WO2017111160A1 - Matériau composite d'activation cellulaire, procédé de fabrication d'un matériau composite d'activation cellulaire et kit de culture cellulaire - Google Patents

Matériau composite d'activation cellulaire, procédé de fabrication d'un matériau composite d'activation cellulaire et kit de culture cellulaire Download PDF

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Publication number
WO2017111160A1
WO2017111160A1 PCT/JP2016/088700 JP2016088700W WO2017111160A1 WO 2017111160 A1 WO2017111160 A1 WO 2017111160A1 JP 2016088700 W JP2016088700 W JP 2016088700W WO 2017111160 A1 WO2017111160 A1 WO 2017111160A1
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Prior art keywords
composite material
cell
cells
cell activation
culture
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PCT/JP2016/088700
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English (en)
Japanese (ja)
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土屋　利江
惠珍 朴
伊藤　博
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大日精化工業株式会社
土屋　利江
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Priority to JP2017558327A priority Critical patent/JP6680802B2/ja
Publication of WO2017111160A1 publication Critical patent/WO2017111160A1/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified 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
    • A61L27/14—Macromolecular materials
    • A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
    • A61L27/24—Collagen
    • 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
    • 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
    • C12N11/00—Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof
    • C12N11/02—Enzymes or microbial cells immobilised on or in an organic carrier
    • C12N11/04—Enzymes or microbial cells immobilised on or in an organic carrier entrapped within the carrier, e.g. gel or hollow fibres

Definitions

  • the present invention relates to a composite material for cell activation, a method for producing a composite material for cell activation, a cell culture kit, and use of the composite material for cell activation.
  • Patent Document 1 In growth culture and differentiation induction culture, for example, a factor such as cytokine is added to the cultured cells in order to promote the proliferation and differentiation induction of cultured animal cells (Patent Document 1).
  • a medium capable of maintaining the immunosuppressive function of transplanted mesenchymal stem cells is known (Patent Document 2).
  • animal cells are cultured while promoting proliferation in the presence of an extracellular matrix derived from mesenchymal cells, epithelial cells, cartilage precursor cells, or fibroblasts (Patent Document 3).
  • Patent Document 4 Specific immune rejection can be exempted by transplanting a dried product excellent in biocompatibility obtained by drying the produced extracellular matrix
  • Patent Document 5 an implant composed of chondrocytes and an extracellular matrix for treating sports disorders, osteoarthritis and the like is known (Patent Document 5).
  • Patent Document 6 a porous support for well growing and surviving cells that are uniformly and stably held and engrafted is known (Patent Document 6). Furthermore, a method for activating cells for transplantation in an electrostatic field atmosphere (Patent Document 7), a method for increasing only the production of extracellular matrix without growing cells (Patent Document 8), etc. are known. It has been.
  • the present invention has been made in view of such problems of the prior art, and the problem is to effectively activate cultured cells and cells of damaged tissues both in vivo and in vitro. It is an object of the present invention to provide a cell activation composite material that can be easily handled and can promote proliferation and differentiation induction. Moreover, the place made into the subject of this invention is providing the manufacturing method of said composite material for cell activation. Furthermore, the subject of the present invention is to provide a cell culture kit using the above-mentioned cell activation composite material, and the use of the above-mentioned cell activation composite material for promoting the growth of cultured cells and culturing them. There is to do.
  • the present inventors have lyophilized a composite material precursor obtained by supporting cells inside a porous support made of a biocompatible material. As a result, the present invention has been completed.
  • the following composite material for cell activation is provided.
  • a cell activation composite material obtained by freeze-drying a composite material precursor in which cells are supported inside a porous support made of a biocompatible material.
  • the composite material for cell activation according to [1] including the support and freeze-dried cells that are lyophilized from the cells and supported in the pores of the support.
  • the support is a sponge-like support.
  • a freeze-dried cell mass in which five or more spherical freeze-dried cells are collected is observed [2] to [2]
  • the manufacturing method of the composite material for cell activation shown below is provided.
  • a method for producing a composite material for cell activation comprising: a step (1) for obtaining a composite material precursor; and a step (2) for freeze-drying the obtained composite material precursor.
  • the following cell culture kit is provided.
  • a cell culture kit comprising the composite material for cell activation according to any one of [1] to [8].
  • a culture container is further provided, and the culture container has a culture cell storage unit that stores the culture cell, and the cell at a position that directly or indirectly contacts the culture cell stored in the culture cell storage unit.
  • the composite material for cell activation according to the present invention is capable of effectively activating cultured cells and cells of damaged tissues and promoting their proliferation and differentiation induction both in vivo and in vitro. It is easy. Further, according to the method for producing a composite material for cell activation of the present invention, the composite material for cell activation can be produced. Furthermore, according to the present invention, it is possible to provide a cell culture kit using the above-mentioned cell activation composite material, and use of the above-mentioned cell activation composite material for cultivating cells while promoting the growth of cultured cells. it can.
  • FIG. 3 is an electron micrograph showing the microstructure of the cell activation composite material of Example 1.
  • FIG. 3 is an electron micrograph showing the microstructure of the cell activation composite material of Example 2.
  • FIG. 4 is an electron micrograph showing the microstructure of the cell activation composite material of Example 3.
  • composite material for cell activation of the present invention (hereinafter also simply referred to as “composite material”) is obtained by lyophilizing a composite material precursor in which cells are supported inside a porous support made of a biocompatible material. It is obtained. The details will be described below.
  • factors that activate live cells and promote their proliferation and differentiation are intracellular. It is included as an ingredient. And it is thought that the intracellular component (various factors) which flowed out from the cell whose cell membrane was broken by freeze-drying is supported in the pores of the support existing around. For this reason, when the composite material of the present invention is arranged so as to come into contact with cultured cells or damaged tissues, various factors carried in the pores are relatively quickly released from the support at the beginning of the culture start or the healing start. Released.
  • the composite material of the present invention is such that intracellular components including freeze-dried cells and various factors are carried inside the porous support (more specifically, in the pores), the support It is easier to handle than lyophilized cells themselves that do not have Furthermore, since the cell activation is not inhibited by the support, it is possible to effectively activate cultured cells and cells of damaged tissues to promote their proliferation and differentiation induction.
  • the porous support constituting the composite material of the present invention is formed of a biocompatible material.
  • the biocompatible material may be biodegradable or non-biodegradable.
  • the biocompatible material may be a natural product or a synthetic product. These materials may be chemically modified with a functional group such as a sulfate group.
  • biocompatible materials include proteins such as gelatin, collagen, elastin, fibroin and albumin; polysaccharides such as hyaluronic acid, cellulose, chitin, chitosan, starch, pectin, alginic acid, agarose and agar And derivatives thereof. Moreover, it is also possible to use the commercial item which used these natural products as a raw material.
  • biocompatible materials specific examples of synthetic materials include peptides, polylactic acid, polyglycolic acid, poly- ⁇ -caprolactone and other polymers; copolymers of monomers constituting these polymers; ceramics and the like Can be mentioned. These biocompatible materials can be used singly or in combination of two or more.
  • biodegradable materials are preferably subjected to insolubilization treatment such as crosslinking.
  • insolubilization treatment such as crosslinking.
  • biodegradable materials such as gelatin, collagen, and synthetic peptides because of their adhesiveness to tissues.
  • synthetic peptide a synthetic peptide synthesized organically, a synthetic peptide produced by genetic engineering, or the like can be used. More specifically, in addition to artificial collagen and artificial gelatin imitating natural peptides, peptides having novel physical properties can be used.
  • a large number of pores for holding freeze-dried cells are formed inside the support.
  • As the support in addition to a sponge-like support, non-woven fabric, woven fabric, or the like can be used.
  • the pore diameter is preferably 5 to 1000 ⁇ m, more preferably 10 to 500 ⁇ m.
  • a cell suspension obtained by suspending cells in a liquid medium that has been brought into contact with a support to produce the composite material of the present invention enters the pores. It's easy to do.
  • the cells are unlikely to flow out of the pores, but the liquid medium is preferable because it flows out easily.
  • the composite material of the present invention is used for effectively activating cultured cells and damaged tissue cells to promote their proliferation and differentiation induction.
  • the shape maintaining period of the support is preferably a period until cell proliferation and differentiation induction are completed (in vivo, a period until treatment and repair of the affected area are completed).
  • the shape maintaining period of the support can be adjusted by introducing intermolecular crosslinks into the biodegradable material.
  • the biodegradable material is a protein
  • it can be crosslinked by chemical crosslinking, physical crosslinking, or crosslinking using an enzyme.
  • a chemical crosslinking agent such as formalin, glutaraldehyde, ethylene glycol diglycidyl ether, polyethylene glycol diglycidyl ether, polyglycerol polyglycidyl ether, glycerol polyglycidyl ether, hexamethylene diisocyanate can be used.
  • biodegradable material For physical crosslinking, ⁇ rays, ultraviolet rays, or the like can be used. In addition, an enzyme such as transglutaminase can be used for crosslinking using an enzyme.
  • a dehydrating agent such as carbodiimide, a Ca salt, or the like in addition to the chemical crosslinking agent.
  • the size and shape of the support are not particularly limited, and may be a size and a shape corresponding to the application.
  • Specific examples of the shape of the support include a cube, a cylinder, and a sheet.
  • Freeze-dried cells are formed by freeze-drying the cells carried inside the porous support.
  • Examples of cells include non-human embryonic stem cell lines, human embryonic stem cell lines, induced pluripotent stem cells, spermatogonial stem cells, embryonic stem cells, mesenchymal stem cells, endothelial stem cells, hematopoietic stem cells, neural stem cells, neural crest stem cells, and gastrointestinal stem cells.
  • Stem cells such as bone marrow, adipose tissue, placental tissue, umbilical cord tissue, umbilical cord blood, and dental pulp; human dermal fibroblasts, endothelial cells, gastrointestinal epithelial cells, mammary epithelial cells, melanocytes , Lung epithelial cells, renal epithelial cells, keratinocytes, urothelial cells, hepatocytes, corneal cells, lens cells, osteoblasts, bone cells, odontoblasts, chondrocytes, ligament cells, tendon cells, pituitary gland Cells, salivary gland cells, adrenal cells, exocrine pancreatic cells, endocrine pancreatic cells, Leydig cells, adipocytes, fibroblasts, retinal cells, dopaminergic neurons, GABAergic neurons, glutamatergic Uron, cholinergic neuron, adrenergic neuron, astrocyte, oligodendrocyte, Schwann cell, smooth
  • components other than freeze-dried cells and intracellular components may be further supported in the pores of the support.
  • other components include a medium-containing component, hyaluronic acid, a chemically modified product of hyaluronic acid, iron sulfate (FeSO 4 ), chondroitin sulfate, and various cytokines.
  • the porous support constituting the composite material of the present invention is manufactured using a raw material collected from a breeding-managed animal such as prion or a contamination-free synthetic material. Is preferred.
  • the composite material of the present invention is preferably substantially free of viable bacteria. By substantially not containing viable bacteria, it can be suitably used for culturing cultured cells and transplanting to a site in a living body (such as a human or non-human animal) in which cells or tissues are defective. .
  • substantially free of viable bacteria means that microorganisms (bacteria or fungi, etc.) are detected when tested according to the “sterility test method” defined in the Japanese Pharmacopoeia General Test Method. Means not.
  • the composite material may be produced by an aseptic operation.
  • a composite material is transplanted into a site in a living body (such as a human or a non-human animal) in which cells or tissues are defective.
  • a living body such as a human or a non-human animal
  • various factors released from the composite material can be effectively acted on surrounding cells, and cell proliferation and differentiation induction can be promoted to promote healing.
  • the composite material of the present invention utilizes various factors that promote cell proliferation and differentiation induction, not the cultured cells themselves. For this reason, the composite material of the present invention is not subject to so-called regenerative medicine regulation and can be easily spread widely.
  • the method for producing a composite material for cell activation of the present invention includes a step of obtaining a composite material precursor by supporting cells in pores of a porous support made of a biocompatible material (step (1)), A step of freeze-drying the obtained composite material precursor (step (2)). The details will be described below.
  • step (1) cells are supported in the pores of the porous support described above to obtain a composite material precursor.
  • a cell suspension in which cells are suspended in a liquid medium is brought into contact with the support, and the cell suspension enters (soaks) into the pores.
  • the number of cells in the composite material precursor is preferably 10 4 or more, and more preferably 10 7 or more, per apparent volume of 125 mm 3 of the support.
  • step (2) the composite material precursor obtained in step (1) is freeze-dried.
  • the composite material for cell activation of this invention can be obtained.
  • the predetermined time is preferably a short time, and is preferably within 60 minutes. If it exceeds 60 minutes, undesirable effects may appear on the cells.
  • the composite material precursor can be freeze-dried without using a water-soluble freeze-drying protective agent such as dimethyl sulfoxide (DMSO) or glycerin that is generally used for freeze-drying cells.
  • DMSO dimethyl sulfoxide
  • the composite material precursor is preferably frozen at ⁇ 50 ° C. or lower, and more preferably at ⁇ 80 ° C. or lower.
  • Freeze-drying can be performed using, for example, a general freezer or a cryogenic freezer such as a deep freezer.
  • the composite material precursor is frozen using a cooling medium (liquefied gas) such as carbon dioxide ( ⁇ 78.5 ° C.), liquid nitrogen ( ⁇ 196 ° C.), liquid helium ( ⁇ 269 ° C.), and then dried.
  • a cooling medium liquefied gas
  • lyophilization using an inert gas liquefied gas such as nitrogen or helium is preferable because it has little influence on cells and high safety.
  • it may be frozen by lowering the temperature step by step, or it may be frozen by lowering the temperature at once, but freezing the precursor of the composite material by lowering the temperature at once promotes proliferation and differentiation induction.
  • the composite material superior in ability to be obtained can be obtained. Specifically, since the ratio of frozen cells whose original shape is maintained without breaking the cell membrane can be increased, the composite material has a higher ability to promote proliferation and differentiation induction. After freezing, it is preferable to immediately sublimate water under reduced pressure and dry, as long as the composite material precursor is frozen during drying by sublimation. It may be dried using a freeze dryer, or may be quickly put into a sealed container after freezing and dried under reduced pressure.
  • the cell culture kit of the present invention comprises the above-described composite material for cell activation of the present invention.
  • the above-described composite material can effectively act on cultured cells and activate the cultured cells to promote proliferation. For this reason, by using this composite material, it is possible to construct a cell culture kit capable of culturing various cultured cells that have been slow to grow and difficult to culture.
  • the configuration of the cell culture kit of the present invention can be the same as that of a general conventional cell culture kit except that the above-described composite material is provided.
  • the size (diameter) of a well, the number of wells per one cell culture plate, etc. are not specifically limited.
  • the diameter of the well is, for example, 0.64 to 35 mm.
  • the number of wells per cell culture plate is, for example, 4 to 96.
  • a petri dish as shown in FIG. 2 can also be used as the culture container 20. That is, a cell culture kit can be obtained by housing the composite material in the culture vessel 20.
  • the size (diameter) of the petri dish is not particularly limited. The diameter of the petri dish is, for example, 35 to 150 mm. Commercially available cell culture plates and petri dishes can be used.
  • the cultured cells and the medium are accommodated in the well 5 and a composite material is disposed so as to be in contact with the accommodated cultured cells and medium. That's fine.
  • the cultured cells and the medium are accommodated in the culture container 20 and the composite material is brought into contact with the accommodated cultured cells and medium. What is necessary is just to arrange.
  • FIG. 3 is a schematic view showing one embodiment of the cell culture kit of the present invention.
  • the cell culture kit 100 of the embodiment shown in FIG. 3 includes a culture container 50 having a cultured cell storage unit 35 and a composite material storage unit 45.
  • the cultured cell storage unit 35 is a part (space) that stores the cultured cells 31 and the culture medium 33.
  • the composite material housing 45 is a portion (space) in which the composite material 43 is disposed.
  • the composite material 43 accommodated in the composite material accommodating portion 45 indirectly with the cultured cells 31 accommodated in the cultured cell accommodating portion 35 with the diaphragm 47 constituting the bottom of the composite material accommodating portion 45 interposed therebetween. Contact.
  • the diaphragm 47 is a film in which various components and medium components flowing out from the composite material 43 are permeated, but the placed composite material 43 is formed with a hole of a size that does not fall downward. With the configuration as shown in FIG. 3, various factors released from the composite material can be cultured while acting on cultured cells without applying an excessive load, and promoting proliferation and differentiation induction.
  • the size of the composite material to be used may be appropriately set according to the size of the well or petri dish of the cell culture plate.
  • the composite material may be manufactured in advance to have a desired size in consideration of the size of a well or the like, or may be cut to have a desired size.
  • Examples of cells (cultured cells) that can be cultured using the cell culture kit of the present invention include the same cells as those used to form the above-mentioned freeze-dried cells. Among these, it is particularly effective when culturing slow-growing cells such as mesenchymal stem cells and primary cells.
  • Example 1 The cross-linked gelatin sponge was aseptically cut into a cylindrical shape having a diameter of 6 mm and a height of 5 mm. A suspension of V79 cells (fibroblasts) (10 6 cells / 50 ⁇ L, MEM medium + 2 mmol / L glutamine) was placed on a cylindrical gelatin sponge and allowed to stand for 30 minutes. So that a composite material precursor was obtained. In addition, as V79 cell, what was obtained from JCRB cell bank (cell registration number JCRB0603, Lot No. 10012012) was used. After instantly freezing the composite material precursor using liquid nitrogen, it was transferred to a lyophilizer (trade name “FD-5N”, manufactured by EYELA Co., Ltd.) and immediately evacuated and lyophilized to obtain a composite material. .
  • a lyophilizer trade name “FD-5N”, manufactured by EYELA Co., Ltd.
  • a part of the obtained composite material was cut aseptically to prepare two sample pieces for sterility test in a cylindrical shape (diameter 6 mm, height 1 mm).
  • a sterility test was performed according to the “sterility test method” defined in the Japanese Pharmacopoeia General Test Method. Specifically, the prepared sample pieces were placed in a liquid thioglycolic acid medium and a soybean / casein / digest medium, respectively, and cultured for 14 days. As a result, since no growth of microorganisms was observed in any of the media, the composite material was judged to be substantially sterile.
  • the sterility test was performed on all manufactured composite materials in the same manner as described above, and it was confirmed that all the composite materials were substantially sterile.
  • Example 2 The composite material precursor obtained in Example 1 was frozen in a deep freezer (product number “CLN-30UW”, manufactured by Nippon Freezer Co., Ltd.) at ⁇ 80 ° C. and then freeze-dried in the same manner as in Example 1 to form a composite. Obtained material.
  • Example 3 The composite material precursor obtained in Example 1 was placed in a freezer (product number “MDF-MU300H-P”, manufactured by Panasonic Hercare) at ⁇ 20 ° C., and further frozen in stages using liquid nitrogen. . Subsequently, it lyophilized similarly to Example 1 and the composite material was obtained.
  • Comparative Example 1 The cylindrical gelatin sponge (without lyophilized cells) used in Example 1 was used as the material of Comparative Example 1.
  • Example 4 A composite material was obtained in the same manner as in Example 1 except that a suspension of V79 cells (fibroblasts) (10 5 cells / 50 ⁇ L, MEM medium + 2 mmol / L glutamine) was used.
  • Example 5 The cross-linked gelatin sponge was aseptically cut into a cylindrical shape having a diameter of 6 mm and a height of 5 mm.
  • a suspension of hMSC cells human mesenchymal stem cells (10 6 cells / 50 ⁇ L, MEM medium + 2 mmol / L glutamine) was placed on a cylindrical gelatin sponge and allowed to stand for 30 minutes.
  • a composite material precursor was obtained by soaking into a gelatin sponge.
  • hMSC cells those obtained from Lonza were used.
  • the composite material precursor was instantly frozen using liquid nitrogen, then transferred to a lyophilizer, and immediately evacuated and vacuum-dried to obtain a composite material.
  • Comparative Example 2 The cylindrical gelatin sponge (without lyophilized cells) used in Example 5 was used as the material of Comparative Example 2.
  • Example 6 A composite material was obtained in the same manner as in Example 5 except that a suspension of hMSC cells (human mesenchymal stem cells) (10 5 cells / 50 ⁇ L, MEM medium + 2 mmol / L glutamine) was used.
  • hMSC cells human mesenchymal stem cells
  • hMSC cells 4 ⁇ 10 3 cells were suspended in 2 mL of liquid medium (Lonza, MSCGM Bulletkit). The obtained suspension was placed in each well of four cell culture plates (24 wells) and cultured for 18 hours in a carbon dioxide incubator. After culturing, the composite material of Example 5 is put into a culture insert (pore size: 8 ⁇ m, manufactured by Falcon) having a hole that does not drop the composite material (Evaluation Example 11), and the material of Comparative Example 2 is used. The plate was divided into four plates (Evaluation Example 12), the composite material of Example 6 (Evaluation Example 13), and nothing (Evaluation Example 14), and cultured in a carbon dioxide incubator for 6 days.
  • the number of cells was counted using a reagent for measuring the number of living cells (trade name “WST-8” (Cell Count reagent SF), manufactured by Hanai Chemical Co., Ltd.), and the average value per well was calculated. The results are shown in Table 4.
  • the composite material for cell activation of the present invention is useful as a material for effectively activating cultured cells and cells of damaged tissues and promoting their proliferation and differentiation induction.

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Abstract

La présente invention concerne un matériau composite d'activation cellulaire facile à manipuler, qui peut activer, efficacement, à la fois in vivo et in vitro, des cellules cultivées ou des cellules provenant d'un tissu lésé et favoriser la prolifération et la différenciation de celles-ci. Le matériau composite d'activation cellulaire est obtenu par lyophilisation d'un précurseur de matière composite, dans lequel des cellules sont supportées au sein d'un corps support poreux comprenant un matériau biocompatible, tel que la gélatine ou le collagène. Le matériau composite d'activation cellulaire comprend ledit corps support ainsi que des cellules lyophilisées, qui sont des cellules qui sont lyophilisées et supportées dans les pores du corps support. Le matériau biocompatible est un matériau bioabsorbable, tel que la gélatine ou le collagène.
PCT/JP2016/088700 2015-12-24 2016-12-26 Matériau composite d'activation cellulaire, procédé de fabrication d'un matériau composite d'activation cellulaire et kit de culture cellulaire WO2017111160A1 (fr)

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2020110114A (ja) * 2019-01-15 2020-07-27 東レ・メディカル株式会社 セルカルチャーインサート

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996015818A1 (fr) * 1994-11-22 1996-05-30 Tissue Engineering, Inc. Mousses biopolymeres contenant des matieres particulaires en matrice extracellulaire
JP2008136396A (ja) * 2006-11-30 2008-06-19 Yokohama City Univ 軟骨細胞の培養方法

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996015818A1 (fr) * 1994-11-22 1996-05-30 Tissue Engineering, Inc. Mousses biopolymeres contenant des matieres particulaires en matrice extracellulaire
JP2008136396A (ja) * 2006-11-30 2008-06-19 Yokohama City Univ 軟骨細胞の培養方法

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2020110114A (ja) * 2019-01-15 2020-07-27 東レ・メディカル株式会社 セルカルチャーインサート
JP7265243B2 (ja) 2019-01-15 2023-04-26 ネッパジーン株式会社 セルカルチャーインサート

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