Scaffold material for cell culture, resin film formed from scaffold material for cell culture, and cell culture vessel

Abstract

Provided is a resin film having excellent cell extendibility in cell culture. This resin film is formed by a scaffold material that is for cell culture and that contains a synthetic resin, the resin film having a compressive modulus of 5.5 GPa or more at a frequency of 1 Hz as measured using a nanoindenter device in ion-exchanged water in accordance with ISO14577-1 after being immersed in ion-exchanged water at 37°C for 24 hours.

Description

Technical Field

[0001] The present invention relates to a resin film formed of a scaffold material for cell culture, which is used for culturing cells. The present invention also relates to a cell culture container including the resin film.

Background Art

[0002] In research and development in academic fields, drug discovery fields, regenerative medicine fields, etc., animal cells such as human, mouse, rat, pig, cow, and monkey cells are used. As scaffold materials used for culturing animal cells, adhesive proteins such as laminin and vitronectin, and natural polymer materials such as Matrigel derived from mouse sarcoma are used.

[0003] Also, as shown in Non-Patent Document 1 and Patent Documents 1 and 2 below, scaffold materials using synthetic resins are also known.

[0004] In Non-Patent Document 1 below, a poly(vinyl alcohol-vinyl acetal-itaconic acid) copolymer obtained by condensing fibronectin is used as a scaffold material for culturing human iPS cells and ES cells.

[0005] In Patent Document 1 below, a scaffold material using DMAEMA as a cation, acrylic acid as an anion, and a nucleic acid or heparin is disclosed.

[0006] In Patent Document 2 below, a scaffold material using a cell adhesion substrate containing arylsulfatase protein is disclosed.

Prior Art Documents

Non-Patent Documents

[0007]

Non-Patent Document 1

Patent Documents

[0008]

Patent Document 1

Patent Document 2

Summary of the Invention

Problems to be Solved by the Invention

[0009] In the culture of animal cells, cell aggregates may be seeded on a scaffold material for culture. When a natural polymer material is used as the scaffold material, the natural polymer material is expensive, and because it is a natural-derived substance, there are problems such as large variations and potential safety problems due to animal-derived components.

[0010] Therefore, various scaffold materials using synthetic polymers as described in Non-Patent Document 1, Patent Document 1, and Patent Document 2 mentioned above have been proposed.

[0011] However, in the cell scaffold material described in Non-Patent Document 1, when it does not contain fibronectin, that is, in the case of only a synthetic polymer, there is a problem that the cells contract and float.

[0012] In addition, in the cell scaffold material described in Patent Document 1, there is a problem that it has high hydrophilicity due to ionic properties and causes contraction of cell aggregates and cell aggregation.

[0013] Furthermore, in the cell scaffold material described in Patent Document 2, because of its high hydrophilicity, it was easy to swell. Therefore, there was a problem that the seeded stem cells were easily detached from the scaffold material during culture and the cell adhesion was low.

[0014] An object of the present invention is to provide a resin film excellent in cell extensibility during cell culture, and a cell culture container provided with the above resin film.

Means for Solving the Problems

[0015] The resin film according to the present invention is a resin film formed of a scaffold material for cell culture containing a synthetic resin, and after being immersed in ion-exchanged water at 37 °C for 24 hours, the compression elastic modulus at a frequency of 1 Hz measured using a nanoindentation device in the ion-exchanged water in accordance with ISO14577-1 is 5.5 GPa or more.

[0016] In a specific aspect of the resin film according to the present invention, the water contact angle in ion-exchanged water at 16 °C is less than 145° and 100° or more.

[0017] In another specific aspect of the resin film according to the present invention, the number average molecular weight of the synthetic resin is 4.0×10 4 or more and 150×10 4 or less.

[0018] In still another specific aspect of the resin film according to the present invention, the synthetic resin includes a polyvinyl alcohol derivative or a poly(meth)acrylate derivative.

[0019] In a more limited specific aspect of the resin film according to the present invention, the polyvinyl alcohol derivative is polyvinyl acetal.

[0020] The cell culture container according to the present invention includes a container body and the resin film according to the present invention, and the resin film is disposed on the surface of the container body.

Advantages of the Invention

[0021] Since the compression elastic modulus of the resin film according to the present invention is within the above-specified range, the stretchability of cells during cell culture can be enhanced.

Brief Description of the Drawings

[0022]

Figure 1

Figure 2

Figure 3

Figure 4

Mode for Carrying Out the Invention

[0023] Hereinafter, the details of the present invention will be described.

[0024] The resin film according to the present invention is used for culturing cells. The resin film according to the present invention is formed of a scaffold material for cell culture containing a synthetic resin. Further, the resin film according to the present invention has a compressive elastic modulus of 5.5 GPa or more at a frequency of 1 Hz measured in accordance with ISO 14577-1 using a nanoindenter device in the ion-exchanged water after being immersed in the ion-exchanged water at 37°C for 24 hours.

[0025] Since the resin film according to the present invention has a compressive elastic modulus measured in ion-exchanged water within the above-specified range after being immersed in the ion-exchanged water at 37°C for 24 hours, the stretchability of cell aggregates when used for cell culture can be enhanced.

[0026] The evaluation of the stretchability of cell aggregates can be performed by obtaining the shape factor (SF).

[0027] Here, the shape factor (SF) is a shape evaluation coefficient of the region in the plan view of the cell aggregate after culturing cells, and SF = 4×π×(planar area of cell aggregate) / (length of outer peripheral edge of cell aggregate) 2 and is obtained by

[0028] As shown in Figure 2, if the above SF is 1, the planar shape of the cell aggregate becomes circular. The smaller the SF, the farther it is from a circle, the less likely the cell aggregate is to shrink, and the higher the stretchability of the cells. For example, in the case of the star shape shown in Figure 2, SF is 0.3, and it can be said that the stretchability is superior to that of the circle with SF = 1.

[0029] (Compressive elastic modulus) In the resin film according to the present invention, as described above, after being immersed in ion-exchanged water at 37°C for 24 hours, the compressive elastic modulus at a frequency of 1 Hz measured in accordance with ISO 14577-1 using a nanoindenter device in the ion-exchanged water is within the above-specified range. The compressive elastic modulus can be measured by the following method. First, a resin film formed of a scaffold material for cell culture is placed in a beaker filled with ion-exchanged water, and then the beaker is placed in a thermostatic bath at 37°C and left for 24 hours. The immersed resin film is measured at a frequency of 1 Hz in accordance with ISO 14577-1 using a nanoindenter device (Triboindenter, manufactured by Hysitron) in ion-exchanged water. Regarding the analysis method of the compressive elastic modulus, it is calculated according to the following formula.

[0030] Compressive elastic modulus = √π × (slope of the load-displacement curve in the elastic region) / (2 × √(contact projected area))

[0031] Here, the elastic region refers to the region where the slope of the load-displacement curve is constant. The contact projected area refers to the area where the indenter and the sample are in contact.

[0032] For the indenter, Berkovich (triangular pyramid type, tip diameter R of several 100 nm) can be used, and the indentation depth can be 50 nm.

[0033] When the compressive elastic modulus at this time is 5.5 GPa or more, the stretchability of cells during cell culture can be enhanced.

[0034] The above compressive elastic modulus is preferably greater than 6.0 GPa, more preferably greater than 6.5 GPa. When the compressive elastic modulus is within the above range, the stretchability of cells can be enhanced more effectively.

[0035] The upper limit of the above compressive elastic modulus is not particularly limited, but is, for example, 15 GPa or less.

[0036] The above compression elastic modulus can be adjusted within the above range, for example, by adjusting the type and number average molecular weight of the synthetic resin X described later. For example, as the synthetic resin X described later, by selecting a resin with low hydrophilicity, increasing the number average molecular weight, forming a crosslinked structure between molecules, etc., swelling in water is suppressed, and it becomes easier to increase the compression elastic modulus.

[0037] (Water contact angle) The resin film according to the present invention preferably has a water contact angle in ion-exchanged water at 16°C of less than 145° and 100° or more. When the water contact angle is within the above range, the stretchability of cells can be more effectively enhanced.

[0038] The measurement of the water contact angle can be performed by the following method. First, the resin film formed of the scaffold material for cell culture is immersed in ion-exchanged water at 16°C for 24 hours. Next, using a contact angle meter (DMo-601, manufactured by Kyowa Interface Science Co., Ltd.), 1 μl of air ejected from a syringe is brought into contact with the above resin film. The water contact angle can be measured by fitting the contact angle of the air by the tangent method.

[0039] The range of the above water contact angle is preferably less than 140°, and also preferably 110° or more. When the water contact angle is within the above range, the stretchability of cells can be enhanced more effectively.

[0040] The above water contact angle can be decreased, for example, in the synthetic resin X described later, by increasing the number of hydrophobic functional groups. Also, the above water contact angle can be increased in the synthetic resin X by increasing the number of hydrophilic functional groups.

[0041] (Synthetic resin X) The scaffold material for cell culture used in the present invention contains a synthetic resin. This synthetic resin is hereinafter referred to as synthetic resin X.

[0042] The number average molecular weight of the above synthetic resin X is preferably 4.0×10 4or more, more preferably 5.0×10 4 or more, and preferably 150×10 4 or less, more preferably 100×10 4 or less. When the number average molecular weight is at least the above lower limit, it becomes easier to adjust the compression elastic modulus within the above range. When the number average molecular weight is at most the above upper limit, the cell stretchability during cell culture can be enhanced more effectively.

[0043] The measurement of the number average molecular weight of the synthetic resin X can be performed, for example, by the following method. The synthetic resin X is dissolved in THF to prepare a 0.2% solution. Next, using a gel permeation chromatography (GPC) measuring device (APC system, manufactured by Waters), it is evaluated under the following measurement conditions.

[0044] Column: HSPgel HR MB-M 6.0×150 mm, flow rate: 0.5 mL / min, column temperature: 40°C

[0045] Injection volume: 10 μl, detector: RI, PDA, standard sample: polystyrene

[0046] The content of the synthetic resin X in the scaffold material for cell culture is preferably 90% by weight or more, more preferably 95% by weight or more, still more preferably 97.5% by weight or more, particularly preferably 99% by weight or more, and most preferably 100% by weight.

[0047] In addition to the synthetic resin X, the scaffold material for cell culture may contain a polymer such as a resin different from the synthetic resin X. The synthetic resin X is not particularly limited as long as the compression elastic modulus of the scaffold material for cell culture can be within the above specific range.

[0048] Examples of the synthetic resin X include polyolefin derivatives, polyether derivatives, polyvinyl alcohol derivatives, polyester derivatives, poly(meth)acrylate esters derivatives, epoxy resin derivatives, polyamide derivatives, polyimide derivatives, polyurethane derivatives, polycarbonate derivatives, cellulose derivatives, and polypeptide derivatives.

[0049] From the viewpoint of effectively exerting the effects of the present invention, the synthetic resin X is preferably a polyvinyl alcohol derivative or a poly(meth)acrylate ester derivative. The polyvinyl alcohol derivative is a synthetic resin synthesized using at least polyvinyl alcohol as a raw material. The poly(meth)acrylate ester derivative is a synthetic resin synthesized using at least acrylate ester as a monomer.

[0050] The synthetic resin X preferably has a polyvinyl acetal skeleton or a poly(meth)acrylate ester skeleton. In this case, the synthetic resin X may be a resin having a polyvinyl acetal skeleton, a resin having a poly(meth)acrylate ester skeleton, or a resin having both a polyvinyl acetal skeleton and a poly(meth)acrylate ester skeleton.

[0051] From the viewpoint of effectively exerting the effects of the present invention, the synthetic resin X is preferably a resin having a polyvinyl acetal skeleton or a resin having both a polyvinyl acetal skeleton and a poly(meth)acrylate ester skeleton. The synthetic resin X is preferably a resin having at least a polyvinyl acetal skeleton.

[0052] <Synthetic resin X having a polyvinyl acetal skeleton> The scaffold material for cell culture preferably contains a synthetic resin X having a polyvinyl acetal skeleton.

[0053] In this specification, the "synthetic resin X having a polyvinyl acetal skeleton" may be referred to as "polyvinyl acetal resin X".

[0054] Therefore, polyvinyl acetal resin X is a resin having a polyvinyl acetal skeleton.

[0055] Polyvinyl acetal resin X has an acetal group, an acetyl group, and a hydroxyl group in its side chain.

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

[0057] The aldehyde used for acetalizing polyvinyl alcohol to obtain polyvinyl acetal resin X is not particularly limited. Examples of the aldehyde include aldehydes having 1 to 10 carbon atoms. The aldehyde may have a linear aliphatic group, a cyclic aliphatic group, or an aromatic group. The aldehyde may be a linear 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, etc. Only one kind of the aldehyde may be used, or two or more kinds may be used in combination.

[0059] The above aldehyde is preferably formaldehyde, acetaldehyde, propionaldehyde, butyraldehyde, or pentanaldehyde, and more preferably butyraldehyde. Therefore, the above polyvinyl acetal skeleton is preferably a polyvinyl butyral skeleton. The polyvinyl acetal resin X is preferably a polyvinyl butyral resin.

[0060] Incidentally, the blending amount of the above aldehyde can be appropriately set according to the target amount of acetal groups. From the viewpoint of enhancing the efficiency of the acetalization reaction and easily removing unreacted aldehyde, based on 100 mol% of polyvinyl alcohol, the addition amount of the above aldehyde is preferably 60 mol% or more, more preferably 65 mol% or more, preferably 95 mol% or less, and more preferably 90 mol% or less.

[0061] The number average molecular weight (Mn) of the polyvinyl acetal resin X is preferably 15,000 or more and preferably 1,000,000 or less. When the above Mn is within the above lower limit and the above upper limit, the strength of the resin film formed by the scaffold material for cell culture can be increased.

[0062] The degree of acetalization of the polyvinyl acetal resin X (in the case of polyvinyl butyral resin, it is the degree of butyralization) is preferably 60 mol% or more, more preferably 65 mol% or more, preferably 90 mol% or less, and more preferably 85 mol% or less. When the above degree of acetalization is at least the above lower limit, the cell adhesion can be further enhanced, and the cells can grow efficiently. When the above degree of acetalization is at most the above upper limit, the solubility in the solvent can be improved.

[0063] The glass transition temperature of the polyvinyl acetal resin X is preferably 40 °C or higher, more preferably 45 °C or higher, and still more preferably 50 °C or higher. By setting the glass transition temperature to the above lower limit or higher, it becomes easier to further adjust the compression elastic modulus of the resin film within the above range. The upper limit of the above glass transition temperature is not particularly limited, and may be, for example, 300 °C or lower. The above glass transition temperature can be measured using, for example, a differential scanning calorimeter.

[0064] The degree of acetylation (acetyl group amount) of the above polyvinyl acetal resin X is preferably 0.0001 mol% or more and preferably 5 mol% or less.

[0065] The hydroxyl group content (hydroxyl group amount) of the above polyvinyl acetal resin X is preferably 1 mol% or more, more preferably 10 mol% or more, preferably 80 mol% or less, and more preferably 60 mol% or less.

[0066] The degree of acetalization, degree of acetylation, and hydroxyl group content of the above polyvinyl acetal resin X 1 can be measured by 1H-NMR (nuclear magnetic resonance spectrum).

[0067] The above polyvinyl acetal resin X may be a polyvinyl acetal resin X synthesized using polyvinyl alcohol, or may be a copolymer of a polyvinyl acetal resin and a monomer. As such a monomer, a vinyl compound is preferably used. Examples of the vinyl compound include ethylene, allylamine, vinyl pyrrolidone, maleic anhydride, maleimide, itaconic acid, (meth)acrylic acid, vinylamine, and (meth)acrylic acid ester. Only one kind of the above vinyl compound may be used, or two or more kinds may be used in combination.

[0068] From the viewpoint of further enhancing the adhesiveness of cells, the above polyvinyl acetal resin X preferably has a Bronsted basic group or a Bronsted acidic group, and more preferably has a Bronsted basic group. That is, it is preferable that a part of the polyvinyl acetal resin X is modified with a Bronsted basic group or a Bronsted acidic group, and it is more preferable that a part of the polyvinyl acetal resin X is modified with a Bronsted basic group.

[0069] The above Bronsted basic group is a general term for functional groups that can receive a hydrogen ion H + from other substances. Examples of the above 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.

[0070] The above 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, it may have only one of these structural units, or two or more of them.

[0071] The above polyvinyl acetal resin X preferably has a structural unit having an imine structure. The above imine structure refers to a structure having a C=N bond. The above polyvinyl acetal resin X preferably has an imine structure in the side chain. In this case, the above imine 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. Note that having the above imine structure in the side chain also includes having the above imine structure in the graft chain of the polyvinyl acetal resin X. Examples of the structural unit having the above imine structure include the structural units represented by the following formula (11) or the following formula (12).

[0072]

Chemical formula

[0073] In the above formula (11), R1 represents a group having an imine structure.

[0074]

Chemical formula

[0075] In the above formula (12), R1 represents a group having an imine structure, and R2 represents an alkylene group. The number of carbon atoms of the above alkylene group is preferably 1 or more, preferably 12 or less, and more preferably 5 or less. When the number of carbon atoms of the above alkylene group is within the above lower limit and the above upper limit, the strength of the resin film formed by the scaffold material for cell culture can be increased.

[0076] Examples of the above alkylene group include linear alkylene groups such as methylene group, ethylene group, trimethylene group, tetramethylene group, pentamethylene group, hexamethylene group, octamethylene group, and decamethylene group; branched alkylene groups such as methylmethylene group, methylethylene group, 1-methylpentylene group, and 1,4-dimethylbutylene group; and cyclic alkylene groups such as cyclopropylene group, cyclobutylene group, and cyclohexylene group. The above alkylene group is preferably a linear alkyl group such as methylene group, ethylene group, trimethylene group, or tetramethylene group, and more preferably methylene group or ethylene group.

[0077] Examples of R1 in the above formula (11) and R1 in the above formula (12) include the group represented by the following formula (13).

[0078]

Chemical formula

[0079] In the above formula (13), R3 represents a hydrogen atom or a hydrocarbon group having 1 to 18 carbon atoms, and R4 represents a hydrocarbon group having 1 to 18 carbon atoms.

[0080] Examples of the hydrocarbon group include saturated hydrocarbon groups, unsaturated hydrocarbon groups, and aromatic hydrocarbon groups. The hydrocarbon group may consist of only one of saturated hydrocarbon groups, unsaturated hydrocarbon groups, and aromatic hydrocarbon groups, or may be a combination of two or more thereof.

[0081] Examples of the saturated hydrocarbon group include methyl group, ethyl group, n-propyl group, isopropyl group, n-butyl group, iso-butyl group, sec-butyl group, tert-butyl group, pentyl group, hexyl group, heptyl group, 2-ethylhexyl group, octyl group, nonyl group, decyl group, undecyl group, dodecyl group, tridecyl group, tetradecyl group, pentadecyl group, octadecyl group, etc. The saturated hydrocarbon group is preferably a methyl group, an ethyl group, an n-propyl group, or an n-butyl group.

[0082] Examples of the aromatic hydrocarbon group include phenyl group, toluyl group, xylyl group, t-butylphenyl group, and benzyl group, etc.

[0083] The structural unit having the imine structure preferably has the structure represented by the above formula (11), and in the above formula (13), R3 is a hydrogen atom, a methyl group, or an ethyl group, and R4 is a methyl group, an ethyl group, or a propyl group.

[0084] In the above polyvinyl acetal resin X, the content of the structural unit having the imine structure is preferably 0.1 mol% or more, more preferably 1.0 mol% or more, preferably 20.0 mol% or less, and more preferably 15.0 mol% or less. When the content is within the above lower limit and the above upper limit, the adhesion of cell aggregates can be further enhanced.

[0085] In the above polyvinyl acetal resin X, the ratio of the content of the structural unit having an imine structure to the degree of acetalization (content of the structural unit having an imine structure / degree of acetalization) is preferably 0.001 or more, and preferably 0.5 or less. When the above ratio (content of the structural unit having an imine structure / degree of acetalization) is within the above lower limit and the above upper limit, the strength of the resin film formed by the scaffold material for cell culture can be increased, and the adhesiveness of the cell mass can also be increased.

[0086] The above polyvinyl acetal resin X preferably has a structural unit having an imide structure. The structural unit having an imide structure is preferably a structural unit having an imino group (=NH).

[0087] The above polyvinyl acetal resin X preferably has the above imino group in the side chain. In this case, the above imino group may be directly bonded to the 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.

[0088] The above polyvinyl acetal resin X preferably has a structural unit having an amine structure. The amine group in the above amine structure may be a primary amine group, a secondary amine group, a tertiary amine group, or a quaternary amine group.

[0089] The structural unit having the above amine structure may be a structural unit having an amide structure. The above amide structure refers to a structure having -C(=O)-NH-.

[0090] The above polyvinyl acetal resin X preferably has the above amine structure or the above amide structure in the side chain. In this case, the above amine structure or the above amide structure may be directly bonded to the 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.

[0091] Note that having the above amine structure or amide structure in the side chain also includes having the above amine structure or amide structure in the graft chain of the polyvinyl acetal resin X.

[0092] From the viewpoint of enhancing the adhesion of cell aggregates, the amine group in the above amine structure is preferably a primary amine group (-NH2).

[0093] The structural unit having the above amine structure is preferably a structure represented by the following formula (21).

[0094] [Chemical formula]

[0095] The structural unit having the above amide structure is preferably a structure represented by the following formula (31).

[0096] [Chemical formula]

[0097] In the above formula (31), R1 represents a hydrogen atom or a hydrocarbon group having 1 to 10 carbon atoms. Examples of the hydrocarbon group include an alkyl group, an alkenyl group, a cycloalkyl group, and a cycloalkenyl group.

[0098] In the above polyvinyl acetal resin X, the content of the structural unit having the above amine structure or amide structure is preferably 0.1 mol% or more, more preferably 0.5 mol% or more, preferably 20 mol% or less, and more preferably 10 mol% or less, respectively. When the content is at least the above lower limit, the addition characteristics can be enhanced. When the content is at most the above upper limit, the polyvinyl acetal resin powder can be easily separated by the precipitation method.

[0099] In the above polyvinyl acetal resin X, in a total of 100 mol% of the content of the structural unit having an imine structure and the content of the structural unit having an amine structure, the content of the structural unit having an imine structure is preferably 0.5 mol% or more, more preferably 5 mol% or more, preferably 99.5 mol% or less, and more preferably 90 mol% or less. Further, in the above polyvinyl acetal resin X, in a total of 100 mol% of the content of the structural unit having an imine structure and the content of the structural unit having an amide structure, the content of the structural unit having an imine structure is preferably 0.5 mol% or more, more preferably 5 mol% or more, preferably 99.5 mol% or less, and more preferably 90 mol% or less. When the content of the structural unit having an imine structure is at least the above lower limit, the viscosity stability over time can be made sufficient. When the content of the structural unit having an imine structure is at most the above upper limit, the cell adhesiveness can be further enhanced.

[0100] In the above polyvinyl acetal resin X, the total content of the structural unit having an imine structure, the structural unit having an amine structure, and the structural unit having an imide structure is preferably 0.1 mol% or more, more preferably 1 mol% or more, preferably 30 mol% or less, and more preferably 10 mol% or less. Further, in the above polyvinyl acetal resin X, the total content of the structural unit having an imine structure, the structural unit having an amide structure, and the structural unit having an imide structure is preferably 0.1 mol% or more, more preferably 1 mol% or more, preferably 30 mol% or less, and more preferably 10 mol% or less. When the total content is at least the above lower limit and at most the above upper limit, the cell adhesiveness can be further enhanced.

[0101] 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 can be measured by 1 H-NMR (nuclear magnetic resonance spectrum).

[0102] The above-mentioned Brønsted acidic group is a general term for functional groups that can transfer hydrogen ions H + to other substances.

[0103] Examples of the Brønsted acidic 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, and salts thereof. The Brønsted acidic group is preferably a carboxyl group.

[0104] Examples of the method for modifying the above-mentioned polyvinyl acetal resin X with the above-mentioned Brønsted acidic group include a method of copolymerizing the above-mentioned polyvinyl alcohol with the above-mentioned itaconic acid or (meth)acrylic acid, and a method of introducing a Brønsted acidic group into the side chain of the above-mentioned polyvinyl alcohol.

[0105] The method for producing the polyvinyl acetal resin X having a Brønsted basic group or a Brønsted acidic group is not particularly limited. For example, the polyvinyl acetal resin X having a structural unit having an imine structure can be produced, for example, by the following method (1), (2), (3) or (4). The above-mentioned polyvinyl acetal resin X may be an acetalized product of polyvinyl alcohol having a structural unit having an amine structure or an amide structure.

[0106] (1) A polyvinyl acetate is obtained by copolymerizing the monomer having the above-mentioned imine structure and vinyl acetate. The obtained polyvinyl acetate is saponified to obtain polyvinyl alcohol. The obtained polyvinyl alcohol is acetalized by a conventionally known method.

[0107] (2) An imine structure is introduced by acetalizing polyvinyl alcohol having a structural unit having an amine structure or an amide structure by a conventionally known method.

[0108] (3) Polyvinyl alcohol having an imine structure obtained by post-modifying polyvinyl alcohol having a structural unit having an amine structure or an amide structure is acetalized by a conventionally known method.

[0109] (4) Introduce an imine structure by modifying the polyvinyl acetal resin.

[0110] Among the above (1) to (4), it is particularly preferable to obtain a polyvinyl acetal resin X having a structural unit having an imine structure by the method of (2).

[0111] In the methods of the above (1) to (4), a polyvinyl acetal resin X having a structural unit having an imine structure can be preferably obtained by a method of adding an excessive amount of an aldehyde or an acid catalyst used for acetalization.

[0112] In the method of adding an excessive amount of the above aldehyde, it is preferable to add 70 to 150 parts by weight of the aldehyde with respect to 100 parts by weight of polyvinyl alcohol having a structural unit having an amine structure or an amide structure. In particular, as the aldehyde, acetaldehyde, propionaldehyde, n-butyl aldehyde, isobutyl aldehyde, n-valeraldehyde, and phenyl aldehyde are preferable.

[0113] In the method of adding an excessive amount of the above acid catalyst, it is preferable to add the acid catalyst in an amount of 0.5% by weight or more of the whole. Further, it is preferable to add 5.0 to 70.0 parts by weight of the acid catalyst with respect to 100 parts by weight of polyvinyl alcohol having a structural unit having an amine structure or an amide structure. In particular, as the acid catalyst, hydrochloric acid, nitric acid, sulfuric acid, and p-toluenesulfonic acid are preferable.

[0114] The above acetalization can be carried out using known methods. The above acetalization is preferably carried out in an aqueous solvent, a mixed solvent of an organic solvent compatible with water, or an organic solvent. Examples of the organic solvent compatible with water include alcohol-based organic solvents. Examples of the organic solvent include alcohol-based organic solvents, aromatic organic solvents, aliphatic ester solvents, ketone solvents, lower paraffin solvents, ether solvents, amide solvents, amine solvents, and the like. Only one kind of the above organic solvent may be used, or two or more kinds may be used in combination.

[0115] Examples of the alcohol-based organic solvent include methanol, ethanol, n-propanol, isopropanol, n-butanol, tert-butanol, and the like.

[0116] Examples of the aromatic organic solvent include xylene, toluene, ethylbenzene, methyl benzoate, and the like.

[0117] Examples of the aliphatic ester solvent include methyl acetate, ethyl acetate, butyl acetate, methyl propionate, ethyl propionate, methyl butyrate, ethyl butyrate, methyl acetoacetate, ethyl acetoacetate, and the like.

[0118] Examples of the ketone solvent include acetone, methyl ethyl ketone, methyl isobutyl ketone, cyclohexanone, methylcyclohexanone, benzophenone, acetophenone, and the like.

[0119] Examples of the lower paraffin solvent include hexane, pentane, octane, cyclohexane, decane, and the like.

[0120] Examples of the ether solvent include diethyl ether, tetrahydrofuran, ethylene glycol dimethyl ether, ethylene glycol diethyl ether, propylene glycol diethyl ether, and the like.

[0121] Examples of the amide solvents include N,N-dimethylformamide, N,N-dimethyltetroamide, N-methylpyrrolidone, acetanilide, and the like.

[0122] Examples of the amine solvents include ammonia, trimethylamine, triethylamine, n-butylamine, di-n-butylamine, tri-n-butylamine, aniline, N-methylaniline, N,N-dimethylaniline, pyridine, and the like.

[0123] From the viewpoints of solubility in the resin and simplicity during purification, the organic solvent is preferably ethanol, n-propanol, isopropanol, or tetrahydrofuran.

[0124] The acetalization is preferably carried out in the presence of an acid catalyst. The acid catalyst is not particularly limited, and examples include mineral acids such as sulfuric acid, hydrochloric acid, nitric acid, and phosphoric acid; carboxylic acids such as formic acid, acetic acid, and propionic acid; and sulfonic acids such as methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, and p-toluenesulfonic acid. The acid catalyst may be used alone or in combination of two or more. The acid catalyst is preferably hydrochloric acid, nitric acid, or sulfuric acid, and more preferably hydrochloric acid.

[0125] <Synthetic resin X having a poly(meth)acrylate ester skeleton> The scaffold material for cell culture preferably contains a synthetic resin X having a poly(meth)acrylate ester skeleton.

[0126] In this specification, "synthetic resin X having a poly(meth)acrylate ester skeleton" may be referred to as "poly(meth)acrylate ester resin X".

[0127] Therefore, poly(meth)acrylate ester resin X is a resin having a poly(meth)acrylate ester skeleton.

[0128] The above poly(meth)acrylate resin X is obtained by polymerizing a (meth)acrylate or by polymerizing a (meth)acrylate and the above other monomer.

[0129] Examples of the above (meth)acrylate include alkyl (meth)acrylates, cyclic alkyl (meth)acrylates, aryl (meth)acrylates, (meth)acrylamides, polyethylene glycol (meth)acrylates, phosphorylcholine (meth)acrylates, and the like.

[0130] Examples of the above alkyl (meth)acrylates 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, isotetradecyl (meth)acrylate, and the like.

[0131] The above alkyl (meth)acrylates may be substituted with substituents such as alkoxy groups having 1 to 3 carbon atoms and tetrahydrofurfuryl groups. Examples of such alkyl (meth)acrylates include methoxyethyl acrylate, tetrahydrofurfuryl acrylate, and the like.

[0132] Examples of the above cyclic alkyl (meth)acrylates include cyclohexyl (meth)acrylate, isobornyl (meth)acrylate, and the like.

[0133] Examples of the above aryl (meth)acrylates include phenyl (meth)acrylate, benzyl (meth)acrylate, and the like.

[0134] Examples of the above (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.

[0135] Examples of the above 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.

[0136] Examples of the above (meth)acryloylphosphorylcholine include 2-(meth)acryloyloxyethyl phosphorylcholine.

[0137] As another monomer copolymerized with the (meth)acrylate 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)acrylate ester. The above vinyl compound may be used alone or in combination of two or more.

[0138] In this specification, "(meth)acryl" means "acryl" or "methacryl", and "(meth)acrylate" means "acrylate" or "methacrylate".

[0139] The number average molecular weight (Mn) of the poly(meth)acrylate resin X is preferably 60×10 4 or more, preferably 90×10 4 or less.

[0140] In the poly(meth)acrylate resin X, the total of the structures derived from (meth)acrylic acid alkyl esters, the structures derived from (meth)acrylic acid cyclic alkyl esters, and the structures derived from (meth)acrylic acid aryl esters contained in the constituent units is preferably 50% by weight or more, more preferably 60% by weight or more, and still more preferably 70% by weight or more. By setting the proportion of these hydrophobic (meth)acrylate-derived structures to be not less than the above lower limit, it becomes easier to further adjust the compression elastic modulus and the water contact angle of the resin film within the above ranges.

[0141] The upper limit of the total of the structures derived from (meth)alkyl acrylate alkyl esters, the structures derived from (meth)acrylic acid cyclic alkyl esters, and the structures derived from (meth)acrylic acid aryl esters contained in the constituent units of the poly(meth)acrylate resin X is not particularly limited and may be 100% by weight or less.

[0142] The poly(meth)acrylate resin X may contain a structure derived from a polyfunctional (meth)acrylate in the constituent units. By containing a structure derived from a polyfunctional (meth)acrylate in the constituent units, the structure derived from the polyfunctional (meth)acrylate becomes a crosslinking point, and it becomes easier to further adjust the compression elastic modulus of the resin film within the above range.

[0143] The proportion of the structure derived from the polyfunctional (meth)acrylate contained in the constituent units of the poly(meth)acrylate resin X is preferably 0.1% by weight or more and preferably 30% by weight or less.

[0144] The glass transition temperature of the poly(meth)acrylate resin X is preferably 40 °C or higher, more preferably 45 °C or higher, still more preferably 50 °C or higher. By setting the glass transition temperature to the above lower limit or higher, it becomes easier to further adjust the compression elastic modulus of the resin film within the above range. The upper limit of the glass transition temperature is not particularly limited, and may be, for example, 300 °C or lower. The glass transition temperature can be measured, for example, using a differential scanning calorimeter.

[0145] <Synthetic resin X having a polyvinyl acetal skeleton and a poly(meth)acrylate skeleton> The scaffold material for cell culture preferably contains a synthetic resin X having a polyvinyl acetal skeleton and a poly(meth)acrylate skeleton. The synthetic resin X is preferably a composite resin having a polyvinyl acetal skeleton and a poly(meth)acrylate skeleton.

[0146] The synthetic resin X having a polyvinyl acetal skeleton and a poly(meth)acrylate skeleton can be synthesized, for example, by appropriately combining the above-described synthesis methods of the polyvinyl acetal resin X and the poly(meth)acrylate resin X.

[0147] The synthetic resin X having a polyvinyl acetal skeleton and a poly(meth)acrylate skeleton is preferably a synthetic resin X in which the above-described (meth)acrylate ester is graft copolymerized onto the above-described polyvinyl acetal resin.

[0148] The number average molecular weight (Mn) of the synthetic resin X having a polyvinyl acetal skeleton and a poly(meth)acrylate skeleton is preferably 60×10 4 or more, preferably 90×10 4 or less. When the Mn is within the above lower limit and the above upper limit, the strength of the resin film formed by the scaffold material for cell culture can be increased.

[0149] Incidentally, the synthetic resin X may be crosslinked. The scaffold material for cell culture containing the crosslinked synthetic resin X can effectively suppress water swelling and enhance its strength. By using a crosslinking agent, the synthetic resin X can be crosslinked.

[0150] Examples of the crosslinking agent include polyalcohols, polycarboxylic acids, hydroxycarboxylic acids, metal soaps, and polysaccharides.

[0151] Examples of the above polyalcohols include ethylene glycol, propylene glycol, butanediol, pentanediol, hexanediol, heptanediol, octanediol, nonanediol, decanediol, dodecanediol, undecanediol, diethylene glycol, triethylene glycol, tetraethylene glycol, polyethylene glycol, catechol, pyrogallol, diboronic acid, methylenediboronic acid, ethylenediboronic acid, propylenediboronic acid, phenylenediboronic acid, biphenyldiboronic acid, bisphenol derivatives, and the like.

[0152] Examples of the above polycarboxylic acids include oxalic acid, malonic acid, succinic acid, glutaric acid, adipic acid, pimelic acid, suberic acid, azelaic acid, sebacic acid, phthalic acid, poly(meth)acrylic acid, and the like.

[0153] Examples of the above hydroxycarboxylic acids include glycolic acid, lactic acid, tartronic acid, glyceric acid, hydroxybutyric acid, malic acid, tartaric acid, citramalic acid, citric acid, isocitric acid, leucic acid, mevalonic acid, pantothenic acid, ricinoleic acid, lysineradic acid, cerebric acid, quinic acid, shikimic acid, hydroxybenzoic acid, salicylic acid, creosote acid, vanillic acid, syringic acid, pyrocatechuic acid, resorcylic acid, protocatechuic acid, gentisic acid, orsellinic acid, gallic acid, mandelic acid, benzoic acid, atrolactic acid, melilotate, phloretate, coumaric acid, umbellic acid, caffeic acid, ferulic acid, sinapic acid, hydroxystearic acid, and the like.

[0154] Examples of the metal soap include salts of fatty acids such as stearic acid, lauric acid, ricinoleic acid, and octylic acid, and metals such as lithium, sodium, magnesium, calcium, barium, zinc, and aluminum.

[0155] Examples of the polysaccharides include pectin, guar gum, xanthan gum, tamarind gum, carrageenan, propylene glycol, carboxymethylcellulose, amylose, amylopectin, glycogen, cellulose, chitin, agarose, carrageenan, heparin, hyaluronic acid, xyloglucan, and glucomannanic acid.

[0156] (Scaffolding material for cell culture) The scaffold material for cell culture constituting the resin film 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 scaffold material for cell culture 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, it is most preferable that the scaffold material for cell culture is 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.

[0157] The above-mentioned scaffold material for cell culture may contain components other than the synthetic resin X. Examples of such components include polyolefin resins, polyether resins, polyvinyl alcohol resins, polyesters, epoxy resins, polyamide resins, polyimide resins, polyurethane resins, polycarbonate resins, polysaccharides, cellulose, polypeptides, and synthetic peptides.

[0158] From the viewpoint of effectively exerting the effects of the present invention, the content of components other than synthetic resin X is preferably as low as possible. In 100% by weight of the scaffold material for cell culture, the content of these components is preferably 20% by weight or less, more preferably 10% by weight or less, even more preferably 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 material for cell culture does not contain any components other than synthetic resin X.

[0159] The scaffold material for cell culture preferably contains substantially no animal-derived raw materials. By not containing any animal-derived raw materials, it is possible to provide a scaffold material for cell culture that is highly safe and has little variation in quality during production. Here, "substantially free of animal-derived raw materials" means that the content of animal-derived raw materials in the scaffold material for cell culture is 3% by weight or less. The scaffold material for cell culture preferably contains 1% by weight or less, and more preferably 0% by weight, of animal-derived raw materials. In other words, the scaffold material for cell culture more preferably contains no animal-derived raw materials at all.

[0160] (Cell culture using scaffold materials for cell culture) The above-mentioned scaffold material for cell culture is used for culturing cells. The above-mentioned scaffold material for cell culture is used as a scaffold for cells when culturing the cells. Therefore, a resin film formed from the scaffold material for cell culture of the present invention is used for culturing cells and is also used as a scaffold for the cells when culturing the cells. It is particularly preferable that cell clusters are seeded on the resin film formed from the above-mentioned scaffold material for cell culture. Cell clusters can be obtained by adding a cell release agent to a cell culture vessel containing cells in a confluent state to detach the cells, and then homogenizing the detached cells by pipetting. The cell release agent is not particularly limited, but is preferably an ethylenediamine / phosphate buffer solution. The size of the cell clusters is preferably 50 μm to 200 μm. Note that the resin film formed from the scaffold material for cell culture of the present invention may be used for seeding and culturing cells in a form other than cell clusters.

[0161] The cells include animal cells from humans, mice, rats, pigs, cows, monkeys, etc. The cells also include somatic cells, such as stem cells, progenitor cells, and mature cells. The somatic cells may be cancer cells.

[0162] Examples of the mature cells include nerve cells, cardiac muscle cells, retinal cells, and hepatic cells.

[0163] 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.

[0164] (Shape of scaffold material for cell culture) The resin film of the present invention is formed from a scaffold material for cell culture. The resin film is formed using a scaffold material for cell culture. The resin film is preferably a membrane-like scaffold material for cell culture. The resin film is preferably a membrane-like product of a scaffold material for cell culture.

[0165] The present specification also provides particles, fibers, porous bodies, or films containing the above-mentioned scaffold material for cell culture. In this case, the shape of the scaffold material for cell culture is not particularly limited, and may be particles, fibers, porous bodies, or films. Note that the particles, fibers, porous bodies, or films may contain components other than the above-mentioned scaffold material for cell culture.

[0166] A film containing the scaffold material for cell culture is preferably used for flat culture (two-dimensional culture) of cells, and particles, fibers, or porous bodies containing the scaffold material for cell culture are preferably used for three-dimensional culture of cells.

[0167] (Cell culture carrier) The present invention may also be a carrier for cell culture in which the resin film is disposed on the surface of the carrier. The carrier for cell culture can be obtained, for example, by disposing the resin film on the surface of the carrier by coating or the like. The shape of the carrier may be particles, fibers, a porous body, or a film. That is, the carrier for cell culture may be in the shape of particles, fibers, a porous body, or a film. The carrier for cell culture may also include components other than the carrier and the resin film.

[0168] (Cell culture container) The cell culture vessel according to the present invention comprises a vessel body and a resin film formed from the above-mentioned scaffold material for cell culture, with the resin film being disposed on the surface of the vessel body.

[0169] FIG. 1 is a front cross-sectional view that schematically shows a cell culture vessel according to one embodiment of the present invention.

[0170] The cell culture vessel 1 comprises a vessel body 2 and a resin film 3 formed from a scaffold material for cell culture. The resin film 3 is disposed on a surface 2a of the vessel body 2. The resin film 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 such as cell clumps on the surface of the resin film 3, cells can be cultured on a plate.

[0171] 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, a resin film 3 formed from the cell culture scaffold material may be disposed on the surface of the second container body.

[0172] The container body may be a conventionally known container body (container). The shape and size of the container body are not particularly limited.

[0173] Examples of the container body include a cell culture plate having one or more wells (holes) and a cell culture flask. The number of wells in the plate is not particularly limited. Examples of the number of wells include, but are not limited to, 2, 4, 6, 12, 24, 48, 96, 384, etc. The shape of the well is not particularly limited and includes, for example, a perfect circle, an ellipse, a triangle, a square, a rectangle, a pentagon, etc. The shape of the bottom surface of the well is not particularly limited and includes, for example, a flat bottom, a round bottom, unevenness, etc.

[0174] The material of the container body is not particularly limited and includes resins, metals, and inorganic materials. Examples of the resin include polystyrene, polyethylene, polypropylene, polycarbonate, polyester, polyisoprene, cycloolefin polymer, polyimide, polyamide, polyamideimide, (meth)acrylic resin, epoxy resin, silicone, etc. Examples of the metal include stainless steel, copper, iron, nickel, aluminum, titanium, gold, silver, platinum, etc. Examples of the inorganic material include silicon oxide (glass), aluminum oxide, titanium oxide, zirconium oxide, iron oxide, silicon nitride, etc.

[0175] The present invention will be described in more detail below with reference to Examples and Comparative Examples. The present invention is not limited only to these Examples. Note that the following Examples 6 to 8 are reference examples.

[0176] The following were prepared as materials for the cell culture scaffold material.

[0177] Note that the content ratio of the structural units in the obtained synthetic resin was measured by 1H-NMR (nuclear magnetic resonance spectrum) after dissolving the synthetic resin in DMSO-d6 (dimethyl sulfoxide). The content ratios of the respective structural units of the synthetic resin are shown in Table 1. 1 1H-NMR (nuclear magnetic resonance spectrum). The content ratios of the respective structural units of the synthetic resin are shown in Table 1.

[0178] In Example 1, polyvinyl acetal resin X1 (synthetic resin X1) was used.

[0179] Synthesis of synthetic resin X1: 2700 mL of ion-exchanged water and 300 parts by weight of polyvinyl alcohol with an average degree of polymerization of 1700 and a degree of saponification of 97 mol% were placed in a reactor equipped with a stirrer, and the mixture was heated and dissolved while stirring to obtain a solution. 35 wt% hydrochloric acid was added to the obtained solution as a catalyst so that the hydrochloric acid concentration was 0.2 wt%. Next, the temperature was adjusted to 15°C, and 22 parts by weight of n-butylaldehyde was added while stirring. Next, 148 parts by weight of n-butylaldehyde was added, and white particulate polyvinyl butyral resin was precipitated. 15 minutes after the precipitation, 35 wt% hydrochloric acid was added so that the hydrochloric acid concentration was 1.8 wt%, and the mixture was heated to 50°C and maintained 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 polyvinyl butyral resin (number average molecular weight 11×10 4 The resulting product had an acetalization degree (butyralization degree) of 65 mol%, a hydroxyl group content of 34 mol%, and an acetylation degree of 3 mol%.

[0180] In Examples 2 to 5, polyvinyl acetal resins X2 to X5 were used, and in Examples 6 to 8, poly(meth)acrylic acid ester resins X6 to X8 were used.

[0181] Polyvinyl acetal resin X2: The test was carried out in the same manner as in Example 1, except that polyvinyl alcohol having an average degree of polymerization of 2500 and a degree of saponification of 97 mol% was used, and acetaldehyde was used instead of n-butylaldehyde. 4 The resulting product had an acetalization degree of 66 mol%, a hydroxyl group content of 33 mol%, and an acetylation degree of 3 mol%.

[0182] Polyvinyl acetal resin X3: The test was carried out in the same manner as in Example 1, except that polyvinyl alcohol having an average polymerization degree of 850, a saponification degree of 97 mol%, and an amine modification degree of 2 mol% was used. 4 The resulting copolymer had an acetalization degree of 77 mol%, a hydroxyl group content of 20 mol%, an acetylation degree of 1 mol%, and an amine modification degree of 2 mol%.

[0183] Polyvinyl acetal resin X4: 90 parts by weight of the same polyvinyl butyral resin as in Example 1 was dissolved in tetrahydrofuran to give a 30% by weight solution, to which 10 parts by weight of Coronate L (manufactured by Tosoh Corporation) as a crosslinking agent was added, and the mixture was heated at 80°C for 5 hours to obtain a polyvinyl butyral resin (number average molecular weight 100 × 10 4 ) was obtained.

[0184] Polyvinyl acetal resin X5: 90 parts by weight of the same polyvinyl butyral resin as in Example 1 was dissolved in tetrahydrofuran to give a 30% by weight solution, and 0.1 parts by weight of Irgacure 184 as an initiator and 10 parts by weight of 1,6-hexanediol diacrylate were added thereto, followed by graft polymerization to obtain a polyvinyl butyral resin (number average molecular weight 76×10 4 ) was obtained.

[0185] Poly(meth)acrylate resin X6: 25 parts by weight of butyl methacrylate, 70 parts by weight of methyl methacrylate, and 5 parts by weight of 1,6-hexanediol diacrylate were mixed to obtain a (meth)acrylic monomer solution. 0.1 parts by weight of Irgacure 184 (manufactured by BASF) was dissolved in the obtained (meth)acrylic monomer solution and applied to a PET film. The coated material was exposed to light at a wavelength of 365 nm at an integrated dose of 2000 mJ / cm using a UV conveyor device "ECS301G1" manufactured by Eye Graphics Co., Ltd. at 25°C. 2 A poly(meth)acrylic acid ester resin solution was obtained by irradiating the solution with 1000 uV at 1000 uV. The poly(meth)acrylic acid ester resin solution was dried in vacuum at 80°C for 3 hours to obtain a poly(meth)acrylic acid ester resin. The number average molecular weight of the obtained poly(meth)acrylic acid ester resin was 60 × 10 4 It was.

[0186] Poly(meth)acrylate resin X7: Poly(meth)acrylic acid ester resin X7 (number average molecular weight 78×10) was prepared in the same manner as poly(meth)acrylic acid ester resin X6, except that 45 parts by weight of butyl methacrylate, 50 parts by weight of methyl methacrylate, and 5 parts by weight of 1,6-hexanediol diacrylate were used as the (meth)acrylic monomers. 4 ) was obtained.

[0187] Poly(meth)acrylate resin X8: Poly(meth)acrylic acid ester resin X8 (number average molecular weight 90×10) was prepared in the same manner as poly(meth)acrylic acid ester resin X6, except that 65 parts by weight of butyl methacrylate, 30 parts by weight of methyl methacrylate, and 5 parts by weight of 1,6-hexanediol diacrylate were used as the (meth)acrylic monomers. 4 ) was obtained.

[0188] Acrylic resins Y1 to Y4 were used in Comparative Examples 1 to 4. In Table 1, these resins are listed with resin numbers X1 to X8 and Y1 to Y4.

[0189] Synthesis of poly(meth)acrylate resin Y1: Poly(meth)acrylic acid ester resin Y1 (number average molecular weight 20×10) was prepared in the same manner as poly(meth)acrylic acid ester resin X6, except that 95 parts by weight of butyl methacrylate and 5 parts by weight of 1,6-hexanediol diacrylate were used as the (meth)acrylic monomers. 4 ) was obtained.

[0190] Synthesis of poly(meth)acrylate resin Y2: Poly(meth)acrylic acid ester resin Y2 (number average molecular weight 12×10) was prepared in the same manner as poly(meth)acrylic acid ester resin X6, except that 80 parts by weight of butyl methacrylate and 15 parts by weight of lauryl acrylate were used as the (meth)acrylic monomers. 4 ) was obtained.

[0191] Synthesis of poly(meth)acrylate resin Y3: Poly(meth)acrylic acid ester resin Y3 (number average molecular weight 23×10) was prepared in the same manner as poly(meth)acrylic acid ester resin X6, except that 50 parts by weight of butyl methacrylate and 50 parts by weight of methacrylic acid were used as the (meth)acrylic monomers. 4 ) was obtained.

[0192] Synthesis of poly(meth)acrylate resin Y4: Poly(meth)acrylic acid ester resin Y4 (number average molecular weight 25×10) was prepared in the same manner as poly(meth)acrylic acid ester resin X6, except that 35 parts by weight of butyl methacrylate and 65 parts by weight of methacrylic acid were used as the (meth)acrylic monomers. 4 ) was obtained.

[0193] In Examples 1 to 8 and Comparative Examples 1 to 4, the above-mentioned resin was dissolved in 1-butanol to prepare a 5 wt % resin solution. 100 μL of this resin solution was dropped onto the culture area of the cell culture vessel and allowed to soak in. The solution was then left to stand at room temperature for 1 hour and then dried at 60°C for 15 hours to prepare a cell culture vessel having a resin film formed from the cell culture scaffold material in the culture area.

[0194] The compressive elastic modulus of the resin films of the above-mentioned Examples 1 to 8 and Comparative Examples 1 to 4 was measured in the following manner.

[0195] Measurement method: First, a cell culture vessel having a resin film formed from the above-mentioned cell culture scaffold material was placed in a beaker filled with ion-exchanged water, and the beaker was then placed in a thermostatic bath at 37°C and left for 24 hours. The submerged cell culture vessel was removed with ion-exchanged water remaining in the culture area, and the compressive modulus of the resin film was measured using a nanoindenter (Triboindenter, manufactured by Hysitron) according to ISO 14577-1. The compressive modulus was calculated according to the following formula:

[0196] Compressive elastic modulus = √π × (slope of the load-displacement curve in the elastic region) / (2 × √(projected contact area))

[0197] Here, the elastic region refers to the region where the slope of the load-displacement curve is constant, and the contact projection area refers to the area where the indenter and the sample come into contact.

[0198] The indenter used was a Berkovich (triangular pyramid type, tip diameter R number 100 nm), and the indentation depth was set to 50 nm.

[0199] (Contact angle in water) The resin films formed using each of the scaffold materials for cell culture in the above Examples and Comparative Examples were immersed in ion-exchanged water at 16°C for 24 hours. Next, using a contact angle meter (DMo-601, Kyowa Interface Co., Ltd.), 1 μl of air dispensed from a syringe was brought into contact with the resin film. The underwater contact angle was measured by fitting the air contact angle using the tangent method.

[0200] The compressive modulus and water contact angle values are shown in Table 1 below.

[0201] (Evaluation of cell extensibility) Stem cell seeding and culture: One mL of phosphate-buffered saline was added to the resulting cell culture vessel, which was then left to stand in an incubator at 37°C for 1 hour. After removing the phosphate-buffered saline from the dish, 1.0 × 10 h-iPS cells 253G1 were added. 5 The cells were seeded and cultured in an incubator at 37°C and 5% CO2 concentration in the presence of 1 mL of TeSR E8 medium (STEM CELL) and 10 μM of ROCK-inhibitor (Y27632).

[0202] evaluation: SF rating As described above, the shape factor (SF) was evaluated. For the evaluation, the cell clusters were viewed in plan using a phase-contrast microscope (Olympus, IX73). In the plan view of the cell cluster, SF = 4 × π × (plane area of cell cluster / length of outer periphery of cell cluster). 2 The SF value was calculated using the following equation: Figure 3 is a photograph showing the planar shape of the cell aggregate when SF≈0.2, and Figure 4 is a photograph showing the planar shape when SF≈1.

[0203] The results are shown in Table 1 below.

[0204] As is clear from Table 1, in Comparative Examples 1 to 4, the SF values (shape factors) 24 hours after seeding were extremely high at 0.7 to 0.9. In contrast, in the resin films of Examples 1 to 8, the SF values were 0.4 or less, indicating excellent cell extensibility. In particular, in Examples 1, 3, 4, and 5, the SF values were even lower at 0.3 or less, indicating even more excellent extensibility.

[0205] [Table 1] [Explanation of Reference Numerals]

[0206] 1... Container for cell culture 2... Container body 2a... Surface 3... Resin film

Claims

1. A scaffold material for cell culture containing a synthetic resin having a polyvinyl acetal skeleton, wherein the degree of acetalization of the synthetic resin having the polyvinyl acetal skeleton is 60 mol% or more and 90 mol% or less, after being immersed in ion-exchanged water at 37 °C for 24 hours, the compressive elastic modulus at a frequency of 1 Hz measured in accordance with ISO 14577-1 using a nanoindentation device in the ion-exchanged water is 5.5 GPa or more and 9.50 GPa or less, A scaffold material for cell culture having a water contact angle in ion-exchanged water at 16 °C of less than 145° and 100° or more.

2. The scaffold material for cell culture according to claim 1, having a water contact angle in ion-exchanged water at 16 °C of less than 140°.

3. The number average molecular weight of the synthetic resin is 4.0×10 ４ or more and 150×10 ４ or less. The scaffold material for cell culture according to claim 1 or 2.

4. A resin film used for culturing cells, The resin film formed of the scaffold material for cell culture according to any one of claims 1 to 3.

5. Comprising a container body and the resin film according to claim 4, A cell culture container in which the resin film is disposed on the surface of the container body.

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