WO2016159721A1 - 중간엽 줄기세포에서 단백질의 대량 생산 방법 - Google Patents
중간엽 줄기세포에서 단백질의 대량 생산 방법 Download PDFInfo
- Publication number
- WO2016159721A1 WO2016159721A1 PCT/KR2016/003421 KR2016003421W WO2016159721A1 WO 2016159721 A1 WO2016159721 A1 WO 2016159721A1 KR 2016003421 W KR2016003421 W KR 2016003421W WO 2016159721 A1 WO2016159721 A1 WO 2016159721A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- mesenchymal stem
- stem cell
- cells
- culture
- stem cells
- Prior art date
Links
- 210000002901 mesenchymal stem cell Anatomy 0.000 title claims abstract description 212
- 108090000623 proteins and genes Proteins 0.000 title claims abstract description 125
- 102000004169 proteins and genes Human genes 0.000 title claims abstract description 124
- 238000000034 method Methods 0.000 title claims abstract description 51
- 230000003779 hair growth Effects 0.000 claims abstract description 47
- 238000004519 manufacturing process Methods 0.000 claims abstract description 37
- 239000000203 mixture Substances 0.000 claims abstract description 36
- 238000012258 culturing Methods 0.000 claims abstract description 26
- 230000036560 skin regeneration Effects 0.000 claims abstract description 26
- 239000002537 cosmetic Substances 0.000 claims abstract description 20
- 201000004384 Alopecia Diseases 0.000 claims abstract description 19
- 230000003676 hair loss Effects 0.000 claims abstract description 19
- 208000024963 hair loss Diseases 0.000 claims abstract description 19
- 238000011282 treatment Methods 0.000 claims abstract description 18
- 230000037303 wrinkles Effects 0.000 claims abstract description 18
- 239000008194 pharmaceutical composition Substances 0.000 claims abstract description 17
- 230000003658 preventing hair loss Effects 0.000 claims abstract description 12
- 238000004113 cell culture Methods 0.000 claims description 167
- 210000004027 cell Anatomy 0.000 claims description 149
- 210000000130 stem cell Anatomy 0.000 claims description 143
- 230000001737 promoting effect Effects 0.000 claims description 30
- 239000010931 gold Substances 0.000 claims description 20
- 229910052737 gold Inorganic materials 0.000 claims description 20
- 102000008186 Collagen Human genes 0.000 claims description 19
- 108010035532 Collagen Proteins 0.000 claims description 19
- 229920001436 collagen Polymers 0.000 claims description 19
- 210000004381 amniotic fluid Anatomy 0.000 claims description 16
- 210000001185 bone marrow Anatomy 0.000 claims description 13
- 210000004700 fetal blood Anatomy 0.000 claims description 12
- -1 TGFβ1 Proteins 0.000 claims description 11
- 210000001691 amnion Anatomy 0.000 claims description 11
- 210000002950 fibroblast Anatomy 0.000 claims description 10
- 230000006872 improvement Effects 0.000 claims description 10
- 239000012679 serum free medium Substances 0.000 claims description 10
- 102100040896 Growth/differentiation factor 15 Human genes 0.000 claims description 9
- 108090000379 Fibroblast growth factor 2 Proteins 0.000 claims description 8
- 102000004374 Insulin-like growth factor binding protein 3 Human genes 0.000 claims description 8
- 108090000965 Insulin-like growth factor binding protein 3 Proteins 0.000 claims description 8
- 102000004375 Insulin-like growth factor-binding protein 1 Human genes 0.000 claims description 8
- 108090000957 Insulin-like growth factor-binding protein 1 Proteins 0.000 claims description 8
- 102000004369 Insulin-like growth factor-binding protein 4 Human genes 0.000 claims description 8
- 108090000969 Insulin-like growth factor-binding protein 4 Proteins 0.000 claims description 8
- 102100020880 Kit ligand Human genes 0.000 claims description 8
- 102100028123 Macrophage colony-stimulating factor 1 Human genes 0.000 claims description 8
- 101710127797 Macrophage colony-stimulating factor 1 Proteins 0.000 claims description 8
- 230000037319 collagen production Effects 0.000 claims description 8
- 102000015271 Intercellular Adhesion Molecule-1 Human genes 0.000 claims description 7
- 108010064593 Intercellular Adhesion Molecule-1 Proteins 0.000 claims description 7
- 101710177504 Kit ligand Proteins 0.000 claims description 7
- 102100039364 Metalloproteinase inhibitor 1 Human genes 0.000 claims description 7
- 102100035194 Placenta growth factor Human genes 0.000 claims description 7
- 108090000381 Fibroblast growth factor 4 Proteins 0.000 claims description 6
- 102100028071 Fibroblast growth factor 7 Human genes 0.000 claims description 6
- 108090000385 Fibroblast growth factor 7 Proteins 0.000 claims description 6
- 102000004883 Insulin-like growth factor-binding protein 6 Human genes 0.000 claims description 6
- 108090001014 Insulin-like growth factor-binding protein 6 Proteins 0.000 claims description 6
- 108010025020 Nerve Growth Factor Proteins 0.000 claims description 6
- 108090000742 Neurotrophin 3 Proteins 0.000 claims description 6
- 102000003683 Neurotrophin-4 Human genes 0.000 claims description 6
- 108090000099 Neurotrophin-4 Proteins 0.000 claims 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 claims description 6
- 108010049976 Bone Morphogenetic Protein 5 Proteins 0.000 claims description 5
- 108010049870 Bone Morphogenetic Protein 7 Proteins 0.000 claims description 5
- 102100022526 Bone morphogenetic protein 5 Human genes 0.000 claims description 5
- 102100022544 Bone morphogenetic protein 7 Human genes 0.000 claims description 5
- 101710155857 C-C motif chemokine 2 Proteins 0.000 claims description 5
- 102100021943 C-C motif chemokine 2 Human genes 0.000 claims description 5
- 102100032367 C-C motif chemokine 5 Human genes 0.000 claims description 5
- 108010055166 Chemokine CCL5 Proteins 0.000 claims description 5
- 108010017080 Granulocyte Colony-Stimulating Factor Proteins 0.000 claims description 5
- 102000004372 Insulin-like growth factor binding protein 2 Human genes 0.000 claims description 5
- 108090000964 Insulin-like growth factor binding protein 2 Proteins 0.000 claims description 5
- 102000003815 Interleukin-11 Human genes 0.000 claims description 5
- 108090000177 Interleukin-11 Proteins 0.000 claims description 5
- 108010002616 Interleukin-5 Proteins 0.000 claims description 5
- 108090001005 Interleukin-6 Proteins 0.000 claims description 5
- 102000004889 Interleukin-6 Human genes 0.000 claims description 5
- 102100026262 Metalloproteinase inhibitor 2 Human genes 0.000 claims description 5
- 102100033733 Tumor necrosis factor receptor superfamily member 1B Human genes 0.000 claims description 5
- 101710187830 Tumor necrosis factor receptor superfamily member 1B Proteins 0.000 claims description 5
- 108010073919 Vascular Endothelial Growth Factor D Proteins 0.000 claims description 5
- 102100038234 Vascular endothelial growth factor D Human genes 0.000 claims description 5
- 108090000723 Insulin-Like Growth Factor I Proteins 0.000 claims description 4
- 102000006747 Transforming Growth Factor alpha Human genes 0.000 claims description 4
- 101800004564 Transforming growth factor alpha Proteins 0.000 claims description 4
- 108060008682 Tumor Necrosis Factor Proteins 0.000 claims description 4
- 108010049955 Bone Morphogenetic Protein 4 Proteins 0.000 claims description 3
- 102100024505 Bone morphogenetic protein 4 Human genes 0.000 claims description 3
- 102000004219 Brain-derived neurotrophic factor Human genes 0.000 claims description 3
- 108090000715 Brain-derived neurotrophic factor Proteins 0.000 claims description 3
- 102000034615 Glial cell line-derived neurotrophic factor Human genes 0.000 claims description 3
- 108091010837 Glial cell line-derived neurotrophic factor Proteins 0.000 claims description 3
- 108090001061 Insulin Proteins 0.000 claims description 3
- 102000004218 Insulin-Like Growth Factor I Human genes 0.000 claims description 3
- 108010002350 Interleukin-2 Proteins 0.000 claims description 3
- 108090001007 Interleukin-8 Proteins 0.000 claims description 3
- 102000004890 Interleukin-8 Human genes 0.000 claims description 3
- 102000000852 Tumor Necrosis Factor-alpha Human genes 0.000 claims description 3
- 229940125396 insulin Drugs 0.000 claims description 3
- 102100036170 C-X-C motif chemokine 9 Human genes 0.000 claims description 2
- 101710085500 C-X-C motif chemokine 9 Proteins 0.000 claims description 2
- 101000595923 Homo sapiens Placenta growth factor Proteins 0.000 claims description 2
- 102000000588 Interleukin-2 Human genes 0.000 claims description 2
- 102000005789 Vascular Endothelial Growth Factors Human genes 0.000 claims 3
- 108010019530 Vascular Endothelial Growth Factors Proteins 0.000 claims 3
- 102100024785 Fibroblast growth factor 2 Human genes 0.000 claims 2
- 102100028072 Fibroblast growth factor 4 Human genes 0.000 claims 1
- 102100039619 Granulocyte colony-stimulating factor Human genes 0.000 claims 1
- 108010041834 Growth Differentiation Factor 15 Proteins 0.000 claims 1
- 102100021866 Hepatocyte growth factor Human genes 0.000 claims 1
- 101000898034 Homo sapiens Hepatocyte growth factor Proteins 0.000 claims 1
- 101001076408 Homo sapiens Interleukin-6 Proteins 0.000 claims 1
- 101001001487 Homo sapiens Phosphatidylinositol-glycan biosynthesis class F protein Proteins 0.000 claims 1
- 101000868152 Homo sapiens Son of sevenless homolog 1 Proteins 0.000 claims 1
- 102100023915 Insulin Human genes 0.000 claims 1
- 108010031374 Tissue Inhibitor of Metalloproteinase-1 Proteins 0.000 claims 1
- 108010031372 Tissue Inhibitor of Metalloproteinase-2 Proteins 0.000 claims 1
- 108010017843 platelet-derived growth factor A Proteins 0.000 claims 1
- 239000001963 growth medium Substances 0.000 abstract description 52
- 239000003102 growth factor Substances 0.000 abstract description 49
- 102000004127 Cytokines Human genes 0.000 abstract description 20
- 108090000695 Cytokines Proteins 0.000 abstract description 20
- 238000011069 regeneration method Methods 0.000 abstract description 7
- 230000008929 regeneration Effects 0.000 abstract description 5
- 230000002500 effect on skin Effects 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 69
- 238000012360 testing method Methods 0.000 description 46
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 36
- 210000004209 hair Anatomy 0.000 description 31
- 239000000126 substance Substances 0.000 description 29
- 229940068196 placebo Drugs 0.000 description 27
- 239000000902 placebo Substances 0.000 description 27
- 230000000694 effects Effects 0.000 description 26
- 230000012010 growth Effects 0.000 description 26
- 238000003860 storage Methods 0.000 description 26
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 25
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 22
- 239000002609 medium Substances 0.000 description 21
- 239000000523 sample Substances 0.000 description 21
- 238000005138 cryopreservation Methods 0.000 description 20
- 239000013641 positive control Substances 0.000 description 20
- 239000012153 distilled water Substances 0.000 description 19
- 229960003632 minoxidil Drugs 0.000 description 19
- 210000002966 serum Anatomy 0.000 description 19
- 239000006143 cell culture medium Substances 0.000 description 18
- ZFMITUMMTDLWHR-UHFFFAOYSA-N Minoxidil Chemical compound NC1=[N+]([O-])C(N)=CC(N2CCCCC2)=N1 ZFMITUMMTDLWHR-UHFFFAOYSA-N 0.000 description 17
- 230000029663 wound healing Effects 0.000 description 16
- 210000003780 hair follicle Anatomy 0.000 description 15
- 238000011081 inoculation Methods 0.000 description 15
- 239000012091 fetal bovine serum Substances 0.000 description 14
- 230000001965 increasing effect Effects 0.000 description 14
- 241001465754 Metazoa Species 0.000 description 13
- 239000007788 liquid Substances 0.000 description 13
- 210000003491 skin Anatomy 0.000 description 13
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 12
- 210000001519 tissue Anatomy 0.000 description 12
- 241000699666 Mus <mouse, genus> Species 0.000 description 11
- 229910052757 nitrogen Inorganic materials 0.000 description 11
- 230000008569 process Effects 0.000 description 11
- OIRDTQYFTABQOQ-KQYNXXCUSA-N adenosine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O OIRDTQYFTABQOQ-KQYNXXCUSA-N 0.000 description 10
- 241000699670 Mus sp. Species 0.000 description 9
- 108010073929 Vascular Endothelial Growth Factor A Proteins 0.000 description 9
- 102100039037 Vascular endothelial growth factor A Human genes 0.000 description 9
- 230000004663 cell proliferation Effects 0.000 description 9
- 238000000338 in vitro Methods 0.000 description 9
- 238000011534 incubation Methods 0.000 description 9
- 238000005259 measurement Methods 0.000 description 9
- 230000004083 survival effect Effects 0.000 description 9
- 102100032236 Tumor necrosis factor receptor superfamily member 11B Human genes 0.000 description 8
- 230000015572 biosynthetic process Effects 0.000 description 8
- 230000003833 cell viability Effects 0.000 description 8
- 238000007796 conventional method Methods 0.000 description 8
- 230000005764 inhibitory process Effects 0.000 description 8
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 8
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 7
- 210000000988 bone and bone Anatomy 0.000 description 7
- 230000012292 cell migration Effects 0.000 description 7
- 201000010099 disease Diseases 0.000 description 7
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 7
- 239000003814 drug Substances 0.000 description 7
- 238000007710 freezing Methods 0.000 description 7
- 230000008014 freezing Effects 0.000 description 7
- 230000006870 function Effects 0.000 description 7
- 230000031774 hair cycle Effects 0.000 description 7
- 230000002401 inhibitory effect Effects 0.000 description 7
- 239000000546 pharmaceutical excipient Substances 0.000 description 7
- 238000011002 quantification Methods 0.000 description 7
- 102000003974 Fibroblast growth factor 2 Human genes 0.000 description 6
- 101710194460 Growth/differentiation factor 15 Proteins 0.000 description 6
- 108090000100 Hepatocyte Growth Factor Proteins 0.000 description 6
- 102000003745 Hepatocyte Growth Factor Human genes 0.000 description 6
- 101000669513 Homo sapiens Metalloproteinase inhibitor 1 Proteins 0.000 description 6
- XUMBMVFBXHLACL-UHFFFAOYSA-N Melanin Chemical compound O=C1C(=O)C(C2=CNC3=C(C(C(=O)C4=C32)=O)C)=C2C4=CNC2=C1C XUMBMVFBXHLACL-UHFFFAOYSA-N 0.000 description 6
- 108010035042 Osteoprotegerin Proteins 0.000 description 6
- 102100040247 Tumor necrosis factor Human genes 0.000 description 6
- 230000024245 cell differentiation Effects 0.000 description 6
- 230000003412 degenerative effect Effects 0.000 description 6
- 238000010790 dilution Methods 0.000 description 6
- 239000012895 dilution Substances 0.000 description 6
- 229940079593 drug Drugs 0.000 description 6
- 238000002474 experimental method Methods 0.000 description 6
- 238000002493 microarray Methods 0.000 description 6
- XXUPLYBCNPLTIW-UHFFFAOYSA-N octadec-7-ynoic acid Chemical compound CCCCCCCCCCC#CCCCCCC(O)=O XXUPLYBCNPLTIW-UHFFFAOYSA-N 0.000 description 6
- 239000000049 pigment Substances 0.000 description 6
- 230000001105 regulatory effect Effects 0.000 description 6
- 230000014616 translation Effects 0.000 description 6
- ATRRKUHOCOJYRX-UHFFFAOYSA-N Ammonium bicarbonate Chemical compound [NH4+].OC([O-])=O ATRRKUHOCOJYRX-UHFFFAOYSA-N 0.000 description 5
- 229910000013 Ammonium bicarbonate Inorganic materials 0.000 description 5
- 102100031092 C-C motif chemokine 3 Human genes 0.000 description 5
- 101710155856 C-C motif chemokine 3 Proteins 0.000 description 5
- 239000002126 C01EB10 - Adenosine Substances 0.000 description 5
- 102000003969 Fibroblast growth factor 4 Human genes 0.000 description 5
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 5
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 5
- 102000004230 Neurotrophin 3 Human genes 0.000 description 5
- 108010082093 Placenta Growth Factor Proteins 0.000 description 5
- 102100037596 Platelet-derived growth factor subunit A Human genes 0.000 description 5
- 101710103506 Platelet-derived growth factor subunit A Proteins 0.000 description 5
- 206010052428 Wound Diseases 0.000 description 5
- 208000027418 Wounds and injury Diseases 0.000 description 5
- 229960005305 adenosine Drugs 0.000 description 5
- 235000012538 ammonium bicarbonate Nutrition 0.000 description 5
- 239000001099 ammonium carbonate Substances 0.000 description 5
- 229940098773 bovine serum albumin Drugs 0.000 description 5
- 230000002338 cryopreservative effect Effects 0.000 description 5
- 238000012136 culture method Methods 0.000 description 5
- 230000006378 damage Effects 0.000 description 5
- 229940098448 fibroblast growth factor 7 Drugs 0.000 description 5
- 230000003834 intracellular effect Effects 0.000 description 5
- 229940032018 neurotrophin 3 Drugs 0.000 description 5
- 229940097998 neurotrophin 4 Drugs 0.000 description 5
- 238000003786 synthesis reaction Methods 0.000 description 5
- 239000003981 vehicle Substances 0.000 description 5
- 238000012795 verification Methods 0.000 description 5
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 4
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 4
- 238000011740 C57BL/6 mouse Methods 0.000 description 4
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 4
- 102000004269 Granulocyte Colony-Stimulating Factor Human genes 0.000 description 4
- 101000645296 Homo sapiens Metalloproteinase inhibitor 2 Proteins 0.000 description 4
- 206010028980 Neoplasm Diseases 0.000 description 4
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 description 4
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 4
- 238000003556 assay Methods 0.000 description 4
- 230000030833 cell death Effects 0.000 description 4
- 230000010261 cell growth Effects 0.000 description 4
- 239000003153 chemical reaction reagent Substances 0.000 description 4
- 238000011161 development Methods 0.000 description 4
- 230000018109 developmental process Effects 0.000 description 4
- 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 4
- 229960003957 dexamethasone Drugs 0.000 description 4
- 230000004069 differentiation Effects 0.000 description 4
- 239000003085 diluting agent Substances 0.000 description 4
- 210000002889 endothelial cell Anatomy 0.000 description 4
- 235000019253 formic acid Nutrition 0.000 description 4
- 230000001976 improved effect Effects 0.000 description 4
- 230000001939 inductive effect Effects 0.000 description 4
- 230000002757 inflammatory effect Effects 0.000 description 4
- 229940100601 interleukin-6 Drugs 0.000 description 4
- 238000004949 mass spectrometry Methods 0.000 description 4
- 239000000463 material Substances 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- 230000000284 resting effect Effects 0.000 description 4
- 230000028327 secretion Effects 0.000 description 4
- 230000000638 stimulation Effects 0.000 description 4
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 4
- 230000000007 visual effect Effects 0.000 description 4
- 102000001301 EGF receptor Human genes 0.000 description 3
- 108060006698 EGF receptor Proteins 0.000 description 3
- 102000009024 Epidermal Growth Factor Human genes 0.000 description 3
- 102000007072 Nerve Growth Factors Human genes 0.000 description 3
- 102000046299 Transforming Growth Factor beta1 Human genes 0.000 description 3
- 101800002279 Transforming growth factor beta-1 Proteins 0.000 description 3
- 239000004480 active ingredient Substances 0.000 description 3
- 230000033115 angiogenesis Effects 0.000 description 3
- 201000011510 cancer Diseases 0.000 description 3
- 238000006243 chemical reaction Methods 0.000 description 3
- 239000003795 chemical substances by application Substances 0.000 description 3
- 230000000052 comparative effect Effects 0.000 description 3
- 230000008878 coupling Effects 0.000 description 3
- 238000010168 coupling process Methods 0.000 description 3
- 238000005859 coupling reaction Methods 0.000 description 3
- 230000013020 embryo development Effects 0.000 description 3
- 239000000839 emulsion Substances 0.000 description 3
- 230000014509 gene expression Effects 0.000 description 3
- 230000002068 genetic effect Effects 0.000 description 3
- 210000003958 hematopoietic stem cell Anatomy 0.000 description 3
- 230000006698 induction Effects 0.000 description 3
- 230000000670 limiting effect Effects 0.000 description 3
- 210000004185 liver Anatomy 0.000 description 3
- 239000006210 lotion Substances 0.000 description 3
- 210000004072 lung Anatomy 0.000 description 3
- 238000000691 measurement method Methods 0.000 description 3
- 238000013508 migration Methods 0.000 description 3
- 239000003226 mitogen Substances 0.000 description 3
- 210000001616 monocyte Anatomy 0.000 description 3
- 239000003900 neurotrophic factor Substances 0.000 description 3
- 239000003921 oil Substances 0.000 description 3
- 235000019198 oils Nutrition 0.000 description 3
- 210000002997 osteoclast Anatomy 0.000 description 3
- 230000002093 peripheral effect Effects 0.000 description 3
- 239000000843 powder Substances 0.000 description 3
- 238000002360 preparation method Methods 0.000 description 3
- 239000003755 preservative agent Substances 0.000 description 3
- 108090000765 processed proteins & peptides Proteins 0.000 description 3
- 230000035755 proliferation Effects 0.000 description 3
- 239000012488 sample solution Substances 0.000 description 3
- 239000004094 surface-active agent Substances 0.000 description 3
- 208000024891 symptom Diseases 0.000 description 3
- 229940099456 transforming growth factor beta 1 Drugs 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- ZKRFOXLVOKTUTA-KQYNXXCUSA-N 9-(5-phosphoribofuranosyl)-6-mercaptopurine Chemical compound O[C@@H]1[C@H](O)[C@@H](COP(O)(O)=O)O[C@H]1N1C(NC=NC2=S)=C2N=C1 ZKRFOXLVOKTUTA-KQYNXXCUSA-N 0.000 description 2
- 241000894006 Bacteria Species 0.000 description 2
- 102000001902 CC Chemokines Human genes 0.000 description 2
- 108010040471 CC Chemokines Proteins 0.000 description 2
- 239000006147 Glasgow's Minimal Essential Medium Substances 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- 101000893549 Homo sapiens Growth/differentiation factor 15 Proteins 0.000 description 2
- 102000004877 Insulin Human genes 0.000 description 2
- 239000007760 Iscove's Modified Dulbecco's Medium Substances 0.000 description 2
- 102000007330 LDL Lipoproteins Human genes 0.000 description 2
- 108010007622 LDL Lipoproteins Proteins 0.000 description 2
- 108010058398 Macrophage Colony-Stimulating Factor Receptor Proteins 0.000 description 2
- 241000124008 Mammalia Species 0.000 description 2
- 229930182555 Penicillin Natural products 0.000 description 2
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 2
- 206010060862 Prostate cancer Diseases 0.000 description 2
- 208000000236 Prostatic Neoplasms Diseases 0.000 description 2
- 210000001744 T-lymphocyte Anatomy 0.000 description 2
- 102000004142 Trypsin Human genes 0.000 description 2
- 108090000631 Trypsin Proteins 0.000 description 2
- 108060008683 Tumor Necrosis Factor Receptor Proteins 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 230000009471 action Effects 0.000 description 2
- 230000003698 anagen phase Effects 0.000 description 2
- 239000003963 antioxidant agent Substances 0.000 description 2
- 230000006907 apoptotic process Effects 0.000 description 2
- 238000003491 array Methods 0.000 description 2
- 210000003719 b-lymphocyte Anatomy 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 230000004071 biological effect Effects 0.000 description 2
- 239000011616 biotin Substances 0.000 description 2
- 229960002685 biotin Drugs 0.000 description 2
- 235000020958 biotin Nutrition 0.000 description 2
- 230000000903 blocking effect Effects 0.000 description 2
- 210000000601 blood cell Anatomy 0.000 description 2
- 230000017531 blood circulation Effects 0.000 description 2
- 239000012888 bovine serum Substances 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 239000007853 buffer solution Substances 0.000 description 2
- 239000002775 capsule Substances 0.000 description 2
- 239000004202 carbamide Substances 0.000 description 2
- 229910052799 carbon Inorganic materials 0.000 description 2
- 239000000969 carrier Substances 0.000 description 2
- 230000032823 cell division Effects 0.000 description 2
- 238000005119 centrifugation Methods 0.000 description 2
- 239000012141 concentrate Substances 0.000 description 2
- 235000008504 concentrate Nutrition 0.000 description 2
- 239000006071 cream Substances 0.000 description 2
- 230000035617 depilation Effects 0.000 description 2
- 210000001671 embryonic stem cell Anatomy 0.000 description 2
- 230000002708 enhancing effect Effects 0.000 description 2
- 238000009472 formulation Methods 0.000 description 2
- 239000003205 fragrance Substances 0.000 description 2
- 238000011990 functional testing Methods 0.000 description 2
- 239000000499 gel Substances 0.000 description 2
- 235000011187 glycerol Nutrition 0.000 description 2
- 210000003494 hepatocyte Anatomy 0.000 description 2
- 210000000987 immune system Anatomy 0.000 description 2
- 238000001727 in vivo Methods 0.000 description 2
- 208000015181 infectious disease Diseases 0.000 description 2
- 102000028416 insulin-like growth factor binding Human genes 0.000 description 2
- 108091022911 insulin-like growth factor binding Proteins 0.000 description 2
- 230000016507 interphase Effects 0.000 description 2
- 210000003734 kidney Anatomy 0.000 description 2
- 238000002372 labelling Methods 0.000 description 2
- 208000019423 liver disease Diseases 0.000 description 2
- 210000002540 macrophage Anatomy 0.000 description 2
- HQKMJHAJHXVSDF-UHFFFAOYSA-L magnesium stearate Chemical compound [Mg+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O HQKMJHAJHXVSDF-UHFFFAOYSA-L 0.000 description 2
- 210000002752 melanocyte Anatomy 0.000 description 2
- 235000013336 milk Nutrition 0.000 description 2
- 239000008267 milk Substances 0.000 description 2
- 210000004080 milk Anatomy 0.000 description 2
- 210000005036 nerve Anatomy 0.000 description 2
- 229940049954 penicillin Drugs 0.000 description 2
- 238000004445 quantitative analysis Methods 0.000 description 2
- 102000005962 receptors Human genes 0.000 description 2
- 108020003175 receptors Proteins 0.000 description 2
- 230000004044 response Effects 0.000 description 2
- 210000004761 scalp Anatomy 0.000 description 2
- 210000001044 sensory neuron Anatomy 0.000 description 2
- 239000012086 standard solution Substances 0.000 description 2
- 238000009168 stem cell therapy Methods 0.000 description 2
- 229960005322 streptomycin Drugs 0.000 description 2
- 230000035882 stress Effects 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- 239000003826 tablet Substances 0.000 description 2
- 230000007704 transition Effects 0.000 description 2
- 239000012588 trypsin Substances 0.000 description 2
- 102000003298 tumor necrosis factor receptor Human genes 0.000 description 2
- 239000003643 water by type Substances 0.000 description 2
- 210000002268 wool Anatomy 0.000 description 2
- FHVDTGUDJYJELY-UHFFFAOYSA-N 6-{[2-carboxy-4,5-dihydroxy-6-(phosphanyloxy)oxan-3-yl]oxy}-4,5-dihydroxy-3-phosphanyloxane-2-carboxylic acid Chemical compound O1C(C(O)=O)C(P)C(O)C(O)C1OC1C(C(O)=O)OC(OP)C(O)C1O FHVDTGUDJYJELY-UHFFFAOYSA-N 0.000 description 1
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 1
- 208000023275 Autoimmune disease Diseases 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 102100023703 C-C motif chemokine 15 Human genes 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 102000014914 Carrier Proteins Human genes 0.000 description 1
- 108010078791 Carrier Proteins Proteins 0.000 description 1
- 102000000844 Cell Surface Receptors Human genes 0.000 description 1
- 108010001857 Cell Surface Receptors Proteins 0.000 description 1
- PTHCMJGKKRQCBF-UHFFFAOYSA-N Cellulose, microcrystalline Chemical compound OC1C(O)C(OC)OC(CO)C1OC1C(O)C(O)C(OC)C(CO)O1 PTHCMJGKKRQCBF-UHFFFAOYSA-N 0.000 description 1
- 206010009944 Colon cancer Diseases 0.000 description 1
- FBPFZTCFMRRESA-FSIIMWSLSA-N D-Glucitol Natural products OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-FSIIMWSLSA-N 0.000 description 1
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 1
- FBPFZTCFMRRESA-JGWLITMVSA-N D-glucitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-JGWLITMVSA-N 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 241000283074 Equus asinus Species 0.000 description 1
- 241000283073 Equus caballus Species 0.000 description 1
- 239000004386 Erythritol Substances 0.000 description 1
- UNXHWFMMPAWVPI-UHFFFAOYSA-N Erythritol Natural products OCC(O)C(O)CO UNXHWFMMPAWVPI-UHFFFAOYSA-N 0.000 description 1
- 102100028073 Fibroblast growth factor 5 Human genes 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- 229930182566 Gentamicin Natural products 0.000 description 1
- CEAZRRDELHUEMR-URQXQFDESA-N Gentamicin Chemical compound O1[C@H](C(C)NC)CC[C@@H](N)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](NC)[C@@](C)(O)CO2)O)[C@H](N)C[C@@H]1N CEAZRRDELHUEMR-URQXQFDESA-N 0.000 description 1
- 108010054017 Granulocyte Colony-Stimulating Factor Receptors Proteins 0.000 description 1
- 102100039622 Granulocyte colony-stimulating factor receptor Human genes 0.000 description 1
- 241000282412 Homo Species 0.000 description 1
- 101000978376 Homo sapiens C-C motif chemokine 15 Proteins 0.000 description 1
- 101001060267 Homo sapiens Fibroblast growth factor 5 Proteins 0.000 description 1
- 101000617830 Homo sapiens Sterol O-acyltransferase 1 Proteins 0.000 description 1
- 101000798130 Homo sapiens Tumor necrosis factor receptor superfamily member 11B Proteins 0.000 description 1
- 206010021143 Hypoxia Diseases 0.000 description 1
- 102000014429 Insulin-like growth factor Human genes 0.000 description 1
- 101150073396 LTA gene Proteins 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- 235000010643 Leucaena leucocephala Nutrition 0.000 description 1
- 240000007472 Leucaena leucocephala Species 0.000 description 1
- 206010067125 Liver injury Diseases 0.000 description 1
- 208000019693 Lung disease Diseases 0.000 description 1
- 102000043136 MAP kinase family Human genes 0.000 description 1
- 108091054455 MAP kinase family Proteins 0.000 description 1
- 102000009571 Macrophage Inflammatory Proteins Human genes 0.000 description 1
- 108010009474 Macrophage Inflammatory Proteins Proteins 0.000 description 1
- 229930195725 Mannitol Natural products 0.000 description 1
- 102000002274 Matrix Metalloproteinases Human genes 0.000 description 1
- 108010000684 Matrix Metalloproteinases Proteins 0.000 description 1
- 229920000168 Microcrystalline cellulose Polymers 0.000 description 1
- 102000013760 Microphthalmia-Associated Transcription Factor Human genes 0.000 description 1
- 108010050345 Microphthalmia-Associated Transcription Factor Proteins 0.000 description 1
- 101710151805 Mitochondrial intermediate peptidase 1 Proteins 0.000 description 1
- 239000004909 Moisturizer Substances 0.000 description 1
- 206010029113 Neovascularisation Diseases 0.000 description 1
- 208000012902 Nervous system disease Diseases 0.000 description 1
- 208000025966 Neurological disease Diseases 0.000 description 1
- 235000011205 Ocimum Nutrition 0.000 description 1
- 241001529734 Ocimum Species 0.000 description 1
- 102000038030 PI3Ks Human genes 0.000 description 1
- 108091007960 PI3Ks Proteins 0.000 description 1
- 108091008611 Protein Kinase B Proteins 0.000 description 1
- 108090000412 Protein-Tyrosine Kinases Proteins 0.000 description 1
- 102000004022 Protein-Tyrosine Kinases Human genes 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- 241000700159 Rattus Species 0.000 description 1
- 102000040739 Secretory proteins Human genes 0.000 description 1
- 108091058545 Secretory proteins Proteins 0.000 description 1
- DBMJMQXJHONAFJ-UHFFFAOYSA-M Sodium laurylsulphate Chemical compound [Na+].CCCCCCCCCCCCOS([O-])(=O)=O DBMJMQXJHONAFJ-UHFFFAOYSA-M 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 102100021993 Sterol O-acyltransferase 1 Human genes 0.000 description 1
- 108010090804 Streptavidin Proteins 0.000 description 1
- 101000697584 Streptomyces lavendulae Streptothricin acetyltransferase Proteins 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 235000019486 Sunflower oil Nutrition 0.000 description 1
- 101150109894 TGFA gene Proteins 0.000 description 1
- 108700012920 TNF Proteins 0.000 description 1
- GLNADSQYFUSGOU-GPTZEZBUSA-J Trypan blue Chemical compound [Na+].[Na+].[Na+].[Na+].C1=C(S([O-])(=O)=O)C=C2C=C(S([O-])(=O)=O)C(/N=N/C3=CC=C(C=C3C)C=3C=C(C(=CC=3)\N=N\C=3C(=CC4=CC(=CC(N)=C4C=3O)S([O-])(=O)=O)S([O-])(=O)=O)C)=C(O)C2=C1N GLNADSQYFUSGOU-GPTZEZBUSA-J 0.000 description 1
- 101710178443 Tumor necrosis factor receptor superfamily member 11B Proteins 0.000 description 1
- 208000005475 Vascular calcification Diseases 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- TVXBFESIOXBWNM-UHFFFAOYSA-N Xylitol Natural products OCCC(O)C(O)C(O)CCO TVXBFESIOXBWNM-UHFFFAOYSA-N 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 230000001154 acute effect Effects 0.000 description 1
- 239000000443 aerosol Substances 0.000 description 1
- 238000005054 agglomeration Methods 0.000 description 1
- 230000002776 aggregation Effects 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 229940072056 alginate Drugs 0.000 description 1
- 235000010443 alginic acid Nutrition 0.000 description 1
- 229920000615 alginic acid Polymers 0.000 description 1
- 230000000735 allogeneic effect Effects 0.000 description 1
- XAGFODPZIPBFFR-UHFFFAOYSA-N aluminium Chemical compound [Al] XAGFODPZIPBFFR-UHFFFAOYSA-N 0.000 description 1
- 229910052782 aluminium Inorganic materials 0.000 description 1
- 239000003708 ampul Substances 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000002870 angiogenesis inducing agent Substances 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 230000003656 anti-hair-loss Effects 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 210000000628 antibody-producing cell Anatomy 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 230000005775 apoptotic pathway Effects 0.000 description 1
- 238000003149 assay kit Methods 0.000 description 1
- 210000003651 basophil Anatomy 0.000 description 1
- 210000000227 basophil cell of anterior lobe of hypophysis Anatomy 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- 239000011230 binding agent Substances 0.000 description 1
- 108091008324 binding proteins Proteins 0.000 description 1
- 230000033228 biological regulation Effects 0.000 description 1
- 210000004204 blood vessel Anatomy 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 210000002798 bone marrow cell Anatomy 0.000 description 1
- 230000010072 bone remodeling Effects 0.000 description 1
- 210000004556 brain Anatomy 0.000 description 1
- 239000001506 calcium phosphate Substances 0.000 description 1
- 229910000389 calcium phosphate Inorganic materials 0.000 description 1
- 235000011010 calcium phosphates Nutrition 0.000 description 1
- 239000000378 calcium silicate Substances 0.000 description 1
- 229910052918 calcium silicate Inorganic materials 0.000 description 1
- 235000012241 calcium silicate Nutrition 0.000 description 1
- OYACROKNLOSFPA-UHFFFAOYSA-N calcium;dioxido(oxo)silane Chemical compound [Ca+2].[O-][Si]([O-])=O OYACROKNLOSFPA-UHFFFAOYSA-N 0.000 description 1
- 210000001043 capillary endothelial cell Anatomy 0.000 description 1
- 238000007623 carbamidomethylation reaction Methods 0.000 description 1
- 210000000845 cartilage Anatomy 0.000 description 1
- 230000021164 cell adhesion Effects 0.000 description 1
- 230000032677 cell aging Effects 0.000 description 1
- 238000013354 cell banking Methods 0.000 description 1
- 230000022131 cell cycle Effects 0.000 description 1
- 230000025084 cell cycle arrest Effects 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 230000009087 cell motility Effects 0.000 description 1
- 238000002659 cell therapy Methods 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 235000010980 cellulose Nutrition 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000002738 chelating agent Substances 0.000 description 1
- 239000002975 chemoattractant Substances 0.000 description 1
- 230000035605 chemotaxis Effects 0.000 description 1
- 208000029742 colonic neoplasm Diseases 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 239000003636 conditioned culture medium Substances 0.000 description 1
- 238000012790 confirmation Methods 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 239000006059 cover glass Substances 0.000 description 1
- 238000002425 crystallisation Methods 0.000 description 1
- 230000008025 crystallization Effects 0.000 description 1
- 210000004748 cultured cell Anatomy 0.000 description 1
- 238000005520 cutting process Methods 0.000 description 1
- 230000003013 cytotoxicity Effects 0.000 description 1
- 231100000135 cytotoxicity Toxicity 0.000 description 1
- 238000007405 data analysis Methods 0.000 description 1
- 230000034994 death Effects 0.000 description 1
- 238000000354 decomposition reaction Methods 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 230000003111 delayed effect Effects 0.000 description 1
- 230000002951 depilatory effect Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 230000006866 deterioration Effects 0.000 description 1
- 239000008121 dextrose Substances 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 230000037213 diet Effects 0.000 description 1
- 235000005911 diet Nutrition 0.000 description 1
- VHJLVAABSRFDPM-QWWZWVQMSA-N dithiothreitol Chemical compound SC[C@@H](O)[C@H](O)CS VHJLVAABSRFDPM-QWWZWVQMSA-N 0.000 description 1
- 229920001971 elastomer Polymers 0.000 description 1
- 239000003344 environmental pollutant Substances 0.000 description 1
- 210000002615 epidermis Anatomy 0.000 description 1
- 206010015037 epilepsy Diseases 0.000 description 1
- 230000008556 epithelial cell proliferation Effects 0.000 description 1
- 230000008202 epithelial morphogenesis Effects 0.000 description 1
- UNXHWFMMPAWVPI-ZXZARUISSA-N erythritol Chemical compound OC[C@H](O)[C@H](O)CO UNXHWFMMPAWVPI-ZXZARUISSA-N 0.000 description 1
- 235000019414 erythritol Nutrition 0.000 description 1
- 229940009714 erythritol Drugs 0.000 description 1
- 210000003527 eukaryotic cell Anatomy 0.000 description 1
- 230000008175 fetal development Effects 0.000 description 1
- 230000001605 fetal effect Effects 0.000 description 1
- 239000000945 filler Substances 0.000 description 1
- 239000007850 fluorescent dye Substances 0.000 description 1
- 239000006260 foam Substances 0.000 description 1
- 239000011888 foil Substances 0.000 description 1
- 239000012737 fresh medium Substances 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- 239000008273 gelatin Substances 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 235000011852 gelatine desserts Nutrition 0.000 description 1
- 210000004392 genitalia Anatomy 0.000 description 1
- 229960002518 gentamicin Drugs 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 239000008187 granular material Substances 0.000 description 1
- 230000009036 growth inhibition Effects 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 208000019622 heart disease Diseases 0.000 description 1
- 210000003709 heart valve Anatomy 0.000 description 1
- 210000000777 hematopoietic system Anatomy 0.000 description 1
- 231100000234 hepatic damage Toxicity 0.000 description 1
- 230000013632 homeostatic process Effects 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 239000003906 humectant Substances 0.000 description 1
- 230000007062 hydrolysis Effects 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- 230000001146 hypoxic effect Effects 0.000 description 1
- 238000010191 image analysis Methods 0.000 description 1
- 210000002865 immune cell Anatomy 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 230000008595 infiltration Effects 0.000 description 1
- 238000001764 infiltration Methods 0.000 description 1
- 210000004969 inflammatory cell Anatomy 0.000 description 1
- 239000003112 inhibitor Substances 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 208000014674 injury Diseases 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 230000008611 intercellular interaction Effects 0.000 description 1
- 229940074383 interleukin-11 Drugs 0.000 description 1
- 238000001361 intraarterial administration Methods 0.000 description 1
- 238000007918 intramuscular administration Methods 0.000 description 1
- 238000007912 intraperitoneal administration Methods 0.000 description 1
- 238000001990 intravenous administration Methods 0.000 description 1
- 210000002510 keratinocyte Anatomy 0.000 description 1
- 208000017169 kidney disease Diseases 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 210000000265 leukocyte Anatomy 0.000 description 1
- 201000007270 liver cancer Diseases 0.000 description 1
- 230000008818 liver damage Effects 0.000 description 1
- 208000014018 liver neoplasm Diseases 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 238000011866 long-term treatment Methods 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 235000019359 magnesium stearate Nutrition 0.000 description 1
- 239000000845 maltitol Substances 0.000 description 1
- 235000010449 maltitol Nutrition 0.000 description 1
- VQHSOMBJVWLPSR-WUJBLJFYSA-N maltitol Chemical compound OC[C@H](O)[C@@H](O)[C@@H]([C@H](O)CO)O[C@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O VQHSOMBJVWLPSR-WUJBLJFYSA-N 0.000 description 1
- 229940035436 maltitol Drugs 0.000 description 1
- 239000000594 mannitol Substances 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 230000035800 maturation Effects 0.000 description 1
- 239000012092 media component Substances 0.000 description 1
- 210000003593 megakaryocyte Anatomy 0.000 description 1
- HEBKCHPVOIAQTA-UHFFFAOYSA-N meso ribitol Natural products OCC(O)C(O)C(O)CO HEBKCHPVOIAQTA-UHFFFAOYSA-N 0.000 description 1
- 229920000609 methyl cellulose Polymers 0.000 description 1
- 239000001923 methylcellulose Substances 0.000 description 1
- 235000010981 methylcellulose Nutrition 0.000 description 1
- LXCFILQKKLGQFO-UHFFFAOYSA-N methylparaben Chemical compound COC(=O)C1=CC=C(O)C=C1 LXCFILQKKLGQFO-UHFFFAOYSA-N 0.000 description 1
- 235000019813 microcrystalline cellulose Nutrition 0.000 description 1
- 239000008108 microcrystalline cellulose Substances 0.000 description 1
- 229940016286 microcrystalline cellulose Drugs 0.000 description 1
- 239000002480 mineral oil Substances 0.000 description 1
- 235000010446 mineral oil Nutrition 0.000 description 1
- 239000006151 minimal media Substances 0.000 description 1
- 239000011259 mixed solution Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000001333 moisturizer Effects 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 210000000663 muscle cell Anatomy 0.000 description 1
- 210000000653 nervous system Anatomy 0.000 description 1
- 230000004766 neurogenesis Effects 0.000 description 1
- 230000033667 organ regeneration Effects 0.000 description 1
- 230000011164 ossification Effects 0.000 description 1
- 210000000963 osteoblast Anatomy 0.000 description 1
- 230000007310 pathophysiology Effects 0.000 description 1
- 239000008188 pellet Substances 0.000 description 1
- 210000005259 peripheral blood Anatomy 0.000 description 1
- 239000011886 peripheral blood Substances 0.000 description 1
- 150000002978 peroxides Chemical class 0.000 description 1
- 230000001766 physiological effect Effects 0.000 description 1
- 239000002504 physiological saline solution Substances 0.000 description 1
- 210000002826 placenta Anatomy 0.000 description 1
- 230000028742 placenta development Effects 0.000 description 1
- 231100000719 pollutant Toxicity 0.000 description 1
- 239000001267 polyvinylpyrrolidone Substances 0.000 description 1
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 description 1
- 229920000036 polyvinylpyrrolidone Polymers 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- QELSKZZBTMNZEB-UHFFFAOYSA-N propylparaben Chemical compound CCCOC(=O)C1=CC=C(O)C=C1 QELSKZZBTMNZEB-UHFFFAOYSA-N 0.000 description 1
- 229960003415 propylparaben Drugs 0.000 description 1
- 238000003498 protein array Methods 0.000 description 1
- 239000012460 protein solution Substances 0.000 description 1
- 238000004451 qualitative analysis Methods 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 238000006722 reduction reaction Methods 0.000 description 1
- 210000002374 sebum Anatomy 0.000 description 1
- 230000003248 secreting effect Effects 0.000 description 1
- 230000007651 self-proliferation Effects 0.000 description 1
- 230000009758 senescence Effects 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 230000001953 sensory effect Effects 0.000 description 1
- 239000004017 serum-free culture medium Substances 0.000 description 1
- 230000011664 signaling Effects 0.000 description 1
- 238000007390 skin biopsy Methods 0.000 description 1
- 230000008470 skin growth Effects 0.000 description 1
- 210000000329 smooth muscle myocyte Anatomy 0.000 description 1
- 239000001540 sodium lactate Substances 0.000 description 1
- 239000001488 sodium phosphate Substances 0.000 description 1
- 229910000162 sodium phosphate Inorganic materials 0.000 description 1
- 239000000600 sorbitol Substances 0.000 description 1
- 235000010356 sorbitol Nutrition 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 210000000603 stem cell niche Anatomy 0.000 description 1
- 230000004936 stimulating effect Effects 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 210000002536 stromal cell Anatomy 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000002600 sunflower oil Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000008093 supporting effect Effects 0.000 description 1
- 230000002459 sustained effect Effects 0.000 description 1
- 230000002889 sympathetic effect Effects 0.000 description 1
- 239000006188 syrup Substances 0.000 description 1
- 235000020357 syrup Nutrition 0.000 description 1
- 239000000454 talc Substances 0.000 description 1
- 229910052623 talc Inorganic materials 0.000 description 1
- 235000012222 talc Nutrition 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 238000010257 thawing Methods 0.000 description 1
- 239000002562 thickening agent Substances 0.000 description 1
- ANRHNWWPFJCPAZ-UHFFFAOYSA-M thionine Chemical compound [Cl-].C1=CC(N)=CC2=[S+]C3=CC(N)=CC=C3N=C21 ANRHNWWPFJCPAZ-UHFFFAOYSA-M 0.000 description 1
- 230000009772 tissue formation Effects 0.000 description 1
- 230000000699 topical effect Effects 0.000 description 1
- 235000013619 trace mineral Nutrition 0.000 description 1
- 239000011573 trace mineral Substances 0.000 description 1
- 238000002054 transplantation Methods 0.000 description 1
- 230000008733 trauma Effects 0.000 description 1
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 description 1
- RYFMWSXOAZQYPI-UHFFFAOYSA-K trisodium phosphate Chemical compound [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O RYFMWSXOAZQYPI-UHFFFAOYSA-K 0.000 description 1
- 108010064880 trkB Receptor Proteins 0.000 description 1
- 102000015534 trkB Receptor Human genes 0.000 description 1
- 210000004926 tubular epithelial cell Anatomy 0.000 description 1
- 238000004704 ultra performance liquid chromatography Methods 0.000 description 1
- 230000008728 vascular permeability Effects 0.000 description 1
- 230000035899 viability Effects 0.000 description 1
- 210000001835 viscera Anatomy 0.000 description 1
- 230000009278 visceral effect Effects 0.000 description 1
- 239000004034 viscosity adjusting agent Substances 0.000 description 1
- 239000000080 wetting agent Substances 0.000 description 1
- 239000000811 xylitol Substances 0.000 description 1
- 235000010447 xylitol Nutrition 0.000 description 1
- HEBKCHPVOIAQTA-SCDXWVJYSA-N xylitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)CO HEBKCHPVOIAQTA-SCDXWVJYSA-N 0.000 description 1
- 229960002675 xylitol Drugs 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
- A61K35/28—Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/39—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin, cold insoluble globulin [CIG]
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0012—Galenical forms characterised by the site of application
- A61K9/0014—Skin, i.e. galenical aspects of topical compositions
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P17/00—Drugs for dermatological disorders
- A61P17/16—Emollients or protectives, e.g. against radiation
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
- C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/475—Growth factors; Growth regulators
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/475—Growth factors; Growth regulators
- C07K14/49—Platelet-derived growth factor [PDGF]
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/475—Growth factors; Growth regulators
- C07K14/50—Fibroblast growth factor [FGF]
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/52—Cytokines; Lymphokines; Interferons
- C07K14/525—Tumour necrosis factor [TNF]
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/52—Cytokines; Lymphokines; Interferons
- C07K14/54—Interleukins [IL]
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/52—Cytokines; Lymphokines; Interferons
- C07K14/54—Interleukins [IL]
- C07K14/5409—IL-5
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/52—Cytokines; Lymphokines; Interferons
- C07K14/54—Interleukins [IL]
- C07K14/545—IL-1
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/78—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG]
-
- 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
-
- 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
- C12N5/0018—Culture media for cell or tissue culture
- C12N5/0031—Serum-free culture media
Definitions
- the present invention is a mass production method of mesenchymal stem cell-derived protein containing various growth factors and cytokines, mesenchymal stem cell culture containing a large amount of protein produced by the production method, skin regeneration including the culture, wrinkle improvement
- the present invention relates to a cosmetic composition for preventing hair loss, treating hair loss and promoting hair growth, and a pharmaceutical composition for skin regeneration.
- Stem cell therapy products proliferate, select, or otherwise alter the biological properties of cells in vitro by autologous, allogeneic, or xenogeneic cells to restore cell and tissue function. It is defined as a drug that is used for the purpose of treatment, diagnosis, and prevention through a series of actions (more-than-minimal manipulation).
- Stem cell therapies are meant to include stem cells, particularly stem cells, and are actively used for diseases that are difficult to recover from, but difficult to heal, such as neurological disease, heart disease, lung disease, liver disease and cancer. Development is in progress.
- Stem cells are multipotent and can be differentiated into specific cells, so they have great potential as cell therapeutics.However, to date, the survival rate is not high after transplantation and can cause immune rejection. Hard to find.
- Stem cell culture medium is a medium that does not contain cells obtained after culturing cells, and contains various components (eg, cytokines, growth factors, etc.) necessary for cell growth. Stem cell cultures are used to promote cell growth, or are used to isolate specific components. In addition, stem cell cultures are applied to the treatment of various diseases. For example, it has been reported that the human mesenchymal stem cell culture can be applied to the treatment of acute liver disease by using the effect of inhibiting the death of hepatocytes and increasing the regeneration of the hepatocytes. It contains growth factors and proteins that are effective for regeneration and wound healing, and it has been reported to promote wound healing when applied to the wound.
- fat-derived mesenchymal stem cells have been reported to secrete genes and growth factors involved in blood vessel and cell regeneration, and hepatocellular-like cells differentiated from human embryonic stem cells and proteins secreted from these cells have been damaged in the liver. All of them have been reported to restore liver damage.
- the medium used in the above studies has a disadvantage that it is difficult to apply directly to the clinic because it contains bovine serum, which is an animal-derived component, and a buffer solution and indicator components which are less stable.
- bovine serum which is an animal-derived component
- a buffer solution and indicator components which are less stable.
- the concentration of components secreted into the medium is low, and there is a problem that a large amount of stem cell culture is used in the affected part for clinical application, and also to increase the concentration. If the operation such as enrichment, there is a problem that the production cost is significantly increased and economic efficiency is lowered.
- mesenchymal stem cells capable of mass-producing various growth factors and proteins that have not been expressed by the previous culture method or are expressed only in a very small amount in stem cell culture.
- a culture method was developed. Furthermore, the culturing method produces a large amount of useful proteins including growth factors by changing various culture conditions in the serum-free culturing step, instead of artificially manipulating the cells themselves in order to increase the content of proteins containing specific growth factors.
- Optimal culture conditions have been established for this purpose, and the produced mesenchymal stem cell culture solution is expected to be very useful for application to human body for skin regeneration, wrinkle improvement, hair loss prevention, hair loss treatment and hair growth promotion.
- An object of the present invention is a mass production method of mesenchymal stem cell-derived protein comprising various growth factors and cytokines, mesenchymal stem cell culture medium containing a large amount of protein produced by the production method, skin regeneration comprising the culture medium,
- the present invention provides a cosmetic composition for improving wrinkles, preventing hair loss, treating hair loss or promoting hair growth, and a pharmaceutical composition for skin regeneration.
- the mass production method of the mesenchymal stem cell-derived protein of the present invention significantly increases the type and amount of the protein secreted from the mesenchymal stem cells as compared to the conventional method, in particular the mesenchymal stem cell culture prepared by the above method It contains a large amount of cytokines and growth factors, and contains a large amount of collagen, which is a major component of the skin, and also has an excellent effect on promoting collagen production. It has excellent effects on skin regeneration, wrinkle improvement, hair loss prevention, hair loss treatment, and hair growth promotion. There is.
- Figure 1 confirms the optimal cell inoculation density conditions for the production of stem cell secreted proteins.
- Figure 2 compares protein production according to serum-free incubation time (12 hour intervals).
- Figure 3 compares the cell viability according to the freezing storage conditions of mesenchymal stem cells.
- Figure 4 compares the secretion proteins of stem cells according to the cell freezing solution type.
- Figure 5 compares the total protein amount of the stem cell culture obtained in the optimal serum-free culture and storage conditions.
- Figure 6 is a scan image of the antibody array (Antibody array) of stem cell cultures obtained in optimal serum-free culture and storage conditions.
- Figure 7 confirms the collagen synthesis efficacy of the stem cell culture obtained in the optimal serum-free culture and storage conditions.
- Figure 8 compares the wound healing efficacy of stem cell cultures obtained under optimal serum-free culture conditions and storage conditions.
- Figure 9 confirms the hair growth promoting effect of mesenchymal stem cell culture in the hair growth inhibition model, it shows the hair condition of the mouse on day 0 of the test.
- G1 is treated with distilled water control
- G2 is Placebo control
- G3 is test substance
- G4 is positive control (5% Minoxidil).
- Figure 10 confirms the hair growth promoting effect of the mesenchymal stem cell culture in the hair growth inhibition model, it shows the hair condition of the mouse on day 4 of the test.
- G1 is treated with distilled water control
- G2 is Placebo control
- G3 is test substance
- G4 is positive control (5% Minoxidil).
- Figure 11 confirms the hair growth promoting effect of mesenchymal stem cell culture in the hair growth inhibition model, it shows the hair condition of the mouse on day 8 of the test.
- G1 is treated with distilled water control
- G2 is Placebo control
- G3 is test substance
- G4 is positive control (5% Minoxidil).
- Figure 12 graphically shows the hair growth score of mesenchymal stem cell culture in the hair growth inhibition model.
- G1 is treated with distilled water control
- G2 is Placebo control
- G3 is test substance
- G4 is positive control (5% Minoxidil).
- Figure 13 is a comparison of histopathological hair follicles of mice treated with mesenchymal stem cell culture in the hair growth inhibition model.
- G1 is treated with distilled water control
- G2 is Placebo control
- G3 is test substance
- G4 is positive control (5% Minoxidil).
- Figure 14 is a graph comparing the number of hair follicles of mice treated with mesenchymal stem cell culture in the hair growth inhibition model.
- G1 is treated with distilled water control
- G2 is Placebo control
- G3 is test substance
- G4 is positive control (5% Minoxidil).
- G1 is treated with distilled water control
- G2 is Placebo control
- G3 is test substance
- G4 is positive control (5% Minoxidil).
- Figure 16 confirms the hair growth promoting effect of mesenchymal stem cell culture in the growth hair model, it shows the hair condition of the mouse on day 15 of the test.
- G1 is treated with distilled water control
- G2 is Placebo control
- G3 is test substance
- G4 is positive control (5% Minoxidil).
- Figure 17 graphically shows the hair growth score of mesenchymal stem cell culture in the growing hair model.
- G1 is treated with distilled water control
- G2 is Placebo control
- G3 is test substance
- G4 is positive control (5% Minoxidil).
- Figure 18 is a comparison of histopathological hair follicles of mice treated with mesenchymal stem cell culture in the growth hair model.
- G1 is treated with distilled water control
- G2 is Placebo control
- G3 is test substance
- G4 is positive control (5% Minoxidil).
- 19 is a graph comparing the number of hair follicles of mice treated with mesenchymal stem cell culture in the growth hair model.
- G1 is treated with distilled water control
- G2 is Placebo control
- G3 is test substance
- G4 is positive control (5% Minoxidil).
- 21 shows the verification of the optimal serum-free culture conditions in bone marrow-derived mesenchymal stem cell culture.
- the present invention as one embodiment (a) inoculating mesenchymal stem cells at a density of 18,000 to 22,000 cells / cm 2 ; (b) culturing the mesenchymal stem cells in a serum-free medium; And (c) a method for mass production of mesenchymal stem cell-derived protein comprising the step of obtaining a stem cell culture medium after 114 to 126 hours of culture, the mesenchymal stem cells are CRYO in -90 °C to -70 °C cryogenic freezer -GOLD solution, the protein is AR, bFGF, BMP-5, BMP-7, GH, IGFBP-1, IGFBP-2, IGFBP-3, IGFBP-4, SCF, TGF ⁇ , TGF ⁇ 1, VEGF R3, Bulk of mesenchymal stem cell derived proteins, including VEGF-D, ICAM-1, IL-1a, IL-5, MIP-1a, MIP-b, MIP-d, RAN
- the present inventors have optimized the stem cell culture method for mass production of useful proteins such as cytokines, growth factors, etc. from the stem cell culture medium, the stem cell culture solution obtained from the optimized stem cell culture method contains a large amount of useful proteins, In particular, it contained high levels of collagen, and it was confirmed that it shows excellent wound healing ability by improving collagen synthesis in fibroblasts.
- Step (a) is a step of inoculating mesenchymal stem cells in an appropriate number, in particular, inoculating at a density of 18,000 to 22,000 cells / cm 2 .
- stem cell refers to a cell having the ability to differentiate into various cells through a suitable environment and stimulation, and having a self-proliferation ability.
- Stem cells to be used in the present invention is not limited to the kind if the one having differentiation capacity and autoproliferation ability, preferably mesenchymal stem cells (mesenchymal stem cells), more preferably human fat, umbilical cord blood, It may be bone marrow, amniotic fluid or amnion derived mesenchymal stem cells. Most preferably, it may be amniotic fluid-derived mesenchymal stem cells.
- mesenchymal stem cell is a cell that helps to create cartilage, bone, fat, myeloid epilepsy, muscle, nerves, etc., in adults in the bone marrow in general, but the cord blood, It also exists in peripheral blood, fat, amniotic fluid and other tissues, etc., and means mesenchymal stem cells that can be obtained from them, and is not particularly limited thereto.
- Mesenchymal stem cells may behave differently depending on the surrounding microenvironment (stem cell niche) in vivo, such as cell division, differentiation or migration. More specifically, the mesenchymal stem cells may be changed in gene expression patterns by stimulation from the surrounding microenvironment, and thus, the type and amount of the secreted protein may be changed.
- This peripheral microenvironment includes not only the physical environment around the cell, that is, the characteristics of the tissue in which the cell is present, the location of the cell, the state of attachment, etc., but also the chemical environment, i.e., an external cytokine or growth factor.
- In vitro culture of stem cells involves inoculating the stored stem cells, primary cultured cells or subcultured cells into the culture vessel, where the density of the inoculated cells differs in the type and amount of the stem cell secreted protein. I can lose it. This is because the gene expression pattern of stem cells is changed by cell-cell interaction, which is performed directly or indirectly.
- the density of the cells inoculated in the present invention may be 18,000 to 22,000 cells / cm 2 density, preferably 19,000 to 20,000 cells / cm 2 density, more preferably 20,000 cells / cm 2 density.
- the mesenchymal stem cells were inoculated at a density of 5,000 to 25,000 cells / cm 2 to confirm the concentration of the secreted protein, and the highest protein was expressed at a density of 20,000 cells / cm 2 . It was confirmed (FIG. 1). In particular, in the case of 25,000 cells / cm 2 , it was confirmed that the protein concentration is lower even though there are more cells. Therefore, the optimal cell density for mass production of mesenchymal stem cell-derived protein was found to be about 20,000 cells / cm 2 .
- Next step (b) is a step of culturing the mesenchymal stem cells in serum-free medium.
- culture media refers to a medium capable of supporting the growth and survival of stem cells in vitro , and includes all conventional media used in the art suitable for stem cell culture. Include. Depending on the type of cells, medium and culture conditions can be selected.
- the medium used for the culturing is preferably a cell culture minimum medium (CCMM), and generally includes a carbon source, a nitrogen source and a trace element component.
- CCMM cell culture minimum medium
- Such cell culture minimal media include, for example, Dulbecco's Modified Eagle's Medium (DMEM), Minimal Essential Medium (MEM), Basic Medium Eagle (BME), RPMI1640, F-10, F-12, ⁇ -MEM ( ⁇ -Minimal essential) Medium), Glasgow's Minimal Essential Medium (GMEM), and Iscove's Modified Dulbecco's Medium (IMEM), but are not limited thereto.
- the medium may include antibiotics such as penicillin, streptomycin, or gentamicin.
- the medium is characterized in that the serum-free medium (serum-free media).
- Serum is generally essential to the culture medium when culturing cells in vitro.
- the cells are cycle to occur the cell cycle inhibitory stay groups G 0 (cell cycle arrest), a cell aging (senescence) can take place is taking place, or cell death (apoptosis), if this condition persists.
- G 0 cell cycle arrest
- a cell aging a cell aging
- cell death apoptosis
- Serum contains a variety of cytokines and growth factors to enable cell proliferation, but to date serum components are not fully understood. Therefore, serum derived from animals has the risk of causing an immune response or the development of certain diseases, so the possibility of side effects continues to be addressed for human application of cells or cell cultures cultured in a medium containing serum. The situation is becoming difficult, and there is a difficulty in proceeding with the clinical.
- the present invention has been developed a method for mass production of mesenchymal stem cell-derived protein, including the step of culturing mesenchymal stem cells in serum-free medium to obtain a safe cell culture when applied to the human body.
- Step (c) is a step of obtaining the stem cell culture solution after 114 to 126 hours by culturing the mesenchymal stem cells in a serum-free medium.
- stem cells may vary in the type and amount of proteins secreted according to external stimuli during the culturing process, and factors influencing such external stimuli also include culture time. That is, since the type and amount of proteins secreted from stem cells may vary with time, and the content of proteins in the culture medium may also vary, the present inventors compare the amount of protein production in the culture medium according to the culture time of the stem cells. By culturing the mesenchymal stem cells to establish the optimal time conditions for mass production of protein.
- the time for culturing the stem cells to obtain the stem cell culture solution may be 114 to 126 hours, preferably 117 to 123 hours, more preferably 120 hours.
- mesenchymal stem cells were cultured in a serum-free medium for 72 to 144 hours to compare protein production at each time condition (FIG. 2). As a result, it was confirmed that the protein production was maximized in the culture medium cultured with mesenchymal stem cells for about 120 hours.
- the culture medium containing the protein produced from the mesenchymal stem cells is collected from the culture vessel, and the obtaining step is a one-time culture solution, the cells are washed with a buffer and again in a fresh medium Stem cell cultures can be further obtained after 114 to 126 hours after replacement.
- the process of obtaining such stem cell culture solution may be repeated several times, preferably one to three times, but is not limited thereto.
- the stem cell culture medium obtained by the method for mass production of mesenchymal stem cell-derived protein of the present invention was obtained three times in sequence to confirm the protein concentration of the culture medium, and as the number of times obtained was increased in the culture medium. Protein concentration showed a slight decrease, but compared to the mesenchymal stem cell culture cultured by the conventional method was confirmed to include a significantly larger amount of protein (Fig. 5). Therefore, it can be seen that this obtaining process can be repeated several times in the process of culturing mesenchymal stem cells.
- the mass production method of the mesenchymal stem cell-derived protein characterized in that the mesenchymal stem cells are stored in a CRYO-GOLD solution in -90 °C to -70 °C cryogenic freezer.
- the process of storing the cells may be essential unless the cells cultured through primary culture are used directly.
- Stem cells are self-proliferating cells that can grow indefinitely, but in vitro culture is an artificially created environment, except for some cells including cancer cells and embryonic stem cells. It may occur, and there is a limit of passage culture even for mesenchymal stem cells.
- cell culture requires a lot of cost and manpower, it is very important to store cells in the cell culture process so that the cells can be cultured when needed.
- the cells generally mix the cells in cryopreservatives and store them in cryogenic conditions.
- the cryopreservatives serve to prevent the destruction of the cell membrane by crystallization when the cells are frozen.
- the temperature at which cells are frozen can be important, which can be stored in a liquid nitrogen (LN 2 ) tank at -196 ° C or in a cryogenic freezer at a higher temperature.
- LN 2 liquid nitrogen
- the productivity of the stem cell secretion protein according to the type of cryopreservation liquid in the step of storing the mesenchymal stem cells was confirmed.
- the viability of the cells that were cryopreserved in the control group (10% DMSO, 20% FBS and 70% cDMEM), CRYO-GOLD, CRYO-ROS, STEM-CELL BANKER and CellFreezer was determined. Was absent ( Figure 3).
- the protein concentration appeared to be the highest. It was found that the optimum condition to produce (Fig. 4).
- the mass production method of the mesenchymal stem cell-derived protein of the present invention can be applied irrespective of the tissue derived from the mesenchymal stem cell, and various growth factors including cytokines and growth factors useful in mesenchymal stem cells using the above method. Proteins can be produced in large quantities.
- the mesenchymal stem cells are cultivated for the purpose of mass production of proteins including growth factors.
- the protein may be to increase the amount of protein known to be previously secreted from mesenchymal stem cells, or may be newly secreted by the method, although it is not known to be secreted previously.
- the protein produced by the mass production method of mesenchymal stem cell-derived protein of the present invention is AR, bFGF, BMP-5, BMP-7, GH, IGFBP-1, IGFBP-2, IGFBP-3, IGFBP- 4, SCF, TGF ⁇ , TGF ⁇ 1, VEGF R3, VEGF-D, ICAM-1, IL-1a, IL-5, MIP-1a, MIP-b, MIP-d, RANTES, TNF R1, and TNF RII .
- the stem cell culture medium produced through the mass production method of the mesenchymal stem cell-derived protein of the present invention is BDNF, BMP-4, b-NGF, EGF R, FGF-4, FGF-7, GDF-15, GDNF, HGF, IGFBP-6, IGF-I, Insulin, MCSF R, NGF R, NT-3, NT-4, OPG, PDGF-AA, PIGF, SCF R, VEGF, G-CSF, IL-2, IL-6 , IL-8, IL-11, MCP-1, MCSF, MIG, TIMP-1, TIMP-2, TNF ⁇ and TNF ⁇ may be further included.
- FGF-7 Fibroblast growth factor 7
- FGF-7 Fibroblast growth factor 7
- keratinocytes a growth factor for keratinocytes
- epithelial cell proliferation a major factor for normal epithelial cell proliferation. It is also known to be involved in epithelial formation, re-epithelialization of wounds, hair development, and lung organ formation.
- IGFBP-1 Insulin-like growth factor-binding protein 1
- IGFBP-1 Insulin-like growth factor-binding protein 1
- IGFBP-1 Insulin-like growth factor-binding protein 1
- IGFBP-3 Insulin-like growth factor-binding protein 3
- IGFALS insulin-like growth factor acid-labile subunit
- macrophage colony-stimulating factor receptor is a receptor for MCSF, which affects the differentiation of hematopoietic stem cells into macrophages or other related cells, and in the eukaryotic cell virus. Produced in response to infection and associated with placental development.
- NT-4 Neurotrophin-4
- TrkB receptor tyrosine kinase is a neurotrophic factor that is signaled by the TrkB receptor tyrosine kinase as a receptor and is known as a factor necessary for the survival of peripheral sensory sympathetic nerves.
- TGF- ⁇ 1 Transforming growth factor- ⁇ 1
- Transforming growth factor- ⁇ 1 is a protein having a complex function that positively and negatively regulate a number of different growth factors and regulates cell proliferation, differentiation, and various functions for numerous cell types. . Stimulation of bone formation of osteoblasts plays an important role in bone remodeling and plays an important role in wound healing. It is also important for immune system regulation and most immune cells are known to secrete TGF- ⁇ 1.
- bFGF Fibroblast growth factor 2
- bFGF Fibroblast growth factor 2
- EGF-R Extracellular growth factor receptor
- FGF-4 Fibroblast growth factor 4
- FGF-4 Fibroblast growth factor 4
- FGF-4 Fibroblast growth factor 4
- FGF-4 belongs to the FGF family and plays an important role in regulating embryonic development, cell proliferation, and cell differentiation. It acts as a developmental protein, growth factor, and mitogen, and is a protein necessary for the development of normal limbs and heart valves during embryonic development.
- GDF-15 (Growth / differentiation factor 15) belongs to the TGF-superfamily and is also known as TGF-PL, MIC-1, PDF, PLAB, PTGFB. It plays a role in regulating inflammatory and apoptosis pathways in damaged tissues and during disease processes.
- HGF Hepatocyte growth factor
- mesenchymal cells As used herein, the term, HGF (Hepatocyte growth factor) is secreted from various types of mesenchymal cells as a hepatocyte growth factor. It has cell proliferation promoting activity, cell motility promoting activity, epithelial morphogenesis inducing activity and acts as a neurotrophic factor and angiogenesis factor. In addition, it is involved in the formation of visceral organs such as liver, kidney and lung, placenta and skeletal system as a mediator of epithelial-mesenchymal interaction.In adults, it functions as an organ regeneration factor that promotes regeneration of liver, kidney, lung and digestive tract. It is expected to be a long-term treatment for diseases.
- IGFBP-4 Insulin-like growth factor-binding protein 4
- IGFBP-4 circulates in plasma in the form of glycosylated and unglycosylated with IGF and prolongs the half-life of IGF to promote the effect of IGF growth in cell culture. It inhibits or stimulates action. It is known to reduce the proliferation of cancers such as prostate cancer and colon cancer by inhibiting several cancer cells in vivo and ex vivo and acting as a cell death factor against cancer cells.
- IGFBP-6 Insulin-like growth factor-binding protein 6
- IGFBP-6 binds to the cell surface receptors of the protein and interacts with IGF and prolongs the half-life of IGF to inhibit the growth promoting effect of IGF in cell culture. Or stimulating action.
- NT-3 Neurotrophin-3
- NT-3 Neurotrophin-3 is a protein that is particularly present in the brain and peripheral tissues as a neurotrophic factor and contributes to promoting and regulating neurogenesis. It also enhances survival of visceral sensory neurons and hypersoluble sensory neurons. It is known to have a feature that is expressed with FGF5, TGF-1, etc. in the degenerative phase of the hair cycle.
- OPG Tumor necrosis factor receptor superfamily member 11B
- OPG osteoprotegerin
- OCIF osteocalcosogenesis inhibitory factor
- It counteracts against bone destruction (bone tissue absorption, osteoclastogenesis). It inhibits the activity of osteoclasts (osteoclasts) in vitro and promotes apoptosis of osteoclasts.
- Bone homeostasis depends on the local ratio between TNSF11 and TNFRSF11B. It is also known to prevent arterial calcification.
- platelet-derived growth factor subunit A is a growth factor that plays an important role in regulating fetal development, cell proliferation, cell migration, survival, chemotaxis, and the like. It acts as a mitogen against cells of mesenchymal origin and is known to play an important role in wound healing.
- PIGF Prothelial growth factor
- VEGF Vascular endothelial growth factor A
- VEGF Vascular endothelial growth factor A
- G-CSF G-CSF
- G-CSF granulocyte-colony stimulating factor
- ICAM-1 Intercellular Adhesion Molecule 1
- ICAM-1 Intercellular Adhesion Molecule 1
- ICAM-1 Intercellular Adhesion Molecule 1
- IL-6 Interleukin 6
- IL-6 induces the final differentiation of beta cells into antibody-producing cells, acts as muscle cells and forms bone marrow cells. It is known to stop infection and to withstand bacteria.
- IL-11 Interleukin 11
- B lymphocyte differentiation hematopoietic stem cell proliferation and differentiation
- megakaryocyte proliferation and maturation hematopoietic system It also affects the work, skeletal and nervous systems.
- MCP-1 monocyte chemoattractant protein-1
- monocyte chemoattractant protein-1 is a CC chemokine and selectively induces monocytes, lymphocytes, basophils, etc., in response to IL-1a TNF-a low density lipoprotein (LDL), etc. of stromal cells, glomeruli It is produced in endothelial cells, tubular epithelial cells, capillary endothelial cells and smooth muscle cells.
- LDL low density lipoprotein
- macrophage inflammatory protein 1 alpha is a macrophage inflammatory protein alpha1 that is a member of the CC chemokine or beta subfamily.
- MIP-1 alpha monocytes, T cells, B cells are known to act as a chemical attractant for a variety of cells.
- TIMP-1 TIMP metallopeptidase inhibitor 1
- TIMP-1 TIMP metallopeptidase inhibitor 1
- MMPs MMPs
- TIMP-2 TIMP-2 (TIMP metallopeptidase inhibitor 2) is a constitutive genetic component of TIMP and is known to regulate melanocytes by MITF.
- TNF RI Tumor necrosis factor-and TNF receptor I
- TNF RI Tumor necrosis factor-and TNF receptor I
- TNF-associated cytokines are involved in autoimmune diseases and are particularly known to stimulate T cells.
- the mass production method of the mesenchymal stem cell-derived protein of the present invention it is possible to produce a greater amount than the amount secreted in the conventional culture method.
- various types of useful growth factors and proteins including cytokines it may be very effective in producing, purifying and obtaining desired target proteins, and may be used as a composition for improving skin regeneration and wrinkles.
- the total concentration of secreted protein obtained through the production method is not limited thereto, but may be 30 ⁇ g / ml to 70 ⁇ g / ml by BCA measurement.
- the present invention provides a mesenchymal stem cell culture comprising a protein produced by the method for mass-producing a mesenchymal stem cell-derived protein.
- the method for mass production of mesenchymal stem cell derived proteins and the mesenchymal stem cell culture prepared therefrom are as described above.
- the stem cell culture medium may be one containing 5 to 20% by weight of the collagen, preferably 10 to 15% by weight, but is not limited thereto.
- the stem cell culture may be to promote the production of collagen fibroblasts.
- the absolute content of collagen content contained in the stem cell culture solution of the present invention is compared with the stem cell culture solution produced by the conventional method. As a result, 12.4% by weight of the total detected protein is detected as collagen. It was confirmed that the collagen content more than twice the conventional method (Table 6).
- the present invention provides a cosmetic composition for skin regeneration or wrinkle improvement comprising the mesenchymal stem cell culture.
- the mesenchymal stem cell culture is as described above.
- skin regeneration refers to the process of recovery of skin tissue against damage by external and internal causes of the skin.
- the damage caused by the external causes may include ultraviolet rays, external pollutants, wounds, traumas, and the like, and the damage caused by the internal causes may include stress.
- the term “improving wrinkles” means maintaining or enhancing wrinkles and elasticity of the skin.
- the mesenchymal stem cell culture medium obtained by culturing in the conventional method through a wound healing assay wound healing assay
- wound healing assay a wound healing assay that can evaluate the ability to improve skin regeneration and wrinkles with the mesenchymal stem cell culture medium Compared with the remarkable effect was confirmed (Fig. 8).
- the mesenchymal stem cell culture of the present invention contains a large amount of proteins effective for wound healing, from which the composition comprising the mesenchymal stem cell culture of the present invention can be utilized for skin regeneration and wrinkle improvement. It can be seen that.
- the mesenchymal stem cell culture described above may be included in an appropriate amount to achieve the effect described above.
- the cosmetic composition according to the present invention in addition to the mesenchymal stem cell culture medium described above, various components generally used in external preparations for skin, such as water-soluble components, powder components, oils, as necessary, within the range that does not reduce the effect of the present invention.
- Surfactants, moisturizers, viscosity modifiers, preservatives, antioxidants, fragrances, pigments and the like can be combined.
- the formulation of the cosmetic composition according to the present invention can be arbitrarily selected, with mesenchymal stem cell culture medium as an active ingredient, water, physiological saline, glycerol, oil, surfactants, humectants, thickeners, chelating agents, pigments, preservatives and fragrances, etc.
- Such known cosmetic excipients are mixed to form cosmetic compositions in the form of lotions, liquids, emulsions, emulsions, suspensions, tablets and capsules.
- the cosmetic composition may be used to prepare basic cosmetics such as flexible lotion, milk lotion, nourishing cream, massage cream, essence, cleansing foam, cleansing water, pack or body oil, and color cosmetics such as foundation, lipstick, mascara or makeup base. It may also prepare face washes and baths.
- glycerin 1 to 7% by weight of the cosmetic excipient may be included in the cosmetic composition containing mesenchymal stem cell culture, but is not limited thereto. Do not.
- sunflower oil may be included in the cosmetic excipient to provide an antioxidant function to the cosmetic composition, but is not limited thereto.
- the present invention provides a pharmaceutical composition for skin regeneration comprising the mesenchymal stem cell culture.
- the mesenchymal stem cell culture is as described above.
- composition of the present invention has superior wound healing effect than the mesenchymal stem cell culture cultured by the conventional method through a wound healing assay (wound healing assay) (FIG. 8), it may be used as a pharmaceutical composition for skin regeneration.
- wound healing assay wound healing assay
- the throughput of the pharmaceutical composition for skin regeneration used in the present invention should be a pharmaceutically effective amount.
- pharmaceutically effective amount refers to an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment, and an effective dose level may refer to an individual type and severity, age, sex, The type of disease, the activity of the drug, the sensitivity to the drug, the time of administration, the route of administration and the rate of release, the duration of treatment, the factors including the drug used concurrently, and other factors well known in the medical art can be determined. Effective amounts may vary depending on the route of treatment, the use of excipients, and the possibility of use with other agents, as will be appreciated by those skilled in the art.
- compositions for skin regeneration of the present invention can be prepared in pharmaceutical formulations using methods well known in the art to provide rapid, sustained or delayed release of the active ingredient after administration to a mammal.
- compositions of the present invention can be used in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like, oral formulations, external preparations and patches according to a conventional method, the composition It may further comprise suitable carriers, excipients or diluents commonly used in the preparation of the compounds.
- the carrier may include a non-naturally occuring carrier.
- carriers, excipients and diluents that may be included in the pharmaceutical composition of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium Phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.
- diluents or excipients such as fillers, weights, binders, wetting agents, disintegrating agents and surfactants are usually used.
- the present invention provides a method for skin regeneration comprising administering to a subject a pharmaceutical composition comprising the mesenchymal stem cell culture.
- composition comprising the mesenchymal stem cell culture provided in the present invention can be used as an active ingredient of the pharmaceutical composition for skin regeneration, the composition can be used to treat damaged skin.
- mammals including rats, livestock, humans, and the like, having damaged skin.
- the skin regeneration method of the present invention specifically comprises administering a composition comprising the mesenchymal stem cell culture of the present invention in a pharmaceutically effective amount.
- Suitable total daily usage of the composition comprising the mesenchymal stem cell culture can be appropriately determined by one skilled in the art within the scope of correct medical judgment.
- the specific pharmaceutically effective amount of the composition comprising the mesenchymal stem cell culture for a particular animal may vary depending on the type and extent of the reaction to be achieved, the age, body weight, general health, sex or diet of the individual. It may be determined in consideration of the administration time of the composition containing the stem cell culture, the route of administration and the ratio of the composition, the duration of treatment, and the like, and various factors and medicinal fields including the components of the drug or other composition used together or simultaneously. It can vary according to well-known similar factors.
- the administration route and administration method for administering the composition containing the mesenchymal stem cell culture solution are not particularly limited, and the composition including the mesenchymal stem cell culture solution can reach the desired site. Any route of administration and mode of administration may be followed as long as it is available.
- the composition comprising the mesenchymal stem cell culture may be administered orally or parenterally through various routes, and non-limiting examples of the route of administration include oral, rectal, topical, intravenous, intraperitoneal, intramuscular , Intraarterial, transdermal, nasal or inhaled.
- the present invention provides a skin regeneration or wrinkle improvement of the mesenchymal stem cell culture.
- the present invention provides a cosmetic composition for preventing hair loss or promoting hair growth comprising the mesenchymal stem cell culture.
- the present invention provides a pharmaceutical composition for treating hair loss or promoting hair growth, including the mesenchymal stem cell culture.
- the mesenchymal stem cell culture, cosmetic compositions and pharmaceutical compositions are as described above.
- the term "anti-hair loss” means preventing and suppressing the hair loss phenomenon
- hair loss treatment means to improve or alleviate the symptoms of hair loss.
- the hair loss refers to a phenomenon in which the hair falls off from the scalp, or a condition in which the hair is genital or thin, and includes all the symptoms classified as hair loss in the art, which are indirect and indirect causes of its occurrence.
- the hair loss may include all symptoms of hair loss due to poor blood circulation, excessive secretion of sebum by hormones, deterioration of scalp function by peroxides or bacteria, aging, genetic factors, stress, and a combination of these effects.
- hair growth used in the present invention includes not only promoting the production of new hair, but also allowing the existing hair to grow healthy, and is a broad concept including another term used in the art, meaning promotion of wool or hair growth. .
- Human and animal hair will repeat the hair cycle consisting of growing, degenerative and resting periods.
- the effect of delaying the transition from the growth stage of the hair to the degenerative (inhibiting the retardation or extending the growth phase) and the effect of promoting the transition from the resting phase to the growth (growth induction effect ) were evaluated.
- Human hair has different hair cycles of individual hairs, but in the case of mice, all hairs are initially in the same hair cycle, and it is useful for experiments to see changes in hair cycles by artificially adjusting the hair cycles in the same way.
- the C57BL / 6 mouse used in the embodiment of the present invention has the characteristic that the skin color is determined by the amount of melanin pigment in the hair follicle because the melanocytes making the pigment are not present in the epidermis but only in the hair follicle. Since the melanin pigment synthesis in hair follicles only occurs during the growing season, the skin color becomes black during the growing season, and the skin color becomes pink during the degenerative and resting periods when the melanin pigment is not synthesized. Using such characteristics, there is an advantage in that the hair cycle through the skin color can be checked without a skin biopsy. In C57BL / 6 mice, most hairs are at rest and their skin color becomes pink at 4-7 weeks of age. In addition, by removing and removing the hair at the same time, all the hair can be converted to the growth at the same time.
- hairs of the C57BL / 6 mice in the resting state are removed to induce the growth period, and the hair growth is performed by treating the degenerative induction drugs such as dexamethasone (dexamethasone) and a test substance.
- dexamethasone dexamethasone
- a test substance a test substance
- the level and number of hair follicles were observed to assess the degree of retrograde inhibition.
- the mesenchymal stem cell culture solution treatment group has hair loss prevention and hair growth promoting effects as shown in FIGS. 9 to 14.
- the hair growth level and the number of hair follicles are observed by cutting the hairs of the back of the C57BL / 6 mouse in the resting state and treating the test substance. The degree was evaluated. As a result, as can be seen from the comparison of the results of Figs. 15 to 19, it was found that the mesenchymal stem cell culture solution treatment group has a hair growth promoting effect.
- the present invention has revealed that the mesenchymal stem cell culture of the present invention can contribute to hair loss prevention, hair loss treatment, and hair growth promotion through an animal model.
- the present invention provides a method for treating hair loss, comprising administering to a subject a pharmaceutical composition comprising the mesenchymal stem cell culture.
- the present invention provides a use for preventing hair loss, treating hair loss, or promoting hair growth of the mesenchymal stem cell culture.
- the total protein content present in the stem cell culture obtained by various cell inoculation densities was investigated by BCA measurement.
- Human amniotic fluid derived cells (ANGIOCRINE, cat. # HAmnio-01) were inoculated in culture vessels at densities of 5,000, 10,000, 15,000, 20,000, and 25,000 cells / cm 2 , respectively, for 120 hours.
- the total protein content of the stem cell cultures obtained by serum culture was measured. Specific culture conditions and culture process are as follows.
- the culture solution collected in the above process was quantified protein concentration using BCA quantification.
- the specific experimental procedure is as follows.
- protein was extracted from 7 ml of culture. Fill up to 20 ml of 1X PBS (GIBCO, NY, USA), then concentrate the sample by centrifugation for 4 min at 4000 rpm for 4 minutes at 4 ° C using a 20 ml Vivaspin 3K filter (GE Healthcare, Chalfont St. Giles, UK). I was. Concentration proceeded by continuously adding 1 x PBS. When concentrated to about 1 mL, using a 500 ⁇ l Vivaspin 3K filter (GE Healthcare, Chalfont St. Giles, UK) was centrifuged four times for 30 minutes at 14,000 rpm at 4 °C, concentrated to 300 ⁇ l.
- BSA Bovine serum albumin
- the total protein content present in the stem cell culture solution according to the serum-free culture time from inoculation of human amniotic fluid-derived stem cells to the culture medium was collected by BCA measurement. Protein concentration was measured for stem cell cultures obtained by inoculating cells at a density of 20,000 cells / cm 2 in a culture vessel and serum-free cultivation, every 12 hours from 72 hours to 144 hours. Cell type, culture method and protein concentration measurement method were performed in the same manner as in Example 1, and the serum-free incubation time was different.
- the total protein content in the stem cell culture obtained at the time of 72 hours from 84 hours, 96 hours, 108 hours and 120 hours was 26.934 ⁇ g / ml, 28.647 ⁇ g / ml, 37.674 ⁇ g, respectively.
- / Ml, 38.400 ⁇ g / ml, 44.520 ⁇ g / ml was confirmed to increase.
- the total protein content in the stem cell culture obtained when the serum-free incubation time was 132 hours and 144 hours beyond 120 hours was 41.166 ⁇ g / ml and 39.610 ⁇ g / ml, respectively, rather than the stem cell culture obtained at 120 hours. It was confirmed that the decrease (Fig. 2).
- the serum-free incubation time was about 120 hours, it was confirmed that the total content of the total protein secreted by the stem cells.
- Optimal storage conditions were established by comparing the survival rate of stem cells according to cryopreservation conditions of mesenchymal stem cells, specifically cryopreservation liquids.
- 1 ml of cell banking media mixed with 10% DMSO, 20% FBS, and 70% cDMEM medium is added together with 1x10 6 stem cells in a freezing vial- It is stored in a liquid nitrogen tank at 196 ° C., together with various freezing to further improve the storage and cell viability of mesenchymal stem cells and the productivity of stem cell secreted proteins obtained through serum-free culture by thawing the stored stem cells.
- the stock solutions were compared.
- Cryopreservatives are CRYO-GOLD (Revive Organtech, Cat. # 10003), CRYO-ROS (Revive Organtech, Cat. # 10002), STEM-CELL BANKER (Zenoaq, Cat. # BLC-3) and CellFreezer (Genenmed, Cat. # GEN-1000-050) 4 types were used.
- the specific experimental method is as follows.
- the frozen cell tubes were taken out, and dissolved in a 37 ° C. constant temperature water bath while shaking for about 2 minutes. After about 90% of the frozen cells were dissolved, the cells were transferred to a 15 ml tube containing 10 ml growth medium. After centrifugation at 1,000 rpm for 5 minutes, the supernatant was removed and 10 ml growth medium was added to the cell pellet. Then, 20 ⁇ l of cell solution was added to a 96-well plate, and 20 ⁇ l of 0.4% trypan blue was mixed and pipetted well. Then, a cover glass was placed on a hemocytometer and 10 ⁇ l of the mixed solution was added to each gap. The microscope was then magnified 40 times and cell number was measured. At this time, the dead cells stained blue were counted first, and the living cells not stained were counted. Four sections of the grid were counted and the number of viable cells in one grid was averaged. The total live cell count was calculated according to the following formula.
- Living cells Number of living cells in 1 cell of the grid x 2 x 10 4 x Cell solution volume
- Cell viability number of living cells / (number of living cells + number of dead cells) x 100
- mesenchymal stem cells stored in a liquid nitrogen tank for 2 weeks by the known cryopreservation method showed an average cell survival rate of 84.41%.
- Mesenchymal stem cells stored at -80 ° C for 2 weeks with CRYO-GOLD, CRYO-ROS, STEM-CELL BANKER and CellFreezer were thawed and averaged 92.25%, 89.15%, 91.92% and It was confirmed that the cell survival rate of 86.55%, the remaining three cell preservatives except for the CellFreezer showed excellent cell viability compared to the known cryopreservation method (Fig. 3).
- the total protein content in the stem cell cultures harvested after 120 hours serum-free culture after inoculating the leaf stem cells at a density of 20,000 cells / cm 2 was 51.030 ⁇ g / ml, 46.229 ⁇ g / ml, 33.803 ⁇ g / ml and 20.477, respectively. At ⁇ g / ml, it was confirmed that the most protein is secreted from the stem cells stored in CRYO-GOLD solution (Fig. 4).
- the total protein content present in the stem cell culture medium obtained under known serum-free culture conditions and cryopreservation conditions and the stem cell culture medium obtained under optimal serum-free culture conditions and cryopreservation conditions established through the present invention It was investigated and compared by BCA measurement.
- the culture was obtained three times at 120 hour intervals.
- Existing culture medium was inoculated with 10% DMSO + 20% FBS + 70% cDMEM mixed medium in -196 °C liquid nitrogen tank in mesenchymal stem cells in a culture vessel at a density of 10,000 cells / cm 2 , and then serum-free. Stem cell cultures collected at hours, 144 hours, and 216 hours, respectively.
- the new culture medium contained 20,000 cells / cm 2 in mesenchymal stem cells stored in the culture vessel with CRYO-GOLD solution in -80 ° C cryogenic freezer. After inoculation with a serum-free culture means a stem cell culture solution collected at 120 hours, 240 hours, 360 hours respectively.
- the mesenchymal stem cells stored in the conventional cryopreservation method (-196 °C, 10% DMSO + 20% FBS + 70% cDMEM) in a serum-free culture condition inoculated in a culture vessel at a density of 10,000 cells / cm 2
- the total protein content in the stem cell cultures obtained at 72 hours, 144 hours and 216 hours was 4.187 ⁇ g / ml, 4.52 ⁇ g / ml and 2.686 ⁇ g / ml, respectively. It was measured at the ⁇ g / ml level.
- mesenchymal stem cells stored in the newly established cryopreservation method (-80 °C, CRYO-GOLD) were inoculated at 20,000 cells / cm 2 in the same culture vessel at 120 hours, 240 hours, 360
- the total protein content in the stem cell culture obtained at time was 44.52 ⁇ g / ml, 35.69 ⁇ g / ml and 28.74 ⁇ g / ml on average 36.3 ⁇ g / ml, it was confirmed that the protein content was significantly increased (Fig. 5 ).
- binding protein was detected using fluorescence and read by a scanner to quantify the degree of fluorescence.
- the scanned protein array data was quantified by the degree of expression of each protein through data analysis.
- Figure 6 shows a photograph of the scanned antibody array data, the analysis results are shown in Table 2 below.
- Proteins produced in the mesenchymal stem cell culture and storage conditions established through Examples 1 to 5 were quantitatively analyzed.
- human amniotic fluid-derived stem cell cultures obtained under previously known serum-free culture and cryopreservation conditions and human amniotic fluid-derived stem cell cultures obtained under optimal serum-free culture and cryopreservation conditions established through the present invention The content of growth factor was investigated and compared by microarray quantitative analysis.
- Existing culture medium was inoculated with 10% DMSO + 20% FBS + 70% cDMEM mixed medium in -196 °C liquid nitrogen tank at 10 cells / cm 2 inoculated into the culture vessel at a density of 10,000 cells / cm 2 .
- Mean culture medium obtained by mixing the stem cell culture solution obtained at the time, 144 hours and 216 hours, respectively, the new culture medium is 20,000 cells in the culture vessel containing mesenchymal stem cells that were stored with CRYO-GOLD solution in -80 °C cryogenic freezer It refers to a culture solution in which the stem cell culture solution obtained at 120 hours, 240 hours and 360 hours by inoculation at a density of / cm 2 and serum-free culture, respectively.
- microarray used Raybiotech Quantibody Human Growth Factor Array 1 (Cat. # QAH-GF-1).
- the specific experimental method is as follows.
- the slide glass was taken out from Quantibody Human Cytokine Antibody Array Q1000 (RayBiotech, Inc.) and dried at room temperature. Subsequently, cytokine dilutions were prepared at seven different concentrations. Cytokine standard solution or solution was then added to each well of the slide and left at room temperature for 1 to 2 hours, after which the added solution was removed and washed with a wash solution five times. Then, 80 ⁇ l of the antibody complex solution was added to each well and incubated for 1-2 hours. After incubation, the washing solution was washed twice with washing solution, and the washing solution was completely removed.
- the sample was analyzed using a microarray laser scanner, and the cytokine content of the sample was calculated by comparison with the standard solution detection amount.
- human amniotic fluid-derived mesenchymal stem cells that were stored in a conventional cryopreservation method (-196 ° C, 10% DMSO + 20% FBS + 70% cDMEM) were stored in a culture vessel for 10,000 cells / The newly established cryopreservation method of 23 growth factors, which were not detected in the culture medium mixed with the stem cell cultures obtained at 72 hours, 144 hours, and 216 hours in the serum-free culture condition inoculated at a cm 2 density.
- the fat-derived mesenchymal stem cell culture obtained in the known serum-free culture conditions and cryopreservation conditions and the fat-derived mesenchymal stem cells obtained under the optimal serum-free culture conditions and cryopreservation conditions established through the present invention The growth factor content present in the culture was investigated and compared by microarray quantification.
- the sample was inoculated with a 10 mL DMSO + 20% FBS + 70% cDMEM mixed medium in -196 °C liquid nitrogen tank, and then inoculated at a density of 10,000 cells / cm 2 into the culture vessel.
- a culture medium containing the stem cell culture solution obtained at 72 hours, 144 hours, and 216 hours, respectively, by serum-free culture and the new culture medium is a fat-derived mesenchyme stored with CRYO-GOLD solution in -80 °C cryogenic freezer. It refers to a culture medium in which stem cells were inoculated at a density of 20,000 cells / cm 2 in a culture vessel, and then serum-free cultured to obtain stem cell cultures obtained at 120 hours, 240 hours, and 360 hours, respectively.
- the microarray used a Raybiotech Quantibody Human Growth Factor Array 1 (Cat. # QAH-GF-1). Experimental method is the same as the method performed on the human amniotic fluid-derived mesenchymal stem cells.
- fat-derived mesenchymal stem cells stored in a conventional cryopreservation method were 10,000 cells / cm in the culture vessel.
- the newly established cryopreservation method of 6 growth factors among the growth factors not detected in the mixture of stem cell cultures obtained at 72 hours, 144 hours, and 216 hours in serum-free inoculated at 2 densities (- Stem cells obtained at 120 hours, 240 hours, and 360 hours in serum-free culture conditions in which fat-derived mesenchymal stem cells stored at 80 ° C., CRYO-GOLD) were inoculated at the density of 20,000 cells / cm 2 in the same culture vessel.
- the culture solution was present in the mixed culture solution.
- the growth factor content was increased by at least 133% to 8,833% for the remaining 12 growth factors among the total 18 growth factors (Table 4).
- the mass production method of the mesenchymal stem cell-derived protein established in the present invention was confirmed to have an effect of increasing the amount of protein secreted from mesenchymal stem cells or secreting new proteins as compared to the conventional method.
- Example 8 Optimal Serum free Under cultivation and storage conditions Obtained Absolute amount of collagen content in stem cell culture
- Existing culture medium was inoculated with 10% DMSO + 20% FBS + 70% cDMEM mixed medium in -196 °C liquid nitrogen tank at 10 cells / cm 2 inoculated into the culture vessel at a density of 10,000 cells / cm 2 .
- Mean culture medium obtained by mixing the stem cell culture solution obtained at the time, 144 hours and 216 hours, respectively, the new culture medium is 20,000 cells in the culture vessel containing mesenchymal stem cells that were stored with CRYO-GOLD solution in -80 °C cryogenic freezer It refers to a culture solution in which the stem cell culture solution obtained at 120 hours, 240 hours, and 360 hours by inoculation at a density of / cm 2 and serum-free culture, respectively.
- the protein of the sample solution was quantified using the BCA quantification method, and 100 ⁇ g of the sample solution was lyophilized.
- the dried sample was dissolved in 25 ⁇ g of 6 M urea and then reacted at 90 ° C. for 20 minutes.
- 25 ⁇ l of a 0.2 M ammonium bicarbonate solution dissolved in 10 mM ethylenediaminetetraacetic acid, 4% sodium dodecyl sulfate, and 6 M urea buffer solution was added and reacted at 37 ° C. for 30 minutes, followed by acylamide / bisacylamide (40% v / v 29: 1) to gel the sample.
- the dried sample was dissolved in 50 mM sodium phosphate, pH 7.3, 100 ⁇ l, 5 ⁇ l of 1 M dithiothreitol was added and mixed well, followed by reaction at 55 ° C. for 30 minutes.
- the dried sample was then mixed well with 100 ⁇ l of 100 mM ammonium bicarbonate solution, and then 2 ⁇ l of trypsin (1 mg / ml) was added, and the reaction was performed at 37 ° C. for about 18 hours. Then extracted with 200 ⁇ l of 50 mM ammonium bicarbonate solution, 200 ⁇ l of 0.1% (v / v) trifluoroacetic acid, 100 ⁇ l of 0.1% trifluoroacetic acid dissolved in acetonitrile, and 100 ⁇ l of acetonitrile solution for 1 hour each It was. After freezing the extracted sample was completely dried and reacted at 80 °C for 30 minutes to proceed with the decomposition of ammonium bicarbonate.
- the sample was then taken into a column of Oasis SPE (Waters) and desalted using vacuum and dried thoroughly.
- the standard protein peptide was dissolved in 1 ml (400 nM) of 0.1% formic acid to prepare a standard protein peptide solution. Then, 2 ⁇ l of the standard protein peptide protein solution was added to the sample subjected to trypsin hydrolysis (final concentration 10 nM), and then 98 ⁇ l of 0.1% formic acid was added to prepare a final 100 ⁇ l and subjected to mass spectrometry. It was. The conditions at this time are as follows.
- the result obtained by the mass spectrometry is ProteinLynx Global Server (PLGS) Ver. Protein identification and absolute quantification were performed using 3.0. Protein identification was performed using the Human Database (Ver. 3.87) of the International Protein Index, and absolute quantification was performed based on mass value information of standard BSA (SwissProt P2769).
- the stem cell culture solution obtained under the optimal serum-free culture and storage conditions of the present invention had an increase of about 230% collagen content compared to the conventional culture solution (Table 6).
- CCD-98sk human fibroblast cells
- cytotoxicity was not induced at the sample concentration of 0 to 0.5%.
- Standard was set to the concentration of the sample solution for intracellular collagen production test, 0.04% of the test content of adenosine (Adenosine), a functional test material for wrinkle improvement was used as a positive control material.
- the mesenchymal stem cell culture of the present invention not only contains a large amount of collagen, but also has an effect of inducing collagen production of fibroblasts, and thus it can be seen that it is effective in skin regeneration and wrinkle improvement.
- the wound healing efficacy of the mesenchymal stem cell culture obtained in the known serum-free culture and storage conditions and the mesenchymal stem cell culture obtained in the optimal serum-free culture and storage conditions established through the present invention were investigated and compared.
- Human fibroblasts were inoculated with 7.5 x 10 5 cells in a culture vessel, wounded using a microtip, and measured 3 mm each of the culture medium and the new culture medium immediately after the wound was measured by photographing. The treatment was taken and photographed again after 12 hours to close the wound.
- the stem cell culture obtained in the optimal serum-free culture and storage conditions of the present invention showed a wound healing effect about 79% higher than the conventional culture (Fig. 8).
- test animals were treated with a depilatory agent to induce the growth phase at rest, and growth was inhibited by repeated transdermal administration of 0.1% dexamethasone (dexamethanone) for 5 days from 8 days after depilation to further inhibit hair growth. Then, in order to compare the growth promoting effect of the test substance containing 3% of the mesenchymal stem cell culture of the present invention, G1 (distilled water control), G2 (Placebo control), G3 (test substance) and G4 (positive control group)-Minoxidil 5%) was repeated transdermally for 8 days to compare the growth promoting effect with the test substance.
- G1 distilled water control
- G2 Placebo control
- G3 test substance
- G4 positive control group-Minoxidil 5%
- G1 distilled water control group
- G2 Placebo control group
- G3 test substance
- G4 positive control group (5% Minoxidil)
- G1 distilled water control group
- G2 Placebo control group
- G3 test substance
- G4 positive control group (5% Minoxidil)
- the test substance containing the human mesenchymal stem cell culture solution established through the present invention at the same time inducing growth inhibition by administering 0.1% dexamethasone to the growth hair C57BL / 6 mouse model for 5 days under these test conditions.
- 0.1% dexamethasone As a result of repeated daily transdermal administration, it was confirmed that the hair growth was significantly superior to the positive control group, Minoxidil.
- G1 distilled water control
- G2 Placebo control
- G3 test substance
- G4 positive control-Minoxidil 5%
- G1 distilled water control group
- G2 Placebo control group
- G3 test substance
- G4 positive control group (5% Minoxidil)
- G1 distilled water control group
- G2 Placebo control group
- G3 test substance
- G4 positive control group (5% Minoxidil)
- G1 distilled water control group
- G2 Placebo control group
- G3 test substance
- G4 positive control group (5% Minoxidil)
- test material containing the human mesenchymal stem cell culture solution established through the present invention in the growing hair C57BL / 6 mouse model under the present conditions for 14 days repeated transdermal administration and excellent effect on promoting skin growth I could confirm that there is.
- Existing culture medium refers to a culture medium in which the stem cell cultures obtained at 72 hours, 144 hours, and 216 hours after inoculating fat-derived mesenchymal stem cells in a culture vessel at a density of 10,000 cells / cm 2 and then serum-free were cultured.
- the new culture medium refers to a culture solution obtained by inoculating a culture vessel with a density of 20,000 cells / cm 2 and then culturing serum-free and mixing stem cell culture solutions obtained at 120 hours, 240 hours, and 360 hours, respectively.
- the optimal serum-free culture conditions to maximize the content of human mesenchymal stem cell secreted protein established through the present invention is not only for human amniotic mesenchymal stem cells but also human adipose-derived mesenchymal stem cells It could be confirmed that the application is possible.
- Example 14 Bone Marrow Origin Mesenchyme Optimal in Stem Cell Cultures Serum free Culture condition verification
- Human bone marrow-derived mesenchymal stem cells (Human Bone Marrow-Derived Mesenchymal Stem Cell, SCIENCELL, Cat. # 7500) were used to determine whether the optimal serum-free culture conditions of the present invention are applicable to mesenchymal stem cells derived from other tissues.
- the total protein content in the stem cell culture obtained by culturing each of the two serum-free culture conditions was investigated and compared by BCA measurement.
- Existing culture medium means a culture medium in which bone marrow-derived mesenchymal stem cells were inoculated at a density of 10,000 cells / cm 2 , and then serum-free cultured and mixed with the stem cell cultures collected at 72 hours, 144 hours, and 216 hours.
- New culture medium refers to a culture solution in which the stem cell culture solution collected at 120 hours, 240 hours, and 360 hours, respectively, after inoculation at a density of 20,000 cells / cm 2 in a culture vessel, and serum-free.
- the optimal serum-free culture conditions for maximizing the content of human mesenchymal stem cell secreted protein established through the present invention is not only for human amniotic mesenchymal-derived mesenchymal stem cells but also for human bone marrow-derived mesenchymal stem cells. It could be confirmed that the application is possible.
- Example 15 Cord blood origin Mesenchyme Optimal in Stem Cell Cultures Serum free Culture condition verification
- human umbilical cord blood-derived mesenchymal stem cells Human Umblical Mesenchymal Stem Cell, SCIENCELL, Cat. # 7530
- SCIENCELL Human Umblical Mesenchymal Stem Cell
- Existing culture medium refers to a culture medium in which umbilical cord blood-derived mesenchymal stem cells were inoculated at a density of 10,000 cells / cm 2 , and then serum-free cultured and mixed with the stem cell cultures collected at 72 hours, 144 hours, and 216 hours, respectively.
- the new culture medium refers to a culture solution obtained by inoculating a culture vessel at a density of 20,000 cells / cm 2 and then culturing serum-free and mixing stem cell cultures collected at 120 hours, 240 hours, and 360 hours, respectively.
- the optimal serum-free culture conditions for maximizing the content of human mesenchymal stem cell secreted protein established through the present invention is not only for human amniotic fluid-derived mesenchymal stem cells but also for human umbilical cord blood-derived mesenchymal stem cells. It could be confirmed that the application is possible.
- Example 16 Amniotic Origin Mesenchyme Optimal in Stem Cell Cultures Serum free Culture condition verification
- human amnion-derived mesenchymal stem cells Human Amniotic Mesenchymal Stem Cell, SCIENCELL, Cat. # 7501
- SCIENCELL Human Amniotic Mesenchymal Stem Cell
- Existing culture medium refers to a culture medium in which the amnion-derived mesenchymal stem cells were inoculated in a culture vessel at a density of 10,000 cells / cm 2 and then serum-free cultured and mixed with the stem cell cultures collected at 72 hours, 144 hours and 216 hours.
- the culture medium refers to a culture solution in which the stem cell culture solution was collected at 120 hours, 240 hours and 360 hours by inoculating a culture vessel at a density of 20,000 cells / cm 2, and then serum-free.
- the serum-free culture conditions in which human amnion-derived mesenchymal stem cells were inoculated in a culture vessel at a density of 10,000 cells / cm 2 existed in the culture medium containing the stem cell culture medium obtained at 72 hours, 144 hours and 216 hours, respectively.
- the total protein content was 27.269 ⁇ g / mL, and stem cells obtained at 120 hours, 240 hours and 360 hours, respectively, in serum-free culture conditions inoculated with human amnion derived mesenchymal stem cells at a density of 20,000 cells / cm 2 in the same culture vessel.
- the total protein content present in the culture medium mixed with the culture solution was 48.379 ⁇ g / ml was confirmed to improve the protein content of more than 77% (Fig. 23).
- the optimal serum-free culture conditions for maximizing the content of human mesenchymal stem cell secreted protein established through the present invention are applicable to human amniotic membrane-derived mesenchymal stem cells as well as human amniotic membrane-derived mesenchymal stem cells. This could be confirmed.
- the optimal serum-free culture conditions established in the present invention are methods for mass-producing mesenchymal stem cell-derived proteins, as well as amniotic fluid-derived mesenchymal stem cells, fat-derived, bone marrow-derived, cord blood-derived and amniotic membranes. Since it can be applied to the derived mesenchymal stem cells, any mesenchymal stem cells can be applied regardless of their origin, and the mesenchymal stem cell culture obtained through this contains a large amount of various growth factors and cytokines to improve skin regeneration and wrinkles. It can be used as a composition for.
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Genetics & Genomics (AREA)
- Biochemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Gastroenterology & Hepatology (AREA)
- Toxicology (AREA)
- Engineering & Computer Science (AREA)
- Biophysics (AREA)
- Molecular Biology (AREA)
- Biomedical Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Pharmacology & Pharmacy (AREA)
- Animal Behavior & Ethology (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Wood Science & Technology (AREA)
- Immunology (AREA)
- Epidemiology (AREA)
- Developmental Biology & Embryology (AREA)
- General Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Dermatology (AREA)
- Hematology (AREA)
- Virology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Cosmetics (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
Description
Vial | Volume of Diluent (㎕) | Volume and Source of BSA (㎕) | Final BSA Concentration (㎍/㎖) |
A | 0 | 300 of Stock | 2000 |
B | 125 | 375 of Stock | 1500 |
C | 325 | 325 of Stock | 1000 |
D | 175 | 175 of vial B dilution | 750 |
E | 325 | 325 of vial B dilution | 500 |
F | 325 | 325 of vial B dilution | 250 |
G | 325 | 325 of vial B dilution | 125 |
H | 400 | 100 of vial B dilution | 25 |
I | 400 | 0 | 0 = Blank |
순번 | 성장인자 | 기존 ( pg /㎖) | 신규 ( pg /㎖) | 증가율 ( % ) |
1 | AR | 0.0 | 104.7 | 새롭게 발현 |
2 | BDNF | 2.0 | 7.5 | 275 |
3 | bFGF | 0.0 | 84.35 | 새롭게 발현 |
4 | BMP-4 | 41.0 | 237.85 | 480 |
5 | BMP-5 | 0.0 | 983.2 | 새롭게 발현 |
6 | BMP-7 | 0.0 | 251.25 | 새롭게 발현 |
7 | b-NGF | 2.0 | 14.5 | 625 |
8 | EGF R | 112.0 | 1243.8 | 1011 |
9 | FGF-4 | 131.0 | 213.2 | 63 |
10 | FGF-7 | 5.0 | 407.45 | 8049 |
11 | GDF-15 | 1.0 | 336 | 33500 |
12 | GDNF | 2.0 | 50.05 | 2403 |
13 | GH | 0.0 | 48.9 | 새롭게 발현 |
14 | HGF | 7.0 | 1757.95 | 25014 |
15 | IGFBP-1 | 0.0 | 132.4 | 새롭게 발현 |
16 | IGFBP-2 | 0.0 | 54 | 새롭게 발현 |
17 | IGFBP-3 | 0.0 | 43691.75 | 새롭게 발현 |
18 | IGFBP-4 | 0.0 | 4612.25 | 새롭게 발현 |
19 | IGFBP-6 | 2090.0 | 40427.8 | 1834 |
20 | IGF-I | 85.0 | 128.2 | 51 |
21 | Insulin | 46.0 | 253.35 | 451 |
22 | MCSF R | 26.0 | 117.65 | 353 |
23 | NGF R | 18.0 | 46.6 | 159 |
24 | NT-3 | 58.0 | 104.45 | 80 |
25 | NT-4 | 3.0 | 16.8 | 460 |
26 | OPG | 6.0 | 2946.7 | 49012 |
27 | PDGF-AA | 32.0 | 373.9 | 1068 |
28 | PIGF | 6.0 | 223.25 | 3621 |
29 | SCF | 0.0 | 58.55 | 새롭게 발현 |
30 | SCF R | 156.0 | 433.75 | 178 |
31 | TGFa | 0.0 | 23 | 새롭게 발현 |
32 | TGFb1 | 0.0 | 2350.1 | 새롭게 발현 |
33 | VEGF | 116.0 | 4483.7 | 3765 |
34 | VEGF R3 | 0.0 | 37.6 | 새롭게 발현 |
35 | VEGF-D | 0.0 | 33.9 | 새롭게 발현 |
36 | G-CSF | 15 | 587 | 3913 |
37 | ICAM-1 | 0 | 17298 | 새롭게 발현 |
38 | IL-1a | 0 | 25 | 새롭게 발현 |
39 | IL-2 | 5 | 34 | 680 |
40 | IL-5 | 2 | 16 | 새롭게 발현 |
41 | IL-6 | 2 | 5528 | 276400 |
42 | IL-8 | 4 | 473 | 11825 |
43 | IL-11 | 323 | 14643 | 4533 |
44 | MCP-1 | 233 | 4256 | 1827 |
45 | MCSF | 589 | 1816 | 308 |
46 | MIG | 35 | 114 | 326 |
47 | MIP-1a | 0 | 7436 | 새롭게 발현 |
48 | MIP-1b | 0 | 193 | 새롭게 발현 |
49 | MIP-1d | 0 | 131 | 새롭게 발현 |
50 | RANTES | 0 | 141 | 새롭게 발현 |
51 | TIMP-1 | 87405 | 119191 | 136 |
52 | TIMP-2 | 102452 | 288596 | 282 |
53 | TNFa | 12 | 43 | 358 |
54 | TNFb | 26 | 95 | 365 |
55 | TNF R1 | 0 | 5334 | 새롭게 발현 |
56 | TNF RII | 0 | 658 | 새롭게 발현 |
순번 | 성장인자 | 기존 | 신규 | 증가율 (%) |
1 | bFGF | 6.0 | 14.0 | 133.0 |
2 | EGF R | 187.0 | 606.0 | 224.0 |
3 | FGF-4 | 44.0 | 340.0 | 673.0 |
4 | FGF-7 | 0.0 | 159.0 | 새롭게 발현 |
5 | GDF-15 | 1.0 | 23.0 | 2,200.0 |
6 | HGF | 3.0 | 268.0 | 8,833.0 |
7 | IGFBP-1 | 0.0 | 7.0 | 새롭게 발현 |
8 | IGFBP-3 | 0.0 | 55224.0 | 새롭게 발현 |
9 | IGFBP-4 | 252.0 | 2537.0 | 907.0 |
10 | IGFBP-6 | 994.0 | 52997.0 | 5,232.0 |
11 | MCSF R | 0.0 | 54.0 | 새롭게 발현 |
12 | NT-3 | 11.0 | 55.0 | 400.0 |
13 | NT-4 | 0.0 | 18.0 | 새롭게 발현 |
14 | OPG | 134.0 | 1855.0 | 1,284.0 |
15 | PDGF-AA | 7.0 | 253.0 | 3,514.0 |
16 | PIGF | 2.0 | 55.0 | 2,650.0 |
17 | TGFb1 | 0.0 | 553.0 | 새롭게 발현 |
18 | VEGF | 591.0 | 4546.0 | 669.0 |
시간 (분) | A(% v/v) | B(% v/v) | |
1 | 초기 | 97.0 | 3.0 |
2 | 5 | 97.0 | 3.0 |
3 | 300 | 65.0 | 35.0 |
4 | 320 | 20.0 | 80.0 |
5 | 340 | 20.0 | 80.0 |
6 | 355 | 97.0 | 3.0 |
7 | 360 | 97.0 | 3.0 |
기존 | 신규 | |
총 검출된 단백질 (㎍) | 21.94 | 25.12 |
총 검출된 콜라겐 (㎍) | 1.18 | 3.12 |
콜라겐 함량 (%) | 5.42 | 12.4 |
모발 생장 점수 (성별: 암컷) | |||||||||
그룹 | 동물 ID | 점수 | 평균 | S.D. | 총 평균 | 총 S.D. | |||
1차 | 2차 | 3차 | 4차 | ||||||
G1 | 0101 | 2 | 2 | 3 | 4 | 2.75 | 0.96 | 1.70 | 1.01 |
0102 | 1 | 1 | 1 | 2 | 1.25 | 0.50 | |||
0103 | 1 | 2 | 2 | 3 | 2.00 | 0.82 | |||
0104 | 1 | 1 | 0 | 2 | 1.00 | 0.82 | |||
0105 | 3 | 3 | 2 | 4 | 3.00 | 0.82 | |||
0106 | 1 | 2 | 1 | 3 | 1.75 | 0.96 | |||
0107 | 0 | 0 | 0 | 0 | 0.00 | 0.00 | |||
0108 | 3 | 3 | 2 | 4 | 3.00 | 0.82 | |||
0109 | 1 | 1 | 1 | 3 | 1.50 | 1.00 | |||
0110 | 0 | 1 | 0 | 2 | 0.75 | 0.96 | |||
G2 | 0201 | 0 | 1 | 0 | 1 | 0.50 | 0.58 | 1.45 | 1.13 |
0202 | 0 | 1 | 0 | 1 | 0.50 | 0.58 | |||
0203 | 2 | 1 | 2 | 2 | 1.75 | 0.50 | |||
0204 | 2 | 2 | 2 | 3 | 2.25 | 0.50 | |||
0205 | 2 | 1 | 1 | 3 | 1.75 | 0.96 | |||
0206 | 0 | 0 | 0 | 0 | 0.00 | 0.00 | |||
0207 | 4 | 2 | 3 | 4 | 3.25 | 0.96 | |||
0208 | 4 | 2 | 2 | 4 | 3.00 | 1.15 | |||
0209 | 0 | 0 | 0 | 2 | 0.50 | 1.00 | |||
0210 | 1 | 0 | 0 | 3 | 1.00 | 1.41 | |||
G3 | 0301 | 4 | 4 | 4 | 4 | 4.00 | 0.00 | 3.50**/## | 0.37 |
0302 | 3 | 2 | 4 | 4 | 3.25 | 0.96 | |||
0303 | 3 | 2 | 4 | 4 | 3.25 | 0.96 | |||
0304 | 4 | 2 | 4 | 4 | 3.50 | 1.00 | |||
0305 | 4 | 3 | 4 | 4 | 3.75 | 0.50 | |||
0306 | 4 | 2 | 4 | 4 | 3.50 | 1.00 | |||
0307 | 3 | 2 | 4 | 3 | 3.00 | 0.82 | |||
0308 | 4 | 4 | 4 | 4 | 4.00 | 0.00 | |||
0309 | 4 | 3 | 4 | 4 | 3.75 | 0.50 | |||
0310 | 3 | 3 | 3 | 3 | 3.00 | 0.00 | |||
G4 | 0401 | 2 | 2 | 4 | 4 | 3.00 | 1.15 | 2.68** | 0.67 |
0402 | 2 | 2 | 4 | 4 | 3.00 | 1.15 | |||
0403 | 2 | 2 | 4 | 4 | 3.00 | 1.15 | |||
0404 | 1 | 1 | 2 | 3 | 1.75 | 0.96 | |||
0405 | 1 | 2 | 3 | 3 | 2.25 | 0.96 | |||
0406 | 2 | 2 | 3 | 4 | 2.75 | 0.96 | |||
0407 | 3 | 2 | 2 | 4 | 2.75 | 0.96 | |||
0408 | 2 | 1 | 1 | 3 | 1.75 | 0.96 | |||
0409 | 2 | 2 | 2 | 4 | 2.50 | 1.00 | |||
0410 | 4 | 4 | 4 | 4 | 4.00 | 0.00 |
모낭 수 (성별: 암컷) | ||||||||
그룹 | 동물 ID | 수 측정 | 평균 | S.D. | 총 평균 | 총 S.D. | ||
영역 1 | 영역 2 | 영역 3 | ||||||
G1 | 0101 | 73 | 22 | 37 | 44.00 | 26.21 | 54.37 | 23.48 |
0102 | 47 | 33 | 0 | 26.67 | 24.13 | |||
0103 | 61 | 74 | 82 | 72.33 | 10.60 | |||
0104 | 28 | 55 | 37 | 40.00 | 13.75 | |||
0105 | 66 | 98 | 89 | 84.33 | 16.50 | |||
0106 | 60 | 84 | 113 | 85.67 | 26.54 | |||
0107 | 16 | 4 | 19 | 13.00 | 7.94 | |||
0108 | 47 | 62 | 58 | 55.67 | 7.77 | |||
0109 | 82 | 70 | 75 | 75.67 | 60.3 | |||
0110 | 45 | 51 | 43 | 46.33 | 4.16 | |||
G2 | 0201 | 1 | 13 | 13 | 9.00 | 6.93 | 53.87 | 40.05 |
0202 | 35 | 27 | 30 | 30.67 | 4.04 | |||
0203 | 87 | 67 | 63 | 72.33 | 12.86 | |||
0204 | 88 | 72 | 36 | 65.33 | 26.63 | |||
0205 | 85 | 108 | 77 | 90.00 | 16.09 | |||
0206 | 18 | 13 | 5 | 12.00 | 6.56 | |||
0207 | 124 | 110 | 106 | 113.33 | 9.45 | |||
0208 | 140 | 127 | 74 | 113.67 | 34.96 | |||
0209 | 28 | 9 | 8 | 15.00 | 11.27 | |||
0210 | 13 | 24 | 15 | 17.33 | 5.86 | |||
G3 | 0301 | 138 | 136 | 135 | 136.33 | 1.53 | 105.33**/## | 15.52 |
0302 | 81 | 120 | 99 | 100.00 | 19.52 | |||
0303 | 86 | 92 | 115 | 97.67 | 15.31 | |||
0304 | 95 | 117 | 103 | 105.00 | 11.14 | |||
0305 | 112 | 100 | 109 | 107.00 | 6.24 | |||
0306 | 124 | 112 | 126 | 120.67 | 7.57 | |||
0307 | 91 | 85 | 88 | 88.00 | 3.00 | |||
0308 | 156 | 72 | 84 | 104.00 | 45.43 | |||
0309 | 92 | 87 | 59 | 79.33 | 17.79 | |||
0310 | 110 | 124 | 118 | 117.33 | 7.02 | |||
G4 | 0401 | 70 | 96 | 97 | 87.67 | 15.31 | 66.37 | 27.22 |
0402 | 97 | 87 | 93 | 92.33 | 5.03 | |||
0403 | 66 | 65 | 82 | 71.00 | 9.54 | |||
0404 | 53 | 37 | 19 | 36.33 | 17.01 | |||
0405 | 114 | 108 | 103 | 108.33 | 5.51 | |||
0406 | 65 | 52 | 67 | 61.33 | 8.14 | |||
0407 | 78 | 37 | 21 | 45.33 | 29.40 | |||
0408 | 46 | 19 | 24 | 29.67 | 14.36 | |||
0409 | 37 | 48 | 23 | 36.00 | 12.53 | |||
0410 | 95 | 87 | 105 | 95.67 | 9.02 |
모발 생장 점수 (성별: 암컷) | |||||||||
그룹 | 동물 ID | 점수 | 평균 | S.D. | 총 평균 | 총 S.D. | |||
1차 | 2차 | 3차 | 4차 | ||||||
G1 | 0101 | 0 | 0 | 0 | 2 | 0.67 | 1.00 | 1.47 | 1.00 |
0102 | 3 | 2 | 3 | 3 | 2.67 | 0.50 | |||
0103 | 0 | 0 | 0 | 2 | 0.67 | 1.00 | |||
0104 | 3 | 3 | 3 | 4 | 3.33 | 0.50 | |||
0105 | 1 | 0 | 1 | 3 | 1.33 | 1.26 | |||
0106 | 1 | 0 | 1 | 3 | 1.33 | 1.26 | |||
0107 | 1 | 0 | 0 | 2 | 0.67 | 0.96 | |||
0108 | 0 | 0 | 0 | 1 | 0.33 | 0.50 | |||
0109 | 3 | 2 | 2 | 3 | 2.33 | 0.58 | |||
0110 | 2 | 0 | 1 | 3 | 1.33 | 1.29 | |||
G2 | 0201 | 2 | 1 | 3 | 4 | 2.67 | 1.29 | 1.33 | 0.92 |
0202 | 3 | 2 | 3 | 4 | 3.00 | 0.82 | |||
0203 | 1 | 0 | 1 | 3 | 1.33 | 1.26 | |||
0204 | 0 | 0 | 0 | 3 | 1.00 | 1.50 | |||
0205 | 0 | 0 | 0 | 2 | 0.67 | 1.00 | |||
0206 | 1 | 0 | 0 | 3 | 1.00 | 1.41 | |||
0207 | 2 | 1 | 2 | 3 | 2.00 | 0.82 | |||
0208 | 0 | 0 | 0 | 1 | 0.33 | 0.50 | |||
0209 | 0 | 0 | 0 | 2 | 0.67 | 1.00 | |||
0210 | 0 | 0 | 0 | 2 | 0.67 | 1.00 | |||
G3 | 0301 | 3 | 3 | 4 | 4 | 3.67 | 0.58 | 2.53**/## | 0.98 |
0302 | 0 | 0 | 1 | 3 | 1.33 | 1.41 | |||
0303 | 2 | 2 | 3 | 3 | 2.67 | 0.58 | |||
0304 | 1 | 1 | 2 | 3 | 2.00 | 0.96 | |||
0305 | 2 | 1 | 3 | 4 | 2.67 | 1.29 | |||
0306 | 4 | 4 | 4 | 4 | 4.00 | 0.00 | |||
0307 | 4 | 4 | 3 | 4 | 3.67 | 0.50 | |||
0308 | 2 | 1 | 3 | 3 | 2.33 | 0.96 | |||
0309 | 1 | 1 | 1 | 3 | 1.67 | 1.00 | |||
0310 | 1 | 0 | 1 | 3 | 1.33 | 1.26 | |||
G4 | 0401 | 3 | 3 | 4 | 4 | 3.67 | 0.58 | 3.27** | 0.58 |
0402 | 4 | 4 | 4 | 4 | 4.00 | 0.00 | |||
0403 | 2 | 3 | 4 | 3 | 3.33 | 0.82 | |||
0404 | 3 | 3 | 4 | 4 | 3.67 | 0.58 | |||
0405 | 2 | 3 | 3 | 3 | 3.00 | 0.50 | |||
0406 | 3 | 3 | 3 | 4 | 3.33 | 0.50 | |||
0407 | 4 | 4 | 4 | 4 | 4.00 | 0.00 | |||
0408 | 3 | 1 | 3 | 3 | 2.33 | 1.00 | |||
0409 | 2 | 2 | 3 | 3 | 2.67 | 0.58 | |||
0410 | 2 | 1 | 4 | 3 | 2.67 | 1.29 |
모발 생장 점수 (성별: 암컷) | |||||||||
그룹 | 동물 ID | 점수 | 평균 | S.D. | 총 평균 | 총 S.D. | |||
1차 | 2차 | 3차 | 4차 | ||||||
G1 | 0101 | 1 | 1 | 3 | 3 | 2 | 1.15 | 3.05 | 1.07 |
0102 | 4 | 4 | 4 | 4 | 4 | 0.00 | |||
0103 | 1 | 1 | 3 | 2 | 1.75 | 0.96 | |||
0104 | 4 | 4 | 4 | 4 | 4 | 0.00 | |||
0105 | 2 | 2 | 3 | 3 | 2.5 | 0.58 | |||
0106 | 4 | 4 | 4 | 4 | 4 | 0.00 | |||
0107 | 3 | 3 | 4 | 4 | 3.5 | 0.58 | |||
0108 | 1 | 1 | 1 | 2 | 1.25 | 0.50 | |||
0109 | 4 | 4 | 4 | 4 | 4 | 0.00 | |||
0110 | 4 | 3 | 4 | 3 | 3.5 | 0.58 | |||
G2 | 0201 | 4 | 3 | 4 | 4 | 3.75 | 0.50 | 2.80 | 0.90 |
0202 | 4 | 4 | 4 | 4 | 4 | 0.00 | |||
0203 | 2 | 2 | 4 | 3 | 2.75 | 0.96 | |||
0204 | 1 | 1 | 3 | 2 | 1.75 | 0.96 | |||
0205 | 1 | 1 | 2 | 2 | 1.5 | 0.58 | |||
0206 | 2 | 2 | 3 | 4 | 2.75 | 0.96 | |||
0207 | 4 | 4 | 4 | 4 | 4 | 0.00 | |||
0208 | 1 | 1 | 3 | 3 | 2 | 1.15 | |||
0209 | 2 | 2 | 4 | 3 | 2.75 | 0.96 | |||
0210 | 2 | 2 | 4 | 3 | 2.75 | 0.96 | |||
G3 | 0301 | 4 | 4 | 4 | 4 | 4 | 0.00 | 3.60**/## | 0.46 |
0302 | 2 | 3 | 3 | 3 | 2.75 | 0.50 | |||
0303 | 4 | 4 | 4 | 4 | 4 | 0.00 | |||
0304 | 4 | 4 | 4 | 4 | 4 | 0.00 | |||
0305 | 4 | 3 | 4 | 4 | 3.75 | 0.50 | |||
0306 | 4 | 3 | 4 | 4 | 3.75 | 0.50 | |||
0307 | 4 | 4 | 4 | 4 | 4 | 0.00 | |||
0308 | 4 | 3 | 4 | 3 | 3.5 | 0.58 | |||
0309 | 3 | 2 | 4 | 3 | 3 | 0.82 | |||
0310 | 3 | 2 | 4 | 4 | 3.25 | 0.96 | |||
G4 | 0401 | 4 | 4 | 4 | 4 | 4 | 0.00 | 3.78** | 0.22 |
0402 | 4 | 3 | 4 | 4 | 3.75 | 0.50 | |||
0403 | 3 | 4 | 4 | 3 | 3.5 | 0.58 | |||
0404 | 4 | 3 | 4 | 3 | 3.5 | 0.58 | |||
0405 | 4 | 3 | 4 | 4 | 3.75 | 0.50 | |||
0406 | 4 | 3 | 4 | 3 | 3.5 | 0.58 | |||
0407 | 4 | 4 | 4 | 4 | 4 | 0.00 | |||
0408 | 4 | 3 | 4 | 4 | 3.75 | 0.50 | |||
0409 | 4 | 4 | 4 | 4 | 4 | 0.00 | |||
0410 | 4 | 4 | 4 | 4 | 4 | 0.00 |
모낭 수 (성별: 암컷) | ||||||||
그룹 | 동물 ID | 수 측정 | 평균 | S.D. | 총 평균 | 총 S.D. | ||
영역 1 | 영역 2 | 영역 3 | ||||||
G1 | 0101 | 108 | 86 | 115 | 103.00 | 15.13 | 87.80 | 26.77 |
0102 | 87 | 121 | 119 | 109.00 | 19.08 | |||
0103 | 57 | 74 | 76 | 69.00 | 10.44 | |||
0104 | 129 | 137 | 124 | 130.00 | 6.56 | |||
0105 | 91 | 87 | 105 | 94.33 | 9.45 | |||
0106 | 40 | 51 | 48 | 46.33 | 5.69 | |||
0107 | 112 | 120 | 100 | 110.67 | 10.07 | |||
0108 | 26 | 47 | 61 | 44.67 | 17.62 | |||
0109 | 78 | 73 | 76 | 75.67 | 2.52 | |||
0110 | 31 | 106 | 149 | 95.33 | 59.72 | |||
G2 | 0201 | 114 | 104 | 132 | 116.67 | 14.19 | 103.57 | 23.18 |
0202 | 133 | 130 | 136 | 133.00 | 3.00 | |||
0203 | 132 | 130 | 154 | 138.67 | 13.32 | |||
0204 | 124 | 105 | 117 | 115.33 | 9.61 | |||
0205 | 111 | 79 | 137 | 109.00 | 29.05 | |||
0206 | 104 | 98 | 84 | 95.33 | 10.26 | |||
0207 | 113 | 98 | 107 | 106.00 | 7.55 | |||
0208 | 73 | 91 | 94 | 86.00 | 11.36 | |||
0209 | 66 | 71 | 64 | 67.00 | 3.61 | |||
0210 | 75 | 56 | 75 | 68.67 | 10.97 | |||
G3 | 0301 | 145 | 116 | 92 | 117.67 | 26.54 | 130.63**/## | 28.45 |
0302 | 135 | 121 | 104 | 120.00 | 15.52 | |||
0303 | 167 | 156 | 171 | 164.67 | 7.77 | |||
0304 | 143 | 133 | 215 | 163.67 | 44.74 | |||
0305 | 164 | 172 | 142 | 159.33 | 15.53 | |||
0306 | 166 | 167 | 108 | 147.00 | 33.78 | |||
0307 | 139 | 102 | 138 | 126.33 | 21.08 | |||
0308 | 91 | 45 | 60 | 65.33 | 23.46 | |||
0309 | 129 | 125 | 103 | 119.00 | 14.00 | |||
0310 | 98 | 158 | 114 | 123.33 | 31.07 | |||
G4 | 0401 | 232 | 209 | 133 | 191.33 | 51.81 | 104.70 | 44.25 |
0402 | 48 | 135 | 140 | 107.67 | 51.73 | |||
0403 | 92 | 55 | 60 | 69.00 | 20.07 | |||
0404 | 135 | 155 | 159 | 149.67 | 12.86 | |||
0405 | 45 | 50 | 69 | 54.67 | 12.66 | |||
0406 | 90 | 94 | 64 | 82.67 | 16.29 | |||
0407 | 71 | 79 | 67 | 72.33 | 6.11 | |||
0408 | 149 | 126 | 116 | 130.33 | 16.92 | |||
0409 | 121 | 164 | 131 | 138.67 | 22.50 | |||
0410 | 53 | 41 | 58 | 50.67 | 8.74 |
Claims (14)
- (a) 중간엽 줄기세포를 18,000 내지 22,000 세포/cm2의 밀도로 접종하는 단계;(b) 상기 중간엽 줄기세포를 무혈청 배지에서 배양하는 단계; 및(c) 배양 114 내지 126 시간 경과 후 줄기세포 배양액을 수득하는 단계를 포함하는 중간엽 줄기세포 유래 단백질의 대량 생산방법으로,상기 중간엽 줄기세포는 -90 ℃ 내지 -70 ℃ 초저온 냉동고에서 CRYO-GOLD 용액으로 보관한 것이고,상기 단백질은 AR, bFGF, BMP-5, BMP-7, GH, IGFBP-1, IGFBP-2, IGFBP-3, IGFBP-4, SCF, TGFα, TGFβ1, VEGF R3, VEGF-D, ICAM-1, IL-1a, IL-5, MIP-1a, MIP-b, MIP-d, RANTES, TNF R1, 및 TNF RII를 포함하는 것인, 생산방법.
- 제1항에 있어서, 상기 중간엽 줄기세포는 양수, 지방, 골수, 제대혈, 또는 양막 유래 중간엽 줄기세포인 것인, 생산방법.
- 제1항에 있어서, 상기 중간엽 줄기세포는 양수 유래 중간엽 줄기세포인 것인, 생산방법.
- 제1항에 있어서, 상기 단백질은 BDNF, BMP-4, b-NGF, EGF R, FGF-4, FGF-7, GDF-15, GDNF, HGF, IGFBP-6, IGF-I, Insulin, MCSF R, NGF R, NT-3, NT-4, OPG, PDGF-AA, PIGF, SCF R, VEGF, G-CSF, IL-2, IL-6, IL-8, IL-11, MCP-1, MCSF, MIG, TIMP-1, TIMP-2, TNFα, 및 TNFβ를 추가로 포함하는 것인, 생산방법.
- 제1항에 있어서, 상기 단계 (c)는 114 내지 126 시간 간격으로 1 회 내지 3 회 줄기세포 배양액을 수득하는 것인, 생산방법.
- 제1항에 있어서, 상기 생산방법은 분비 단백질의 총 농도가 BCA 측정법으로 30㎍/㎖ 내지 70㎍/㎖인 것인, 생산방법.
- (a) 중간엽 줄기세포를 20,000 세포/cm2의 밀도로 접종하는 단계;(b) 상기 중간엽 줄기세포를 무혈청 배지에서 배양하는 단계; 및(c) 배양 120 시간 경과 후 줄기세포 배양액을 수득하는 단계를 포함하는 중간엽 줄기세포 유래 단백질의 대량 생산방법으로,상기 중간엽 줄기세포는 -80 ℃ 초저온 냉동고에서 CRYO-GOLD 용액으로 보관한 것이고,상기 단백질은 AR, bFGF, BMP-5, BMP-7, GH, IGFBP-1, IGFBP-2, IGFBP-3, IGFBP-4, SCF, TGFα, TGFβ1, VEGF R3, VEGF-D, ICAM-1, IL-1a, IL-5, MIP-1a, MIP-b, MIP-d, RANTES, TNF R1, 및 TNF RII를 포함하는 것인, 생산방법.
- 제1항 내지 제7항 중 어느 한 항의 생산방법으로 생산된 단백질을 포함하는, 중간엽 줄기세포 배양액.
- 제8항에 있어서, 상기 배양액은 콜라겐을 10 내지 15 중량%로 함유하는 것인, 중간엽 줄기세포 배양액.
- 제8항에 있어서, 상기 배양액은 섬유아세포의 콜라겐 생성을 촉진시키는 것인, 중간엽 줄기세포 배양액.
- 제8항의 중간엽 줄기세포 배양액을 포함하는 피부 재생 또는 주름 개선용 화장료 조성물.
- 제8항의 중간엽 줄기세포 배양액을 포함하는 피부 재생용 약학 조성물.
- 제8항의 중간엽 줄기세포 배양액을 포함하는 탈모 방지 또는 발모 촉진용 화장료 조성물.
- 제8항의 중간엽 줄기세포 배양액을 포함하는 탈모 치료 또는 발모 촉진용 약학 조성물.
Priority Applications (4)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US15/562,382 US11077145B2 (en) | 2015-04-03 | 2016-04-01 | Method for mass producing proteins in mesenchymal stem cells |
JP2018503447A JP6609034B2 (ja) | 2015-04-03 | 2016-04-01 | 間葉幹細胞でのタンパク質量産方法 |
EP16773502.6A EP3279212A4 (en) | 2015-04-03 | 2016-04-01 | Method for mass producing proteins in mesenchymal stem cells |
HK18103531.0A HK1244017A1 (zh) | 2015-04-03 | 2018-03-13 | 在間充質幹細胞中批量生產蛋白質的方法 |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
KR10-2015-0047774 | 2015-04-03 | ||
KR1020150047774A KR101566450B1 (ko) | 2015-04-03 | 2015-04-03 | 중간엽 줄기세포에서 단백질의 대량 생산 방법 |
Publications (1)
Publication Number | Publication Date |
---|---|
WO2016159721A1 true WO2016159721A1 (ko) | 2016-10-06 |
Family
ID=54600764
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/KR2016/003421 WO2016159721A1 (ko) | 2015-04-03 | 2016-04-01 | 중간엽 줄기세포에서 단백질의 대량 생산 방법 |
Country Status (6)
Country | Link |
---|---|
US (1) | US11077145B2 (ko) |
EP (1) | EP3279212A4 (ko) |
JP (1) | JP6609034B2 (ko) |
KR (1) | KR101566450B1 (ko) |
HK (1) | HK1244017A1 (ko) |
WO (1) | WO2016159721A1 (ko) |
Cited By (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN108236614A (zh) * | 2016-12-23 | 2018-07-03 | 吟斯蔻碧株式会社 | 用于促进生发的生物移植用移植物 |
WO2020036418A1 (ko) * | 2018-08-17 | 2020-02-20 | 고려대학교 산학협력단 | 태반유래 세포 조건화 배지에서의 모낭세포 기능강화 및 대량생산 방법 |
US10758571B1 (en) | 2019-04-09 | 2020-09-01 | Combangio, Inc. | Processes for making and using a mesenchymal stem cell derived secretome |
CN111712227A (zh) * | 2017-12-14 | 2020-09-25 | 绿十字生命健康有限公司 | 用于缓解特应性皮炎、脱发和创伤或者减少皮肤皱纹的化妆品组合物和药物组合物 |
Families Citing this family (9)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN106191127B (zh) * | 2016-02-22 | 2020-01-14 | 华南生物医药研究院 | 干细胞生物活性组合物及其制备方法与应用 |
KR102018609B1 (ko) * | 2017-04-26 | 2019-09-06 | 이장호 | 중간엽 줄기세포 배양액을 포함하는 세포 또는 조직의 동결보존용 조성물 |
KR102323056B1 (ko) * | 2019-05-02 | 2021-11-09 | 에스씨엠생명과학 주식회사 | hPL 함유 배지에서 배양된 중간엽 줄기세포의 배양액을 포함하는 화장료 조성물 |
CN112972493A (zh) * | 2019-12-12 | 2021-06-18 | 郝志号 | 一种抗衰老剂的制备方法及其应用 |
CN114081861B (zh) * | 2021-11-19 | 2023-10-31 | 和泓尚医(成都)生物科技有限公司 | 一种干细胞外泌体与复合肽抗衰制剂及其制备方法和应用 |
CN114480267A (zh) * | 2022-01-21 | 2022-05-13 | 济南万泉生物技术有限公司 | TGF-β2与Val混合刺激因子促进间充质干细胞表达更多细胞外基质 |
CN114672455A (zh) * | 2022-03-25 | 2022-06-28 | 中山大学 | 一种利用多能干细胞诱导骨髓基质细胞的方法 |
CN115287259B (zh) * | 2022-08-02 | 2024-04-05 | 南京生航生物技术有限公司 | 一种培养脂肪间充质干细胞专用无血清培养基及其应用 |
CN115466724A (zh) * | 2022-10-21 | 2022-12-13 | 唐颐控股(深圳)有限公司 | 一种用于人间充质干细胞扩增中的最优因子值的筛选方法及应用 |
Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
KR20100105166A (ko) * | 2009-03-20 | 2010-09-29 | 주식회사 스템메디언스 | 양수 내 태아 유래 중간엽줄기세포를 이용한 피부 상태개선용 조성물 |
KR20100105165A (ko) * | 2009-03-20 | 2010-09-29 | 주식회사 스템메디언스 | 양수 내 태아 유래 중간엽줄기세포를 이용한 성장인자의 대량생산 방법 |
KR20130006123A (ko) * | 2011-07-08 | 2013-01-16 | 경북대학교병원 | 인간 양수줄기세포 배양배지 및 이를 이용한 양수줄기세포 배양방법 |
Family Cites Families (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6255112B1 (en) * | 1998-06-08 | 2001-07-03 | Osiris Therapeutics, Inc. | Regulation of hematopoietic stem cell differentiation by the use of human mesenchymal stem cells |
US8524494B2 (en) * | 2008-01-30 | 2013-09-03 | Histogen, Inc. | Low oxygen tension and BFGF generates a multipotent stem cell from a fibroblast in vitro |
US20110274670A1 (en) * | 2008-06-13 | 2011-11-10 | Myeong Jin Nam | Mesenchymal stem cells which express human hepatic growth factor,manufacturing method thereof, and use thereof as therapeutic agent for liver diseases |
CA2829586C (en) * | 2011-03-11 | 2021-03-02 | Children's Medical Center Corporation | Methods and compositions relating to mesenchymal stem cell exosomes |
-
2015
- 2015-04-03 KR KR1020150047774A patent/KR101566450B1/ko active IP Right Grant
-
2016
- 2016-04-01 JP JP2018503447A patent/JP6609034B2/ja active Active
- 2016-04-01 WO PCT/KR2016/003421 patent/WO2016159721A1/ko active Application Filing
- 2016-04-01 US US15/562,382 patent/US11077145B2/en active Active
- 2016-04-01 EP EP16773502.6A patent/EP3279212A4/en not_active Withdrawn
-
2018
- 2018-03-13 HK HK18103531.0A patent/HK1244017A1/zh unknown
Patent Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
KR20100105166A (ko) * | 2009-03-20 | 2010-09-29 | 주식회사 스템메디언스 | 양수 내 태아 유래 중간엽줄기세포를 이용한 피부 상태개선용 조성물 |
KR20100105165A (ko) * | 2009-03-20 | 2010-09-29 | 주식회사 스템메디언스 | 양수 내 태아 유래 중간엽줄기세포를 이용한 성장인자의 대량생산 방법 |
KR20130006123A (ko) * | 2011-07-08 | 2013-01-16 | 경북대학교병원 | 인간 양수줄기세포 배양배지 및 이를 이용한 양수줄기세포 배양방법 |
Non-Patent Citations (3)
Title |
---|
"CRYO-GOLD, Chemically defined Slow Freezing Medium", INTERNET MATERIAL, 11 March 2015 (2015-03-11), XP055319995 * |
SALAZAR, K. D. ET AL.: "Mesenchymal Stem Cells Produce Wnt Isoforms and TGF-Betal that Mediate Proliferation and Procollagen Expression by Lung Fibroblasts", AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY, vol. 297, no. 5, 2009, pages 1002 - 1011, XP055320023 * |
See also references of EP3279212A4 * |
Cited By (10)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN108236614A (zh) * | 2016-12-23 | 2018-07-03 | 吟斯蔻碧株式会社 | 用于促进生发的生物移植用移植物 |
JP2018102926A (ja) * | 2016-12-23 | 2018-07-05 | インスコビー・インコーポレイテッドInscobee,Inc. | 生体植立用インプラント及びその製造方法 |
US10993904B2 (en) | 2016-12-23 | 2021-05-04 | Inscobee Inc. | Transplantation implant for promoting hair growth |
CN108236614B (zh) * | 2016-12-23 | 2021-07-20 | 吟斯蔻碧株式会社 | 用于促进生发的生物移植用移植物 |
CN111712227A (zh) * | 2017-12-14 | 2020-09-25 | 绿十字生命健康有限公司 | 用于缓解特应性皮炎、脱发和创伤或者减少皮肤皱纹的化妆品组合物和药物组合物 |
WO2020036418A1 (ko) * | 2018-08-17 | 2020-02-20 | 고려대학교 산학협력단 | 태반유래 세포 조건화 배지에서의 모낭세포 기능강화 및 대량생산 방법 |
US10758571B1 (en) | 2019-04-09 | 2020-09-01 | Combangio, Inc. | Processes for making and using a mesenchymal stem cell derived secretome |
US10881693B2 (en) | 2019-04-09 | 2021-01-05 | Combangio, Inc. | Processes for making and using a mesenchymal stem cell derived secretome |
US11129853B2 (en) | 2019-04-09 | 2021-09-28 | Combangio, Inc. | Processes for making and using a mesenchymal stem cell derived secretome |
US11654160B2 (en) | 2019-04-09 | 2023-05-23 | Combangio, Inc. | Processes for making and using a mesenchymal stem cell derived secretome |
Also Published As
Publication number | Publication date |
---|---|
EP3279212A4 (en) | 2018-09-05 |
EP3279212A1 (en) | 2018-02-07 |
JP6609034B2 (ja) | 2019-11-20 |
JP2018512176A (ja) | 2018-05-17 |
US20180280441A1 (en) | 2018-10-04 |
KR101566450B1 (ko) | 2015-11-05 |
HK1244017A1 (zh) | 2018-07-27 |
US11077145B2 (en) | 2021-08-03 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
WO2016159721A1 (ko) | 중간엽 줄기세포에서 단백질의 대량 생산 방법 | |
EP3953327A1 (en) | Melanocortin-4 receptor agonists | |
WO2016006885A1 (ko) | 자극된 줄기세포 배양액의 발모 촉진능 및 이의 용도 | |
WO2020251181A1 (ko) | 줄기세포 유래 엑소좀 생성 촉진 및 줄기세포능 증가용 조성물 | |
WO2016108669A1 (ko) | Rgd 모티프 함유 펩티드 또는 이의 단편을 포함하는 화상 및 녹내장 치료, 피부 주름 개선 및 발모 촉진용 조성물 | |
WO2019004758A2 (ko) | 피부투과 촉진용 펩타이드가 결합된 융합 단백질을 포함하는 피부개선용 화장료 조성물 | |
WO2019168237A1 (ko) | 신규 화합물 및 이를 유효성분으로 함유하는 섬유증 또는 비알코올성 지방간염의 예방, 개선 또는 치료용 조성물 | |
WO2017179767A1 (ko) | 지방줄기세포에서 신경줄기세포, 신경세포 및 가바성 신경세포로의 분화 유도 방법, 및 인간 골수 유래 중간엽 줄기세포로부터 성장인자를 다량 분비하는 인간 줄기세포의 분화 유도 방법 | |
WO2011142514A1 (ko) | Pias3을 유효성분으로 포함하는 암 또는 면역질환의 예방 또는 치료용 조성물 | |
WO2011118954A2 (ko) | 사람 하비갑개 유래 중간엽 기질세포로부터 연골, 골, 신경세포 또는 지방세포를 분화시키는 방법 | |
WO2017047996A1 (ko) | 나노그를 도입한 양수 내 태아 유래 중간엽 줄기세포를 이용한 육모 촉진용 조성물의 제조방법 | |
WO2018199482A1 (en) | Composition for cryopreservation of cells or tissues including mesenchymal stem cell culture medium | |
WO2017082512A1 (ko) | 발모 조절 중추 세포인 모유두 세포의 증식과 생존 및 세포 활성화 촉진 작용을 지닌 기능성 조절 항체 및 이의 용도 | |
WO2018135907A1 (ko) | 슈반 세포 전구체 및 이로부터 분화된 슈반 세포의 제조 방법 | |
WO2018026212A2 (ko) | 섬유증 질환 모델의 제조방법 및 섬유증 질환 모델의 용도 | |
WO2011111914A1 (ko) | 합토글로빈 유전자가 도입된 혈관내피 전구세포 및 이를 포함하는 혈관형성 촉진용 조성물 | |
WO2016178511A1 (ko) | Kai1 폴리펩타이드 또는 이를 코딩하는 유전자를 포함하는 혈관신생 억제용 조성물 및 이의 용도 | |
WO2023277584A1 (ko) | 셀레늄 화합물을 유효성분으로 포함하는 노화세포 제거용 조성물 | |
WO2022075780A1 (ko) | 과민성 방광의 예방 또는 치료용 약학적 조성물 | |
WO2009151207A1 (ko) | 인간 간성장인자를 발현하는 중간엽 줄기세포, 그의 제조방법 및 그의 간질환 치료제로서의 용도 | |
WO2013162330A1 (ko) | 키메라 중간엽 줄기세포군, 그의 제조방법 및 편도줄기세포를 이용하여 부갑상선 호르몬을 생산하는 방법 | |
WO2022139493A1 (ko) | TGF-β 신호전달을 억제할 수 있는 신규한 펩타이드 및 이의 용도 | |
WO2020149538A1 (ko) | 클로날 줄기세포를 포함하는 아토피 피부염 예방 또는 치료용 약학적 조성물 | |
WO2014038882A1 (ko) | 신규 화합물 및 이의 인터루킨-1베타 또는 인터루킨-6 저해 용도 | |
WO2022045824A1 (en) | Phenyl alkyl carbamate compounds for use in preventing or treating neurodegenerative disease |
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: 16773502 Country of ref document: EP Kind code of ref document: A1 |
|
WWE | Wipo information: entry into national phase |
Ref document number: 15562382 Country of ref document: US |
|
REEP | Request for entry into the european phase |
Ref document number: 2016773502 Country of ref document: EP |
|
ENP | Entry into the national phase |
Ref document number: 2018503447 Country of ref document: JP Kind code of ref document: A |
|
NENP | Non-entry into the national phase |
Ref country code: DE |