WO2012133900A1 - 細胞シート移植治具及びその利用方法 - Google Patents
細胞シート移植治具及びその利用方法 Download PDFInfo
- Publication number
- WO2012133900A1 WO2012133900A1 PCT/JP2012/059003 JP2012059003W WO2012133900A1 WO 2012133900 A1 WO2012133900 A1 WO 2012133900A1 JP 2012059003 W JP2012059003 W JP 2012059003W WO 2012133900 A1 WO2012133900 A1 WO 2012133900A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- cell
- cell sheet
- jig
- cultured
- sheet
- Prior art date
Links
- 238000002054 transplantation Methods 0.000 title claims abstract description 74
- 238000000034 method Methods 0.000 title claims description 57
- 210000004027 cell Anatomy 0.000 claims abstract description 169
- 210000004748 cultured cell Anatomy 0.000 claims abstract description 98
- 230000021164 cell adhesion Effects 0.000 claims description 86
- 239000000758 substrate Substances 0.000 claims description 33
- 238000004113 cell culture Methods 0.000 claims description 29
- 102000004169 proteins and genes Human genes 0.000 claims description 19
- 108090000623 proteins and genes Proteins 0.000 claims description 19
- 229920001477 hydrophilic polymer Polymers 0.000 claims description 17
- 229920000208 temperature-responsive polymer Polymers 0.000 claims description 15
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 14
- 239000000499 gel Substances 0.000 claims description 6
- 230000003313 weakening effect Effects 0.000 claims description 5
- 102000008186 Collagen Human genes 0.000 claims description 3
- 108010035532 Collagen Proteins 0.000 claims description 3
- 102000009123 Fibrin Human genes 0.000 claims description 3
- 108010073385 Fibrin Proteins 0.000 claims description 3
- BWGVNKXGVNDBDI-UHFFFAOYSA-N Fibrin monomer Chemical compound CNC(=O)CNC(=O)CN BWGVNKXGVNDBDI-UHFFFAOYSA-N 0.000 claims description 3
- 102000016359 Fibronectins Human genes 0.000 claims description 3
- 108010067306 Fibronectins Proteins 0.000 claims description 3
- 108010010803 Gelatin Proteins 0.000 claims description 3
- 102000007547 Laminin Human genes 0.000 claims description 3
- 108010085895 Laminin Proteins 0.000 claims description 3
- 229920001436 collagen Polymers 0.000 claims description 3
- 229950003499 fibrin Drugs 0.000 claims description 3
- 239000008273 gelatin Substances 0.000 claims description 3
- 229920000159 gelatin Polymers 0.000 claims description 3
- 235000019322 gelatine Nutrition 0.000 claims description 3
- 235000011852 gelatine desserts Nutrition 0.000 claims description 3
- 230000004956 cell adhesive effect Effects 0.000 claims description 2
- 239000004615 ingredient Substances 0.000 claims 1
- 238000010030 laminating Methods 0.000 claims 1
- 239000011148 porous material Substances 0.000 abstract 1
- 210000001519 tissue Anatomy 0.000 description 20
- 229920000642 polymer Polymers 0.000 description 19
- -1 polyethylene Polymers 0.000 description 12
- 239000000178 monomer Substances 0.000 description 10
- 239000011557 critical solution Substances 0.000 description 9
- 239000011248 coating agent Substances 0.000 description 8
- 238000000576 coating method Methods 0.000 description 8
- 239000002609 medium Substances 0.000 description 8
- 238000000338 in vitro Methods 0.000 description 7
- 239000004698 Polyethylene Substances 0.000 description 6
- 210000004102 animal cell Anatomy 0.000 description 6
- 238000007334 copolymerization reaction Methods 0.000 description 6
- 229920001971 elastomer Polymers 0.000 description 6
- 239000000463 material Substances 0.000 description 6
- 229920000573 polyethylene Polymers 0.000 description 6
- 230000010069 protein adhesion Effects 0.000 description 6
- 229920001577 copolymer Polymers 0.000 description 5
- 238000012258 culturing Methods 0.000 description 5
- 230000002107 myocardial effect Effects 0.000 description 5
- 238000007796 conventional method Methods 0.000 description 4
- 230000009089 cytolysis Effects 0.000 description 4
- 239000003814 drug Substances 0.000 description 4
- 239000000806 elastomer Substances 0.000 description 4
- 238000005516 engineering process Methods 0.000 description 4
- 210000002919 epithelial cell Anatomy 0.000 description 4
- 239000000203 mixture Substances 0.000 description 4
- 210000000056 organ Anatomy 0.000 description 4
- 229920002635 polyurethane Polymers 0.000 description 4
- 239000004814 polyurethane Substances 0.000 description 4
- 241000282412 Homo Species 0.000 description 3
- 210000002216 heart Anatomy 0.000 description 3
- 229920001519 homopolymer Polymers 0.000 description 3
- 230000003100 immobilizing effect Effects 0.000 description 3
- 238000001727 in vivo Methods 0.000 description 3
- 229920002050 silicone resin Polymers 0.000 description 3
- 210000004927 skin cell Anatomy 0.000 description 3
- 238000001179 sorption measurement Methods 0.000 description 3
- 238000002560 therapeutic procedure Methods 0.000 description 3
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 2
- 241000282472 Canis lupus familiaris Species 0.000 description 2
- 241000282326 Felis catus Species 0.000 description 2
- 241000283973 Oryctolagus cuniculus Species 0.000 description 2
- 241001494479 Pecora Species 0.000 description 2
- 239000004743 Polypropylene Substances 0.000 description 2
- 241000700159 Rattus Species 0.000 description 2
- XUIMIQQOPSSXEZ-UHFFFAOYSA-N Silicon Chemical compound [Si] XUIMIQQOPSSXEZ-UHFFFAOYSA-N 0.000 description 2
- 241000282887 Suidae Species 0.000 description 2
- 239000004809 Teflon Substances 0.000 description 2
- 229920006362 Teflon® Polymers 0.000 description 2
- QYKIQEUNHZKYBP-UHFFFAOYSA-N Vinyl ether Chemical class C=COC=C QYKIQEUNHZKYBP-UHFFFAOYSA-N 0.000 description 2
- 150000003926 acrylamides Chemical class 0.000 description 2
- 239000000853 adhesive Substances 0.000 description 2
- 230000001070 adhesive effect Effects 0.000 description 2
- 239000007864 aqueous solution Substances 0.000 description 2
- 210000004413 cardiac myocyte Anatomy 0.000 description 2
- 238000003851 corona treatment Methods 0.000 description 2
- 230000006837 decompression Effects 0.000 description 2
- 238000004090 dissolution Methods 0.000 description 2
- 238000010894 electron beam technology Methods 0.000 description 2
- 210000005175 epidermal keratinocyte Anatomy 0.000 description 2
- 239000012894 fetal calf serum Substances 0.000 description 2
- 210000002950 fibroblast Anatomy 0.000 description 2
- 230000005251 gamma ray Effects 0.000 description 2
- 238000004898 kneading Methods 0.000 description 2
- 239000002184 metal Substances 0.000 description 2
- 229910052751 metal Inorganic materials 0.000 description 2
- 150000002739 metals Chemical class 0.000 description 2
- 238000009832 plasma treatment Methods 0.000 description 2
- 229920000139 polyethylene terephthalate Polymers 0.000 description 2
- 239000005020 polyethylene terephthalate Substances 0.000 description 2
- 229920005597 polymer membrane Polymers 0.000 description 2
- 238000006116 polymerization reaction Methods 0.000 description 2
- 229920001155 polypropylene Polymers 0.000 description 2
- 230000001737 promoting effect Effects 0.000 description 2
- 229920005989 resin Polymers 0.000 description 2
- 239000011347 resin Substances 0.000 description 2
- 239000005060 rubber Substances 0.000 description 2
- 210000002966 serum Anatomy 0.000 description 2
- 229910052710 silicon Inorganic materials 0.000 description 2
- 239000010703 silicon Substances 0.000 description 2
- 210000003491 skin Anatomy 0.000 description 2
- 239000007779 soft material Substances 0.000 description 2
- 230000007847 structural defect Effects 0.000 description 2
- 210000003556 vascular endothelial cell Anatomy 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- SEFVRKXJJPMVHQ-YUMQZZPRSA-N (2s)-2-[[2-[[(2s)-2-[(2-aminoacetyl)amino]-5-(diaminomethylideneamino)pentanoyl]amino]acetyl]amino]butanedioic acid Chemical compound NC(N)=NCCC[C@H](NC(=O)CN)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(O)=O SEFVRKXJJPMVHQ-YUMQZZPRSA-N 0.000 description 1
- RGNVSYKVCGAEHK-GUBZILKMSA-N (3s)-3-[[2-[[(2s)-2-[(2-aminoacetyl)amino]-5-(diaminomethylideneamino)pentanoyl]amino]acetyl]amino]-4-[[(1s)-1-carboxy-2-hydroxyethyl]amino]-4-oxobutanoic acid Chemical compound NC(N)=NCCC[C@H](NC(=O)CN)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(O)=O RGNVSYKVCGAEHK-GUBZILKMSA-N 0.000 description 1
- NNRFRJQMBSBXGO-CIUDSAMLSA-N (3s)-3-[[2-[[(2s)-2-amino-5-(diaminomethylideneamino)pentanoyl]amino]acetyl]amino]-4-[[(1s)-1-carboxy-2-hydroxyethyl]amino]-4-oxobutanoic acid Chemical compound NC(N)=NCCC[C@H](N)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(O)=O NNRFRJQMBSBXGO-CIUDSAMLSA-N 0.000 description 1
- NIXOWILDQLNWCW-UHFFFAOYSA-M Acrylate Chemical compound [O-]C(=O)C=C NIXOWILDQLNWCW-UHFFFAOYSA-M 0.000 description 1
- IYMAXBFPHPZYIK-BQBZGAKWSA-N Arg-Gly-Asp Chemical compound NC(N)=NCCC[C@H](N)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(O)=O IYMAXBFPHPZYIK-BQBZGAKWSA-N 0.000 description 1
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 102000018697 Membrane Proteins Human genes 0.000 description 1
- 108010052285 Membrane Proteins Proteins 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 102000035195 Peptidases Human genes 0.000 description 1
- 239000004372 Polyvinyl alcohol Substances 0.000 description 1
- 229920002125 Sokalan® Polymers 0.000 description 1
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 210000002821 alveolar epithelial cell Anatomy 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 108010072041 arginyl-glycyl-aspartic acid Proteins 0.000 description 1
- 108010089975 arginyl-glycyl-aspartyl-serine Proteins 0.000 description 1
- 210000002449 bone cell Anatomy 0.000 description 1
- 239000001768 carboxy methyl cellulose Substances 0.000 description 1
- 235000010948 carboxy methyl cellulose Nutrition 0.000 description 1
- 239000008112 carboxymethyl-cellulose Substances 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 235000010980 cellulose Nutrition 0.000 description 1
- 210000001612 chondrocyte Anatomy 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 210000004087 cornea Anatomy 0.000 description 1
- 208000021921 corneal disease Diseases 0.000 description 1
- 210000000399 corneal endothelial cell Anatomy 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 210000001047 desmosome Anatomy 0.000 description 1
- 108010007093 dispase Proteins 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 108010019407 glycyl-arginyl-glycyl-aspartic acid Proteins 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 210000005003 heart tissue Anatomy 0.000 description 1
- 230000002440 hepatic effect Effects 0.000 description 1
- 210000003494 hepatocyte Anatomy 0.000 description 1
- 210000005260 human cell Anatomy 0.000 description 1
- 239000000017 hydrogel Substances 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 210000001822 immobilized cell Anatomy 0.000 description 1
- 230000009545 invasion Effects 0.000 description 1
- 239000000644 isotonic solution Substances 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 210000002901 mesenchymal stem cell Anatomy 0.000 description 1
- 210000005033 mesothelial cell Anatomy 0.000 description 1
- 210000003098 myoblast Anatomy 0.000 description 1
- 208000031225 myocardial ischemia Diseases 0.000 description 1
- 210000004738 parenchymal cell Anatomy 0.000 description 1
- 210000002379 periodontal ligament Anatomy 0.000 description 1
- 210000004694 pigment cell Anatomy 0.000 description 1
- 229920002401 polyacrylamide Polymers 0.000 description 1
- 239000004584 polyacrylic acid Substances 0.000 description 1
- 229920002338 polyhydroxyethylmethacrylate Polymers 0.000 description 1
- 229920002451 polyvinyl alcohol Polymers 0.000 description 1
- 235000019422 polyvinyl alcohol Nutrition 0.000 description 1
- 229920000036 polyvinylpyrrolidone Polymers 0.000 description 1
- 239000001267 polyvinylpyrrolidone Substances 0.000 description 1
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 description 1
- 230000002829 reductive effect Effects 0.000 description 1
- 230000001172 regenerating effect Effects 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 230000000717 retained effect Effects 0.000 description 1
- 239000000790 retinal pigment Substances 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000012679 serum free medium Substances 0.000 description 1
- 210000002027 skeletal muscle Anatomy 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 210000000130 stem cell Anatomy 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000001356 surgical procedure Methods 0.000 description 1
- 210000002437 synoviocyte Anatomy 0.000 description 1
- 239000012588 trypsin Substances 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
- A61F2/00—Filters implantable into blood vessels; Prostheses, i.e. artificial substitutes or replacements for parts of the body; Appliances for connecting them with the body; Devices providing patency to, or preventing collapsing of, tubular structures of the body, e.g. stents
- A61F2/02—Prostheses implantable into the body
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/38—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
- A61L27/52—Hydrogels or hydrocolloids
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M25/00—Means for supporting, enclosing or fixing the microorganisms, e.g. immunocoatings
- C12M25/02—Membranes; Filters
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M33/00—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
- A61F2230/00—Geometry of prostheses classified in groups A61F2/00 - A61F2/26 or A61F2/82 or A61F9/00 or A61F11/00 or subgroups thereof
- A61F2230/0002—Two-dimensional shapes, e.g. cross-sections
- A61F2230/0004—Rounded shapes, e.g. with rounded corners
- A61F2230/0006—Rounded shapes, e.g. with rounded corners circular
Definitions
- the present invention relates to a transplantation jig for cultured cell sheets in the fields of biology, medicine, and the like, and a method for using the same.
- animal cell culture technology has made significant progress, and research and development for animal cells has been extended to various fields.
- the target animal cells can be used not only to commercialize the original cells, but also to produce their products, and to design effective drugs by analyzing the cells and their cell surface proteins. It is also being practiced to regenerate the patient's own cells outside the living body, or to improve the function of the cell and then return it to the living body for treatment.
- techniques for culturing animal cells and techniques for evaluation, analysis, and utilization are one field that researchers are paying attention to.
- many animal cells including human cells are adhesion-dependent. That is, when culturing animal cells in vitro, it is necessary to attach them to the substrate surface once. And it is necessary to peel the cultured cells while keeping the cultured form on the surface of the base material without peeling them apart.
- organ transplants that attempt to replace difficult-to-treat organs with other people's organs have become common in recent years.
- the target organs are very diverse, such as skin, cornea, kidney, liver, heart, etc., and the progress after surgery has improved remarkably, and has already been established as a medical technology.
- corneal transplantation an eye bank was established in Japan about 50 years ago and transplantation activities began. However, the number of donors is still small, and there are about 20,000 patients who need corneal transplants in Japan alone, whereas it is said that there are only about 1/10 of the 2000 patients who can actually perform transplantation treatment. Yes.
- corneal transplantation the current situation is that the next medical technology is required due to the shortage of donors. Under such circumstances, a technique for culturing and transplanting normal cells of a patient to a desired size has been developed.
- the upper limit of skin cells is determined on a cell culture support having a base surface coated with a polymer having an upper or lower critical solution temperature of 0 to 80 ° C.
- a skin cell culture method characterized by culturing at or below the critical dissolution temperature or above the lower critical dissolution temperature and then peeling the cultured skin cells above the upper critical solution temperature or below the lower critical solution temperature is described. Yes.
- cells are detached from the culture substrate coated with the temperature-responsive polymer by temperature.
- this method has poor releasability, and the obtained cell sheet has many structural defects. Therefore, it has been difficult to apply the method described in Japanese Patent Laid-Open No. 05-192138 to the construction of myocardial tissue in vitro.
- myocardial tissue cells are cultured on a cell culture support whose surface is coated with a temperature-responsive polymer to obtain a myocardial cell sheet.
- a temperature-responsive polymer to obtain a myocardial cell sheet.
- Japanese Patent Application Laid-Open No. 2005-176812 discloses a technique relating to a cell sheet transplantation jig having a cell adhesion surface.
- the cultured cells can be detached, and then the detached cultured cells can be attached again.
- the jig shown here is for stacking cell sheets, and does not have a sufficient function as a method for transplanting to the curved surface to be transplanted as described above.
- an object of the present invention is to provide a novel cell sheet transplantation jig based on a completely different idea from the prior art. Moreover, an object of this invention is to provide the utilization method.
- the present inventors have studied and developed from various angles. As a result, the periphery of the transplanted part can be sucked and fixed by the suction hole of the cell sheet transplanting jig, and then the cell sheet existing on the cell sheet capturing surface of the cell sheet transplanting jig can be uniformly contacted by the transplanted part. I found out that Moreover, it discovered that the peeled cultured cell was made to adhere again to a transplanted part by weakening the adhesive force of the cell adhesion surface of the cell sheet transplant jig
- a jig having a plane for transplanting sheet-like cultured cells, (1) a surface portion for capturing the cell sheet while maintaining the sheet-like shape in the same direction plane; A cell sheet transplanting jig having a suction hole for sucking and fixing a transplanted part around.
- B The cell sheet transplantation jig according to (a), wherein the cell sheet transplantation jig has a flat plate shape.
- C The cell sheet transplantation jig according to any one of (a) and (b), wherein the cell sheet transplantation jig is an elastic body.
- the surface portion that captures the cell sheet is a cell adhesion surface for peeling the cells cultured on the surface of the cell culture substrate, and then attaching the peeled cultured cells to another place again.
- the hydrophilic polymer is a hydrous gel.
- the cell sheet transplantation jig having a convex cell adhesion surface described in the present invention is used, cultured cells in an arbitrary range on the cell culture substrate can be efficiently detached, and the detached cultured cells can be removed. It comes to adhere easily again. Therefore, the cultured cells can be easily moved to a place where the cells are desired to move, and can be moved accurately.
- the cell sheet transplanting jig provided with the suction hole shown in Example 1 is shown.
- (a) is a view seen from the top, and (b) is a view seen from the bottom.
- the cell sheet transplanting jig provided with the suction hole shown in Example 1 is shown.
- (a) is a view seen from the top, and (b) is a view seen from the bottom.
- the cell sheet transplanting jig provided with the suction hole shown in Example 1 is shown.
- (a) is a view seen from the top, and (b) is a view seen from the bottom.
- the cell sheet transplanting jig provided with the suction hole shown in Example 1 is shown.
- Example 1 shows a jig for threading and fixing a cell sheet in Example 1.
- FIG. Example 1 shows a jig capable of transplanting three cell sheets in a single transplant.
- Example 1 shows a jig capable of transplanting six cell sheets in a single transplant.
- Example 2 shows a transplantation jig made of a soft material.
- Example 2 shows that FIG. 11 is soft. In Example 2, it is shown that the thing of FIG. 11 can closely_contact
- Example 3 shows a transplantation jig that can be housed in a tube.
- the present invention relates to a transplantation jig for reliably transplanting a culture sheet cultured in a sheet shape to a transplanted part.
- the jig is provided with a suction hole for fixing the transplanted part.
- the suction hole is applied to the transplanted portion and suctioned, whereby the curved surface, for example, the surface of the living tissue is changed into a flat shape, which is convenient for the subsequent cell sheet transplantation step.
- the number of suction holes in the jig is not particularly limited, but it is preferable that there are three or more suction holes on the same plane for the purpose of the present invention.
- the shape of the suction hole, the distance from other suction holes, and the position with respect to the plane are not particularly limited, and the conditions may be appropriately changed according to the target transplanted part. Further, the strength of the degree of suction is not particularly limited as long as the transplanted portion is fixed on a flat surface, and the conditions may be appropriately changed according to the situation of the transplanted portion. At that time, if the suction is strong, the transplanted portion is denatured, which is not preferable as the method of the present invention.
- a means for suction is not particularly limited, but a decompression pump is usually used.
- the material of the present transplantation jig is not particularly limited, and examples thereof include polyurethane, polyethylene elastomer, silicone resin, Teflon (registered trademark), rubber, polyethylene, polypropylene, polyethylene terephthalate, and metals. It is not particularly limited. Among them, polyurethane, polyethylene elastomer, and silicon resin are moderately soft elastic bodies that can cope with curved structures, and are advantageous in the present invention.
- the transplantation jig of the present invention has a surface for capturing a cell sheet in a plane in the same direction as the suction hole described above.
- the material of the capture surface is not particularly limited, and examples thereof include polyurethane, polyethylene elastomer, silicone resin, Teflon (registered trademark), rubber, polyethylene, polypropylene, polyethylene terephthalate, metals, and the like. It is not a thing. Among them, polyurethane, polyethylene elastomer, and silicon resin are moderately soft elastic bodies that can cope with curved structures, and are advantageous in the present invention. At this time, the material of the suction hole portion described above may or may not be the same.
- the capturing method is not particularly limited, and examples thereof include a method of slightly sucking, a method of fixing using a thread and the like, and a method of attaching with a cell adhesive substance.
- a method of slightly sucking it is only necessary to provide suction holes on the cell sheet capturing surface.
- the number of suction holes in the jig is not particularly limited, but three or more on the same plane for the purpose of the present invention. It is preferable that there is a suction hole.
- the shape of the suction holes, the distance from other suction holes, and the position with respect to the plane are not particularly limited, and the conditions may be appropriately changed according to the target cell sheet.
- the strength of the suction level is not particularly limited as long as the cell sheet is fixed on a flat surface, and the conditions may be appropriately changed according to the state of the cell sheet. In that case, if it attracts
- a means for suction is not particularly limited, but a decompression pump is usually used.
- the transplantation jig of the present invention includes (1) a surface portion for capturing a cell sheet while maintaining a sheet-like shape on a plane in the same direction, and (2) a suction hole for sucking and fixing the transplanted portion around the surface portion.
- the plane may be a jig that maintains its shape, or may be a jig that can be housed in a tube and can be taken in and out as needed.
- the cell sheet can be used with an endoscope, and the cell sheet can be transplanted with less invasion to the affected part of the living body.
- the cell sheet capturing surface may be composed of, for example, one or more of cell adhesion protein, cell adhesion peptide, or hydrophilic polymer.
- the cell adhesion protein include one or more of fibrin gel, fibronectin, laminin, collagen, gelatin and the like.
- the cell adhesion peptide include one or more of RGD peptide, RGDS peptide, GRGD peptide, GRGDS peptide and the like.
- the method for immobilizing the cell adhesion protein and the cell adhesion peptide on the cell adhesion surface is not particularly limited, but physical adsorption by applying a cell adhesion protein or cell adhesion peptide aqueous solution known as a conventional method may be performed.
- the amount of the cell adhesion protein and cell adhesion peptide immobilized on the cell sheet capture surface is not particularly limited as long as it is sufficient to immobilize the cells to be transferred. volume of the 0.005 / cm 2 or more, preferably 0.01 [mu] g / cm 2 or more, more preferably 0.02 .mu.g / cm 2 or more.
- Measurement of the amount of immobilized cell adhesion protein and cell adhesion peptide may be in accordance with a conventional method, for example, a method of directly measuring a cell adhesion surface using FT-IR-ATR, a cell adhesion protein labeled in advance, a cell Examples thereof include a labeled cell adhesion protein immobilized on the cell adhesion surface by immobilizing the adhesive peptide by the same method, a method inferred from the amount of cell adhesion peptide, and any method may be used.
- the hydrophilic polymer used in the present invention may be either a homopolymer or a copolymer.
- a hydrogel such as polyacrylamide, polydimethylacrylamide, polyacrylic acid and salts thereof, polyhydroxyethyl methacrylate, polyhydroxyethyl acrylate, polyvinyl alcohol, polyvinyl pyrrolidone, cellulose, carboxymethyl cellulose, or the water content thereof varies depending on the temperature.
- temperature-responsive gel etc. are mentioned, it does not restrict
- the hydrophilic polymer used in the present invention may be a temperature responsive polymer.
- the temperature-responsive polymer may be either a homopolymer or a copolymer, and examples of such a polymer include polymers described in JP-A-2-21865. Specifically, for example, it can be obtained by homopolymerization or copolymerization of the following monomers.
- the monomer that can be used include a (meth) acrylamide compound, an N- (or N, N-di) alkyl-substituted (meth) acrylamide derivative, or a vinyl ether derivative. Two or more of these can be used.
- copolymerization with monomers other than the above monomers, grafting or copolymerization of polymers, or a mixture of polymers and copolymers may be used. It is also possible to crosslink within a range that does not impair the original properties of the polymer.
- the method for coating the surface of the base material with various polymers is not particularly limited.
- the method described in JP-A-2-21865 may be used. That is, such coating is performed by applying a substrate and the above monomer or polymer to one of electron beam irradiation (EB), ⁇ -ray irradiation, ultraviolet irradiation, plasma treatment, corona treatment, organic polymerization reaction, or physical application such as coating and kneading.
- EB electron beam irradiation
- ⁇ -ray irradiation ultraviolet irradiation
- plasma treatment corona treatment
- organic polymerization reaction or physical application such as coating and kneading.
- the amount of the hydrophilic polymer immobilized on the cell adhesion surface is not particularly limited as long as the amount sufficient to attach the cells to be moved is immobilized, but the amount immobilized is 0.5 ⁇ g / cm 2 or more, preferably 1.0 ⁇ g / cm 2 or more, more preferably 1.5 ⁇ g / cm 2 or more.
- the amount of the hydrophilic polymer immobilized may be measured by a conventional method, for example, a method of directly measuring the cell adhesion surface using FT-IR-ATR, a method of immobilizing a previously labeled hydrophilic polymer by the same method and cell adhesion. A method inferred from the amount of labeled hydrophilic polymer immobilized on the surface can be mentioned, and any method may be used.
- the cell adhesion surface in the cell sheet transplantation jig used in the present invention is not particularly limited as long as it is determined at any time according to the size of the cultured cell to be moved and the size of the cultured cell sheet. Moreover, what is necessary is just to determine at any time according to the magnitude
- the cell sheet capture surface is convex
- the cell adhesion surface of the moving jig when the cell adhesion surface of the moving jig is brought closer to the cultured cells on the surface of the cell culture substrate, first, the most convex part of the cell adhesion surface can contact. .
- the cell adhesion surface When the cell adhesion surface is further brought closer, the cell adhesion surface spreads and contacts the entire cell culture surface to be peeled around the most convex part.
- air bubbles do not enter between the cell adhesion surface and the cultured cell, and the cell adhesion surface and the cultured cell are brought into close contact with each other.
- the cultured cells can be detached.
- the shape of the convex portion of the cell adhesion surface is not particularly limited, and any location on the cell adhesion surface may be convex as compared with other locations.
- the most convex part of the cell adhesion surface should be at the center of the cell adhesion surface.
- the center of the cell adhesion surface here means the center of the cell adhesion surface if the most convex part is a dot or a surface having the center as the center, and the most convex part is linear or it. In the case of a planar shape centering on, it means a center line portion including the center of the cell adhesion surface.
- the convex cell contact surface is brought into contact with the cultured cell surface from the convex portion.
- the cell contact surface of the convex portion first comes into contact with the cultured cell surface, and further the cell adhesion surface approaches the cultured cell surface, so that the cell adhesion surface changes into a flat shape and comes into contact.
- the dimension of the convex part is preferably in the range of 0.5 mm to 5 mm, preferably in the range of 0.8 mm to 3 mm, more preferably as the height of the most convex part in the range used for cell adhesion.
- the range of 1.0 mm to 2.5 mm is good, and the range of 1.2 mm to 2.0 mm is most preferred.
- the height of the convex portion is 0.5 mm or less, the cell adhesion surface is not different from that when it is flat, and it is not preferable because not all the cells to be moved can be detached, and when the height of the convex portion is 5 mm or more. Distortion when the convex cell adhesion surface finally changes to a flat surface, and in some cases, pressure to make the convex cell adhesion surface flat are undesirably loaded on the cultured cells.
- the ratio of the convex area to the whole cell adhesion surface is not particularly limited, but the ratio of the convex area is preferably in the range of 40% to 100%, preferably Is preferably in the range of 50% to 100%, more preferably in the range of 70% to 100%, and most preferably in the range of 80% to 100%.
- the ratio of the area of the convex portion is 40% or less, there are many cases where air bubbles enter between the cell adhesion surface and the cultured cells, which is not preferable as the present invention.
- the shape of the convex portion in the present invention is not particularly limited to the shape when observed from the cell adhesion surface side and the shape when observed perpendicular to the cell adhesion surface, and when it is vertical
- the entire cell adhesion surface to be used may be convex continuously or stepwise.
- Examples of cells used in the present invention include corneal epithelial cells, corneal endothelial cells, retinal pigment cells, epidermal keratinocytes, oral mucosal cells, conjunctival epithelial cells, cardiomyocytes, fibroblasts, vascular endothelial cells, hepatocytes, Examples include skeletal muscle myoblasts, mesenchymal stem cells, alveolar epithelial cells, mesothelial cells, chondrocytes, synoviocytes, bone cells, periodontal ligament cells, other stem cells, or a mixture of two or more. However, the type is not restricted at all.
- the origin of the cells is not particularly limited, and examples include humans, dogs, cats, rabbits, rats, pigs, sheep, and the like.
- humans are used. It is desirable to use cells derived from them.
- the culture medium for cell culture in the present invention is not particularly limited as long as it is a commonly used medium for cells to be cultured. However, when the obtained cultured cells are used for human therapy, the components of the medium to be used are Those with a clear origin or those recognized as pharmaceuticals are desirable.
- the shape of the culture substrate in the present invention is not particularly limited, and examples thereof include dishes, multiplates, flasks, cell inserts, and flat membranes.
- the present invention peels the cultured cells from the cell culture substrate by attaching the cultured cells on the cell culture substrate to the cell adhesion surface provided in the cell sheet transplant jig,
- a method for moving cultured cells characterized in that the peeled cultured cells are attached again to a specific location by weakening the adhesion between the cell adhesion surface and the cultured cells.
- the cultured cells on the cell culture substrate can be easily detached, and the detached cultured cells are simply attached again. Therefore, it has been found that the cultured cells can be easily moved to the place where the cells are desired to move and can be transplanted accurately.
- the cultured cells in the present invention In order to move the cultured cells in the present invention, it is necessary to first attach the cultured cells on the cell culture substrate to the cell adhesion surface provided on the cell sheet transplantation jig.
- the attachment method is not limited in any way, but since the cell sheet transplantation jig in the present invention has a cell adhesion surface, the portion is placed on the cultured cell to be moved and left to stand. good.
- a cell adhesion protein or cell adhesion peptide is used on the cell adhesion surface of the present invention, the cultured cells adhere to the cell sheet transplantation jig via the cell adhesion protein and cell adhesion peptide.
- the cultured cells physically adhere to the hydrophilicity of the hydrophilic polymer or the hydrophilic / hydrophobic nature of the cell adhesion surface polymer layer surface.
- a load may be applied to the extent that the cultured cells are not burdened, or a sufficient time may be taken until adhesion.
- an operation for promoting adhesion of cultured cells such as increasing or decreasing the amount of medium and changing the culture temperature, may be used in combination.
- the attaching operation may be automatically performed using a Z-stage that can be operated in the vertical direction.
- the cultured cells attached to the cell sheet transplantation jig by the above-described method can be freely moved to a desired location and transplanted by moving together with the cell sheet transplantation jig. At that time, it is better that the cultured cells are moved aseptically in order to prevent the cultured cells from being contaminated. Further, the moving operation may be performed under humidification so that the attached cells are not dried.
- the cultured cells moved by the above-described method are placed on a place where they are to be reattached and reattached.
- the method of reattaching is not particularly limited, but usually, after the cultured cells that have moved are attached to the place where they want to reattach, the cell adhesion surface of the cell sheet transplantation jig and the cultured cells are attached. The operation is completed by weakening and separating the cell sheet transplantation jig from the cultured cells.
- a method of peeling the cell adhesion surface and the cultured cells together on the cell adhesion surface of the cell sheet transplantation jig may be used.
- the amino acid adheres more strongly than the adhesion between the cell adhesion protein and the cell.
- a method of adding a peptide, a protein, etc., and a method of adding a medium sufficiently.
- the cell adhesion surface of the cell sheet transplanting jig is a hydrophilic polymer
- a method of sufficiently adding a medium to weaken the water absorption power of the hydrophilic polymer, by changing the surface of the cell adhesion surface polymer layer to sufficiently hydrophilic The cultured cells can be detached.
- it in order to promote the attachment to the place where it wants to reattach, it is used in combination with applying a load to the extent that the cultured cells are not burdened, taking enough time to attach, and changing the culture temperature. You may do it.
- the place to be transplanted in the present invention is not particularly limited, and may be, for example, on the surface of a culture substrate, the surface of an in vivo tissue, the surface of an in vitro tissue, another cultured cell, or another cultured cell sheet described later.
- Examples of the in vivo tissue surface and in vitro tissue surface herein include, but are not limited to, humans, dogs, cats, rabbits, rats, pigs, and sheep.
- Another cultured cell includes corneal epithelial cells, epidermal keratinocytes, oral mucosal cells, conjunctival epithelial cells, cardiomyocytes, fibroblasts, vascular endothelial cells, hepatic parenchymal cells, or a mixture of two or more.
- the type is not limited at all, but when the cultured cells of the present invention are used for human therapy, it is desirable to use human-derived cells.
- the cultured cells can be peeled off from the surface of the cell culture substrate just by changing the culture temperature as shown in International Application Publication No. WO02 / 08387.
- the peeling operation, moving operation, and reattachment operation can be performed easily and accurately.
- the temperature-responsive polymer coated on the substrate surface has an upper critical solution temperature or a lower critical solution temperature of 0 ° C. to 80 ° C., more preferably 20 ° C. to 50 ° C. in an aqueous solution. If the upper critical lysis temperature or the lower critical lysis temperature exceeds 80 ° C., the cells may die, which is not preferable. Further, if the upper critical lysis temperature or the lower critical lysis temperature is lower than 0 ° C., the cell growth rate is generally extremely reduced or cells are killed, which is also not preferable.
- the temperature-responsive polymer used in the present invention may be either a homopolymer or a copolymer.
- a polymer examples include polymers described in JP-A-2-21865. Specifically, for example, it can be obtained by homopolymerization or copolymerization of the following monomers.
- the monomer that can be used include a (meth) acrylamide compound, an N- (or N, N-di) alkyl-substituted (meth) acrylamide derivative, or a vinyl ether derivative. Two or more of these can be used.
- copolymerization with monomers other than the above monomers, grafting or copolymerization of polymers, or a mixture of polymers and copolymers may be used.
- the method for coating the surface of the base material with the temperature-responsive polymer is not particularly limited.
- the method described in JP-A-2-21865 may be used. That is, such coating is performed by applying a substrate and the above monomer or polymer to one of electron beam irradiation (EB), ⁇ -ray irradiation, ultraviolet irradiation, plasma treatment, corona treatment, organic polymerization reaction, or physical application such as coating and kneading. It can be performed by, for example, mechanical adsorption.
- the coating amount of the temperature-responsive polymer is preferably in the range of 0.4 to 4.5 ⁇ g / cm 2 , preferably 0.7 to 3.5 ⁇ g / cm 2 , more preferably 0.9 to 3.0 ⁇ g / cm 2. cm 2 .
- the coating amount is less than 0.2 ⁇ g / cm 2 , the cells on the polymer are hardly detached even when a stimulus is applied, and the working efficiency is remarkably deteriorated.
- it is 4.5 ⁇ g / cm 2 or more, it is difficult for cells to adhere to the region, and it becomes difficult to sufficiently attach the cells.
- the present invention in order to exfoliate the cultured cells in a sheet form and stack the cultured cell sheets using a cell sheet transplanting jig, or to transplant the cultured cell sheets to an in vivo tissue or an in vitro tissue.
- the cells must be cultured on a cell culture substrate coated with a temperature-responsive polymer, and the cultured cells must be peeled into a sheet.
- the temperature of the medium is not more than that temperature when the polymer coated on the surface of the culture substrate has an upper critical solution temperature, and particularly if it is more than that temperature if the polymer has a lower critical solution temperature. Not limited.
- the culture conditions other than the temperature may be in accordance with conventional methods and are not particularly limited.
- the medium to be used may be a medium to which serum such as known fetal calf serum (FCS) is added, or a serum-free medium to which such serum is not added.
- FCS fetal calf serum
- the cultured cell sheet in order to peel and recover the cultured cell sheet from the cell culture substrate coated with the temperature-responsive polymer, the cultured cell sheet is attached to the cell sheet transplanting jig, and the temperature of the culture substrate surface is adjusted. Separation can be achieved by setting the temperature to the upper critical solution temperature or higher or the lower critical solution temperature or lower. It should be noted that peeling the cultured cell sheet can be performed in a culture solution in which cells are cultured or in another isotonic solution, and can be selected according to the purpose.
- the cultured cell sheet peeled off from the cell culture substrate coated with the temperature-responsive polymer in the present invention and obtained by using a cell sheet transplanting jig is a proteolytic enzyme represented by dispase, trypsin, etc.
- the cell-substrate basement membrane-like protein formed during culture is not damaged by the enzyme, and the cell-cell desmosome structure is retained, resulting in structural defects. There is little and high strength.
- a cell sheet transplantation jig it is possible to accurately stack the cultured cell sheets or move them accurately to the affected tissue. For these reasons, for example, at the time of transplantation of a cultured cell sheet, it can be adhered to the affected tissue well and accurately, and an efficient treatment can be performed.
- the method for fixing the cultured cell sheet and the living tissue shown in the present invention is not particularly limited, and the cultured cell sheet and the living tissue may be sutured, or the cultured cell sheet shown in the present invention is a living body.
- the cultured cell sheet attached to the affected part does not need to be sutured to the living body side so as to be engrafted with the tissue quickly.
- the cell sheet transplantation jig having a suction hole of the present invention By using the cell sheet transplantation jig having a suction hole of the present invention, cultured cells in an arbitrary range on the cell culture substrate can be efficiently detached, and the detached cultured cells can be easily transplanted again. Become. For this reason, the cultured cells can be easily transplanted to a place where the cultured cells are desired to be transferred, and can be transplanted accurately.
- Example 1 As specific examples of the cell sheet transplanting jig according to the present invention, four types of shapes with different arrangements of suction holes were designed (FIGS. 1 to 4).
- (A) shows the entire image of the jig, and (b) shows the suction hole and the bottom surface with the surface for capturing the cell sheet.
- FIG. 5 shows specific dimensions of the jig shown in FIG.
- FIG. 6 shows an actual prototype of the jig designed in FIG. This can be improved so that it is easy to transplant a cell sheet by applying a pattern by two kinds of methods as shown in FIG.
- FIG. 8 shows a jig that can be fixed by threading the cell sheet.
- FIG. 9 specifically shows a jig capable of transplanting three cell sheets in a single transplant.
- FIG. 10 specifically shows a jig capable of transplanting six cell sheets by one transplant.
- Example 2 The cell sheet transplanting jig according to the present invention was molded using a soft silicone resin. The external appearance from the upper surface of the entire transplanted jig is shown in FIG.
- the cell sheet capturing portion (the surface on the opposite side of the figure) has the suction holes as shown in the present invention.
- FIG. 12 shows that the product of the present invention is a soft material. As a result, it was found that the product of the present invention can be adhered to the spherical pig heart surface (FIG. 13).
- Example 3 A surface portion for capturing a cell sheet while maintaining a sheet-like shape and (2) a surface portion having a suction hole for sucking and fixing a transplanted portion around the cell sheet is housed in the tube.
- the transplant jig which can take in and out the said surface part as needed was shown concretely.
- the cell sheet transplantation jig described in the present invention is used, cultured cells in an arbitrary range on the cell culture substrate can be efficiently detached, and the detached cultured cells can be easily transplanted. For this reason, the cultured cells can be easily transplanted to a place where the cultured cells are desired to be transferred, and can be transplanted accurately. Furthermore, if a cell culture substrate surface coated with a temperature-responsive polymer is used, a cultured cell sheet with extremely high engraftment on living tissue can be obtained.
- the cultured cell sheet obtained by this method is strongly expected to have clinical applications such as corneal transplantation, skin transplantation, corneal disease treatment, ischemic heart disease treatment and the like. Therefore, the present invention is extremely useful in the fields of medicine, biology, etc., such as cell engineering and medical engineering.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Biomedical Technology (AREA)
- General Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Wood Science & Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Veterinary Medicine (AREA)
- Oral & Maxillofacial Surgery (AREA)
- Transplantation (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Genetics & Genomics (AREA)
- General Engineering & Computer Science (AREA)
- Medicinal Chemistry (AREA)
- Epidemiology (AREA)
- Biochemistry (AREA)
- Dermatology (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Sustainable Development (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Molecular Biology (AREA)
- Dispersion Chemistry (AREA)
- Botany (AREA)
- Cell Biology (AREA)
- Immunology (AREA)
- Cardiology (AREA)
- Heart & Thoracic Surgery (AREA)
- Vascular Medicine (AREA)
- Materials For Medical Uses (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Apparatus Associated With Microorganisms And Enzymes (AREA)
Abstract
Description
(a) シート状の培養細胞を移植するための平面を有する治具であって、同一方向の平面において(1)細胞シートをシート状の形状を保たせながら捕捉する面部分と(2)その周囲に被移植部を吸引固定するための吸引孔を有する細胞シート移植治具。
(b) 細胞シート移植治具が平板状である、(a)記載の細胞シート移植治具。
(c) 細胞シート移植治具が弾性体である、(a),(b)のいずれか記載の細胞シート移植治具。
(d) 細胞シートを捕捉する面部分が、細胞培養基材表面上に培養させた細胞を剥離させ、その後、その剥離させた培養細胞を再び他の場所へ付着させるための細胞接着面である、(a)~(c)のいずれか1つに記載の細胞シート移植治具。
(e) 細胞シートを捕捉する面部分に、細胞シートを吸引固定するための吸引孔を有する、(a)~(d)のいずれか1つに記載の細胞シート移植治具。
(f) 細胞シートを捕捉する面部分が凸状の弾性体であって、細胞培養基材表面と接する際、当該細胞接着面の凸部先端部から接し始め、最終的に細胞培養基材の任意の表面全体と接触することのできる、(a)~(e)請求項1~5のいずれか1つに記載の細胞シート移植治具。
(g) 細胞シートを捕捉する面部分に細胞接着性タンパク質、細胞接着性ペプチド、或いは親水性ポリマーの1種、もしくは2種以上を付着させたものである、(a)~(f)のいずれか1つに記載の細胞シート移植治具。
(h) 細胞接着性タンパク質がフィブリンゲル、フィブロネクチン、ラミニン、コラーゲン、ゼラチンの1種、もしくは2種以上からなるものである、(g)記載の細胞シート移植治具。
(i) 親水性ポリマーが含水ゲルである、(g)記載の細胞シート移植治具。
(j) 親水性ポリマーが温度応答性ポリマーである、(g)記載の細胞シート移植治具。
(k) (1)(a)~(j)のいずれかに記載の細胞シート移植治具に設けた細胞シートを捕捉する面に細胞培養基材上の培養細胞を付着させることで培養細胞シートを細胞培養基材上から剥離させ、(2)当該細胞シート移植治具を被移植部まで移動し、次に(3)当該細胞シート移植治具に設けた吸引孔で被移植部を吸引固定し、(4)その固定された被移植部へ細胞シート捕捉面に付着した細胞シートを細胞接着面と培養細胞との付着力を弱めることで剥離させた培養細胞シートを被移植面へ付着させることを特徴とする細胞シート移植治具の利用方法。
(l) (k)(2)において、当該細胞シート移植治具を別の培養細胞シート上に置き、2以上の細胞シートを積層化させた後、当該細胞シート移植治具を被移植部まで移動することを特徴とする(k)記載の細胞シート移植治具の利用方法。
(m) 被移植部が生体組織内である、(k),(l)のいずれか1つに記載の細胞シート移植治具の利用方法。
(n) 被移植部が組織の一部或いは全部を損傷もしくは欠損した部分である、(k)~(m)のいずれか1つに記載の細胞シート移植治具の利用方法。
(o) 組織の一部或いは全部を損傷もしくは欠損した患部に対し、(a)~(j)のいずれかに記載の細胞シート移植治具を用いてシート状の培養細胞を生体組織内に移植することを特徴とする治療法。
本発明である細胞シート移植治具の具体例として、吸引孔の配置の異なる4種類の形状を設計した(図1~4)。それぞれ(a)に治具の全体像を、(b)に吸引孔、並びに細胞シートを捕捉する面のある底面を示す。図5に図3に示す治具の具体的な寸法を示す。図6に図2で設計した治具を実際に試作した。このものは、図7に示されるように2種類の方法で柄を付け、細胞シートを移植し易いように改良できる。さらに、図8では細胞シートに糸を掛けて固定できる治具を示した。また、図9には1度の移植で3個の細胞シートを移植できる治具を具体的に示した。さらに、図10には1度の移植で6個の細胞シートを移植できる治具を具体的に示した。
本発明である細胞シート移植治具を柔らかな素材であるシリコン樹脂を使って成型加工した。作製した移植治具の全体の上面からの外観を図11に示す。細胞シート捕捉部(図の反対側の面)には、本発明で示すところの吸引孔を有している。本発明品が柔らかな材質であることを図12に示す。その結果、球面状のブタの心臓表面に対しても本発明品が密着できることが分かった(図13)。
本発明である(1)細胞シートをシート状の形状を保たせながら捕捉する面部分と(2)その周囲に被移植部を吸引固定するための吸引孔を有する面部分を管内に収納し、必要に応じ当該面部分を出し入れできる移植治具を具体的に示した。
Claims (14)
- シート状の培養細胞を移植するための平面を有する治具であって、同一方向の平面において(1)細胞シートをシート状の形状を保たせながら捕捉する面部分と(2)その周囲に被移植部を吸引固定するための吸引孔を有する、細胞シート移植治具。
- 細胞シート移植治具が平板状である、請求項1記載の細胞シート移植治具。
- 細胞シート移植治具が弾性体である、請求項1、2のいずれか1項記載の細胞シート移植治具。
- 細胞シートを捕捉する面部分が、細胞培養基材表面上に培養させた細胞を剥離させ、その後、その剥離させた培養細胞を再び他の場所へ付着させるための細胞接着面である、請求項1~3のいずれか1項記載の細胞シート移植治具。
- 細胞シートを捕捉する面部分に、細胞シートを吸引固定するための吸引孔を有する、請求項1~4のいずれか1項記載の細胞シート移植治具。
- 細胞シートを捕捉する面部分が凸状の弾性体であって、細胞培養基材表面と接する際、当該細胞接着面の凸部先端部から接し始め、最終的に細胞培養基材の任意の表面全体と接触することのできる、請求項1~5のいずれか1項記載の細胞シート移植治具。
- 細胞シートを捕捉する面部分に細胞接着性タンパク質、細胞接着性ペプチド、或いは親水性ポリマーの1種、もしくは2種以上を付着させたものである、請求項1~6のいずれか1項記載の細胞シート移植治具。
- 細胞接着性タンパク質がフィブリンゲル、フィブロネクチン、ラミニン、コラーゲン、ゼラチンの1種、もしくは2種以上からなるものである、請求項7記載の細胞シート移植治具。
- 親水性ポリマーが含水ゲルである、請求項7記載の細胞シート移植治具。
- 親水性ポリマーが温度応答性ポリマーである、請求項7記載の細胞シート移植治具。
- (1)請求項1~9記載の細胞シート移植治具に設けた細胞シートを捕捉する面に細胞培養基材上の培養細胞を付着させることで培養細胞シートを細胞培養基材上から剥離させ、(2)当該細胞シート移植治具を被移植部まで移動し、次に(3)当該細胞シート移植治具に設けた吸引孔で被移植部を吸引固定し、(4)その固定された被移植部へ細胞シート捕捉面に付着した細胞シートを細胞接着面と培養細胞との付着力を弱めることで剥離させた培養細胞シートを被移植面へ付着させることを特徴とする細胞シート移植治具の利用方法。
- 請求項11(2)において、当該細胞シート移植治具を別の培養細胞シート上に置き、2以上の細胞シートを積層化させた後、当該細胞シート移植治具を被移植部まで移動することを特徴とする請求項11記載の細胞シート移植治具の利用方法。
- 被移植部が生体組織内である、請求項11、12のいずれか1項記載の細胞シート移植治具の利用方法。
- 被移植部が組織の一部或いは全部を損傷もしくは欠損した部分である、請求項11~13のいずれか1項記載の細胞シート移植治具の利用方法。
Priority Applications (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
EP12763447.5A EP2692852A4 (en) | 2011-03-31 | 2012-04-02 | Device for cell layer transport and method for its use |
JP2013507845A JP5943907B2 (ja) | 2011-03-31 | 2012-04-02 | 細胞シート移植治具及びその利用方法 |
US14/008,769 US20150032223A1 (en) | 2011-03-31 | 2012-04-02 | Cell sheet transplantation device and method for using the same |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP2011-092461 | 2011-03-31 | ||
JP2011092461A JP5816452B2 (ja) | 2011-03-31 | 2011-03-31 | 細胞シート移植治具及びその利用方法 |
Publications (1)
Publication Number | Publication Date |
---|---|
WO2012133900A1 true WO2012133900A1 (ja) | 2012-10-04 |
Family
ID=46931587
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/JP2012/059003 WO2012133900A1 (ja) | 2011-03-31 | 2012-04-02 | 細胞シート移植治具及びその利用方法 |
Country Status (4)
Country | Link |
---|---|
US (1) | US20150032223A1 (ja) |
EP (1) | EP2692852A4 (ja) |
JP (2) | JP5816452B2 (ja) |
WO (1) | WO2012133900A1 (ja) |
Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2015070832A (ja) * | 2013-10-02 | 2015-04-16 | 國立中央大學 | 細胞培養用製品及びその製造方法 |
JP2015070804A (ja) * | 2013-10-02 | 2015-04-16 | テルモ株式会社 | シート状細胞培養物の剥離方法および製造方法ならびにこれらに用いる容器 |
WO2021024943A1 (ja) * | 2019-08-02 | 2021-02-11 | 積水化学工業株式会社 | 細胞培養用足場材及び細胞培養用容器 |
Families Citing this family (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP5816452B2 (ja) * | 2011-03-31 | 2015-11-18 | 株式会社セルシード | 細胞シート移植治具及びその利用方法 |
WO2014192909A1 (ja) | 2013-05-31 | 2014-12-04 | iHeart Japan株式会社 | ハイドロゲルを組み込んだ積層化細胞シート |
US20190233788A1 (en) * | 2016-07-05 | 2019-08-01 | Korea Advanced Institute Of Science And Technology | Production Method For And Use Of Polymer Thin-Film Culture Plat For Production Method For And Application Of Cell Sheet |
WO2018106414A1 (en) | 2016-12-07 | 2018-06-14 | Mayo Foundation For Medical Education And Research | Methods and materials for using fibrin supports for retinal pigment epithelium transplantation |
KR102109455B1 (ko) * | 2017-07-28 | 2020-05-13 | 주식회사 아모라이프사이언스 | 세포 배양 플레이트 설치장치 |
Citations (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JPH02211865A (ja) | 1989-02-10 | 1990-08-23 | Kao Corp | 細胞培養支持体材料 |
JPH05192138A (ja) | 1992-01-22 | 1993-08-03 | Kao Corp | 皮膚細胞培養法及び培養皮膚 |
WO2002008387A1 (fr) | 2000-07-21 | 2002-01-31 | Cellseed Inc. | Feuille cellulaire du type muscle cardiaque, construction tridimensionnelle, tissu du type muscle cardiaque et procede de production associe |
JP2005176812A (ja) | 2003-12-15 | 2005-07-07 | Mitsuo Okano | 培養細胞移動治具及びその利用方法 |
JP2008043239A (ja) * | 2006-08-11 | 2008-02-28 | Dainippon Printing Co Ltd | 細胞転写用部材 |
JP2010075081A (ja) * | 2008-09-25 | 2010-04-08 | Nitto Denko Corp | 細胞シート搬送治具 |
JP2010088416A (ja) * | 2008-10-09 | 2010-04-22 | Cellseed Inc | 培養細胞移動治具及びその利用方法 |
JP2010094114A (ja) * | 2008-10-14 | 2010-04-30 | Cellseed Inc | 培養細胞移動治具、及びその利用方法 |
Family Cites Families (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
DD284843A5 (de) * | 1989-09-25 | 1990-11-28 | Veb Kombinat Polygraph "Werner Lamberz",Dd | Saugscheiben zum ausrichten von bogen |
US7385591B2 (en) * | 2001-03-31 | 2008-06-10 | Microsoft Corporation | Out-of-vocabulary word determination and user interface for text input via reduced keypad keys |
JP5698900B2 (ja) * | 2009-03-19 | 2015-04-08 | テルモ株式会社 | 細胞培養物移送器具 |
JP5816452B2 (ja) * | 2011-03-31 | 2015-11-18 | 株式会社セルシード | 細胞シート移植治具及びその利用方法 |
-
2011
- 2011-03-31 JP JP2011092461A patent/JP5816452B2/ja not_active Expired - Fee Related
-
2012
- 2012-04-02 US US14/008,769 patent/US20150032223A1/en not_active Abandoned
- 2012-04-02 JP JP2013507845A patent/JP5943907B2/ja not_active Expired - Fee Related
- 2012-04-02 WO PCT/JP2012/059003 patent/WO2012133900A1/ja active Application Filing
- 2012-04-02 EP EP12763447.5A patent/EP2692852A4/en not_active Withdrawn
Patent Citations (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JPH02211865A (ja) | 1989-02-10 | 1990-08-23 | Kao Corp | 細胞培養支持体材料 |
JPH05192138A (ja) | 1992-01-22 | 1993-08-03 | Kao Corp | 皮膚細胞培養法及び培養皮膚 |
WO2002008387A1 (fr) | 2000-07-21 | 2002-01-31 | Cellseed Inc. | Feuille cellulaire du type muscle cardiaque, construction tridimensionnelle, tissu du type muscle cardiaque et procede de production associe |
JP2005176812A (ja) | 2003-12-15 | 2005-07-07 | Mitsuo Okano | 培養細胞移動治具及びその利用方法 |
JP2008043239A (ja) * | 2006-08-11 | 2008-02-28 | Dainippon Printing Co Ltd | 細胞転写用部材 |
JP2010075081A (ja) * | 2008-09-25 | 2010-04-08 | Nitto Denko Corp | 細胞シート搬送治具 |
JP2010088416A (ja) * | 2008-10-09 | 2010-04-22 | Cellseed Inc | 培養細胞移動治具及びその利用方法 |
JP2010094114A (ja) * | 2008-10-14 | 2010-04-30 | Cellseed Inc | 培養細胞移動治具、及びその利用方法 |
Non-Patent Citations (1)
Title |
---|
See also references of EP2692852A4 |
Cited By (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2015070832A (ja) * | 2013-10-02 | 2015-04-16 | 國立中央大學 | 細胞培養用製品及びその製造方法 |
JP2015070804A (ja) * | 2013-10-02 | 2015-04-16 | テルモ株式会社 | シート状細胞培養物の剥離方法および製造方法ならびにこれらに用いる容器 |
US9902941B2 (en) | 2013-10-02 | 2018-02-27 | National Central University | Method for manufacturing a cell culturing article |
US10336986B2 (en) | 2013-10-02 | 2019-07-02 | National Central University | Cell culturing article and method for manufacturing thereof |
WO2021024943A1 (ja) * | 2019-08-02 | 2021-02-11 | 積水化学工業株式会社 | 細胞培養用足場材及び細胞培養用容器 |
Also Published As
Publication number | Publication date |
---|---|
JPWO2012133900A1 (ja) | 2014-07-28 |
JP5943907B2 (ja) | 2016-07-05 |
JP5816452B2 (ja) | 2015-11-18 |
US20150032223A1 (en) | 2015-01-29 |
JP2014075979A (ja) | 2014-05-01 |
EP2692852A1 (en) | 2014-02-05 |
EP2692852A4 (en) | 2014-10-01 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
JP5943907B2 (ja) | 細胞シート移植治具及びその利用方法 | |
JP4486359B2 (ja) | 培養細胞移動治具及びその利用方法 | |
JP5080848B2 (ja) | 細胞培養支持体とその製造方法 | |
US20100184182A1 (en) | Method for preparing biological tissue | |
US20100216242A1 (en) | Cell culture support and production method and uses thereof | |
JP2016039806A (ja) | サイトカイン産生細胞シートとその利用方法 | |
JP2003038170A (ja) | 前眼部関連細胞シート、3次元構造体、及びそれらの製造法 | |
US20100184222A1 (en) | Method for preparing biological tissue | |
US9440004B2 (en) | Method for preparing biological tissue | |
JP2006320304A (ja) | 密閉系細胞培養容器及びそれを利用した細胞培養方法 | |
US20130101659A1 (en) | Cell sheet for myocardial regeneration, method of producing the same, and method of using the same | |
JP6468714B2 (ja) | 培養容器及びそれを利用した複数の積層細胞シートの作製方法 | |
JP5731728B2 (ja) | 密閉系細胞培養容器及びそれを利用した細胞培養方法 | |
JP2015204838A (ja) | 肝組織細胞機能の長期維持方法 | |
JP2010094114A (ja) | 培養細胞移動治具、及びその利用方法 | |
JP5926764B2 (ja) | 培養細胞移動治具、及びその利用方法 | |
JP5815052B2 (ja) | 培養細胞移動治具及びその利用方法 | |
JP2010088416A (ja) | 培養細胞移動治具及びその利用方法 | |
JP5252828B2 (ja) | 上皮系細胞培養方法 | |
EP3298124A1 (en) | Cell culture substrate for rapid release and re-plating | |
JP2008035834A (ja) | 細胞の培養方法、細胞培養アレイ装置、細胞培養物及び再生医用生体材料 | |
JP6452304B2 (ja) | 細胞シート培養基材、細胞シート培養基材複合物、及び細胞シート/培養基材複合の製造方法 | |
JP5837530B2 (ja) | 密閉系細胞培養容器及びそれを利用した細胞培養方法 | |
JP6111013B2 (ja) | 腱細胞シート及びその製造方法 | |
JP6835266B2 (ja) | 細胞培養容器及び細胞積層体の作製方法 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
121 | Ep: the epo has been informed by wipo that ep was designated in this application |
Ref document number: 12763447 Country of ref document: EP Kind code of ref document: A1 |
|
ENP | Entry into the national phase |
Ref document number: 2013507845 Country of ref document: JP Kind code of ref document: A |
|
NENP | Non-entry into the national phase |
Ref country code: DE |
|
WWE | Wipo information: entry into national phase |
Ref document number: 2012763447 Country of ref document: EP |
|
WWE | Wipo information: entry into national phase |
Ref document number: 14008769 Country of ref document: US |