Scaffolding material for cell culture and cell culture vessel

Abstract

To provide cell culture scaffolding materials that have excellent fixation of cells after seeding and can increase a cell proliferation rate.SOLUTION: Provided is a cell culture scaffolding material containing synthetic resin, having a zeta potential at pH 7 of -60 mV or higher, and having a plurality of pores with an average diameter of 10 nm or more and 5 μm or less.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to a scaffold for cell culture used for culturing cells. The present invention also relates to a cell culture vessel using the scaffold for cell culture. [Background technology]

[0002] In recent years, cell medicine and next-generation medicine using stem cells have been attracting attention. In particular, human pluripotent stem cells (hPSCs), such as human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), as well as differentiated cells derived from them, are expected to be applied to drug discovery and regenerative medicine. To achieve such applications, it is necessary to safely and reproducibly culture and proliferate pluripotent stem cells and differentiated cells.

[0003] In particular, in industrial applications of regenerative medicine, it is necessary to handle a large amount of stem cells, and therefore it is necessary to support the proliferation of pluripotent stem cells using natural polymer materials, synthetic polymer materials, or feeder cells. For this reason, various culture methods using scaffolds made of natural polymer materials, synthetic polymer materials, etc. have been investigated.

[0004] For example, Patent Document 1 listed below discloses a carrier for cell culture which comprises a molded product made of a polyvinyl acetal compound or a molded product made of the polyvinyl acetal compound and a water-soluble polysaccharide, and in which the degree of acetalization of the polyvinyl acetal compound is 20 to 60 mol %.

[0005] The following Patent Document 2 discloses a composition including a first fibrous polymer scaffold material, in which the fibers of the first fibrous polymer scaffold material are aligned. It is described that the fibers constituting the first fibrous polymer scaffold material are made of an aliphatic polyester such as polyglycolic acid or polylactic acid.

[0006] Furthermore, the following Patent Document 3 discloses a cell culture method for maintaining the undifferentiated state of pluripotent stem cells, the method comprising a step of culturing the pluripotent stem cells on a culture vessel having a surface coated with a polyrotaxane block copolymer. [Prior art documents] [Patent documents]

[0007] [Patent Document 1] JP 2006-314285 A [Patent Document 2] Special Publication No. 2009-524507 [Patent Document 3] JP 2017-23008 A Summary of the Invention [Problem to be solved by the invention]

[0008] Incidentally, when a scaffold is made of a natural polymer material, the adhesion of cells after seeding can be improved. In particular, it is known that the adhesion of cells after seeding is very high when adhesive proteins such as laminin and vitronectin or Matrigel derived from mouse sarcoma are used as natural polymer materials. On the other hand, natural polymer materials are expensive, have large lot-to-lot variations due to being naturally derived substances, and have safety concerns due to animal-derived components.

[0009] In contrast, scaffolds using synthetic resins are easier to handle, less expensive, have smaller lot-to-lot variations, and are safer than scaffolds using natural polymeric materials. However, in scaffolds using synthetic resins such as those in Patent Documents 1 to 3, the synthetic resins used are highly hydrophilic, which can lead to excessive swelling of the synthetic resins. In addition, the synthetic resins have a lower affinity for cells than natural polymeric materials, which can lead to cell masses peeling off during culture. In this way, scaffolds using synthetic resins have a lower fixation property after seeding of cells, and cells may not grow sufficiently.

[0010] An object of the present invention is to provide a scaffold for cell culture and a vessel for cell culture that are excellent in terms of cell fixation after seeding and that can increase the proliferation rate of cells. [Means for solving the problem]

[0011] The cell culture scaffold material according to the present invention contains a synthetic resin, has a zeta potential of -60 mV or more at pH 7, and has a plurality of pores with an average diameter of 10 nm or more and 5 μm or less.

[0012] In a particular aspect of the cell culture scaffold according to the present invention, the zeta potential at pH 7 is +30 mV or less.

[0013] In another particular aspect of the cell culture scaffold according to the present invention, the elastic modulus at 25° C. measured by a nanoindentation method is 0.01 GPa or more.

[0014] In still another specific aspect of the scaffold for cell culture according to the present invention, the synthetic resin contains 0.2 mol % or more and 20 mol % or less of a Bronsted basic group in its constituent units.

[0015] In still another particular aspect of the scaffold for cell culture according to the present invention, the water swelling rate is 100% or less.

[0016] In still another particular aspect of the scaffold for cell culture according to the present invention, the scaffold is substantially free of animal-derived raw materials.

[0017] In still another specific aspect of the scaffold for cell culture according to the present invention, the synthetic resin contains a vinyl polymer.

[0018] In still another specific aspect of the scaffold for cell culture according to the present invention, the synthetic resin contains at least a polyvinyl alcohol derivative or a poly(meth)acrylic resin.

[0019] In still another specific aspect of the scaffold for cell culture according to the present invention, the synthetic resin includes a polyvinyl acetal resin.

[0020] In yet another specific aspect of the cell culture scaffold according to the present invention, the number of pores per unit area is 0.01 pores / μm 2 More than 1000 pieces / μm 2 The following is the result.

[0021] The cell culture vessel according to the present invention includes a cell culture scaffold material constructed according to the present invention in at least a part of a cell culture region. Effect of the Invention

[0022] According to the present invention, it is possible to provide a scaffold for cell culture and a vessel for cell culture which are excellent in terms of cell fixation after seeding and can increase the proliferation rate of cells. [Brief description of the drawings]

[0023] [Figure 1] FIG. 1 is a schematic front cross-sectional view showing a cell culture vessel according to an embodiment of the present invention. [Diagram 2] 1 is a SEM image at a magnification of 1000 times of the scaffold for cell culture obtained in Example 1. [Diagram 3] 1 is an SEM image at a magnification of 10,000 times of the scaffold for cell culture obtained in Example 1. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0024] The present invention will be clarified below by describing specific embodiments of the present invention with reference to the drawings.

[0025] The cell culture scaffold according to the present invention is a scaffold containing a synthetic resin. The cell culture scaffold has a zeta potential of -60 mV or more at pH 7. The cell culture scaffold has a plurality of pores with an average diameter of 10 nm or more and 5 μm or less.

[0026] Since the scaffold for cell culture of the present invention has the above-mentioned configuration, it has excellent adhesion to cells after seeding and can increase the proliferation rate of the cells.

[0027] Conventional cell culture scaffolds using natural polymer materials can increase the fixation of cells after seeding, but they are expensive, have large lot-to-lot variations due to being naturally derived substances, and have safety concerns due to animal-derived components. On the other hand, scaffolds using synthetic resins may cause cell masses to peel off during culture because the synthetic resins swell excessively or have low affinity with cells. Therefore, scaffolds using synthetic resins may have low fixation of cells after seeding, and cells may not grow sufficiently.

[0028] In contrast, the scaffold for cell culture according to the present invention has a zeta potential at pH 7 of -60 mV or more.

[0029] Furthermore, the present inventors have focused on the structure of such a cell culture scaffold and found that the cell proliferation rate can be increased by having a plurality of pores with an average diameter of 10 nm or more and 5 μm or less.

[0030] Although the reason for this is unclear, by forming a structure in which a plurality of pores having an average diameter within a specific range as described above are provided, it is possible to form both cell adhesion parts, which are parts where no pores are provided, and cell non-adhesion parts, which are parts where pores are provided. Therefore, it is possible for cells to move freely while maintaining the fixation of cells in the cell adhesion parts. Therefore, it is believed that it is possible to enhance the migration ability of cells and the proliferation ability of cells.

[0031] Therefore, the scaffold for cell culture of the present invention can enhance adhesiveness to cells after seeding, thereby increasing the proliferation rate of the cells.

[0032] Furthermore, since the present invention can use synthetic resins as described above, it is easier to handle, less expensive, has less variation between lots, and is safer than scaffolding materials made from natural polymer materials.

[0033] It is preferable that the scaffold for cell culture of the present invention is substantially free of animal-derived raw materials. In this case, the scaffold for cell culture can be made even safer. Here, "substantially free of animal-derived raw materials" means that the content of animal-derived raw materials in the scaffold for cell culture is 3% by weight or less. However, it is more preferable that the scaffold for cell culture of the present invention is completely free of animal-derived raw materials.

[0034] In the cell culture scaffold of the present invention, the average pore size is preferably 50 nm or more, more preferably 200 nm or more, even more preferably 500 nm or more, and preferably 4.5 μm or less, more preferably 4 μm or less, even more preferably 3.5 μm or less. When the average pore size is within the above range, the fixation of cells after seeding can be further improved, and the proliferation rate of the cells can be further improved.

[0035] In the cell culture scaffold of the present invention, the number of pores per unit area is preferably 0.01 pores / μm 2 More preferably, 0.1 particles / μm 2 More preferably, 1 particle / μm 2 More preferably, 10 particles / μm 2 More than 1000 / μm 2 Less than or equal to 500 / μm 2 Less than 100 / μm, more preferably 2 When the number of pores per unit area is within the above range, the fixation of cells after seeding can be further improved, and the proliferation rate of the cells can be further increased.

[0036] In this specification, the average diameter and number of pores can be measured by a scanning electron microscope (SEM), a transmission electron microscope (TEM), or the like. In this case, the average value of the pore diameters of any 100 pores confirmed by observation with a microscope such as a scanning electron microscope (SEM), a transmission electron microscope (TEM), or the like can be regarded as the average diameter of the pores. The number of pores per unit area can be obtained by dividing the number of pores in an observation area (5 μm×5 μm) by the area of ​​the observation area. The number of pores can be, for example, the average value of the number of pores obtained by repeating the same operation 10 times. In addition, when measuring the average diameter and number of pores, pores with a pore diameter of 10 nm or more and 5 μm or less are judged to be pores.

[0037] In addition, the multiple pores can be formed, for example, by applying a solution containing a synthetic resin and an alcohol onto a substrate, freezing it with liquid nitrogen or the like, and then freeze-drying it. In this case, it is considered that the pores are formed by the parts from which the alcohol has been removed. In addition, when a synthetic resin that is difficult to expand, such as a polyvinyl acetal resin, is used, the size of the pores can be made more uniform, so that pores with a controlled average diameter can be formed more easily.

[0038] The average diameter and number of pores can be controlled by adjusting the type and amount of alcohol added, or by adding a small amount of water to the alcohol.

[0039] In the present invention, a scaffold for cell culture may be formed by crushing a synthetic resin having a plurality of pores formed therein and disposing the crushed synthetic resin on a substrate. A scaffold for cell culture may also be formed in the form of a particle or fiber having a plurality of pores.

[0040] In the present invention, the zeta potential of the cell culture scaffold at pH 7 is preferably -40 mV or more, more preferably -35 mV or more, even more preferably -30 mV or more, preferably +30 mV or less, more preferably less than +30 mV, even more preferably less than +25 mV. In this case, the adhesiveness of cells can be further increased.

[0041] The zeta potential of a cell culture scaffold is the potential difference between the surface of the cell culture scaffold, which is the interface, and a bulk portion in the solution sufficiently distant from the surface of the cell culture scaffold when the cell culture scaffold is placed in a solution. The value of the zeta potential in this specification can be determined by appropriately selecting a method such as a streaming potential method, an electrophoresis method, or an electroosmosis method depending on the shape of the cell culture scaffold.

[0042] For example, when the cell culture scaffold is a membrane, it can be measured by a streaming potential method, and when the cell culture scaffold is a fiber or particle, it can be measured by an electroosmosis method.

[0043] The streaming potential method can be measured, for example, by a zeta potential measuring device manufactured by Anton Paar. Specifically, the measurement can be performed in a dilute KCl solution by the method described in the Examples below. The electrophoretic method can be measured, for example, by light scattering in water using a zeta potential / particle size / molecular weight measuring system (manufactured by Otsuka Electronics Co., Ltd.).

[0044] The zeta potential can be increased by increasing the content of cationic functional groups, such as amino groups, in the synthetic resin, and can be decreased by increasing the content of anionic functional groups, such as carboxyl groups, in the synthetic resin.

[0045] The elastic modulus of the cell culture scaffold of the present invention measured by nanoindentation at 25° C. is preferably 0.01 GPa or more, more preferably 0.05 GPa or more, even more preferably 0.1 GPa or more, particularly preferably 0.5 GPa or more, preferably 5 GPa or less, more preferably 3 GPa or less, and even more preferably 1 GPa or less. When the elastic modulus is within the above range, the adhesiveness of cells can be further improved.

[0046] The elastic modulus can be measured, for example, by using a nanoindenter (Hysitron, Tryboindenter, manufactured by Bruker Corp.) In this measurement, a Berkovich (triangular pyramid-shaped) indenter with a tip diameter R of 100 nm is used, and a single indentation measurement is performed in air at room temperature, with the indentation depth being 50 nm.

[0047] The cell culture scaffold of the present invention has a water swelling rate of preferably 100% or less, more preferably 50% or less, and even more preferably 40% or less. In this case, the fixation of cells after seeding can be further improved. The lower limit of the water swelling rate is not particularly limited, but can be, for example, 0.5%. The water swelling rate can be measured, for example, by the following method.

[0048] First, the scaffold for cell culture is immersed in water at 25°C for 24 hours. Next, the scaffold for cell culture is removed from the water and the water remaining in the pores is drained by placing it on a water-absorbent material, etc. Next, the weight of the sample before and after immersion is measured, and the water swelling rate = (sample weight after immersion - sample weight before immersion) / (sample weight before immersion) x 100 (%) is calculated.

[0049] The water swelling rate can be reduced, for example, by increasing the hydrophobic functional groups of the synthetic resin, decreasing the number average molecular weight, or the like.

[0050] [Synthetic resin] The cell culture scaffold of the present invention contains a synthetic resin (hereinafter, may be referred to as synthetic resin X). In this specification, the term "structural unit" refers to a repeating unit of a monomer constituting the synthetic resin. In addition, when the synthetic resin has a graft chain, it includes a repeating unit of a monomer constituting the graft chain.

[0051] The synthetic resin X preferably contains 0.2 mol% or more of the Bronsted basic group in the structural unit, more preferably 2 mol% or more, more preferably 30 mol% or less, more preferably 20 mol% or less, and even more preferably 15 mol% or less. In this case, the effect of the present invention can be more effectively exhibited. The Bronsted basic group and the like will be described later.

[0052] (vinyl polymer) The synthetic resin X is preferably a vinyl polymer. Incidentally, a vinyl polymer is a polymer of a compound having a vinyl group or a vinylidene group. When the synthetic resin X is a vinyl polymer, it is possible to more easily suppress swelling of the scaffold for cell culture in water. Examples of the vinyl polymer include polyvinyl alcohol derivatives, poly(meth)acrylic acid esters, polyvinylpyrrolidone, polystyrene, ethylene-vinyl acetate copolymers, and the like. Furthermore, the vinyl polymer is preferably a polyvinyl alcohol derivative or a poly(meth)acrylic acid ester from the viewpoint of more easily increasing adhesiveness to cells.

[0053] (Synthetic resin with polyvinyl acetal structure) The cell culture scaffold preferably contains a synthetic resin having a polyvinyl acetal skeleton. The synthetic resin X preferably contains a synthetic resin having a polyvinyl acetal skeleton.

[0054] In this specification, the "synthetic resin having a polyvinyl acetal skeleton" may be referred to as "polyvinyl acetal resin X."

[0055] The polyvinyl acetal resin X preferably has an acetal group, an acetyl group, and a hydroxyl group in the side chain. However, the polyvinyl acetal resin X may not have an acetyl group, for example. For example, the polyvinyl acetal resin X may not have an acetyl group by bonding all of the acetyl groups of the polyvinyl acetal resin X to a linker described later.

[0056] The synthesis of the polyvinyl acetal resin X includes at least a step of acetalizing polyvinyl alcohol with an aldehyde.

[0057] The aldehyde used in the acetalization of polyvinyl alcohol to obtain the polyvinyl acetal resin X is not particularly limited. The aldehyde may be, for example, an aldehyde having 1 to 10 carbon atoms. The aldehyde may have a chain aliphatic group, a cyclic aliphatic group, or an aromatic group. The aldehyde may be a chain aldehyde or a cyclic aldehyde.

[0058] Examples of the aldehyde include formaldehyde, acetaldehyde, propionaldehyde, butyraldehyde, pentanal, hexanal, heptanal, octanal, nonanal, decanal, acrolein, benzaldehyde, cinnamaldehyde, perillaldehyde, formylpyridine, formylimidazole, formylpyrrole, formylpiperidine, formyltriazole, formyltetrazole, formylindole, formylisoindole, formylpurine, formylbenzimidazole, formylbenzotriazole, formylquinoline, formylisoquinoline, formylquinoxaline, formylcinnoline, formylpteridine, formylfuran, formyloxolane, formyloxane, formylthiophene, formylthiolane, formylthiane, formyladenine, formylguanine, formylcytosine, formylthymine, and formyluracil. The aldehydes may be used alone or in combination of two or more.

[0059] The aldehyde is preferably formaldehyde, acetaldehyde, propionaldehyde, butylaldehyde, or pentanal, and more preferably butylaldehyde. Therefore, the polyvinyl acetal skeleton is preferably a polyvinyl butyral skeleton. The polyvinyl acetal resin X is preferably a synthetic resin having a polyvinyl butyral skeleton, and more preferably a polyvinyl butyral resin.

[0060] A vinyl compound may be copolymerized in the polyvinyl acetal resin X. That is, the polyvinyl acetal resin X may be a copolymer of a structural unit of the polyvinyl acetal resin and a vinyl compound. In the present invention, the polyvinyl acetal resin copolymerized with a vinyl compound is also referred to as the polyvinyl acetal resin.

[0061] The vinyl compound is a compound having a vinyl group (H2C=CH-). The vinyl compound may be a polymer having a structural unit having a vinyl group.

[0062] The copolymer may be a block copolymer of a polyvinyl acetal resin and a vinyl compound, or a graft copolymer in which a vinyl compound is grafted onto a polyvinyl acetal resin. The copolymer is preferably a graft copolymer.

[0063] The copolymer can be synthesized, for example, by the following methods (1) to (3): (1) a method of synthesizing a polyvinyl acetal resin using polyvinyl alcohol copolymerized with a vinyl compound, (2) a method of synthesizing a polyvinyl acetal resin using polyvinyl alcohol and polyvinyl alcohol copolymerized with a vinyl compound, and (3) a method of graft copolymerizing a vinyl compound onto a polyvinyl acetal resin before graft copolymerization.

[0064] Examples of the vinyl compound include ethylene, allylamine, vinylpyrrolidone, vinylimidazole, maleic anhydride, maleimide, itaconic acid, (meth)acrylic acid, vinylamine, and (meth)acrylic acid ester, etc. These vinyl compounds may be used alone or in combination of two or more.

[0065] From the viewpoint of further enhancing the adhesiveness of cells, the polyvinyl acetal resin X preferably has a Brønsted basic group or a Brønsted acidic group, more preferably has a Brønsted basic group. That is, it is preferable that a part of the polyvinyl acetal resin X is modified with a Brønsted basic group or a Brønsted acidic group, and more preferably that a part of the polyvinyl acetal resin X is modified with a Brønsted basic group.

[0066] The polyvinyl acetal resin X preferably has a Brønsted basic group or a Brønsted acidic group in a part thereof, and more preferably has a Brønsted basic group. In that case, the adhesiveness of cells can be further improved. The Brønsted basic group or the Brønsted acidic group may be present in a part of the polyvinyl acetal resin X, and in that case, a monomer having a Brønsted basic group or a Brønsted acidic group may be copolymerized or grafted. The degree of modification in the Brønsted basic group is preferably 0.2 mol% or more, more preferably 2 mol% or more, preferably 30 mol% or less, more preferably 20 mol% or less, and even more preferably 15 mol% or less. If the degree of modification by the Brønsted basic group is within the above specific range, the adhesiveness of cells can be further improved.

[0067] The Bronsted basic group is a general term for a functional group that can receive a hydrogen ion H+ from another substance. Examples of the Bronsted basic group include amine-based basic groups such as a substituent having an imine structure, a substituent having an imide structure, a substituent having an amine structure, and a substituent having an amide structure. Examples of the Bronsted basic group include, but are not limited to, conjugated amine functional groups such as hydroxyamino group, urea group, guanidine, and biguanide; heterocyclic amino functional groups such as piperazine, piperidine, pyrrolidine, 1,4-diazabicyclo[2.2.2]octane, hexamethylenetetraamine, morpholine, pyridine, pyridazine, pyrimidine, pyrazine, pyrrole, azatropyridene, pyridone, imidazole, benzimidazole, benzotriazole, pyrazole, oxazole, imidazoline, triazole, thiazole, thiazine, tetrazole, indole, isoindole, purine, quinoline, isoquinoline, quinazoline, quinoxaline, cinnoline, pteridine, carbazole, acridine, adenine, guanine, cytosine, thymine, uracil, and melamine; cyclic pyrrole functional groups such as porphyrin, chlorine, and choline, and derivatives thereof.

[0068] Examples of the Bronsted acid group include a carboxyl group, a sulfonic acid group, a maleic acid group, a sulfinic acid group, a sulfenic acid group, a phosphoric acid group, a phosphonic acid group, or salts thereof. The Bronsted acid group is preferably a carboxyl group.

[0069] The polyvinyl acetal resin X preferably has a structural unit having an imine structure, a structural unit having an imide structure, a structural unit having an amine structure, or a structural unit having an amide structure. In this case, the polyvinyl acetal resin X may have only one type of these structural units or two or more types of them.

[0070] The polyvinyl acetal resin X may have a structural unit having an imine structure. The imine structure refers to a structure having a C=N bond. In particular, the polyvinyl acetal resin X preferably has an imine structure in a side chain.

[0071] The polyvinyl acetal resin X may have a structural unit having an imide structure. The structural unit having an imide structure is preferably a structural unit having an imino group (=NH).

[0072] The polyvinyl acetal resin X preferably has an imino group in a side chain. In this case, the imino group may be directly bonded to a carbon atom constituting the main chain of the polyvinyl acetal resin X, or may be bonded to the main chain via a linking group such as an alkylene group.

[0073] The polyvinyl acetal resin X may have a structural unit having an amine structure. The amine group in the amine structure may be a primary amine group, a secondary amine group, a tertiary amine group, or a quaternary amine group.

[0074] The structural unit having an amine structure may be a structural unit having an amide structure, which refers to a structure having -C(=O)-NH-.

[0075] The polyvinyl acetal resin X preferably has an amine structure or an amide structure in a side chain. In this case, the amine structure or the amide structure may be directly bonded to a carbon atom constituting the main chain of the polyvinyl acetal resin X, or may be bonded to the main chain via a linking group such as an alkylene group.

[0076] The content of the structural unit having an imine structure, the content of the structural unit having an imide structure, the content of the structural unit having an amine structure, and the content of the structural unit having an amide structure are 1 It can be measured by H-NMR (nuclear magnetic resonance spectroscopy).

[0077] (Synthetic resin having a poly(meth)acrylic acid ester skeleton) The cell culture scaffold of the present invention preferably contains a synthetic resin having a poly(meth)acrylic acid ester skeleton. The synthetic resin X preferably contains a synthetic resin having a poly(meth)acrylic acid ester skeleton.

[0078] In this specification, the "synthetic resin having a poly(meth)acrylic acid ester skeleton" may be referred to as "poly(meth)acrylic acid ester resin X".

[0079] Therefore, the poly(meth)acrylic acid ester resin X is a resin having a poly(meth)acrylic acid ester skeleton.

[0080] The poly(meth)acrylic acid ester resin X can be obtained by polymerization of a (meth)acrylic acid ester or by copolymerization of a (meth)acrylic acid ester with another monomer.

[0081] Examples of the (meth)acrylic acid ester include (meth)acrylic acid alkyl esters, (meth)acrylic acid cyclic alkyl esters, (meth)acrylic acid aryl esters, (meth)acrylamides, (meth)acrylic acid polyethylene glycols, and (meth)acrylic acid phosphorylcholine.

[0082] Examples of (meth)acrylic acid alkyl esters include methyl (meth)acrylate, ethyl (meth)acrylate, n-propyl (meth)acrylate, isopropyl (meth)acrylate, n-butyl (meth)acrylate, isobutyl (meth)acrylate, t-butyl (meth)acrylate, n-octyl (meth)acrylate, isooctyl (meth)acrylate, 2-ethylhexyl (meth)acrylate, nonyl (meth)acrylate, isononyl (meth)acrylate, decyl (meth)acrylate, isodecyl (meth)acrylate, lauryl (meth)acrylate, stearyl (meth)acrylate, and isotetradecyl (meth)acrylate.

[0083] The (meth)acrylic acid alkyl ester may be substituted with a substituent such as an alkoxy group having 1 to 3 carbon atoms and a tetrahydrofurfuryl group. Examples of such (meth)acrylic acid alkyl ester include methoxyethyl acrylate and tetrahydrofurfuryl acrylate.

[0084] Examples of the (meth)acrylic acid cyclic alkyl ester include cyclohexyl (meth)acrylate and isobornyl (meth)acrylate.

[0085] Examples of the (meth)acrylic acid aryl ester include phenyl (meth)acrylate and benzyl (meth)acrylate.

[0086] Examples of (meth)acrylamides include (meth)acrylamide, N-isopropyl(meth)acrylamide, N-tert-butyl(meth)acrylamide, N,N'-dimethyl(meth)acrylamide, (3-(meth)acrylamidopropyl)trimethylammonium chloride, 4-(meth)acryloylmorpholine, 3-(meth)acryloyl-2-oxazolidinone, N-[3-(dimethylamino)propyl](meth)acrylamide, N-(2-hydroxyethyl)(meth)acrylamide, N-methylol(meth)acrylamide, and 6-(meth)acrylamidohexanoic acid.

[0087] Examples of polyethylene glycol (meth)acrylates include methoxy-polyethylene glycol (meth)acrylate, ethoxy-polyethylene glycol (meth)acrylate, hydroxy-polyethylene glycol (meth)acrylate, methoxy-diethylene glycol (meth)acrylate, ethoxy-diethylene glycol (meth)acrylate, hydroxy-diethylene glycol (meth)acrylate, methoxy-triethylene glycol (meth)acrylate, ethoxy-triethylene glycol (meth)acrylate, and hydroxy-triethylene glycol (meth)acrylate.

[0088] Examples of the (meth)acrylate phosphorylcholine include 2-(meth)acryloyloxyethyl phosphorylcholine.

[0089] As another monomer to be copolymerized with the (meth)acrylic acid ester, a vinyl compound is preferably used. Examples of the vinyl compound include ethylene, allylamine, vinylpyrrolidone, maleic anhydride, maleimide, itaconic acid, (meth)acrylic acid, vinylamine, and (meth)acrylic acid ester. Only one type of vinyl compound may be used, or two or more types may be used in combination.

[0090] In this specification, "(meth)acrylic" means "acrylic" or "methacrylic", and "(meth)acrylate" means "acrylate" or "methacrylate".

[0091] The poly(meth)acrylic acid ester resin X preferably has a Brønsted basic group or a Brønsted acidic group in part, similar to the polyvinyl acetal resin X. In that case, the extensibility can be further improved. The Brønsted basic group may be part of the poly(meth)acrylic acid ester resin X, and in that case, a monomer having a Brønsted basic group may be copolymerized or grafted. The degree of modification in the Brønsted basic group is preferably 0.2 mol% or more, more preferably 2 mol% or more, preferably 30 mol% or less, more preferably 20 mol% or less, and even more preferably 15 mol% or less. If the degree of modification by the Brønsted basic group is within the above specific range, the adhesiveness of the cells can be further improved.

[0092] (Synthetic resin having a peptide portion) The cell culture scaffold of the present invention preferably contains a synthetic resin having a peptide portion. The synthetic resin X preferably contains a synthetic resin having a peptide portion. By using a synthetic resin having a peptide portion as a material for the cell culture scaffold, the cell can be more excellent in fixation after seeding, and the cell proliferation rate can be further increased.

[0093] The synthetic resin having a peptide portion can be obtained by reacting a synthetic resin, a linker, and a peptide. The synthetic resin having a peptide portion is preferably a peptide-containing polyvinyl acetal resin having a polyvinyl acetal resin portion, a linker portion, and a peptide portion, and more preferably a peptide-containing polyvinyl butyral resin having a polyvinyl butyral resin portion, a linker portion, and a peptide portion. Therefore, the synthetic resin having the polyvinyl acetal skeleton (polyvinyl acetal resin X) is preferably a peptide-containing polyvinyl acetal resin, and more preferably a peptide-containing polyvinyl butyral resin. Only one type of synthetic resin having a peptide portion may be used, or two or more types may be used in combination.

[0094] The peptide portion is preferably composed of 3 or more amino acids, more preferably 4 or more amino acids, even more preferably 5 or more amino acids, preferably 10 or less amino acids, and more preferably 6 or less amino acids. When the number of amino acids constituting the peptide portion is equal to or more than the above lower limit and equal to or less than the above upper limit, the adhesiveness to cells after seeding can be further increased, and the proliferation rate of the cells can be further increased.

[0095] The peptide portion preferably has a cell-adhesive amino acid sequence. The cell-adhesive amino acid sequence refers to an amino acid sequence whose cell-adhesive activity has been confirmed by the phage display method, the Sepharose bead method, or the plate coating method. As the phage display method, for example, the method described in "The Journal of Cell Biology, Volume 130, Number 5, September 1995 pp. 1189-1196" can be used. As the Sepharose bead method, for example, the method described in "Protein, Nucleic Acid, Enzyme, Vol. 45, No. 15 (2000) 2477" can be used. As the plate coating method, for example, the method described in "Protein, Nucleic Acid, Enzyme, Vol. 45, No. 15 (2000) 2477" can be used.

[0096] Examples of the cell adhesive amino acid sequences include the RGD sequence (Arg-Gly-Asp), YIGSR sequence (Tyr-Ile-Gly-Ser-Arg), PDSGR sequence (Pro-Asp-Ser-Gly-Arg), HAV sequence (His-Ala-Val), ADT sequence (Ala-Asp-Thr), QAV sequence (Gln-Ala-Val), LDV sequence (Leu-Asp-Val), IDS sequence (Ile-Asp-Ser), REDV sequence (Arg-Glu-Asp-Val), IDAPS sequence (Ile-Asp-Ala-Pro-Ser), KQAGDV sequence (Lys-Gln-Ala-Gly-Asp-Val), and TDE sequence (Thr-Asp-Glu). Further, examples of the cell adhesive amino acid sequence include the sequences described in "Pathophysiology, Vol. 9, No. 7, pp. 527-535, 1990" and "Osaka Prefectural Maternal and Child Medical Center Journal, Vol. 8, No. 1, pp. 58-66, 1992." The peptide portion may have only one type of cell adhesive amino acid sequence, or two or more types.

[0097] The cell adhesive amino acid sequence preferably has at least any one of the above-mentioned cell adhesive amino acid sequences, more preferably has at least an RGD sequence, a YIGSR sequence, or a PDSGR sequence, and even more preferably has at least an RGD sequence represented by the following formula (1). In this case, the adhesiveness to cells after seeding can be further increased, and the cell proliferation rate can be further increased.

[0098] Arg-Gly-Asp-X...Formula (1)

[0099] In the above formula (1), X represents Gly, Ala, Val, Ser, Thr, Phe, Met, Pro, or Asn.

[0100] The peptide portion may be linear or have a cyclic peptide backbone. The cyclic peptide backbone is a cyclic backbone composed of a plurality of amino acids. From the viewpoint of more effectively exerting the effects of the present invention, the cyclic peptide backbone is preferably composed of 4 or more amino acids, more preferably composed of 5 or more amino acids, and preferably composed of 10 or less amino acids.

[0101] In the synthetic resin having a peptide portion, the content of the peptide portion is preferably 0.1 mol% or more, more preferably 1 mol% or more, even more preferably 5 mol% or more, and particularly preferably 10 mol% or more. In the synthetic resin having a peptide portion, the content of the peptide portion is preferably 60 mol% or less, more preferably 50 mol% or less, even more preferably 35 mol% or less, and particularly preferably 25 mol% or less. When the content of the peptide portion is within the above range, the adhesiveness to cells after seeding can be further improved, and the cell proliferation rate can be further improved. In addition, when the content of the peptide portion is equal to or less than the above upper limit, the production cost can be reduced. The content of the peptide portion (mol%) is the amount of substance of the peptide portion relative to the sum of the amounts of substances of each structural unit constituting the synthetic resin having a peptide portion.

[0102] The content of the above peptide portion can be measured by FT-IR or LC-MS.

[0103] In the synthetic resin having a peptide portion, the synthetic resin portion and the peptide portion are preferably bonded via a linker. That is, the synthetic resin having a peptide portion is preferably a synthetic resin having a peptide portion and a linker portion. The above linkers may be used alone or in combination of two or more types.

[0104] The linker is preferably a compound having a functional group capable of condensing with the carboxyl group or amino group of the peptide. Examples of the functional group capable of condensing with the carboxyl group or amino group of the peptide include a carboxyl group, a thiol group, and an amino group. From the viewpoint of reacting well with the peptide, the linker is preferably a compound having a carboxyl group. The above-mentioned vinyl compound can also be used as the linker.

[0105] Examples of the linker having a carboxyl group include (meth)acrylic acid and acrylamide containing a carboxyl group, etc. By using a carboxylic acid (carboxylic acid monomer) having a polymerizable unsaturated group as the linker having a carboxyl group, the carboxylic acid monomer can be polymerized by graft polymerization when reacting the linker with a synthetic resin, so that the number of carboxyl groups that can be reacted with a peptide can be increased.

[0106] (Other resins) The cell culture scaffold may contain a polymer other than the above-mentioned synthetic resins, such as polyvinylpyrrolidone, polystyrene, ethylene-vinyl acetate copolymer, polyolefin resin, polyether resin, polyvinyl alcohol resin, polyester, epoxy resin, polyamide resin, polyimide resin, polyurethane resin, polycarbonate resin, etc.

[0107] [Details of other cell culture scaffolds] The cell culture scaffold according to the present invention contains the synthetic resin X. From the viewpoint of effectively exerting the effects of the present invention and increasing productivity, the content of the synthetic resin X in 100% by weight of the cell culture scaffold is preferably 90% by weight or more, more preferably 95% by weight or more, even more preferably 97.5% by weight or more, particularly preferably 99% by weight or more, and most preferably 100% by weight (total amount). Therefore, the cell culture scaffold is most preferably the synthetic resin X. When the content of the synthetic resin X is equal to or more than the lower limit, the effects of the present invention can be exerted even more effectively.

[0108] The above-mentioned cell culture scaffold may contain components other than the above-mentioned synthetic resin X. Examples of the components other than the above-mentioned synthetic resin X include polysaccharides, cellulose, synthetic peptides, and the like.

[0109] In order to effectively exert the effects of the present invention, the content of components other than the synthetic resin X is preferably as small as possible. In 100% by weight of the scaffold for cell culture, the content of the components is preferably 10% by weight or less, more preferably 5% by weight or less, even more preferably 2.5% by weight or less, particularly preferably 1% by weight or less, and most preferably 0% by weight (not contained). Therefore, it is most preferable that the scaffold for cell culture does not contain any components other than the synthetic resin X.

[0110] The cell culture scaffold according to the present invention preferably does not contain any raw materials derived from animals. By not containing any raw materials derived from animals, a cell culture scaffold having high safety and little variation in quality during production can be provided.

[0111] (Cell culture using scaffolds for cell culture) The scaffold for cell culture according to the present invention is used for culturing cells. The scaffold for cell culture according to the present invention is used as a scaffold for cells when culturing the cells.

[0112] The above cells include animal cells such as human, mouse, rat, pig, cow, and monkey cells. The above cells also include somatic cells, such as stem cells, progenitor cells, and mature cells. The above somatic cells may be cancer cells.

[0113] Examples of the stem cells include mesenchymal stem cells (MSCs), iPS cells, ES cells, Muse cells, embryonic cancer cells, embryonic germ stem cells, and mGS cells.

[0114] Examples of the mature cells include nerve cells, cardiomyocytes, retinal cells, and hepatic cells.

[0115] (Shape of scaffold for cell culture) The shape of the scaffold for cell culture of the present invention is not particularly limited, and may be a particle, a fiber, a porous body, or a film. The particle, fiber, porous body, or film may contain components other than the scaffold for cell culture.

[0116] When the scaffold for cell culture has a film shape, it is preferably used for plate culture (two-dimensional culture).When the scaffold for cell culture has a particle, fiber, or porous shape, it is preferably used for three-dimensional culture of cells.

[0117] (Cell culture container) The present invention also relates to a cell culture vessel comprising the above-mentioned cell culture scaffold in at least a part of a cell culture region. Fig. 1 is a schematic front cross-sectional view showing a cell culture vessel according to one embodiment of the present invention.

[0118] The cell culture vessel 1 comprises a vessel body 2 and a cell culture scaffold 3. The cell culture scaffold 3 is disposed on a surface 2a of the vessel body 2. The cell culture scaffold 3 is disposed on a bottom surface of the vessel body 2. By adding a liquid medium to the cell culture vessel 1 and seeding cells on the surface of the cell culture scaffold 3, the cells can be cultured on a surface or three-dimensionally.

[0119] The container body may include a second container body such as a cover glass on the bottom surface of the first container body. The first container body and the second container body may be separable. In this case, the scaffold for cell culture may be disposed on the surface of the second container body.

[0120] The container body may be a conventionally known container body (container). The shape and size of the container body are not particularly limited.

[0121] The container body may be a cell culture plate or a cell culture flask having one or more wells (holes). The number of wells in the plate is not particularly limited. The number of wells may be, but is not particularly limited to, 2, 4, 6, 12, 24, 48, 96, 384, etc. The shape of the well may be, but is not particularly limited to, a perfect circle, an ellipse, a triangle, a square, a rectangle, a pentagon, etc. The shape of the bottom of the well may be, but is not particularly limited to, a flat bottom, a round bottom, an uneven bottom, etc.

[0122] The material of the container body is not particularly limited, but examples thereof include resins, metals, and inorganic materials. Examples of the resins include polystyrene, polyethylene, polypropylene, polycarbonate, polyester, polyisoprene, cycloolefin polymers, polyimide, polyamide, polyamideimide, (meth)acrylic resins, epoxy resins, and silicone. Examples of the metals include stainless steel, copper, iron, nickel, aluminum, titanium, gold, silver, and platinum. Examples of the inorganic materials include silicon oxide (glass), aluminum oxide, titanium oxide, zirconium oxide, iron oxide, and silicon nitride. EXAMPLES

[0123] The present invention will now be described in more detail with reference to specific examples and comparative examples, although the present invention is not limited to the following examples.

[0124] The following synthetic resin X1 was synthesized as a raw material for a scaffold for cell culture.

[0125] In a reactor equipped with a stirrer, 2700 mL of ion-exchanged water and 300 parts by weight of amine-modified polyvinyl alcohol with an average degree of polymerization of 800, an amine modification degree of 1 mol%, and a saponification degree of 97.5 mol% were added, and the mixture was heated and dissolved while stirring to obtain a solution. To the obtained solution, 35% by weight hydrochloric acid was added as a catalyst so that the hydrochloric acid concentration was 0.2% by weight. Next, the temperature was adjusted to 15°C, and 22 parts by weight of n-butylaldehyde were added while stirring. Next, 148 parts by weight of n-butylaldehyde were added to precipitate a white particulate polyvinyl butyral resin. 15 minutes after the precipitation, 35% by weight hydrochloric acid was added so that the hydrochloric acid concentration was 1.8% by weight, and then the mixture was heated to 50°C and held at 50°C for 2 hours. Next, the solution was cooled and neutralized, and the polyvinyl butyral resin was washed with water and dried to obtain a synthetic resin X1, which is a polyvinyl butyral resin.

[0126] The obtained polyvinyl butyral resin (synthetic resin X1) had an average degree of polymerization of 800, a hydroxyl group amount of 20 mol %, an amine group amount of 1 mol %, an acetylation degree of 1 mol %, and a butyralization degree of 78 mol %.

[0127] The content of structural units in the obtained synthetic resin was measured by dissolving the synthetic resin in DMSO-D6 (dimethyl sulfoxide) and then 1 The measurements were performed using H-NMR (nuclear magnetic resonance spectroscopy).

[0128] (Examples 1 to 10 and Comparative Examples 2 and 3) In Examples 1 and 2, the synthetic resin X1 was used, and in Examples 3 to 7, 9 and 10, the synthetic resins X2 to X8 were used, respectively.

[0129] Synthetic resin X2 was obtained by blending 5 mol% of dimethylaminoacrylamide as a monomer with polyvinyl butyral resin having a butyralization degree of 68 mol%, an acetylation degree of 1 mol%, and a hydroxyl group amount of 26 mol%. The synthesis conditions were as follows. That is, the polyvinyl butyral resin was dissolved in THF at a concentration of 20% by weight, and 5 parts by weight of dimethylaminoacrylamide was added. After that, 3 parts by weight of Irgacure184 (manufactured by IGM Resins BV) was dissolved as an initiator, and UV irradiation was performed for 20 minutes using a UV polymerization tester.

[0130] Moreover, synthetic resin X3 was obtained in the same manner as synthetic resin X2, except that the content of dimethylaminoacrylamide was changed from 5 mol % to 20 mol % as shown in Table 1.

[0131] Synthetic resin X4 was obtained in the same manner as synthetic resin X2, except that diethylaminoacrylamide was used as the monomer and its content was 28 mol %.

[0132] Synthetic resin X5 was obtained in the same manner as synthetic resin X2, except that vinylimidazole was used as the monomer and its content was 13 mol %.

[0133] For synthetic resin X6, a synthetic resin that is a polyvinyl acetal resin was obtained in the same manner as in Example 1, except that polyvinyl alcohol with an average degree of polymerization of 2400 and a degree of saponification of 97 mol% was used, and acetaldehyde was used instead of n-butylaldehyde (n-BA). The obtained polyvinyl acetal resin was mixed with 5 mol% of dimethylaminoacrylamide as a monomer. The synthesis conditions were as follows. That is, the obtained polyvinyl acetal resin was dissolved in THF at a concentration of 20% by weight, 5 parts by weight of dimethylaminoacrylamide was added, and then 3 parts by weight of an initiator (Irgacure184) was dissolved and irradiated with UV for 20 minutes using a UV polymerization tester.

[0134] Synthetic resin X7 was prepared as follows. First, 81.4 parts by weight of butyl methacrylate and 18.6 parts by weight of hydroxyethyl methacrylate were mixed to obtain a (meth)acrylic monomer solution. Next, 0.5 parts by weight of Irgacure 184 (manufactured by BASF) was dissolved in the obtained (meth)acrylic monomer solution and applied onto a PET film. The applied material was exposed to light with a wavelength of 365 nm at an integrated light intensity of 2000 mJ / cm using a UV conveyor device "ECS301G1" manufactured by iGraphics Co., Ltd. at 25°C. 2 A poly(meth)acrylic acid ester resin solution was obtained by irradiating with 1000 uV at 400 uV for 3 hours. The poly(meth)acrylic acid ester resin solution was dried in vacuum at 80° C. for 3 hours to obtain synthetic resin X7 as a poly(meth)acrylic acid ester resin.

[0135] For synthetic resin X8, except that polyvinyl alcohol with an average degree of polymerization of 800 and a degree of saponification of 1 mol% was used, a synthetic resin that was a polyvinyl acetal resin was obtained in the same manner as in Example 1. 95 parts by weight of the obtained polyvinyl acetal resin and 5 parts by weight of acrylic acid (linker) were dissolved in 300 parts by weight of THF, and reacted for 20 minutes under ultraviolet irradiation in the presence of a photoradical polymerization initiator, to graft copolymerize the polyvinyl acetal resin and acrylic acid to form a linker portion.

[0136] A linear peptide having an amino acid sequence of Gly-Arg-Gly-Asp-Ser (5 amino acid residues, indicated as GRGDS in Table 1) was prepared as a peptide. 5 parts by weight of this peptide and 5 parts by weight of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (condensing agent) were added to 90 parts by weight of methanol not containing either calcium or magnesium to prepare a peptide-containing liquid. 100 parts by weight of this peptide-containing liquid and 97 parts by weight of a polyvinyl acetal resin having a linker portion formed thereon were added to 300 parts by weight of methanol, and reacted to dehydrate and condense the carboxyl group of the linker portion and the amino group of Arg of the peptide. After the reaction was completed, the mixture was vacuum dried at 60°C for 1 hour to volatilize the methanol. In this way, synthetic resin X8, which is a peptide-containing polyvinyl acetal resin having a polyvinyl acetal resin portion, a linker portion, and a peptide portion, was prepared.

[0137] In Comparative Example 2, synthetic resin X1 was used, and in Example 8 and Comparative Example 3, a polyvinyl butyral resin having the composition shown in Table 1 was used.

[0138] Fabrication of cell culture vessels; (Examples 1 to 10, Comparative Example 3) In Examples 1 to 10 and Comparative Example 3, the synthetic resin shown in Table 1 was first dissolved in a butanol / methanol mixed solvent (9:1) at a concentration of 5 wt %. 40 μL of the resulting solution was cast into a polystyrene dish with a diameter of 22 mm, and then immersed in liquid nitrogen to freeze together with the solvent. After that, the dish was freeze-dried for two days using a freeze dryer to obtain a cell culture vessel equipped with a cell culture scaffold having pores.

[0139] The average pore size of the cell culture scaffold was determined using a scanning electron microscope (SEM). Figure 2 is an SEM image of the cell culture scaffold obtained in Example 1 at a magnification of 1000 times, and Figure 3 is an SEM image of the cell culture scaffold obtained in Example 1 at a magnification of 10000 times.

[0140] (Comparative Examples 1 and 2) In Comparative Example 1, a polystyrene dish itself was used as the cell culture vessel.

[0141] In Comparative Example 2, synthetic resin X1 was first dissolved in a butanol-methanol mixed solvent (9:1) at a concentration of 5 wt%. 40 μL of the resulting solution was cast onto a φ22 mm polystyrene dish, and then dried in an oven at 45°C for 6 hours without freezing to form a flat membrane. In Comparative Examples 1 and 2, no pores were present whose average diameter could be measured.

[0142] Zeta potential measurement; Using a zeta potential measuring device (model number SURPASS 3, manufactured by Anton Paar), the zeta potential at pH 7 of the cell culture scaffold materials of Examples 1 to 10 and Comparative Examples 1 to 3 was determined. First, a 0.1 mM KCl solution was prepared. Next, the resin film of the scaffold material was set in a cell, and then a pH electrode and a conductivity meter were set. The zeta potential at each pH was measured by titrating from pH 9 to 3 using HCl and NaOH solutions.

[0143] In addition, in Comparative Example 3, the zeta potential was below the lower limit of measurement, and therefore it was deemed impossible to measure.

[0144] Water swelling rate; To determine the water swelling rate of each scaffold, the scaffold was removed from the cell culture vessel and immersed in water at 25°C for 24 hours. After immersion, the sample was placed on a Kimtowel for 1 minute to drain the water from the pores, and then the weight was measured. The weights of the samples before and after immersion were measured, and the water swelling rate was calculated as follows: (sample weight after immersion - sample weight before immersion) / (sample weight before immersion) x 100 (%).

[0145] Surface elastic modulus measurement; A nanoindenter (Hysitron, Triboindenter, manufactured by Bruker) was used to determine the surface elastic modulus at 25° C. of the cell culture scaffolds of Examples 1 to 10 and Comparative Examples 1 to 3. A Berkovich (triangular pyramid) indenter with a tip diameter R of 100 nm was used, and a single indentation measurement was performed in air at room temperature, with the measurement being performed at an indentation depth of 50 nm.

[0146] Average pore size: The surface of the cell culture scaffold was observed using a scanning electron microscope (SEM), and the average pore size of 100 pores randomly selected within the observation area (5 μm × 5 μm) was calculated.

[0147] Cell seeding and culture; The following liquid medium and ROCK (Rho-associated kinase) specific inhibitor were prepared.

[0148] TeSR E8 medium (STEM CELL) ROCK Inhibitor (Y27632)

[0149] 1 mL of phosphate buffered saline was added to the obtained cell culture vessel, and the vessel was allowed to stand in an incubator at 37° C. for 1 hour, after which the phosphate buffered saline was removed from the cell culture vessel.

[0150] A colony of h-iPS cells 253G1 that had reached a confluent state was placed on a φ35 mm dish, 1 mL of 0.5 mM ethylenediamine / phosphate buffer solution was added, and the dish was left to stand at room temperature for 2 minutes. After removing the ethylenediamine / phosphate buffer solution, cell clusters were obtained that were broken into pieces of 50 μm to 200 μm in size by pipetting with 1 mL of TeSR E8 medium. The obtained cell clusters (cell count 0.2 × 105 cells) were clamp-seeded into the above cell culture vessel.

[0151] At the time of seeding, 1.5 mL of liquid medium and a ROCK-specific inhibitor were added to the cell culture vessel to a final concentration of 10 μM, and the cells were cultured in an incubator at 37°C and a CO2 concentration of 5%. After that, the liquid in the dish was replaced with 1.5 mL of fresh liquid medium every 24 hours, and the culture was continued for 5 days. After the culture, 1 mL of Thermo-Fisher Triple Express was added to the dish and left to stand at 37°C for 2 minutes, after which the cells were collected and the cell count was measured using an EVE auto cell counter.

[0152] The results are shown in Table 1 below.

[0153] [Table 1] [Explanation of symbols]

[0154] 1...Cell culture container 2…Container body 2a…Surface 3. Scaffold for cell culture

Claims

1. Contains synthetic resin, the synthetic resin contains at least a polyvinyl alcohol derivative or a poly(meth)acrylic resin; The synthetic resin contains 0.2 mol % or more and 20 mol % or less of a Bronsted basic group in a constituent unit, The zeta potential at pH 7 is −60 mV or more; A scaffold for cell culture having a plurality of pores with an average diameter of 10 nm or more and 5 μm or less.

2. The scaffold for cell culture according to claim 1 , having a zeta potential of +30 mV or less at pH 7.

3. 3. The scaffold for cell culture according to claim 1, having an elastic modulus of 0.01 GPa or more at 25°C as measured by a nanoindentation method.

4. The scaffold for cell culture according to any one of claims 1 to 3, which has a water swelling rate of 100% or less.

5. The scaffold for cell culture according to any one of claims 1 to 4, which is substantially free of animal-derived raw materials.

6. The cell culture scaffold according to any one of claims 1 to 5, wherein the synthetic resin comprises a synthetic resin having a polyvinyl acetal skeleton.

7. The number of pores per unit area is 0.01 pores / μm ２ More than 1000 pieces / μm ２ The scaffold for cell culture according to any one of claims 1 to 6, wherein:

8. A cell culture vessel comprising the scaffold for cell culture according to any one of claims 1 to 7 in at least a part of a cell culture region.

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