WO2021200545A1 - 幹細胞の染色体安定化剤 - Google Patents
幹細胞の染色体安定化剤 Download PDFInfo
- Publication number
- WO2021200545A1 WO2021200545A1 PCT/JP2021/012557 JP2021012557W WO2021200545A1 WO 2021200545 A1 WO2021200545 A1 WO 2021200545A1 JP 2021012557 W JP2021012557 W JP 2021012557W WO 2021200545 A1 WO2021200545 A1 WO 2021200545A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- stem cells
- nmn
- cells
- chromosomal
- medium
- Prior art date
Links
- 210000000130 stem cell Anatomy 0.000 title claims abstract description 73
- 239000003381 stabilizer Substances 0.000 title claims abstract description 30
- 210000000349 chromosome Anatomy 0.000 title abstract description 28
- 238000000034 method Methods 0.000 claims abstract description 21
- 239000001963 growth medium Substances 0.000 claims abstract description 19
- 238000012258 culturing Methods 0.000 claims abstract description 18
- 150000003839 salts Chemical class 0.000 claims abstract description 17
- 239000004480 active ingredient Substances 0.000 claims abstract description 13
- FZAQROFXYZPAKI-UHFFFAOYSA-N anthracene-2-sulfonyl chloride Chemical compound C1=CC=CC2=CC3=CC(S(=O)(=O)Cl)=CC=C3C=C21 FZAQROFXYZPAKI-UHFFFAOYSA-N 0.000 claims abstract description 12
- 239000012453 solvate Substances 0.000 claims abstract description 10
- 210000001778 pluripotent stem cell Anatomy 0.000 claims abstract description 8
- 230000002759 chromosomal effect Effects 0.000 claims description 29
- 210000002901 mesenchymal stem cell Anatomy 0.000 claims description 21
- 210000001671 embryonic stem cell Anatomy 0.000 claims description 5
- 210000003958 hematopoietic stem cell Anatomy 0.000 claims description 5
- 210000004263 induced pluripotent stem cell Anatomy 0.000 claims description 5
- 238000012136 culture method Methods 0.000 claims description 4
- 230000006641 stabilisation Effects 0.000 claims description 4
- 238000011105 stabilization Methods 0.000 claims description 4
- 230000000087 stabilizing effect Effects 0.000 claims description 4
- 239000003795 chemical substances by application Substances 0.000 claims 1
- 210000004027 cell Anatomy 0.000 description 47
- DAYLJWODMCOQEW-TURQNECASA-N NMN zwitterion Chemical compound NC(=O)C1=CC=C[N+]([C@H]2[C@@H]([C@H](O)[C@@H](COP(O)([O-])=O)O2)O)=C1 DAYLJWODMCOQEW-TURQNECASA-N 0.000 description 44
- 239000002609 medium Substances 0.000 description 34
- 239000007640 basal medium Substances 0.000 description 17
- 238000012360 testing method Methods 0.000 description 17
- 208000031404 Chromosome Aberrations Diseases 0.000 description 14
- 206010008805 Chromosomal abnormalities Diseases 0.000 description 13
- -1 for example Chemical compound 0.000 description 7
- 239000000047 product Substances 0.000 description 7
- 239000002253 acid Substances 0.000 description 6
- 230000000694 effects Effects 0.000 description 6
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 6
- 208000037051 Chromosomal Instability Diseases 0.000 description 5
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 5
- 238000004458 analytical method Methods 0.000 description 5
- 210000003855 cell nucleus Anatomy 0.000 description 5
- XJMOSONTPMZWPB-UHFFFAOYSA-M propidium iodide Chemical compound [I-].[I-].C12=CC(N)=CC=C2C2=CC=C(N)C=C2[N+](CCC[N+](C)(CC)CC)=C1C1=CC=CC=C1 XJMOSONTPMZWPB-UHFFFAOYSA-M 0.000 description 5
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 4
- 206010028980 Neoplasm Diseases 0.000 description 4
- 239000003112 inhibitor Substances 0.000 description 4
- 239000000463 material Substances 0.000 description 4
- 230000002062 proliferating effect Effects 0.000 description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- 102000004877 Insulin Human genes 0.000 description 3
- 108090001061 Insulin Proteins 0.000 description 3
- WMFOQBRAJBCJND-UHFFFAOYSA-M Lithium hydroxide Chemical compound [Li+].[OH-] WMFOQBRAJBCJND-UHFFFAOYSA-M 0.000 description 3
- 241001465754 Metazoa Species 0.000 description 3
- 102000015532 Nicotinamide phosphoribosyltransferase Human genes 0.000 description 3
- 108010064862 Nicotinamide phosphoribosyltransferase Proteins 0.000 description 3
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 description 3
- 230000005856 abnormality Effects 0.000 description 3
- 238000004113 cell culture Methods 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 230000004069 differentiation Effects 0.000 description 3
- 239000003814 drug Substances 0.000 description 3
- 229940125396 insulin Drugs 0.000 description 3
- 230000001172 regenerating effect Effects 0.000 description 3
- 210000001519 tissue Anatomy 0.000 description 3
- GHOKWGTUZJEAQD-ZETCQYMHSA-N (D)-(+)-Pantothenic acid Chemical compound OCC(C)(C)[C@@H](O)C(=O)NCCC(O)=O GHOKWGTUZJEAQD-ZETCQYMHSA-N 0.000 description 2
- HZAXFHJVJLSVMW-UHFFFAOYSA-N 2-Aminoethan-1-ol Chemical compound NCCO HZAXFHJVJLSVMW-UHFFFAOYSA-N 0.000 description 2
- IOJUJUOXKXMJNF-UHFFFAOYSA-N 2-acetyloxybenzoic acid [3-(nitrooxymethyl)phenyl] ester Chemical compound CC(=O)OC1=CC=CC=C1C(=O)OC1=CC=CC(CO[N+]([O-])=O)=C1 IOJUJUOXKXMJNF-UHFFFAOYSA-N 0.000 description 2
- PMYDPQQPEAYXKD-UHFFFAOYSA-N 3-hydroxy-n-naphthalen-2-ylnaphthalene-2-carboxamide Chemical compound C1=CC=CC2=CC(NC(=O)C3=CC4=CC=CC=C4C=C3O)=CC=C21 PMYDPQQPEAYXKD-UHFFFAOYSA-N 0.000 description 2
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 2
- FERIUCNNQQJTOY-UHFFFAOYSA-N Butyric acid Chemical compound CCCC(O)=O FERIUCNNQQJTOY-UHFFFAOYSA-N 0.000 description 2
- RGHNJXZEOKUKBD-SQOUGZDYSA-N D-gluconic acid Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C(O)=O RGHNJXZEOKUKBD-SQOUGZDYSA-N 0.000 description 2
- 239000006145 Eagle's minimal essential medium Substances 0.000 description 2
- VZCYOOQTPOCHFL-OWOJBTEDSA-N Fumaric acid Chemical compound OC(=O)\C=C\C(O)=O VZCYOOQTPOCHFL-OWOJBTEDSA-N 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- AFVFQIVMOAPDHO-UHFFFAOYSA-N Methanesulfonic acid Chemical compound CS(O)(=O)=O AFVFQIVMOAPDHO-UHFFFAOYSA-N 0.000 description 2
- DFPAKSUCGFBDDF-UHFFFAOYSA-N Nicotinamide Chemical compound NC(=O)C1=CC=CN=C1 DFPAKSUCGFBDDF-UHFFFAOYSA-N 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- 102000004357 Transferases Human genes 0.000 description 2
- 108090000992 Transferases Proteins 0.000 description 2
- 108090001012 Transforming Growth Factor beta Proteins 0.000 description 2
- 102000004887 Transforming Growth Factor beta Human genes 0.000 description 2
- 208000037280 Trisomy Diseases 0.000 description 2
- 150000007513 acids Chemical class 0.000 description 2
- 210000004102 animal cell Anatomy 0.000 description 2
- 235000010323 ascorbic acid Nutrition 0.000 description 2
- 239000011668 ascorbic acid Substances 0.000 description 2
- 229960005070 ascorbic acid Drugs 0.000 description 2
- 239000002585 base Substances 0.000 description 2
- WPYMKLBDIGXBTP-UHFFFAOYSA-N benzoic acid Chemical compound OC(=O)C1=CC=CC=C1 WPYMKLBDIGXBTP-UHFFFAOYSA-N 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 239000006143 cell culture medium Substances 0.000 description 2
- 230000003247 decreasing effect Effects 0.000 description 2
- 238000000855 fermentation Methods 0.000 description 2
- 230000004151 fermentation Effects 0.000 description 2
- 238000012744 immunostaining Methods 0.000 description 2
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 2
- 210000003716 mesoderm Anatomy 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 238000010899 nucleation Methods 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 230000035755 proliferation Effects 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 235000018716 sodium selenate Nutrition 0.000 description 2
- 239000011655 sodium selenate Substances 0.000 description 2
- 229960001881 sodium selenate Drugs 0.000 description 2
- 230000004083 survival effect Effects 0.000 description 2
- 238000003786 synthesis reaction Methods 0.000 description 2
- WGTYBPLFGIVFAS-UHFFFAOYSA-M tetramethylammonium hydroxide Chemical compound [OH-].C[N+](C)(C)C WGTYBPLFGIVFAS-UHFFFAOYSA-M 0.000 description 2
- JOXIMZWYDAKGHI-UHFFFAOYSA-N toluene-4-sulfonic acid Chemical compound CC1=CC=C(S(O)(=O)=O)C=C1 JOXIMZWYDAKGHI-UHFFFAOYSA-N 0.000 description 2
- VZCYOOQTPOCHFL-UHFFFAOYSA-N trans-butenedioic acid Natural products OC(=O)C=CC(O)=O VZCYOOQTPOCHFL-UHFFFAOYSA-N 0.000 description 2
- GVJHHUAWPYXKBD-IEOSBIPESA-N α-tocopherol Chemical compound OC1=C(C)C(C)=C2O[C@@](CCC[C@H](C)CCC[C@H](C)CCCC(C)C)(C)CCC2=C1C GVJHHUAWPYXKBD-IEOSBIPESA-N 0.000 description 2
- QBYIENPQHBMVBV-HFEGYEGKSA-N (2R)-2-hydroxy-2-phenylacetic acid Chemical compound O[C@@H](C(O)=O)c1ccccc1.O[C@@H](C(O)=O)c1ccccc1 QBYIENPQHBMVBV-HFEGYEGKSA-N 0.000 description 1
- MIOPJNTWMNEORI-GMSGAONNSA-N (S)-camphorsulfonic acid Chemical compound C1C[C@@]2(CS(O)(=O)=O)C(=O)C[C@@H]1C2(C)C MIOPJNTWMNEORI-GMSGAONNSA-N 0.000 description 1
- BJEPYKJPYRNKOW-REOHCLBHSA-N (S)-malic acid Chemical compound OC(=O)[C@@H](O)CC(O)=O BJEPYKJPYRNKOW-REOHCLBHSA-N 0.000 description 1
- BMYNFMYTOJXKLE-UHFFFAOYSA-N 3-azaniumyl-2-hydroxypropanoate Chemical compound NCC(O)C(O)=O BMYNFMYTOJXKLE-UHFFFAOYSA-N 0.000 description 1
- 108010059616 Activins Proteins 0.000 description 1
- 102000009027 Albumins Human genes 0.000 description 1
- 108010088751 Albumins Proteins 0.000 description 1
- 239000004475 Arginine Substances 0.000 description 1
- 239000005552 B01AC04 - Clopidogrel Substances 0.000 description 1
- 239000005711 Benzoic acid Substances 0.000 description 1
- BVKZGUZCCUSVTD-UHFFFAOYSA-M Bicarbonate Chemical compound OC([O-])=O BVKZGUZCCUSVTD-UHFFFAOYSA-M 0.000 description 1
- HKXCBMWEWUFFHX-YUMQZZPRSA-N C[C@@H]1O[C@H](CO[O](OC)([O]#C)=O)CC1 Chemical compound C[C@@H]1O[C@H](CO[O](OC)([O]#C)=O)CC1 HKXCBMWEWUFFHX-YUMQZZPRSA-N 0.000 description 1
- 208000005623 Carcinogenesis Diseases 0.000 description 1
- GHOKWGTUZJEAQD-UHFFFAOYSA-N Chick antidermatitis factor Natural products OCC(C)(C)C(O)C(=O)NCCC(O)=O GHOKWGTUZJEAQD-UHFFFAOYSA-N 0.000 description 1
- 102000004127 Cytokines Human genes 0.000 description 1
- 108090000695 Cytokines Proteins 0.000 description 1
- RGHNJXZEOKUKBD-UHFFFAOYSA-N D-gluconic acid Natural products OCC(O)C(O)C(O)C(O)C(O)=O RGHNJXZEOKUKBD-UHFFFAOYSA-N 0.000 description 1
- FEWJPZIEWOKRBE-JCYAYHJZSA-N Dextrotartaric acid Chemical compound OC(=O)[C@H](O)[C@@H](O)C(O)=O FEWJPZIEWOKRBE-JCYAYHJZSA-N 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- PIICEJLVQHRZGT-UHFFFAOYSA-N Ethylenediamine Chemical compound NCCN PIICEJLVQHRZGT-UHFFFAOYSA-N 0.000 description 1
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 1
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 1
- 101150021185 FGF gene Proteins 0.000 description 1
- 102000016359 Fibronectins Human genes 0.000 description 1
- 108010067306 Fibronectins Proteins 0.000 description 1
- 102000001267 GSK3 Human genes 0.000 description 1
- 108060006662 GSK3 Proteins 0.000 description 1
- ZWQVYZXPYSYPJD-RYUDHWBXSA-N Glu-Gly-Phe Chemical compound OC(=O)CC[C@H](N)C(=O)NCC(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 ZWQVYZXPYSYPJD-RYUDHWBXSA-N 0.000 description 1
- WHUUTDBJXJRKMK-UHFFFAOYSA-N Glutamic acid Natural products OC(=O)C(N)CCC(O)=O WHUUTDBJXJRKMK-UHFFFAOYSA-N 0.000 description 1
- 102000017286 Histone H2A Human genes 0.000 description 1
- 108050005231 Histone H2A Proteins 0.000 description 1
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- 102100026818 Inhibin beta E chain Human genes 0.000 description 1
- 206010022489 Insulin Resistance Diseases 0.000 description 1
- AHLPHDHHMVZTML-BYPYZUCNSA-N L-Ornithine Chemical compound NCCC[C@H](N)C(O)=O AHLPHDHHMVZTML-BYPYZUCNSA-N 0.000 description 1
- ODKSFYDXXFIFQN-BYPYZUCNSA-P L-argininium(2+) Chemical compound NC(=[NH2+])NCCC[C@H]([NH3+])C(O)=O ODKSFYDXXFIFQN-BYPYZUCNSA-P 0.000 description 1
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 description 1
- KDXKERNSBIXSRK-YFKPBYRVSA-N L-lysine Chemical compound NCCCC[C@H](N)C(O)=O KDXKERNSBIXSRK-YFKPBYRVSA-N 0.000 description 1
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 description 1
- 239000004472 Lysine Substances 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- 229910002651 NO3 Inorganic materials 0.000 description 1
- NHNBFGGVMKEFGY-UHFFFAOYSA-N Nitrate Chemical compound [O-][N+]([O-])=O NHNBFGGVMKEFGY-UHFFFAOYSA-N 0.000 description 1
- AHLPHDHHMVZTML-UHFFFAOYSA-N Orn-delta-NH2 Natural products NCCCC(N)C(O)=O AHLPHDHHMVZTML-UHFFFAOYSA-N 0.000 description 1
- UTJLXEIPEHZYQJ-UHFFFAOYSA-N Ornithine Natural products OC(=O)C(C)CCCN UTJLXEIPEHZYQJ-UHFFFAOYSA-N 0.000 description 1
- GLUUGHFHXGJENI-UHFFFAOYSA-N Piperazine Chemical compound C1CNCCN1 GLUUGHFHXGJENI-UHFFFAOYSA-N 0.000 description 1
- OFOBLEOULBTSOW-UHFFFAOYSA-N Propanedioic acid Natural products OC(=O)CC(O)=O OFOBLEOULBTSOW-UHFFFAOYSA-N 0.000 description 1
- IWYDHOAUDWTVEP-UHFFFAOYSA-N R-2-phenyl-2-hydroxyacetic acid Natural products OC(=O)C(O)C1=CC=CC=C1 IWYDHOAUDWTVEP-UHFFFAOYSA-N 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- KEAYESYHFKHZAL-UHFFFAOYSA-N Sodium Chemical compound [Na] KEAYESYHFKHZAL-UHFFFAOYSA-N 0.000 description 1
- KDYFGRWQOYBRFD-UHFFFAOYSA-N Succinic acid Natural products OC(=O)CCC(O)=O KDYFGRWQOYBRFD-UHFFFAOYSA-N 0.000 description 1
- FEWJPZIEWOKRBE-UHFFFAOYSA-N Tartaric acid Natural products [H+].[H+].[O-]C(=O)C(O)C(O)C([O-])=O FEWJPZIEWOKRBE-UHFFFAOYSA-N 0.000 description 1
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 description 1
- 239000007983 Tris buffer Substances 0.000 description 1
- 229920004890 Triton X-100 Polymers 0.000 description 1
- 239000013504 Triton X-100 Substances 0.000 description 1
- 102000013814 Wnt Human genes 0.000 description 1
- ZYPMNZKYVVSXOJ-FRRDWIJNSA-N [(2s,3s,4s)-2,3,4-triacetyloxy-5-oxopentyl] acetate Chemical compound CC(=O)OC[C@H](OC(C)=O)[C@H](OC(C)=O)[C@H](OC(C)=O)C=O ZYPMNZKYVVSXOJ-FRRDWIJNSA-N 0.000 description 1
- 230000002159 abnormal effect Effects 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 239000000488 activin Substances 0.000 description 1
- 210000000577 adipose tissue Anatomy 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 229940050528 albumin Drugs 0.000 description 1
- 229910052783 alkali metal Inorganic materials 0.000 description 1
- 229910052784 alkaline earth metal Inorganic materials 0.000 description 1
- 229940087168 alpha tocopherol Drugs 0.000 description 1
- BJEPYKJPYRNKOW-UHFFFAOYSA-N alpha-hydroxysuccinic acid Natural products OC(=O)C(O)CC(O)=O BJEPYKJPYRNKOW-UHFFFAOYSA-N 0.000 description 1
- WNROFYMDJYEPJX-UHFFFAOYSA-K aluminium hydroxide Chemical compound [OH-].[OH-].[OH-].[Al+3] WNROFYMDJYEPJX-UHFFFAOYSA-K 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- 235000001014 amino acid Nutrition 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 229910021529 ammonia Inorganic materials 0.000 description 1
- 208000036878 aneuploidy Diseases 0.000 description 1
- 231100001075 aneuploidy Toxicity 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 1
- 229960003121 arginine Drugs 0.000 description 1
- UPABQMWFWCMOFV-UHFFFAOYSA-N benethamine Chemical compound C=1C=CC=CC=1CNCCC1=CC=CC=C1 UPABQMWFWCMOFV-UHFFFAOYSA-N 0.000 description 1
- JUHORIMYRDESRB-UHFFFAOYSA-N benzathine Chemical compound C=1C=CC=CC=1CNCCNCC1=CC=CC=C1 JUHORIMYRDESRB-UHFFFAOYSA-N 0.000 description 1
- SRSXLGNVWSONIS-UHFFFAOYSA-N benzenesulfonic acid Chemical compound OS(=O)(=O)C1=CC=CC=C1 SRSXLGNVWSONIS-UHFFFAOYSA-N 0.000 description 1
- 229940092714 benzenesulfonic acid Drugs 0.000 description 1
- 235000010233 benzoic acid Nutrition 0.000 description 1
- 230000001851 biosynthetic effect Effects 0.000 description 1
- KDYFGRWQOYBRFD-NUQCWPJISA-N butanedioic acid Chemical compound O[14C](=O)CC[14C](O)=O KDYFGRWQOYBRFD-NUQCWPJISA-N 0.000 description 1
- AXCZMVOFGPJBDE-UHFFFAOYSA-L calcium dihydroxide Chemical compound [OH-].[OH-].[Ca+2] AXCZMVOFGPJBDE-UHFFFAOYSA-L 0.000 description 1
- 239000000920 calcium hydroxide Substances 0.000 description 1
- 229910001861 calcium hydroxide Inorganic materials 0.000 description 1
- 230000036952 cancer formation Effects 0.000 description 1
- 229910002091 carbon monoxide Inorganic materials 0.000 description 1
- 150000001732 carboxylic acid derivatives Chemical class 0.000 description 1
- 231100000504 carcinogenesis Toxicity 0.000 description 1
- 230000004663 cell proliferation Effects 0.000 description 1
- OEYIOHPDSNJKLS-UHFFFAOYSA-N choline Chemical compound C[N+](C)(C)CCO OEYIOHPDSNJKLS-UHFFFAOYSA-N 0.000 description 1
- 229960001231 choline Drugs 0.000 description 1
- 231100000244 chromosomal damage Toxicity 0.000 description 1
- GKTWGGQPFAXNFI-HNNXBMFYSA-N clopidogrel Chemical compound C1([C@H](N2CC=3C=CSC=3CC2)C(=O)OC)=CC=CC=C1Cl GKTWGGQPFAXNFI-HNNXBMFYSA-N 0.000 description 1
- 239000005515 coenzyme Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 239000012228 culture supernatant Substances 0.000 description 1
- 238000012217 deletion Methods 0.000 description 1
- 230000037430 deletion Effects 0.000 description 1
- UREBDLICKHMUKA-CXSFZGCWSA-N dexamethasone Chemical compound C1CC2=CC(=O)C=C[C@]2(C)[C@]2(F)[C@@H]1[C@@H]1C[C@@H](C)[C@@](C(=O)CO)(O)[C@@]1(C)C[C@@H]2O UREBDLICKHMUKA-CXSFZGCWSA-N 0.000 description 1
- 229960003957 dexamethasone Drugs 0.000 description 1
- NIJJYAXOARWZEE-UHFFFAOYSA-N di-n-propyl-acetic acid Natural products CCCC(C(O)=O)CCC NIJJYAXOARWZEE-UHFFFAOYSA-N 0.000 description 1
- 235000014113 dietary fatty acids Nutrition 0.000 description 1
- ZBCBWPMODOFKDW-UHFFFAOYSA-N diethanolamine Chemical compound OCCNCCO ZBCBWPMODOFKDW-UHFFFAOYSA-N 0.000 description 1
- HPNMFZURTQLUMO-UHFFFAOYSA-N diethylamine Chemical compound CCNCC HPNMFZURTQLUMO-UHFFFAOYSA-N 0.000 description 1
- 229960001760 dimethyl sulfoxide Drugs 0.000 description 1
- 210000003981 ectoderm Anatomy 0.000 description 1
- 210000001900 endoderm Anatomy 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- 229940012017 ethylenediamine Drugs 0.000 description 1
- 210000002744 extracellular matrix Anatomy 0.000 description 1
- 229930195729 fatty acid Natural products 0.000 description 1
- 239000000194 fatty acid Substances 0.000 description 1
- 150000004665 fatty acids Chemical class 0.000 description 1
- MHMNJMPURVTYEJ-UHFFFAOYSA-N fluorescein-5-isothiocyanate Chemical compound O1C(=O)C2=CC(N=C=S)=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 MHMNJMPURVTYEJ-UHFFFAOYSA-N 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 239000001530 fumaric acid Substances 0.000 description 1
- 235000011087 fumaric acid Nutrition 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 239000000174 gluconic acid Substances 0.000 description 1
- 235000012208 gluconic acid Nutrition 0.000 description 1
- 239000004220 glutamic acid Substances 0.000 description 1
- 235000013922 glutamic acid Nutrition 0.000 description 1
- 159000000011 group IA salts Chemical class 0.000 description 1
- 235000009200 high fat diet Nutrition 0.000 description 1
- 210000003917 human chromosome Anatomy 0.000 description 1
- 125000004029 hydroxymethyl group Chemical group [H]OC([H])([H])* 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 238000002372 labelling Methods 0.000 description 1
- 239000004310 lactic acid Substances 0.000 description 1
- 235000014655 lactic acid Nutrition 0.000 description 1
- 229960000448 lactic acid Drugs 0.000 description 1
- 229960003646 lysine Drugs 0.000 description 1
- VTHJTEIRLNZDEV-UHFFFAOYSA-L magnesium dihydroxide Chemical compound [OH-].[OH-].[Mg+2] VTHJTEIRLNZDEV-UHFFFAOYSA-L 0.000 description 1
- 239000000347 magnesium hydroxide Substances 0.000 description 1
- 229910001862 magnesium hydroxide Inorganic materials 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- VZCYOOQTPOCHFL-UPHRSURJSA-N maleic acid Chemical compound OC(=O)\C=C/C(O)=O VZCYOOQTPOCHFL-UPHRSURJSA-N 0.000 description 1
- 239000011976 maleic acid Substances 0.000 description 1
- 229940098895 maleic acid Drugs 0.000 description 1
- 239000001630 malic acid Substances 0.000 description 1
- 235000011090 malic acid Nutrition 0.000 description 1
- 229940099690 malic acid Drugs 0.000 description 1
- 229960002510 mandelic acid Drugs 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 108010082117 matrigel Proteins 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 229940098779 methanesulfonic acid Drugs 0.000 description 1
- WSFSSNUMVMOOMR-NJFSPNSNSA-N methanone Chemical compound O=[14CH2] WSFSSNUMVMOOMR-NJFSPNSNSA-N 0.000 description 1
- 230000004898 mitochondrial function Effects 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 208000030454 monosomy Diseases 0.000 description 1
- 238000010172 mouse model Methods 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- NFVJNJQRWPQVOA-UHFFFAOYSA-N n-[2-chloro-5-(trifluoromethyl)phenyl]-2-[3-(4-ethyl-5-ethylsulfanyl-1,2,4-triazol-3-yl)piperidin-1-yl]acetamide Chemical compound CCN1C(SCC)=NN=C1C1CN(CC(=O)NC=2C(=CC=C(C=2)C(F)(F)F)Cl)CCC1 NFVJNJQRWPQVOA-UHFFFAOYSA-N 0.000 description 1
- 210000005036 nerve Anatomy 0.000 description 1
- 229960003966 nicotinamide Drugs 0.000 description 1
- 235000005152 nicotinamide Nutrition 0.000 description 1
- 239000011570 nicotinamide Substances 0.000 description 1
- 210000004940 nucleus Anatomy 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 230000008520 organization Effects 0.000 description 1
- 229960003104 ornithine Drugs 0.000 description 1
- WLJNZVDCPSBLRP-UHFFFAOYSA-N pamoic acid Chemical compound C1=CC=C2C(CC=3C4=CC=CC=C4C=C(C=3O)C(=O)O)=C(O)C(C(O)=O)=CC2=C1 WLJNZVDCPSBLRP-UHFFFAOYSA-N 0.000 description 1
- 239000011713 pantothenic acid Substances 0.000 description 1
- 229940055726 pantothenic acid Drugs 0.000 description 1
- 235000019161 pantothenic acid Nutrition 0.000 description 1
- 150000003904 phospholipids Chemical class 0.000 description 1
- 230000000865 phosphorylative effect Effects 0.000 description 1
- 239000003761 preservation solution Substances 0.000 description 1
- MFDFERRIHVXMIY-UHFFFAOYSA-N procaine Chemical compound CCN(CC)CCOC(=O)C1=CC=C(N)C=C1 MFDFERRIHVXMIY-UHFFFAOYSA-N 0.000 description 1
- 229960004919 procaine Drugs 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 239000011435 rock Substances 0.000 description 1
- 238000011076 safety test Methods 0.000 description 1
- 230000003248 secreting effect Effects 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 239000012312 sodium hydride Substances 0.000 description 1
- 229910000104 sodium hydride Inorganic materials 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 210000001988 somatic stem cell Anatomy 0.000 description 1
- 238000009331 sowing Methods 0.000 description 1
- 238000013112 stability test Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
- 239000011975 tartaric acid Substances 0.000 description 1
- 235000002906 tartaric acid Nutrition 0.000 description 1
- ZRKFYGHZFMAOKI-QMGMOQQFSA-N tgfbeta Chemical compound C([C@H](NC(=O)[C@H](C(C)C)NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CC(C)C)NC(=O)CNC(=O)[C@H](C)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](NC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)CCSC)C(C)C)[C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](C)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(=O)N1[C@@H](CCC1)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(C)C)C(O)=O)C1=CC=C(O)C=C1 ZRKFYGHZFMAOKI-QMGMOQQFSA-N 0.000 description 1
- 229960000984 tocofersolan Drugs 0.000 description 1
- 230000005945 translocation Effects 0.000 description 1
- RTKIYFITIVXBLE-QEQCGCAPSA-N trichostatin A Chemical compound ONC(=O)/C=C/C(/C)=C/[C@@H](C)C(=O)C1=CC=C(N(C)C)C=C1 RTKIYFITIVXBLE-QEQCGCAPSA-N 0.000 description 1
- 208000001072 type 2 diabetes mellitus Diseases 0.000 description 1
- MSRILKIQRXUYCT-UHFFFAOYSA-M valproate semisodium Chemical compound [Na+].CCCC(C(O)=O)CCC.CCCC(C([O-])=O)CCC MSRILKIQRXUYCT-UHFFFAOYSA-M 0.000 description 1
- 229960000604 valproic acid Drugs 0.000 description 1
- NLVXSWCKKBEXTG-UHFFFAOYSA-N vinylsulfonic acid Chemical compound OS(=O)(=O)C=C NLVXSWCKKBEXTG-UHFFFAOYSA-N 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- UGZADUVQMDAIAO-UHFFFAOYSA-L zinc hydroxide Chemical compound [OH-].[OH-].[Zn+2] UGZADUVQMDAIAO-UHFFFAOYSA-L 0.000 description 1
- 229940007718 zinc hydroxide Drugs 0.000 description 1
- 229910021511 zinc hydroxide Inorganic materials 0.000 description 1
- 239000002076 α-tocopherol Substances 0.000 description 1
- 235000004835 α-tocopherol Nutrition 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0652—Cells of skeletal and connective tissues; Mesenchyme
- C12N5/0662—Stem cells
- C12N5/0667—Adipose-derived stem cells [ADSC]; Adipose stromal stem cells
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0696—Artificially induced pluripotent stem cells, e.g. iPS
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0603—Embryonic cells ; Embryoid bodies
- C12N5/0606—Pluripotent embryonic cells, e.g. embryonic stem cells [ES]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0634—Cells from the blood or the immune system
- C12N5/0647—Haematopoietic stem cells; Uncommitted or multipotent progenitors
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0652—Cells of skeletal and connective tissues; Mesenchyme
- C12N5/0662—Stem cells
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/10—Cells modified by introduction of foreign genetic material
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2500/00—Specific components of cell culture medium
- C12N2500/05—Inorganic components
- C12N2500/10—Metals; Metal chelators
- C12N2500/20—Transition metals
- C12N2500/24—Iron; Fe chelators; Transferrin
- C12N2500/25—Insulin-transferrin; Insulin-transferrin-selenium
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2500/00—Specific components of cell culture medium
- C12N2500/30—Organic components
- C12N2500/40—Nucleotides, nucleosides or bases
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/11—Epidermal growth factor [EGF]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/115—Basic fibroblast growth factor (bFGF, FGF-2)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/12—Hepatocyte growth factor [HGF]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/135—Platelet-derived growth factor [PDGF]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/15—Transforming growth factor beta (TGF-β)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2533/00—Supports or coatings for cell culture, characterised by material
- C12N2533/50—Proteins
- C12N2533/52—Fibronectin; Laminin
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2533/00—Supports or coatings for cell culture, characterised by material
- C12N2533/90—Substrates of biological origin, e.g. extracellular matrix, decellularised tissue
Definitions
- the present invention relates to a material capable of preventing the occurrence of chromosomal abnormalities in the process of culturing and subculturing stem cells, and a chromosomal stabilizer for stem cells using the material.
- Stem cells represented by pluripotent stem cells are undifferentiated cells having self-renewal ability and capable of differentiating into various cells.
- regenerative medicine has been actively studied in which stem cells and cells induced to differentiate from stem cells are transplanted into damaged tissues of patients to regenerate their functions.
- it is necessary to prepare a large amount of stem cells and their differentiated cells, and therefore, a method for efficiently proliferating stem cells and a method for efficiently differentiating stem cells are being actively developed.
- stem cells may have chromosomal abnormalities if the culture period is long or the number of passages is large.
- Chromosomal abnormalities include aneuploidy abnormalities such as monosomy in which two pairs of chromosomes become one and trisomy in which three chromosomes become one, as well as structures such as translocation, inversion, partial duplication, and partial deletion. There is an abnormality.
- a chromosomal abnormality occurs in a stem cell, not only the proliferative ability and differentiation ability held by the stem cell are lost, but also when the cell or tissue differentiated from the stem cell is used for regenerative medicine or the like, it is mutated into a cancer cell. Or there is a risk of developing a tumor.
- Chromosomal abnormalities may occur in cancer cells and tumors, and in recent years, methods for more accurately detecting chromosomal abnormalities have been reported (see, for example, Patent Documents 1 and 2).
- NMN nicotinamide mononucleotide
- NAD + nicotinamide mononucleotide
- the present invention relates to a material capable of preventing the occurrence of chromosomal abnormalities in the process of culturing and subculturing stem cells, a stem cell chromosomal stabilizer using the material, a stem cell culturing method, and a stem cell chromosomal stabilizing method.
- the purpose is to provide.
- the present invention provides the following stem cell chromosome stabilizer, stem cell culture method, and stem cell chromosome stabilization method.
- a chromosomal stabilizer for stem cells which comprises ⁇ -nicotinamide mononucleotide, a pharmacologically acceptable salt thereof, or a solvate thereof as an active ingredient.
- the chromosomal stabilizer according to the above [1] which is added to a stem cell culture medium at 0.01 to 5 mM in terms of ⁇ -nicotinamide mononucleotide.
- Culture of stem cells which comprises culturing pluripotent stem cells in a culture medium containing ⁇ -nicotinamide mononucleotide, a pharmacologically acceptable salt thereof, or a solvate thereof.
- Method. [5] The culture method according to [4] above, wherein the ⁇ -nicotinamide mononucleotide concentration in the culture medium is 0.01 to 5 mM.
- stem cells are one or more selected from the group consisting of embryonic stem cells, induced pluripotent stem cells, mesenchymal stem cells, and hematopoietic stem cells.
- a method for stabilizing stem cells which comprises culturing stem cells in a culture medium containing ⁇ -nicotinamide mononucleotide, a pharmacologically acceptable salt thereof, or a solvate thereof. ..
- the stem cell chromosomal stabilizer according to the present invention can improve the chromosomal stability of the stem cell and suppress or prevent the occurrence of chromosomal abnormalities by acting on the stem cell. Therefore, by containing the chromosomal stabilizer in the culture medium, the chromosomal stability of the stem cells can be improved, the occurrence of chromosomal abnormalities can be suppressed or prevented, and the stem cells can be cultured more stably. In addition, by using the chromosome stabilizer, the risk of canceration / tumorigenesis of cells or tissues differentiated from stem cells can be reduced.
- (A) is a fluorescence photograph of cells stained with PI in Example 2, and (b) is an anti- ⁇ H2A. It is a fluorescent photograph immunostained with an X antibody, and (c) is a photograph in which (a) and (b) are merged. ⁇ H2A. In all PI-stained cells. It is a graph which shows the ratio of the cell nucleus of X positive.
- stem cells are undifferentiated cells having self-renewal ability and differentiation ability (ability to differentiate into various cell types), for example, ES cells (embryonic stem cells).
- pluripotent stem cells such as iPS cells (induced pluripotent stem cells)
- somatic stem cells such as mesenchymal stem cells, hematopoietic stem cells, nerve stem cells, and skin stem cells can be mentioned.
- the stem cell chromosomal stabilizer according to the present invention (hereinafter, may be referred to as “chromosome stabilizer according to the present invention”) contains NMN (chemical formula: C 11 H 15 N 2 O 8 P) as an active ingredient. It is added to the culture medium when stem cells are cultured and subcultured. By culturing and subculturing stem cells in the presence of NMN, the stability of the stem cells' chromosomes can be improved, and the occurrence of chromosomal abnormalities can be suppressed or prevented.
- NMN chemical formula: C 11 H 15 N 2 O 8 P
- NMN optical isomers
- ⁇ and ⁇ the NMN that is the active ingredient of the chromosomal stabilizer according to the present invention
- ⁇ -NMN CAS number: 1094-61-7
- the structure of ⁇ -NMN is shown below.
- the ⁇ -NMN as the active ingredient may be prepared by any method.
- ⁇ -NMN that has been artificially synthesized by a chemical synthesis method, an enzyme method, a fermentation method, or the like can be used as an active ingredient.
- ⁇ -NMN is a component widely present in a living body
- ⁇ -NMN obtained by extracting and purifying from natural raw materials such as animals, plants and microorganisms can also be used as an active ingredient.
- commercially available purified ⁇ -NMN may be used.
- ⁇ -NMN can be produced by reacting NAM with L-ribose tetraacetate and phosphorylating the obtained nicotinamide mononucleoside.
- ⁇ -NMN can be produced from NAM and 5'-phosphoribosyl-1'-pyrrolic acid (PRPP) by nicotinamide phosphoribosyl transferase (NAMPT).
- PRPP 5'-phosphoribosyl-1'-pyrrolic acid
- NAMPT nicotinamide phosphoribosyl transferase
- ⁇ -NMN can be produced from NAM by utilizing the metabolic system of a microorganism expressing NAMPT.
- the active ingredient of the chromosomal stabilizer according to the present invention may be a pharmacologically acceptable salt of ⁇ -NMN.
- the pharmacologically acceptable salt of ⁇ -NMN may be an inorganic acid salt or an organic acid salt having a basic moiety such as amine.
- the acids constituting such acid salts include acetic acid, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, citric acid, ethenesulfonic acid, fumaric acid, gluconic acid, glutamic acid, hydrobromic acid, hydrochloric acid and ISEthione.
- the pharmacologically acceptable salt of ⁇ -NMN may be an alkaline salt or an organic salt having an acidic moiety such as a carboxylic acid.
- Examples of the base constituting such an acid salt are alkali metal salts or alkaline earth metal salts, such as sodium hydride, potassium hydroxide, calcium hydroxide, aluminum hydroxide, lithium hydroxide, and magnesium hydroxide.
- Zinc hydroxide ammonia, trimethylamonia, triethylammonia, ethylenediamine, lysine, arginine, ornithine, choline, N, N'-dibenzylethylenediamine, chloroprocine, procaine, diethanolamine, N-benzylphenethylamine, diethylamine, piperazin, tris ( Those derived from bases such as hydroxymethyl) -aminomethane and tetramethylammonium hydroxide can be mentioned.
- the active ingredient of the chromosomal stabilizer according to the present invention may be a solvate of a free ⁇ -NMN or a pharmacologically acceptable salt of ⁇ -NMN.
- the solvent for forming the solvate include water, ethanol and the like.
- the chromosomal stabilizer according to the present invention may contain other active ingredients in addition to ⁇ -NMN.
- the other active ingredient used in combination with ⁇ -NMN may be only one kind or a combination of two or more kinds.
- Other active ingredients are known to enhance the survival efficiency and proliferation efficiency of stem cells such as albumin, ascorbic acid, ⁇ -tocopherol, insulin, transferase, sodium selenate, ethanolamine, and Rock inhibitor.
- Ingredients such as valproic acid, dimethylsulfoxide, dexamethasone, butyric acid, tricostatin A, GSK3 inhibitor, BMP inhibitor, Wnt inhibitor, activin, nogin and other components known to enhance the differentiation efficiency of stem cells. It can be appropriately selected from the above and used.
- the culture medium contains the chromosomal stabilizer according to the present invention to improve the chromosomal stability of the stem cells and to proliferate the stem cells while suppressing or preventing the occurrence of chromosomal abnormalities. Can be done.
- the amount of the chromosomal stabilizer according to the present invention contained in the stem cell culture medium is lower than that in the case of culturing in the culture medium not containing the chromosomal stabilizer.
- the amount is not particularly limited as long as the concentration is sufficient to suppress the suppression, and can be appropriately adjusted in consideration of the type of stem cells, the balance with other components of the culture medium, and the like.
- the chromosomal stabilizing effect may be weak, and if an excessive amount of ⁇ -NMN is contained, cell proliferation may be suppressed.
- the content of the chromosomal stabilizer according to the present invention in the culture medium is preferably such that the ⁇ -NMN concentration is 0.01 to 5 mM, and more preferably 0.05 to 2 mM. , 0.1 to 1 mM, more preferably.
- the ⁇ -NMN concentration is within the above range, the chromosome of the stem cell can be sufficiently stabilized.
- Stem cell culture in the presence of the chromosomal stabilizer according to the present invention can be carried out by a conventional method except that the culture medium contains the chromosomal stabilizer according to the present invention.
- the culture medium a medium used for maintaining or proliferating stem cells or a medium used for culturing animal cells can be generally used.
- commercially available culture media for various stem cells can also be used.
- examples of the medium containing the chromosomal stabilizer according to the present invention and used for culturing stem cells include Eagle's minimum essential medium (MEM), Dalbeco's modified Eagle's medium (DMEM), and ⁇ Eagle's minimum essential medium.
- MEM Eagle's minimum essential medium
- DMEM Dalbeco's modified Eagle's medium
- ⁇ Eagle's minimum essential medium ⁇ Eagle's minimum essential medium.
- ⁇ MEM Iscover modified Dalveco medium
- IMDM Iscover modified Dalveco medium
- F-12 medium F-10 medium
- DMEM / F12 medium RPMI-1640 medium
- MSCBM Membrane cell basal medium
- E8 (Essential 8) medium E8 (Essential 8) medium
- TeSR -E8 medium mTeSR1 medium
- MCDB medium MCDB medium and the like
- these culture media may have components known to enhance the survival efficiency and proliferation efficiency of stem cells, and have an action of maintaining an undifferentiated state of stem cells.
- Known components and the like may be appropriately contained. As these components, the above-mentioned ones can be used.
- the culture conditions can generally be the culture conditions for culturing animal cells, and may be appropriately modified as necessary.
- the culture can be performed at a culture temperature of 30 to 40 ° C., a CO 2 concentration of 1 to 10% by volume, and an O 2 concentration of 0.1 to 25% by volume.
- stem cells whose chromosomes are stabilized by the chromosome stabilizer according to the present invention animal-derived stem cells are preferable, mammalian-derived stem cells are more preferable, and human-derived stem cells are even more preferable.
- the stem cells whose chromosomes are stabilized by the chromosomal stabilizer according to the present invention are preferably animal-derived ES cells, iPS cells, or mesenchymal stem cells, and are mammalian-derived ES cells and iPS cells. , Or mesenchymal stem cells, more preferably human-derived ES cells, iPS cells, or mesenchymal stem cells.
- Example 1 The effect of ⁇ -NMN on chromosomes during culture of mesenchymal stem cells was investigated.
- mesenchymal stem cells two donors of human adipose tissue-derived mesenchymal stem cells (ADSC) (product number: PT-5006, manufactured by Lonza) were used.
- ADSC human adipose tissue-derived mesenchymal stem cells
- a medium obtained by mixing DMEM and MCDB medium at a ratio of 1: 1 is used as a basal medium (see Patent No. 5804385), and FGF, PDGF, TGF- ⁇ , HGF, etc. are used in the basal medium.
- the medium to which EGF, phospholipids, fatty acids and the like were added was used as the basal medium.
- the ⁇ -NMN-added medium was prepared by adding ⁇ -NMN to the basal medium so as to have a concentration of 0.25 mM.
- ⁇ Subculture> Mesenchymal stem cells were seeded at 5 ⁇ 10 3 cells / cm 2 in a culture dish previously coated with 2.5 ⁇ g / cm 2 fibronectin, and cultured using a basal medium. One day after sowing, the medium was replaced with basal medium or ⁇ -NMN-added medium. After that, the medium was changed once every 2 or 3 days. Subculture was performed when the cell density in the culture dish reached a confluency of about 80 to 90%.
- the culture supernatant was removed, washed with 1 ⁇ PBS (-), and then the cells were detached using 1 ⁇ Triple select (product number: 12563011, manufactured by Thermo Fisher) to produce 37 ° C. and 5% CO 2 . It was allowed to stand for 4 minutes under the conditions. After the cells were singled by pipetting, basal medium or ⁇ -NMN-added medium was added, and the cells were centrifuged. After suspending the collected cells in each medium, cell counting was performed, and the cells were seeded in a pre-coated culture dish so as to have a concentration of 5 ⁇ 10 3 cells / cm 2. Medium exchange was performed once every 2 or 3 days using basal medium or ⁇ -NMN-added medium.
- Frozen cells were seeded in a coated culture dish at 5 ⁇ 10 3 cells / cm 2 in basal medium or ⁇ -NMN-added medium, and the medium was replaced 1 day after seeding. After that, the medium was changed once every two or three days.
- Subculture was performed once and seeded in a T25 flask at 5 ⁇ 10 3 cells / cm 2. Three days after seeding, the T25 flask was filled with a medium and sent to a chromosomal safety test consignment organization (Japan Genetic Research Institute Co., Ltd.) for chromosome analysis.
- ⁇ Chromosome stability test> The chromosomal status of the mesenchymal stem cells after culturing was confirmed by the G banding method.
- Table 1 shows the measurement results of the number of chromosomes
- Table 2 shows the results of karyotype analysis. The analysis results are described in accordance with the International System of Human Cytogenomic Nomenclature (ISCN) based on the International Code.
- ISCN International System of Human Cytogenomic Nomenclature
- Test group 1 shows mesenchymal stem cells (ADSC donor 1, male) cultured in the basal medium
- test group 2 shows mesenchymal stem cells (ADSC donor 2, female) cultured in the basal medium
- Test Group 3 contains mesenchymal stem cells (ADSC donor 1, male) cultured in ⁇ -NMN-added medium
- Test Group 4 contains mesenchymal stem cells (ADSC donor 2, female) cultured in ⁇ -NMN-added medium. show.
- test groups 1 and 2 In the mesenchymal stem cells cultured in the basal medium, abnormal numbers of chromosomes were observed in both donors (test groups 1 and 2) (Table 1). That is, as a result of measuring the number of chromosomes of 50 cells in each test group, in test group 1, the number of chromosomes increased by 1 from 46 to 47 in 4 cells. Also, in Test Group 2, the number of chromosomes decreased by 1 from 46 to 45 in one cell. On the other hand, in the mesenchymal stem cells cultured in the ⁇ -NMN-added medium, the number of chromosomes was 46 in all the cells of both donors (test groups 3 and 4), which was the normal number of human chromosomes.
- ⁇ -NMN human pluripotent stem cells
- iPSCs human pluripotent stem cells
- 201B7 strain was used, and 5% until the cell density reached about 80% on a 24-well cell culture plate pre-coated with extracellular matrix (Matrigel: Corning). Those cultured at CO 2 and 37 ° C. were subjected to the test.
- cytokines and salts (insulin, transferase, TGF ⁇ , FGF, sodium selenate, ascorbic acid, hydrogen carbonate) known to be necessary for maintenance subculture of iPSC in the basal medium (DMEM-F12)
- NMN (-) A ⁇ -NMN-free group (NMN (-)) in which culture is performed in the basal medium using a basal medium containing sodium) and a ⁇ -NMN-added group in which ⁇ -NMN is added to the basal medium so as to have a final concentration of 1 mM. (NMN (+)) was provided.
- NCX4016 (Sigma-Aldrich, product number SML1669) was added to each of the ⁇ -NMN-free group and the ⁇ -NMN-added group to a final concentration of 5 ⁇ M, and the mixture was incubated for 2 hours to induce chromosomal damage. The cells with damaged nuclear genomic DNA were then subjected to phosphorylated histone H2A. It was detected by labeling with X ( ⁇ H2AX).
- ⁇ H2A The number of X-stained cell nuclei (indicated by circles) was significantly smaller in the ⁇ -NMN-added group (NMN (+)) than in the ⁇ -NMN-free group (NMN ( ⁇ )).
- ⁇ H2A In whole cell nuclei stained with propidium iodide (PI). The proportion of X-positive cell nuclei was statistically significantly lower in the ⁇ -NMN-added group (NMN (+)) than in the ⁇ -NMN-free group (NMN (-)) (test of difference in population ratio, p. ⁇ 0.05, n> 8000).
- PI propidium iodide
- the bar graph shows ⁇ H2A.
- the ratio of X-positive cell nuclei is shown, and the error bars indicate the range of the standard error. This result indicates that in stem cells, the addition of ⁇ -NMN alleviates damage to genomic DNA that causes chromosomal abnormalities.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Genetics & Genomics (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Developmental Biology & Embryology (AREA)
- Cell Biology (AREA)
- Transplantation (AREA)
- Reproductive Health (AREA)
- Gynecology & Obstetrics (AREA)
- Hematology (AREA)
- Rheumatology (AREA)
- Immunology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Molecular Biology (AREA)
- Biophysics (AREA)
- Analytical Chemistry (AREA)
- Physics & Mathematics (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
Description
[1] β-ニコチンアミドモノヌクレオチド若しくはその薬理学的に許容される塩、又はそれらの溶媒和物を有効成分とすることを特徴とする、幹細胞の染色体安定化剤。
[2] 幹細胞の培養培地に、β-ニコチンアミドモノヌクレオチド換算で0.01~5mM添加される、前記[1]の染色体安定化剤。
[3] 胚性幹細胞、人工多能性幹細胞、間葉系幹細胞、及び造血幹細胞からなる群より選択される一種以上の幹細胞の染色体安定化のために用いられる、前記[1]又は[2]の染色体安定化剤。
[4] 多能性幹細胞を、β-ニコチンアミドモノヌクレオチド若しくはその薬理学的に許容される塩、又はそれらの溶媒和物を含有する培養培地中で培養することを特徴とする、幹細胞の培養方法。
[5] 前記培養培地のβ-ニコチンアミドモノヌクレオチド濃度が0.01~5mMである、前記[4]の培養方法。
[6] 前記幹細胞が、胚性幹細胞、人工多能性幹細胞、間葉系幹細胞、及び造血幹細胞からなる群より選択される一種以上である、前記[4]又は[5]の培養方法。
[7] 幹細胞を、β-ニコチンアミドモノヌクレオチド若しくはその薬理学的に許容される塩、又はそれらの溶媒和物を含有する培養培地中で培養することを特徴とする、幹細胞の染色体安定化方法。
間葉系幹細胞の培養時における、染色体に対するβ-NMNの効果を調べた。間葉系幹細胞としては、ヒト脂肪組織由来間葉系幹細胞(ADSC)(製品番号:PT-5006、Lonza社製)の2ドナーを用いた。
予め2.5μg/cm2フィブロネクチンでコーティングした培養用ディッシュに、間葉系幹細胞を5×103cells/cm2となるように播種し、基礎培地を用いて培養した。播種1日後に、基本培地又はβ-NMN添加培地に培地交換した。以降、2又は3日ごとに一回培地交換を実施した。培養用ディッシュ内の細胞密度が約80~90%のコンフルエンシーになった時点で、継代培養を行った。
培養後の間葉系幹細胞について、Gバンド分染法により、染色体の状態を確認した。染色体本数の測定結果を表1、核型分析結果を表2に示す。なお、分析結果は、国際規約に基づく染色体核型記載法(ISCN:International System for Human Cytogenomic Nomenclature)に順じて記載した。
[実施例2]
<培地・培養>
ヒト多分化能性幹細胞(iPSC)としては201B7株を用い、細胞外マトリックス(マトリゲル:コーニング社)で事前にコーティングした24穴の細胞培養用プレートにて80%程度の細胞密度となるまで5%CO2、37℃で培養したものを試験に供した。
NCX4016添加下でインキュベートされた細胞を0.99%ホルムアルデヒドで固定し、0.10%TritonX-100で透過処理をしたのち、抗γH2A.X抗体(サーモフィッシャー社、製品番号14-9865-82)とFITCで蛍光標識された抗マウスIgG抗体(サーモフィッシャー社、製品番号11-4015-82)を用いて免疫染色し、蛍光顕微鏡下で観察することにより、γH2A.Xを検出した。また、同時にすべての細胞の核を観察できるように、ヨウ化プロピジウム(ナカライテスク社、製品番号19174-31)によってDNAを蛍光染色した。
図1(b)に示すように、γH2A.Xで染色された細胞核(丸印で示す)は、β-NMN無添加区(NMN(-))より、β-NMN添加区(NMN(+))で顕著に少なかった。また、図2に示すように、ヨウ化プロピジウム(PI)で染色された全細胞核中のγH2A.X陽性の細胞核の比率は、β-NMN無添加区(NMN(-))より、β-NMN添加区(NMN(+))で統計的に有意に少なかった(母比率の差の検定、p<0.05、n>8000)。図2において、棒グラフは、それぞれの区のγH2A.X陽性の細胞核の比率を示し、エラーバーはその標準誤差の幅を表している。この結果は、幹細胞において、β-NMNの添加により染色体異常の原因となるゲノムDNAへの損傷が緩和されることを示している。
Claims (7)
- β-ニコチンアミドモノヌクレオチド若しくはその薬理学的に許容される塩、又はそれらの溶媒和物を有効成分とすることを特徴とする、幹細胞の染色体安定化剤。
- 幹細胞の培養培地に、β-ニコチンアミドモノヌクレオチド換算で0.01~5mM添加される、請求項1に記載の染色体安定化剤。
- 胚性幹細胞、人工多能性幹細胞、間葉系幹細胞、及び造血幹細胞からなる群より選択される一種以上の幹細胞の染色体安定化のために用いられる、請求項1又は2に記載の染色体安定化剤。
- 多能性幹細胞を、β-ニコチンアミドモノヌクレオチド若しくはその薬理学的に許容される塩、又はそれらの溶媒和物を含有する培養培地中で培養することを特徴とする、幹細胞の培養方法。
- 前記培養培地のβ-ニコチンアミドモノヌクレオチド濃度が0.01~5mMである、請求項4に記載の培養方法。
- 前記幹細胞が、胚性幹細胞、人工多能性幹細胞、間葉系幹細胞、及び造血幹細胞からなる群より選択される一種以上である、請求項4又は5に記載の培養方法。
- 幹細胞を、β-ニコチンアミドモノヌクレオチド若しくはその薬理学的に許容される塩、又はそれらの溶媒和物を含有する培養培地中で培養することを特徴とする、幹細胞の染色体安定化方法。
Priority Applications (7)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202180022602.5A CN115315507A (zh) | 2020-03-30 | 2021-03-25 | 干细胞的染色体稳定剂 |
IL296817A IL296817A (en) | 2020-03-30 | 2021-03-25 | Chromosome stabilizing agent for stem cells |
KR1020227036502A KR20220160611A (ko) | 2020-03-30 | 2021-03-25 | 줄기세포의 염색체 안정화제 |
JP2022512069A JPWO2021200545A1 (ja) | 2020-03-30 | 2021-03-25 | |
EP21780670.2A EP4130250A4 (en) | 2020-03-30 | 2021-03-25 | STABILIZER FOR STEM CELL CHROMOSOMES |
US17/907,491 US20230117880A1 (en) | 2020-03-30 | 2021-03-25 | Chromosome-stabilizing agent for stem cells |
AU2021247767A AU2021247767A1 (en) | 2020-03-30 | 2021-03-25 | Chromosome-stabilizing agent for stem cells |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP2020-060490 | 2020-03-30 | ||
JP2020060490 | 2020-03-30 |
Publications (1)
Publication Number | Publication Date |
---|---|
WO2021200545A1 true WO2021200545A1 (ja) | 2021-10-07 |
Family
ID=77927457
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/JP2021/012557 WO2021200545A1 (ja) | 2020-03-30 | 2021-03-25 | 幹細胞の染色体安定化剤 |
Country Status (8)
Country | Link |
---|---|
US (1) | US20230117880A1 (ja) |
EP (1) | EP4130250A4 (ja) |
JP (1) | JPWO2021200545A1 (ja) |
KR (1) | KR20220160611A (ja) |
CN (1) | CN115315507A (ja) |
AU (1) | AU2021247767A1 (ja) |
IL (1) | IL296817A (ja) |
WO (1) | WO2021200545A1 (ja) |
Citations (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US7737158B2 (en) | 2005-10-11 | 2010-06-15 | Washington University | Processes for regulating blood glucose in a mammal |
JP5804385B2 (ja) | 2010-03-10 | 2015-11-04 | 株式会社ツーセル | 間葉系幹細胞を含む細胞製剤及びその製造方法 |
WO2018143258A1 (ja) | 2017-01-31 | 2018-08-09 | オリエンタル酵母工業株式会社 | 多分化能性幹細胞増殖促進剤 |
JP2019144097A (ja) | 2018-02-20 | 2019-08-29 | 国立研究開発法人産業技術総合研究所 | クロマチンの異常凝縮の検出方法 |
WO2019182044A1 (ja) | 2018-03-22 | 2019-09-26 | オリエンタル酵母工業株式会社 | 多分化能性幹細胞分化促進剤 |
JP2019213537A (ja) | 2015-06-23 | 2019-12-19 | ザイトヴィジョン・ゲゼルシャフト・ミット・ベシュレンクテル・ハフツング | 染色体異常の検出方法 |
Family Cites Families (14)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
EA201490050A1 (ru) * | 2011-06-29 | 2014-07-30 | Зе Дженерэл Хоспитэл Корпорейшн | Композиции и способы повышения биоэнергетического состояния женских зародышевых клеток |
WO2013180395A1 (ko) * | 2012-05-29 | 2013-12-05 | 한국생명공학연구원 | 다능성 줄기세포의 제작, 유지, 증식을 증진하는 대사산물 및 이를 포함하는 조성물과 배양방법 |
WO2014059031A2 (en) * | 2012-10-09 | 2014-04-17 | President And Fellows Of Harvard College | Nad biosynthesis and precursors in the prevention and treatment of inflammation |
WO2014059034A2 (en) * | 2012-10-09 | 2014-04-17 | President And Fellows Of Harvard College | Nad biosynthesis and precursors for the treatment and prevention of cancer and proliferation |
WO2015002892A1 (en) * | 2013-07-01 | 2015-01-08 | Meier Joshua Abraham | Methods of treating diseases by modulating mitochondrial dna deletions |
KR101655383B1 (ko) * | 2013-07-27 | 2016-09-08 | 고려대학교 산학협력단 | 소분자 화합물을 포함하는 만능성 줄기세포의 염색체 안정성 유지용 조성물 |
WO2016149672A1 (en) * | 2015-03-18 | 2016-09-22 | The Regents Of The University Of California | Methods of preventing and reversing stem cell aging |
DK3331894T3 (da) * | 2015-08-05 | 2021-03-22 | Metro Int Biotech Llc | Nicotinamidmononukleotidderivater og anvendelser deraf |
GB2542881B (en) * | 2015-10-02 | 2020-01-01 | Carr Andrew | Crystal forms of ß-nicotinamide mononucleotide |
US20190350960A1 (en) * | 2017-01-04 | 2019-11-21 | President And Fellows Of Harvard College | Modulating nudix homology domain (nhd) with nicotinamide mononucleotide analogs and derivatives of same |
US11286274B2 (en) * | 2017-06-19 | 2022-03-29 | Mitopower Llc | Nicotinamide riboside derivatives and their uses |
EP3647411A4 (en) * | 2017-06-27 | 2021-05-12 | Ajinomoto Co., Inc. | MEDIUM WITH RIBOFLAVIN DERIVATIVE |
CN110996967A (zh) * | 2017-08-10 | 2020-04-10 | 华盛顿大学 | 使用烟酰胺单核苷酸进行治疗的组合物和方法 |
WO2019165372A1 (en) * | 2018-02-26 | 2019-08-29 | President And Fellows Of Harvard College | Compositions of parp14 modulators and/or mutants and therapeutic use thereof |
-
2021
- 2021-03-25 CN CN202180022602.5A patent/CN115315507A/zh active Pending
- 2021-03-25 US US17/907,491 patent/US20230117880A1/en active Pending
- 2021-03-25 AU AU2021247767A patent/AU2021247767A1/en active Pending
- 2021-03-25 IL IL296817A patent/IL296817A/en unknown
- 2021-03-25 KR KR1020227036502A patent/KR20220160611A/ko active Search and Examination
- 2021-03-25 WO PCT/JP2021/012557 patent/WO2021200545A1/ja unknown
- 2021-03-25 JP JP2022512069A patent/JPWO2021200545A1/ja active Pending
- 2021-03-25 EP EP21780670.2A patent/EP4130250A4/en active Pending
Patent Citations (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US7737158B2 (en) | 2005-10-11 | 2010-06-15 | Washington University | Processes for regulating blood glucose in a mammal |
JP5804385B2 (ja) | 2010-03-10 | 2015-11-04 | 株式会社ツーセル | 間葉系幹細胞を含む細胞製剤及びその製造方法 |
JP2019213537A (ja) | 2015-06-23 | 2019-12-19 | ザイトヴィジョン・ゲゼルシャフト・ミット・ベシュレンクテル・ハフツング | 染色体異常の検出方法 |
WO2018143258A1 (ja) | 2017-01-31 | 2018-08-09 | オリエンタル酵母工業株式会社 | 多分化能性幹細胞増殖促進剤 |
JP2019144097A (ja) | 2018-02-20 | 2019-08-29 | 国立研究開発法人産業技術総合研究所 | クロマチンの異常凝縮の検出方法 |
WO2019182044A1 (ja) | 2018-03-22 | 2019-09-26 | オリエンタル酵母工業株式会社 | 多分化能性幹細胞分化促進剤 |
Non-Patent Citations (5)
Title |
---|
FAKOURI NIMA B., HOU YUJUN, DEMAREST TYLER G., CHRISTIANSEN LOUISE S., OKUR MUSTAFA N., MOHANTY JOY G., CROTEAU DEBORAH L., BOHR V: "Toward understanding genomic instability, mitochondrial dysfunction and aging", THE FEBS JOURNAL, WILEY-BLACKWELL PUBLISHING LTD., GB, vol. 286, no. 6, 1 March 2019 (2019-03-01), GB , pages 1058 - 1073, XP055925933, ISSN: 1742-464X, DOI: 10.1111/febs.14663 * |
HIDENORI MATSUO, KOTA ONO, KAYA TOMORI, NAOTAKA YASUDA: "1P-0891 Effect of beta-Nicotinamide mononucleotide in human pluripotent stem cells", ONLINE ABSTRACTS OF CONSORTIUM OF BIOLOGICAL SCIENCES 2017 (CONBIO2017); 26TH FAOBMB CONFERENCE WITH CONBIO2017, KOBE, JAPAN, 6–9 DECEMBER 2017, 30 November 2016 (2016-11-30) - 9 December 2017 (2017-12-09), JP, pages 1P-0891, XP009536506 * |
KIMURA, NAOKO; ISHII, MIKA; ISHIZUKA, MISAKI; OHARA, TAIKI; FUJII, JUNICHI: "beta-NMN supplementation to IVM medium improves oocyte spindle shape and embryo developmental potential in aged mice", THE 109TH MEETING OF THE SOCIETY FOR REPRODUCTION AND DEVELOPMENT, 16 September 2016 (2016-09-16), pages 1 - 1, XP009540775, DOI: 10.14882/jrds.109.0_OR2-5 * |
See also references of EP4130250A4 |
WILK ANNA, HAYAT FAISAL, CUNNINGHAM RICHARD, LI JIANFENG, GARAVAGLIA SILVIA, ZAMANI LEILA, FERRARIS DAVIDE M., SYKORA PETER, ANDRE: "Extracellular NAD+ enhances PARP-dependent DNA repair capacity independently of CD73 activity", SCIENTIFIC REPORTS, vol. 10, no. 1, 1 December 2020 (2020-12-01), XP055925929, DOI: 10.1038/s41598-020-57506-9 * |
Also Published As
Publication number | Publication date |
---|---|
EP4130250A4 (en) | 2024-05-15 |
KR20220160611A (ko) | 2022-12-06 |
US20230117880A1 (en) | 2023-04-20 |
EP4130250A1 (en) | 2023-02-08 |
AU2021247767A1 (en) | 2022-10-20 |
JPWO2021200545A1 (ja) | 2021-10-07 |
IL296817A (en) | 2022-11-01 |
CN115315507A (zh) | 2022-11-08 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
JP6974360B2 (ja) | 多分化能性幹細胞増殖促進剤 | |
Bischoff et al. | Inhibition of myoblast fusion after one round of DNA synthesis in 5-bromodeoxyuridine | |
CN103857789B (zh) | 制备间充质干细胞基础培养基和利用间充质干细胞基础培养基制备细胞治疗产品的方法及用该培养基得到的分化产品 | |
Scharner et al. | The muscle satellite cell at 50: the formative years | |
US11814652B2 (en) | Pluripotent stem cell differentiation-promoting agent | |
Lee et al. | Chondrogenic potential and anti-senescence effect of hypoxia on canine adipose mesenchymal stem cells | |
Yonenaga et al. | Optimal conditions of collagenase treatment for isolation of articular chondrocytes from aged human tissues | |
Kim et al. | Characterization of human fetal cartilage progenitor cells during long-term expansion in a xeno-free medium | |
WO2019109666A1 (zh) | 一种尿液来源细胞的培养方法 | |
Rajput et al. | Expansion of human umbilical cord derived mesenchymal stem cells in regenerative medicine | |
WO2021200545A1 (ja) | 幹細胞の染色体安定化剤 | |
WO2021220731A1 (ja) | 幹細胞用培地及び幹細胞の培養方法 | |
Ariyoshi et al. | 3D spheroid culture models for chondrocytes using polyethylene glycol-coated microfabricated chip | |
US11248210B2 (en) | Method for isolation of stem cells from bone marrow using subfractionation culturing method and proliferation thereof | |
Islam et al. | Functional characterization of cell hybrids generated by induced fusion of primary porcine mesenchymal stem cells with an immortal murine cell line | |
JP2022007610A (ja) | 多能性幹細胞由来フィーダー細胞 | |
CN114807028B (zh) | 一种无血清间充质干细胞培养基及干细胞培养方法 | |
EP4293109A1 (en) | Non-skeletal muscle-derived cells as a source of suspension capable myogenic cells for cultured foods | |
Choi et al. | Effect of essential and nonessential amino acid compositions on the in vitro behavior of human mesenchymal stem cells | |
JP2024079373A (ja) | 細胞培養 | |
Romero Moya | Mitochondria and stem cell function: from somatic cells to iPSC-based disease modeling |
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: 21780670 Country of ref document: EP Kind code of ref document: A1 |
|
ENP | Entry into the national phase |
Ref document number: 2022512069 Country of ref document: JP Kind code of ref document: A |
|
ENP | Entry into the national phase |
Ref document number: 20227036502 Country of ref document: KR Kind code of ref document: A |
|
ENP | Entry into the national phase |
Ref document number: 2021247767 Country of ref document: AU Date of ref document: 20210325 Kind code of ref document: A |
|
NENP | Non-entry into the national phase |
Ref country code: DE |
|
ENP | Entry into the national phase |
Ref document number: 2021780670 Country of ref document: EP Effective date: 20221031 |