or ultrasound of the flexible substrate.
  • the thickness of the flexible substrate is 50-500 ⁇ m.
  • the surface morphology of the flexible substrate is a plane or a three-dimensional structure
  • the three-dimensional structure includes at least one of a microgroove array, a microcone array, and a microcolumn array.
  • the three-dimensional structure has a width of 50 nm-50 ⁇ m, a height of 50 nm-50 ⁇ m, and a pitch of 50 nm-50 ⁇ m.
  • the flexible substrate is at least one of piezoelectric material, photodeformable material composite piezoelectric material, photovoltaic material, upconversion material composite photovoltaic material, photothermal material composite pyroelectric material, magnetocaloric material composite pyroelectric material, and piezoelectric ion gel.
  • the piezoelectric material includes a piezoelectric crystal or a piezoelectric ceramic or a polyvinylidene fluoride ferroelectric polymer or a piezoelectric polymer
  • the piezoelectric crystal includes a quartz crystal or lithium gallate or lithium germanate or titanium germanate or lithium tantalate
  • the piezoelectric ceramic includes barium titanate or lead zirconate titanate or lead metaniobate or lead niobate Barium lithium
  • the polyvinylidene fluoride ferroelectric polymer includes poly(vinylidene fluoride) or poly(vinylidene fluoride-trifluoroethylene) [P(VDF-TrFE) copolymer] or poly(vinylidene fluoride-chlorofluoroethylene) [P(VDF-CFE) copolymer] or poly(vinylidene fluoride-chlorotrifluoroethylene) [P(VDF-CTFE) copolymer] or poly(vinylid
  • the photodeformable material composite piezoelectric material is a combination of the following photodeformable materials and any one of the piezoelectric materials; wherein the photodeformable material includes at least one of a photoisomerization material or a ferroelectric inorganic photodeformable material, and the photoisomerization material includes at least one of azobenzene and its derivatives, spiropyran and its derivatives; the ferroelectric inorganic photodeformable material includes at least one of lead titanate, barium titanate, potassium niobate, lithium niobate, lithium tantalate, bismuth layered perovskite structure ferroelectric, tungsten bronze type ferroelectric, bismuth ferrite, potassium dihydrogen phosphate, triammonium sulfate, Roche salt, and perovskite type organic metal halide ferroelectric.
  • the photovoltaic material includes at least one of the following substances: organic photovoltaic materials of polyacetylene, polythiophene, polyaniline, polypyrrole, and derivatives and copolymers thereof.
  • the up-conversion material composite photovoltaic material is a combination of any one of the following up-conversion materials and the piezoelectric material; wherein the up-conversion material includes at least one of yttrium oxide, yttrium oxysulfide, lanthanum fluoride, sodium yttrium fluoride, and sodium gadolinium fluoride.
  • the photothermal material composite pyroelectric material includes any combination of photothermal material and pyroelectric material; wherein the photothermal material includes at least one of carbon black, carbon nanotubes, graphene, black phosphorus, polydopamine, gold nanorods, and gallium-indium alloy liquid metal; the pyroelectric material includes at least one of polyvinylidene fluoride ferroelectric polymers and perovskite ferroelectric ceramics, and the polyvinylidene fluoride ferroelectric polymers include poly(vinylidene fluoride), poly(vinylidene fluoride-trifluoroethylene) [P(VDF-TrFE) copolymer], poly(vinylidene fluoride-chlorofluoroethylene) [P(VDF-CFE) copolymer], poly(vinylidene fluoride At least one of poly(vinylidene fluoride-trifluoroethylene) [P(VDF-CTFE) copolymer
  • the magnetocaloric material composite pyroelectric material is a combination of any one of the following magnetocaloric materials and the pyroelectric material; wherein the magnetocaloric material includes at least one of Ru-Fe-B alloy, ferroferric oxide, iron, cobalt, nickel, and gadolinium.
  • the piezoelectric ion gel material includes at least one of a polyacrylic acid and polyacrylamide composite gel, a polyacrylic acid and chitosan composite gel, a sodium alginate and polyacrylamide composite gel, a polyacrylic acid and choline composite gel, a sodium alginate and choline composite gel, a methacrylated gelatin and choline composite gel, and a methacrylated hyaluronic acid and choline composite gel.
  • the stem cells include at least one of totipotent stem cells, pluripotent stem cells, induced pluripotent stem cells and adult unipotent stem cells.
  • the totipotent stem cells include embryonic stem cells.
  • the pluripotent stem cells include mesenchymal stem cells.
  • the adult unipotent stem cells include at least one of corneal stem cells, neural stem cells, endothelial progenitor cells and endothelial stem cells.
  • the hydrogel is at least one of the following materials: alginate, chitosan, gelatin and its derivatives, collagen and its derivatives, hyaluronic acid and its derivatives, extracellular matrix proteins and their derivatives, silk fibroin and its derivatives, agarose, carrageenan, dextran, basement membrane matrix, polycaprolactone, polyethylene glycol and its derivatives, propylene glycol and ethylene oxide block polymers, polyvinyl pyrrolidone.
  • the hydrogel layer has a thickness of 50-500 ⁇ m.
  • the exosome-controlled tissue repair material provided in the present application regulates the behavioral functions of cells in damaged tissues and organs in an orderly manner through the sustained and controllable release of stem cell exosome vesicles in the material mediated by an external field, thereby promoting the regeneration and repair of tissues and organs.
  • FIG 2 is a flowchart of the steps of the method for preparing the exosome-controlled tissue repair material provided in the present application, including the following steps S110 to S130. The implementation method of each step is described in detail below.
  • Step S110 preparing the flexible substrate.
  • the step of preparing the flexible substrate specifically includes the following steps: preparing the flexible substrate by a casting method or a spin coating method.
  • Step S120 growing the stem cells on the surface of the flexible substrate.
  • a planar or three-dimensional structure is formed on the surface of the flexible substrate by photolithography, plasma dry etching or mechanical processing, and the three-dimensional structure includes at least one of a microgroove array, a microcone array and a micropillar array.
  • the seeding density of the stem cells is 10 3 -10 6 cells/cm 2 .
  • Step S130 pre-crosslinking the hydrogel on the surface of the flexible substrate so that the stem cells are encapsulated on the surface of the flexible substrate to obtain the exosome-controlled tissue repair material.
  • the cross-linking method is at least one of ionic cross-linking and ultraviolet light cross-linking.
  • the method for preparing exosome-controlled tissue repair materials Compared with existing exosome-controlled tissue repair materials, the method for preparing exosome-controlled tissue repair materials provided in the present application does not require complicated preparation processes and integrated packaging technologies, and the preparation process is simple.
  • the prepared exosome-controlled tissue repair materials can be used for the regeneration and repair of tissues and organs.
  • An exosome-controlled tissue repair material based on an active interface comprises a functional flexible substrate, stem cells adhered and grown on the functional flexible substrate, and a hydrogel for stem cell encapsulation; wherein the functional flexible substrate is made of a piezoelectric material, the stem cells are mesenchymal stem cells, and the hydrogel is sodium alginate cross-linked with calcium ions.
  • the surface morphology of the functional flexible substrate is a flat, structureless surface with a thickness of 50 ⁇ m.
  • Its material is polyvinylidene fluoride that can generate electricity under the mediation of an ultrasonic external field;
  • the inoculated mesenchymal stem cells are mouse bone marrow-derived mesenchymal stem cells;
  • the hydrogel material is sodium alginate cross-linked by calcium ions with a thickness of 50 ⁇ m.
  • the method for preparing the exosome-controlled tissue repair material based on the active interface comprises the following steps:
  • a polyvinylidene fluoride solution with a concentration of 10 w/v% is prepared; the polyvinylidene fluoride solution is cast on a flat surface of a silicon wafer, and dried at 80 degrees Celsius to obtain a functional flexible substrate with a planar surface morphology; the obtained functional flexible substrate is irradiated with a cobalt source at a dose of 15 kGy and an irradiation time of 30 minutes to complete the material sterilization.
  • the cells were seeded on the surface of a sterilized functional flexible substrate at a seeding density of 10 6 cells/cm 2 .
  • 3w/v% sodium alginate hydrogel prepolymer was used to cover the surface of the functional flexible substrate inoculated with stem cells, and then a 0.3M calcium chloride aqueous solution was added dropwise to complete the cross-linking of the encapsulated hydrogel through ionic crosslinking to obtain an exosome-controlled tissue repair material based on an active interface.
  • An exosome-controlled tissue repair material based on an active interface comprises a functional flexible substrate, stem cells adhered and grown on the functional flexible substrate, and a hydrogel for stem cell encapsulation; wherein the functional flexible substrate is composed of a photodeformable material composited with a piezoelectric material, the stem cells are embryonic stem cells, and the hydrogel is methacrylated gelatin.
  • the surface morphology of the functional flexible substrate is a microgroove array structure (microgroove width: 50 ⁇ m, height: 50 ⁇ m, spacing: 50 ⁇ m), the thickness of the substrate material is 500 ⁇ m, and the material is a polyvinylidene fluoride and azobenzene complex that can generate electricity under visible light irradiation; the inoculated embryonic stem cells are mouse embryonic stem cells; the hydrogel material is methacrylated gelatin with a thickness of 500 ⁇ m.
  • the method for preparing the exosome-controlled tissue repair material based on the active interface comprises the following steps:
  • a mixed solution with a polyvinylidene fluoride concentration of 5 w/v% and an azobenzene concentration of 1 w/v% was prepared; the mixed solution was cast on the surface of a microgroove array structure (microgroove width: 50 ⁇ m, height: 50 ⁇ m, spacing: 50 ⁇ m) template obtained by mechanical processing, and dried at 80 degrees Celsius to obtain a functional flexible substrate with a microgroove array surface morphology; the obtained functional flexible substrate was irradiated with a cobalt source at a dose of 15 kGy and an irradiation time of 30 minutes to complete the material sterilization.
  • a microgroove array structure microgroove width: 50 ⁇ m, height: 50 ⁇ m, spacing: 50 ⁇ m
  • the cells were seeded on the surface of a sterilized functional flexible substrate at a seeding density of 10 4 cells/cm 2 .
  • the surface of the functional flexible substrate inoculated with stem cells was covered with a 10 w/v% methacrylated gelatin hydrogel prepolymer containing 0.1 w/v% 2959 photoinitiator, and then reacted under UV cross-linking instrument for 10 minutes.
  • the encapsulated hydrogel was cross-linked by UV light to obtain an exosome-controlled tissue repair material based on an active interface.
  • An exosome-controlled tissue repair material based on an active interface comprises a functional flexible substrate, stem cells adhered and grown on the functional flexible substrate, and a hydrogel for stem cell encapsulation; wherein the functional flexible substrate is made of photovoltaic material, the stem cells are induced pluripotent stem cells, and the hydrogel is sodium alginate cross-linked with calcium ions.
  • the surface morphology of the functional flexible substrate is a flat structureless surface with a thickness of 50 ⁇ m.
  • Its material is poly-3-hexylthiophene (P3HT) that can generate electricity under visible light irradiation;
  • the inoculated mouse embryonic stem cells are mouse fibroblast-derived induced pluripotent stem cells;
  • the hydrogel material is sodium alginate cross-linked by calcium ions with a thickness of 50 ⁇ m.
  • the method for preparing the exosome-controlled tissue repair material based on the active interface comprises the following steps:
  • a P3HT solution with a concentration of 30 mg/mL was prepared; the solution was poured on a flat surface of a silicon wafer and spin-coated at 60 degrees Celsius to obtain a functional flexible substrate with a planar morphology; the obtained functional flexible substrate was irradiated with a cobalt source at a dose of 15 kGy and an irradiation time of 30 minutes to complete the material sterilization.
  • the cell suspension of mouse fibroblast-derived induced pluripotent stem cells obtained by digestion of fetal trypsin was used to seed the cells on the surface of a sterilized functional flexible substrate at a seeding density of 103 cells/cm2.
  • the surface of the functional flexible substrate inoculated with stem cells was covered with 3w/v% sodium alginate hydrogel prepolymer, and then a 0.3M calcium chloride aqueous solution was added to complete the cross-linking of the encapsulated hydrogel through ionic crosslinking to obtain an exosome-controlled tissue repair material based on an active interface.
  • An exosome-controlled tissue repair material based on an active interface comprises a functional flexible substrate, stem cells adhered and grown on the functional flexible substrate, and a hydrogel for stem cell encapsulation; wherein the functional flexible substrate is made of an upconversion material composite photovoltaic material, the stem cells are mesenchymal stem cells, and the hydrogel is sodium alginate cross-linked with calcium ions.
  • the surface morphology of the functional flexible substrate is a flat and structureless surface with a thickness of 100 ⁇ m.
  • Its material is poly-3-hexylthiophene (P3HT) that can generate electricity under visible light irradiation;
  • the inoculated mesenchymal cells are mouse bone marrow-derived mesenchymal stem cells;
  • the hydrogel material is sodium alginate cross-linked by calcium ions with a thickness of 100 ⁇ m.
  • the method for preparing the exosome-controlled tissue repair material based on the active interface comprises the following steps:
  • a mixture solution containing 30 mg/mL P3HT and 0.1 mg/mL sodium gadolinium fluoride was prepared; the solution was poured on a flat surface of a silicon wafer and spin-coated at 60 degrees Celsius to obtain a functional flexible substrate with a planar morphology; the obtained functional flexible substrate was irradiated with a cobalt source at a dose of 15 kGy and an irradiation time of 30 minutes to complete the material sterilization.
  • the cells were seeded on the surface of a sterilized functional flexible substrate at a seeding density of 10 6 cells/cm 2 .
  • the surface of the functional flexible substrate inoculated with stem cells was covered with 3w/v% sodium alginate hydrogel prepolymer, and then 0.3M calcium chloride aqueous solution was added to complete the encapsulated hydrogel cross-linking through ionic cross-linking to obtain an exosome-controlled tissue repair material based on an active interface.
  • An exosome-controlled tissue repair material based on an active interface comprises a functional flexible substrate, stem cells adhered and grown on the functional flexible substrate, and a hydrogel for stem cell encapsulation; wherein the functional flexible substrate is made of a photothermal material composite with a pyroelectric material, the stem cells are mesenchymal stem cells, and the hydrogel is sodium alginate cross-linked with calcium ions.
  • the surface morphology of the functional flexible substrate is a micro-cone array surface, with a micro-cone width of 5 ⁇ m, a height of 5 ⁇ m, a spacing of 5 ⁇ m, and a substrate thickness of 200 ⁇ m.
  • the material is a complex of P(VDF-TrFE) copolymer and polydopamine that can generate electricity under near-infrared light irradiation;
  • the inoculated mesenchymal stem cells are mouse bone marrow-derived mesenchymal stem cells;
  • the hydrogel material is sodium alginate cross-linked by calcium ions, with a thickness of 200 ⁇ m.
  • the method for preparing the exosome-controlled tissue repair material based on the active interface comprises the following steps:
  • a mixed solution containing 10 w/v% polyvinylidene fluoride and 0.1 w/v% polydopamine was prepared; the mixed solution was cast on the surface of a microcone array structure (microcone width: 5 ⁇ m, height: 5 ⁇ m, spacing: 5 ⁇ m) template obtained by plasma dry etching, and dried at 80 degrees Celsius to obtain a functional flexible substrate with a microcone array surface morphology; the obtained functional flexible substrate was irradiated with a cobalt source at a irradiation dose of 15 kGy and an irradiation time of 30 minutes to complete the material sterilization.
  • a microcone array structure microcone width: 5 ⁇ m, height: 5 ⁇ m, spacing: 5 ⁇ m
  • the cells were seeded on the surface of a sterilized functional flexible substrate at a seeding density of 10 6 cells/cm 2 .
  • 3w/v% sodium alginate hydrogel prepolymer was used to cover the surface of the functional flexible substrate inoculated with stem cells, and then a 0.3M calcium chloride aqueous solution was added to complete the cross-linking of the encapsulated hydrogel through ionic cross-linking to obtain an exosome-controlled tissue repair material based on an active interface.
  • An exosome-controlled tissue repair material based on an active interface comprises a functional flexible substrate, stem cells adhered and grown on the functional flexible substrate, and a hydrogel for stem cell encapsulation; wherein the functional flexible substrate is made of a magnetocaloric material composited with a pyroelectric material, the stem cells are mesenchymal stem cells, and the hydrogel is sodium alginate cross-linked with calcium ions.
  • the surface morphology of the functional flexible substrate is a microcolumn array surface, with a microcolumn width of 5 ⁇ m, a height of 5 ⁇ m, a spacing of 5 ⁇ m, and a substrate thickness of 50 ⁇ m.
  • the material is a complex of P(VDF-TrFE) copolymer and ferroferric oxide that can generate electricity under an alternating magnetic field;
  • the inoculated mesenchymal stem cells are mouse bone marrow-derived mesenchymal stem cells;
  • the hydrogel material is sodium alginate cross-linked by calcium ions, with a thickness of 50 ⁇ m.
  • the method for preparing the exosome-controlled tissue repair material based on the active interface comprises the following steps:
  • a mixed solution containing 10 w/v% polyvinylidene fluoride and 5 mg/mL ferrosoferric oxide nanoparticles was prepared; the mixed solution was cast on the surface of a microcolumn array structure (microcolumn width: 5 ⁇ m, height: 5 ⁇ m, spacing: 5 ⁇ m) obtained by photolithography to obtain a functional flexible substrate with a microcolumn array surface morphology; the obtained functional flexible substrate was irradiated with a cobalt source at an irradiation dose of 15 kGy and an irradiation time of 30 minutes to complete the material sterilization.
  • a microcolumn array structure microcolumn width: 5 ⁇ m, height: 5 ⁇ m, spacing: 5 ⁇ m
  • the cells were seeded on the surface of a sterilized functional flexible substrate at a seeding density of 10 6 cells/cm 2 .
  • 3w/v% sodium alginate hydrogel prepolymer was used to cover the surface of the functional flexible substrate inoculated with stem cells, and then a 0.3M calcium chloride aqueous solution was added to complete the cross-linking of the encapsulated hydrogel through ionic cross-linking to obtain an exosome-controlled tissue repair material based on an active interface.
  • An exosome-controlled tissue repair material based on an active interface comprises a functional flexible substrate, stem cells adhered and grown on the functional flexible substrate, and a hydrogel for stem cell encapsulation; wherein the functional flexible substrate is made of a piezoelectric ion gel material, the stem cells are adult unipotent stem cells, and the hydrogel is sodium alginate cross-linked with calcium ions.
  • the surface morphology of the functional flexible substrate is a microcolumn array surface, with a microcolumn width of 50nm, a height of 50nm, a spacing of 50nm, and a substrate thickness of 50 ⁇ m.
  • the material is a complex of polymethyl methacrylate and choline that can generate electricity under ultrasound mediation;
  • the inoculated adult unipotent stem cells are mouse neural stem cells;
  • the hydrogel material is sodium alginate cross-linked by calcium ions, with a thickness of 50 ⁇ m.
  • the method for preparing the exosome-controlled tissue repair material based on the active interface comprises the following steps:
  • An aqueous solution containing 10 w/v% methacrylic acid, 3 w/v% choline chloride and 0.1 w/v% 2959 photoinitiator is prepared; the mixed solution is cast on the surface of a microcolumn array structure (microcolumn width: 50 nm, height: 50 nm, spacing: 50 nm) template obtained by photolithography, and the mixture is reacted for 10 minutes under irradiation of an ultraviolet cross-linking instrument to obtain a functional flexible substrate with a microcolumn array surface morphology; the obtained functional flexible substrate is irradiated by a cobalt source with an irradiation dose of 15 kGy and an irradiation time of 30 minutes to complete the material sterilization.
  • a microcolumn array structure microcolumn width: 50 nm, height: 50 nm, spacing: 50 nm
  • the cells were seeded on the surface of the sterilized functional flexible substrate at a seeding density of 10 6 cells/cm 2 .
  • 3w/v% sodium alginate hydrogel prepolymer was used to cover the surface of the functional flexible substrate inoculated with stem cells, and then a 0.3M calcium chloride aqueous solution was added to complete the cross-linking of the encapsulated hydrogel through ionic cross-linking to obtain an exosome-controlled tissue repair material based on an active interface.

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Abstract

L'invention concerne un matériau de réparation de tissu commandé par programme d'exosome, comprenant : un substrat souple (110), une cellule souche (120) se développant sur la surface du substrat flexible (110), et un hydrogel (130) conçu pour encapsuler la cellule souche (120) sur la surface du substrat flexible (110). La cellule souche (120) peut libérer en continu des vésicules de type exosomes contenant des molécules bioactives. Le substrat souple (110) peut convertir l'énergie de champ externe en électricité et stimuler la cellule souche (120) se développant sur sa surface pour guider la différenciation de manière directionnelle, de telle sorte que la proportion et la teneur de différents types de molécules bioactives dans les vésicules de type exosomes libérées sont régulées et contrôlées, répondant ainsi aux demandes différenciées pour les molécules bioactives par des tissus et des organes à différents stades de régénération et de réparation de ceux-ci. Par comparaison avec des matériaux de réparation de tissu à base de vésicule de type exosome existants, le matériau de réparation de tissu commandé par programme d'exosome, au moyen de la libération continue et contrôlable des vésicules de type exosome de la cellule souche (120) dans un matériau médié par un champ externe, régule et commande de manière ordonnée les fonctions comportementales de cellules au niveau de tissus et d'organes endommagés, favorisant ainsi la régénération et la réparation des tissus et des organes.
PCT/CN2022/130867 2022-11-09 2022-11-09 Matériau de réparation de tissu commandé par programme d'exosome et son procédé de préparation Ceased WO2024098285A1 (fr)

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CN119345434A (zh) * 2024-11-08 2025-01-24 四川大学 一种超声激发的抗菌纤维创面修复材料制备方法和应用
CN119424742A (zh) * 2024-11-04 2025-02-14 山东师范大学 一种用于原位3d打印的海洋多糖基生物墨水及其制备方法与应用

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