KR20170113733A - Composition containing ganglioside GM3 for inducing chondrogenic differentiation, and uses thereof - Google Patents
Composition containing ganglioside GM3 for inducing chondrogenic differentiation, and uses thereof Download PDFInfo
- Publication number
- KR20170113733A KR20170113733A KR1020160035243A KR20160035243A KR20170113733A KR 20170113733 A KR20170113733 A KR 20170113733A KR 1020160035243 A KR1020160035243 A KR 1020160035243A KR 20160035243 A KR20160035243 A KR 20160035243A KR 20170113733 A KR20170113733 A KR 20170113733A
- Authority
- KR
- South Korea
- Prior art keywords
- ganglioside
- stem cells
- mesenchymal stem
- differentiation
- cell
- Prior art date
Links
- PFJKOHUKELZMLE-VEUXDRLPSA-N ganglioside GM3 Chemical compound O[C@@H]1[C@@H](O)[C@H](OC[C@@H]([C@H](O)/C=C/CCCCCCCCCCCCC)NC(=O)CCCCCCCCCCCCC\C=C/CCCCCCCC)O[C@H](CO)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@]2(O[C@H]([C@H](NC(C)=O)[C@@H](O)C2)[C@H](O)[C@H](O)CO)C(O)=O)[C@@H](O)[C@@H](CO)O1 PFJKOHUKELZMLE-VEUXDRLPSA-N 0.000 title claims abstract description 91
- 239000000203 mixture Substances 0.000 title claims abstract description 33
- 230000001939 inductive effect Effects 0.000 title claims abstract description 24
- 230000009816 chondrogenic differentiation Effects 0.000 title description 3
- 210000001612 chondrocyte Anatomy 0.000 claims abstract description 69
- 210000002901 mesenchymal stem cell Anatomy 0.000 claims abstract description 50
- 230000004069 differentiation Effects 0.000 claims abstract description 45
- 208000015100 cartilage disease Diseases 0.000 claims abstract description 21
- 238000000034 method Methods 0.000 claims abstract description 15
- 239000004480 active ingredient Substances 0.000 claims abstract description 13
- 230000036541 health Effects 0.000 claims abstract description 13
- 239000008194 pharmaceutical composition Substances 0.000 claims abstract description 13
- 235000013376 functional food Nutrition 0.000 claims abstract description 12
- 210000001258 synovial membrane Anatomy 0.000 claims description 12
- 210000003321 cartilage cell Anatomy 0.000 claims description 11
- 238000000338 in vitro Methods 0.000 claims description 7
- 238000012258 culturing Methods 0.000 claims description 4
- 230000002265 prevention Effects 0.000 claims description 3
- 230000000694 effects Effects 0.000 abstract description 22
- 230000003848 cartilage regeneration Effects 0.000 abstract description 5
- 230000001737 promoting effect Effects 0.000 abstract description 4
- 230000003449 preventive effect Effects 0.000 abstract description 2
- 230000001225 therapeutic effect Effects 0.000 abstract description 2
- 210000004027 cell Anatomy 0.000 description 45
- 229920002683 Glycosaminoglycan Polymers 0.000 description 22
- 230000014509 gene expression Effects 0.000 description 21
- 108020004414 DNA Proteins 0.000 description 17
- 230000024245 cell differentiation Effects 0.000 description 16
- 210000000845 cartilage Anatomy 0.000 description 14
- 102000055007 Cartilage Oligomeric Matrix Human genes 0.000 description 13
- 150000002270 gangliosides Chemical class 0.000 description 13
- 101710176668 Cartilage oligomeric matrix protein Proteins 0.000 description 12
- 210000001519 tissue Anatomy 0.000 description 12
- 238000004458 analytical method Methods 0.000 description 11
- 239000003814 drug Substances 0.000 description 11
- 230000002441 reversible effect Effects 0.000 description 11
- 102000004887 Transforming Growth Factor beta Human genes 0.000 description 10
- 108090001012 Transforming Growth Factor beta Proteins 0.000 description 10
- ZRKFYGHZFMAOKI-QMGMOQQFSA-N tgfbeta Chemical compound C([C@H](NC(=O)[C@H](C(C)C)NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CC(C)C)NC(=O)CNC(=O)[C@H](C)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](NC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)CCSC)C(C)C)[C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](C)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(=O)N1[C@@H](CCC1)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(C)C)C(O)=O)C1=CC=C(O)C=C1 ZRKFYGHZFMAOKI-QMGMOQQFSA-N 0.000 description 10
- 101100096242 Mus musculus Sox9 gene Proteins 0.000 description 9
- 238000002474 experimental method Methods 0.000 description 9
- 239000002609 medium Substances 0.000 description 9
- 102100036601 Aggrecan core protein Human genes 0.000 description 8
- 108010067219 Aggrecans Proteins 0.000 description 8
- 108010035532 Collagen Proteins 0.000 description 8
- 102000008186 Collagen Human genes 0.000 description 8
- 229920001436 collagen Polymers 0.000 description 8
- 238000010186 staining Methods 0.000 description 8
- 229950003937 tolonium Drugs 0.000 description 8
- HNONEKILPDHFOL-UHFFFAOYSA-M tolonium chloride Chemical compound [Cl-].C1=C(C)C(N)=CC2=[S+]C3=CC(N(C)C)=CC=C3N=C21 HNONEKILPDHFOL-UHFFFAOYSA-M 0.000 description 8
- 102100025751 Mothers against decapentaplegic homolog 2 Human genes 0.000 description 7
- 101710143123 Mothers against decapentaplegic homolog 2 Proteins 0.000 description 7
- 230000026731 phosphorylation Effects 0.000 description 7
- 238000006366 phosphorylation reaction Methods 0.000 description 7
- 238000002360 preparation method Methods 0.000 description 7
- 108090000623 proteins and genes Proteins 0.000 description 7
- 210000000130 stem cell Anatomy 0.000 description 7
- 210000001789 adipocyte Anatomy 0.000 description 6
- 229940079593 drug Drugs 0.000 description 6
- 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 6
- VZSRBBMJRBPUNF-UHFFFAOYSA-N 2-(2,3-dihydro-1H-inden-2-ylamino)-N-[3-oxo-3-(2,4,6,7-tetrahydrotriazolo[4,5-c]pyridin-5-yl)propyl]pyrimidine-5-carboxamide Chemical compound C1C(CC2=CC=CC=C12)NC1=NC=C(C=N1)C(=O)NCCC(N1CC2=C(CC1)NN=N2)=O VZSRBBMJRBPUNF-UHFFFAOYSA-N 0.000 description 5
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 5
- 239000003795 chemical substances by application Substances 0.000 description 5
- 235000013305 food Nutrition 0.000 description 5
- 108020004445 glyceraldehyde-3-phosphate dehydrogenase Proteins 0.000 description 5
- 230000006698 induction Effects 0.000 description 5
- 239000003550 marker Substances 0.000 description 5
- 201000008482 osteoarthritis Diseases 0.000 description 5
- 210000000963 osteoblast Anatomy 0.000 description 5
- 102100025748 Mothers against decapentaplegic homolog 3 Human genes 0.000 description 4
- 101710143111 Mothers against decapentaplegic homolog 3 Proteins 0.000 description 4
- 108090000526 Papain Proteins 0.000 description 4
- 229930040373 Paraformaldehyde Natural products 0.000 description 4
- 239000004365 Protease Substances 0.000 description 4
- 230000004663 cell proliferation 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
- 201000010099 disease Diseases 0.000 description 4
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 4
- 238000009472 formulation Methods 0.000 description 4
- 239000003102 growth factor Substances 0.000 description 4
- 229940055729 papain Drugs 0.000 description 4
- 235000019834 papain Nutrition 0.000 description 4
- 229920002866 paraformaldehyde Polymers 0.000 description 4
- 239000000546 pharmaceutical excipient Substances 0.000 description 4
- 102000004169 proteins and genes Human genes 0.000 description 4
- 230000001172 regenerating effect Effects 0.000 description 4
- 150000003839 salts Chemical class 0.000 description 4
- 239000000243 solution Substances 0.000 description 4
- 229940124597 therapeutic agent Drugs 0.000 description 4
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 3
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 3
- 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 3
- 238000012413 Fluorescence activated cell sorting analysis Methods 0.000 description 3
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 3
- DNIAPMSPPWPWGF-UHFFFAOYSA-N Propylene glycol Chemical compound CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 description 3
- 108091005735 TGF-beta receptors Proteins 0.000 description 3
- 244000299461 Theobroma cacao Species 0.000 description 3
- 102000016715 Transforming Growth Factor beta Receptors Human genes 0.000 description 3
- 238000009825 accumulation Methods 0.000 description 3
- 230000009471 action Effects 0.000 description 3
- SQVRNKJHWKZAKO-UHFFFAOYSA-N beta-N-Acetyl-D-neuraminic acid Natural products CC(=O)NC1C(O)CC(O)(C(O)=O)OC1C(O)C(O)CO SQVRNKJHWKZAKO-UHFFFAOYSA-N 0.000 description 3
- 210000001185 bone marrow Anatomy 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 230000006378 damage Effects 0.000 description 3
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 3
- 239000000839 emulsion Substances 0.000 description 3
- 210000004700 fetal blood Anatomy 0.000 description 3
- 239000012091 fetal bovine serum Substances 0.000 description 3
- 239000000796 flavoring agent Substances 0.000 description 3
- 235000013373 food additive Nutrition 0.000 description 3
- 239000002778 food additive Substances 0.000 description 3
- 230000006870 function Effects 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 239000012528 membrane Substances 0.000 description 3
- 108020004999 messenger RNA Proteins 0.000 description 3
- 229940049954 penicillin Drugs 0.000 description 3
- 239000002953 phosphate buffered saline Substances 0.000 description 3
- 239000000843 powder Substances 0.000 description 3
- 239000003755 preservative agent Substances 0.000 description 3
- 102000005962 receptors Human genes 0.000 description 3
- 108020003175 receptors Proteins 0.000 description 3
- 125000005629 sialic acid group Chemical group 0.000 description 3
- 229960005322 streptomycin Drugs 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 239000000725 suspension Substances 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 238000001262 western blot Methods 0.000 description 3
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 2
- CDKIEBFIMCSCBB-UHFFFAOYSA-N 1-(6,7-dimethoxy-3,4-dihydro-1h-isoquinolin-2-yl)-3-(1-methyl-2-phenylpyrrolo[2,3-b]pyridin-3-yl)prop-2-en-1-one;hydrochloride Chemical compound Cl.C1C=2C=C(OC)C(OC)=CC=2CCN1C(=O)C=CC(C1=CC=CN=C1N1C)=C1C1=CC=CC=C1 CDKIEBFIMCSCBB-UHFFFAOYSA-N 0.000 description 2
- JKYKXTRKURYNGW-UHFFFAOYSA-N 3,4-dihydroxy-9,10-dioxo-9,10-dihydroanthracene-2-sulfonic acid Chemical compound O=C1C2=CC=CC=C2C(=O)C2=C1C(O)=C(O)C(S(O)(=O)=O)=C2 JKYKXTRKURYNGW-UHFFFAOYSA-N 0.000 description 2
- 102100022464 5'-nucleotidase Human genes 0.000 description 2
- 102000007469 Actins Human genes 0.000 description 2
- 108010085238 Actins Proteins 0.000 description 2
- 208000036487 Arthropathies Diseases 0.000 description 2
- 208000020084 Bone disease Diseases 0.000 description 2
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 2
- 102000000503 Collagen Type II Human genes 0.000 description 2
- 108010041390 Collagen Type II Proteins 0.000 description 2
- 102100037241 Endoglin Human genes 0.000 description 2
- KRHYYFGTRYWZRS-UHFFFAOYSA-N Fluorane Chemical compound F KRHYYFGTRYWZRS-UHFFFAOYSA-N 0.000 description 2
- VZCYOOQTPOCHFL-OWOJBTEDSA-N Fumaric acid Chemical compound OC(=O)\C=C\C(O)=O VZCYOOQTPOCHFL-OWOJBTEDSA-N 0.000 description 2
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 2
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 2
- 102000009465 Growth Factor Receptors Human genes 0.000 description 2
- 108010009202 Growth Factor Receptors Proteins 0.000 description 2
- 102100031573 Hematopoietic progenitor cell antigen CD34 Human genes 0.000 description 2
- 241000282412 Homo Species 0.000 description 2
- 101000678236 Homo sapiens 5'-nucleotidase Proteins 0.000 description 2
- 101000881679 Homo sapiens Endoglin Proteins 0.000 description 2
- 101000777663 Homo sapiens Hematopoietic progenitor cell antigen CD34 Proteins 0.000 description 2
- 101000738771 Homo sapiens Receptor-type tyrosine-protein phosphatase C Proteins 0.000 description 2
- 101000716102 Homo sapiens T-cell surface glycoprotein CD4 Proteins 0.000 description 2
- 101000800116 Homo sapiens Thy-1 membrane glycoprotein Proteins 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- 108090000723 Insulin-Like Growth Factor I Proteins 0.000 description 2
- 208000012659 Joint disease Diseases 0.000 description 2
- MIJPAVRNWPDMOR-ZAFYKAAXSA-N L-ascorbic acid 2-phosphate Chemical compound OC[C@H](O)[C@H]1OC(=O)C(OP(O)(O)=O)=C1O MIJPAVRNWPDMOR-ZAFYKAAXSA-N 0.000 description 2
- 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 2
- 102000018697 Membrane Proteins Human genes 0.000 description 2
- 108010052285 Membrane Proteins Proteins 0.000 description 2
- PVNIIMVLHYAWGP-UHFFFAOYSA-N Niacin Chemical compound OC(=O)C1=CC=CN=C1 PVNIIMVLHYAWGP-UHFFFAOYSA-N 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
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- 108010038512 Platelet-Derived Growth Factor Proteins 0.000 description 2
- 102000010780 Platelet-Derived Growth Factor Human genes 0.000 description 2
- 238000011529 RT qPCR Methods 0.000 description 2
- 102100037422 Receptor-type tyrosine-protein phosphatase C Human genes 0.000 description 2
- 102000013275 Somatomedins Human genes 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- 102100036011 T-cell surface glycoprotein CD4 Human genes 0.000 description 2
- 102100033523 Thy-1 membrane glycoprotein Human genes 0.000 description 2
- 108010009583 Transforming Growth Factors Proteins 0.000 description 2
- 102000009618 Transforming Growth Factors Human genes 0.000 description 2
- 102000004142 Trypsin Human genes 0.000 description 2
- 108090000631 Trypsin Proteins 0.000 description 2
- 230000002159 abnormal effect Effects 0.000 description 2
- 238000002835 absorbance Methods 0.000 description 2
- 210000001188 articular cartilage Anatomy 0.000 description 2
- 238000003149 assay kit Methods 0.000 description 2
- 230000037396 body weight Effects 0.000 description 2
- 229940098773 bovine serum albumin Drugs 0.000 description 2
- 239000002775 capsule Substances 0.000 description 2
- 230000003833 cell viability Effects 0.000 description 2
- 235000019219 chocolate Nutrition 0.000 description 2
- 239000003086 colorant Substances 0.000 description 2
- 235000009508 confectionery Nutrition 0.000 description 2
- 210000004748 cultured cell Anatomy 0.000 description 2
- 231100000135 cytotoxicity Toxicity 0.000 description 2
- 230000003013 cytotoxicity Effects 0.000 description 2
- 230000003412 degenerative effect Effects 0.000 description 2
- 239000003085 diluting agent Substances 0.000 description 2
- 238000001378 electrochemiluminescence detection Methods 0.000 description 2
- -1 ethyl oleate Chemical class 0.000 description 2
- 238000011156 evaluation Methods 0.000 description 2
- 235000019634 flavors Nutrition 0.000 description 2
- 235000015203 fruit juice Nutrition 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- 102000006602 glyceraldehyde-3-phosphate dehydrogenase Human genes 0.000 description 2
- 150000002339 glycosphingolipids Chemical class 0.000 description 2
- 239000008187 granular material Substances 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 238000003119 immunoblot Methods 0.000 description 2
- CGIGDMFJXJATDK-UHFFFAOYSA-N indomethacin Chemical compound CC1=C(CC(O)=O)C2=CC(OC)=CC=C2N1C(=O)C1=CC=C(Cl)C=C1 CGIGDMFJXJATDK-UHFFFAOYSA-N 0.000 description 2
- 239000004615 ingredient Substances 0.000 description 2
- 238000002347 injection Methods 0.000 description 2
- 239000007924 injection Substances 0.000 description 2
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 2
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 2
- 239000008101 lactose Substances 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
- 238000005259 measurement Methods 0.000 description 2
- 210000004379 membrane Anatomy 0.000 description 2
- 210000003205 muscle Anatomy 0.000 description 2
- 210000000944 nerve tissue Anatomy 0.000 description 2
- 239000012457 nonaqueous media Substances 0.000 description 2
- 208000005368 osteomalacia Diseases 0.000 description 2
- 210000002826 placenta Anatomy 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 239000002994 raw material Substances 0.000 description 2
- 206010039073 rheumatoid arthritis Diseases 0.000 description 2
- 229910052711 selenium Inorganic materials 0.000 description 2
- 239000011669 selenium Substances 0.000 description 2
- SQVRNKJHWKZAKO-OQPLDHBCSA-N sialic acid Chemical compound CC(=O)N[C@@H]1[C@@H](O)C[C@@](O)(C(O)=O)OC1[C@H](O)[C@H](O)CO SQVRNKJHWKZAKO-OQPLDHBCSA-N 0.000 description 2
- 230000011664 signaling Effects 0.000 description 2
- 239000000829 suppository Substances 0.000 description 2
- 208000024891 symptom Diseases 0.000 description 2
- 239000006188 syrup Substances 0.000 description 2
- 235000020357 syrup Nutrition 0.000 description 2
- 239000003826 tablet Substances 0.000 description 2
- 239000002562 thickening agent Substances 0.000 description 2
- 239000012588 trypsin Substances 0.000 description 2
- 235000013343 vitamin Nutrition 0.000 description 2
- 239000011782 vitamin Substances 0.000 description 2
- 229940088594 vitamin Drugs 0.000 description 2
- 229930003231 vitamin Natural products 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 239000000080 wetting agent Substances 0.000 description 2
- WZUVPPKBWHMQCE-XJKSGUPXSA-N (+)-haematoxylin Chemical compound C12=CC(O)=C(O)C=C2C[C@]2(O)[C@H]1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-XJKSGUPXSA-N 0.000 description 1
- MTCFGRXMJLQNBG-REOHCLBHSA-N (2S)-2-Amino-3-hydroxypropansäure Chemical compound OC[C@H](N)C(O)=O MTCFGRXMJLQNBG-REOHCLBHSA-N 0.000 description 1
- 239000001100 (2S)-5,7-dihydroxy-2-(3-hydroxy-4-methoxyphenyl)chroman-4-one Substances 0.000 description 1
- FTVWIRXFELQLPI-ZDUSSCGKSA-N (S)-naringenin Chemical compound C1=CC(O)=CC=C1[C@H]1OC2=CC(O)=CC(O)=C2C(=O)C1 FTVWIRXFELQLPI-ZDUSSCGKSA-N 0.000 description 1
- FMKJUUQOYOHLTF-OWOJBTEDSA-N (e)-4-azaniumylbut-2-enoate Chemical compound NC\C=C\C(O)=O FMKJUUQOYOHLTF-OWOJBTEDSA-N 0.000 description 1
- WBYWAXJHAXSJNI-VOTSOKGWSA-M .beta-Phenylacrylic acid Natural products [O-]C(=O)\C=C\C1=CC=CC=C1 WBYWAXJHAXSJNI-VOTSOKGWSA-M 0.000 description 1
- LBLYYCQCTBFVLH-UHFFFAOYSA-N 2-Methylbenzenesulfonic acid Chemical compound CC1=CC=CC=C1S(O)(=O)=O LBLYYCQCTBFVLH-UHFFFAOYSA-N 0.000 description 1
- AZKSAVLVSZKNRD-UHFFFAOYSA-M 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide Chemical compound [Br-].S1C(C)=C(C)N=C1[N+]1=NC(C=2C=CC=CC=2)=NN1C1=CC=CC=C1 AZKSAVLVSZKNRD-UHFFFAOYSA-M 0.000 description 1
- BMYNFMYTOJXKLE-UHFFFAOYSA-N 3-azaniumyl-2-hydroxypropanoate Chemical compound NCC(O)C(O)=O BMYNFMYTOJXKLE-UHFFFAOYSA-N 0.000 description 1
- KYWKJKVQLGANBH-UHFFFAOYSA-N 3-butyl-1-methyl-7h-purine-2,6-dione Chemical compound O=C1N(C)C(=O)N(CCCC)C2=C1NC=N2 KYWKJKVQLGANBH-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
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Natural products OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 1
- 238000009010 Bradford assay Methods 0.000 description 1
- YDNKGFDKKRUKPY-JHOUSYSJSA-N C16 ceramide Natural products CCCCCCCCCCCCCCCC(=O)N[C@@H](CO)[C@H](O)C=CCCCCCCCCCCCCC YDNKGFDKKRUKPY-JHOUSYSJSA-N 0.000 description 1
- 101150013553 CD40 gene Proteins 0.000 description 1
- 102100032912 CD44 antigen Human genes 0.000 description 1
- 101100098985 Caenorhabditis elegans cct-3 gene Proteins 0.000 description 1
- 108700005376 Cartilage Oligomeric Matrix Proteins 0.000 description 1
- WBYWAXJHAXSJNI-SREVYHEPSA-N Cinnamic acid Chemical compound OC(=O)\C=C/C1=CC=CC=C1 WBYWAXJHAXSJNI-SREVYHEPSA-N 0.000 description 1
- 102000029816 Collagenase Human genes 0.000 description 1
- 108060005980 Collagenase Proteins 0.000 description 1
- FCKYPQBAHLOOJQ-UHFFFAOYSA-N Cyclohexane-1,2-diaminetetraacetic acid Chemical compound OC(=O)CN(CC(O)=O)C1CCCCC1N(CC(O)=O)CC(O)=O FCKYPQBAHLOOJQ-UHFFFAOYSA-N 0.000 description 1
- 102000004127 Cytokines Human genes 0.000 description 1
- 108090000695 Cytokines Proteins 0.000 description 1
- 102000053602 DNA Human genes 0.000 description 1
- 101710088194 Dehydrogenase Proteins 0.000 description 1
- FEWJPZIEWOKRBE-JCYAYHJZSA-N Dextrotartaric acid Chemical compound OC(=O)[C@H](O)[C@@H](O)C(O)=O FEWJPZIEWOKRBE-JCYAYHJZSA-N 0.000 description 1
- 206010058314 Dysplasia Diseases 0.000 description 1
- 102000001301 EGF receptor Human genes 0.000 description 1
- 108060006698 EGF receptor Proteins 0.000 description 1
- LVGKNOAMLMIIKO-UHFFFAOYSA-N Elaidinsaeure-aethylester Natural products CCCCCCCCC=CCCCCCCCC(=O)OCC LVGKNOAMLMIIKO-UHFFFAOYSA-N 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 1
- 239000004606 Fillers/Extenders Substances 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- 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 1
- 239000004471 Glycine Substances 0.000 description 1
- WZUVPPKBWHMQCE-UHFFFAOYSA-N Haematoxylin Natural products C12=CC(O)=C(O)C=C2CC2(O)C1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-UHFFFAOYSA-N 0.000 description 1
- 101000868273 Homo sapiens CD44 antigen Proteins 0.000 description 1
- 108010001336 Horseradish Peroxidase Proteins 0.000 description 1
- 102000004877 Insulin Human genes 0.000 description 1
- 108090001061 Insulin Proteins 0.000 description 1
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- SQVRNKJHWKZAKO-PFQGKNLYSA-N N-acetyl-beta-neuraminic acid Chemical compound CC(=O)N[C@@H]1[C@@H](O)C[C@@](O)(C(O)=O)O[C@H]1[C@H](O)[C@H](O)CO SQVRNKJHWKZAKO-PFQGKNLYSA-N 0.000 description 1
- CRJGESKKUOMBCT-VQTJNVASSA-N N-acetylsphinganine Chemical compound CCCCCCCCCCCCCCC[C@@H](O)[C@H](CO)NC(C)=O CRJGESKKUOMBCT-VQTJNVASSA-N 0.000 description 1
- 108010025020 Nerve Growth Factor Proteins 0.000 description 1
- 102000015336 Nerve Growth Factor Human genes 0.000 description 1
- GRYLNZFGIOXLOG-UHFFFAOYSA-N Nitric acid Chemical compound O[N+]([O-])=O GRYLNZFGIOXLOG-UHFFFAOYSA-N 0.000 description 1
- 239000000020 Nitrocellulose Substances 0.000 description 1
- NPGIHFRTRXVWOY-UHFFFAOYSA-N Oil red O Chemical compound Cc1ccc(C)c(c1)N=Nc1cc(C)c(cc1C)N=Nc1c(O)ccc2ccccc12 NPGIHFRTRXVWOY-UHFFFAOYSA-N 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 206010031149 Osteitis Diseases 0.000 description 1
- 229920002230 Pectic acid Polymers 0.000 description 1
- 239000004698 Polyethylene Substances 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- 239000004743 Polypropylene Substances 0.000 description 1
- 229920001213 Polysorbate 20 Polymers 0.000 description 1
- 108010067787 Proteoglycans Proteins 0.000 description 1
- 102000016611 Proteoglycans Human genes 0.000 description 1
- 239000012083 RIPA buffer Substances 0.000 description 1
- 241000700159 Rattus Species 0.000 description 1
- 102000012083 SOX9 Transcription Factor Human genes 0.000 description 1
- 108010036426 SOX9 Transcription Factor Proteins 0.000 description 1
- 229920002472 Starch Polymers 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
- FEWJPZIEWOKRBE-UHFFFAOYSA-N Tartaric acid Natural products [H+].[H+].[O-]C(=O)C(O)C(O)C([O-])=O FEWJPZIEWOKRBE-UHFFFAOYSA-N 0.000 description 1
- 244000269722 Thea sinensis Species 0.000 description 1
- 235000005764 Theobroma cacao ssp. cacao Nutrition 0.000 description 1
- 235000005767 Theobroma cacao ssp. sphaerocarpum Nutrition 0.000 description 1
- AYFVYJQAPQTCCC-UHFFFAOYSA-N Threonine Natural products CC(O)C(N)C(O)=O AYFVYJQAPQTCCC-UHFFFAOYSA-N 0.000 description 1
- 239000004473 Threonine Substances 0.000 description 1
- 102000046299 Transforming Growth Factor beta1 Human genes 0.000 description 1
- 101800002279 Transforming growth factor beta-1 Proteins 0.000 description 1
- 102100040245 Tumor necrosis factor receptor superfamily member 5 Human genes 0.000 description 1
- HSCJRCZFDFQWRP-RDKQLNKOSA-N UDP-D-glucose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)OC1OP(O)(=O)OP(O)(=O)OC[C@@H]1[C@@H](O)[C@@H](O)[C@H](N2C(NC(=O)C=C2)=O)O1 HSCJRCZFDFQWRP-RDKQLNKOSA-N 0.000 description 1
- JRMSLDWZFJZLAS-UHFFFAOYSA-M [7-(dimethylamino)-1,9-dimethylphenothiazin-3-ylidene]-dimethylazanium;chloride Chemical compound [Cl-].CC1=CC(N(C)C)=CC2=[S+]C3=CC(N(C)C)=CC(C)=C3N=C21 JRMSLDWZFJZLAS-UHFFFAOYSA-M 0.000 description 1
- ZMZINYUKVRMNTG-UHFFFAOYSA-N acetic acid;formic acid Chemical compound OC=O.CC(O)=O ZMZINYUKVRMNTG-UHFFFAOYSA-N 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 230000001464 adherent effect Effects 0.000 description 1
- 239000000443 aerosol Substances 0.000 description 1
- 150000001298 alcohols Chemical class 0.000 description 1
- 239000000783 alginic acid Substances 0.000 description 1
- 235000010443 alginic acid Nutrition 0.000 description 1
- 229920000615 alginic acid Polymers 0.000 description 1
- 229960001126 alginic acid Drugs 0.000 description 1
- 150000004781 alginic acids Chemical class 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 208000024429 articular cartilage disease Diseases 0.000 description 1
- 235000010323 ascorbic acid Nutrition 0.000 description 1
- 229960005070 ascorbic acid Drugs 0.000 description 1
- 239000011668 ascorbic acid Substances 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- SRSXLGNVWSONIS-UHFFFAOYSA-N benzenesulfonic acid Chemical compound OS(=O)(=O)C1=CC=CC=C1 SRSXLGNVWSONIS-UHFFFAOYSA-N 0.000 description 1
- 229940092714 benzenesulfonic acid Drugs 0.000 description 1
- 235000013361 beverage Nutrition 0.000 description 1
- 239000011230 binding agent Substances 0.000 description 1
- 238000012742 biochemical analysis Methods 0.000 description 1
- 230000031018 biological processes and functions Effects 0.000 description 1
- 230000033228 biological regulation Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 208000018339 bone inflammation disease Diseases 0.000 description 1
- 235000008429 bread Nutrition 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- 235000001046 cacaotero Nutrition 0.000 description 1
- 229910000019 calcium carbonate Inorganic materials 0.000 description 1
- 235000014171 carbonated beverage Nutrition 0.000 description 1
- 230000022159 cartilage development Effects 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 230000030833 cell death Effects 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 238000002659 cell therapy Methods 0.000 description 1
- 238000003570 cell viability assay Methods 0.000 description 1
- 210000002421 cell wall Anatomy 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 229940106189 ceramide Drugs 0.000 description 1
- ZVEQCJWYRWKARO-UHFFFAOYSA-N ceramide Natural products CCCCCCCCCCCCCCC(O)C(=O)NC(CO)C(O)C=CCCC=C(C)CCCCCCCCC ZVEQCJWYRWKARO-UHFFFAOYSA-N 0.000 description 1
- 235000013351 cheese Nutrition 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 235000013985 cinnamic acid Nutrition 0.000 description 1
- 229930016911 cinnamic acid Natural products 0.000 description 1
- 235000015165 citric acid Nutrition 0.000 description 1
- 229960002424 collagenase Drugs 0.000 description 1
- 239000000084 colloidal system Substances 0.000 description 1
- 238000004040 coloring Methods 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000009833 condensation Methods 0.000 description 1
- 230000005494 condensation Effects 0.000 description 1
- 239000006071 cream Substances 0.000 description 1
- 230000001086 cytosolic effect Effects 0.000 description 1
- 231100000433 cytotoxic Toxicity 0.000 description 1
- 230000001472 cytotoxic effect Effects 0.000 description 1
- 238000002784 cytotoxicity assay Methods 0.000 description 1
- 231100000263 cytotoxicity test Toxicity 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 230000001934 delay Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 235000005911 diet Nutrition 0.000 description 1
- 230000037213 diet Effects 0.000 description 1
- 239000007884 disintegrant Substances 0.000 description 1
- PXEDJBXQKAGXNJ-QTNFYWBSSA-L disodium L-glutamate Chemical compound [Na+].[Na+].[O-]C(=O)[C@@H](N)CCC([O-])=O PXEDJBXQKAGXNJ-QTNFYWBSSA-L 0.000 description 1
- 239000002552 dosage form Substances 0.000 description 1
- 239000003792 electrolyte Substances 0.000 description 1
- YQGOJNYOYNNSMM-UHFFFAOYSA-N eosin Chemical compound [Na+].OC(=O)C1=CC=CC=C1C1=C2C=C(Br)C(=O)C(Br)=C2OC2=C(Br)C(O)=C(Br)C=C21 YQGOJNYOYNNSMM-UHFFFAOYSA-N 0.000 description 1
- 230000007705 epithelial mesenchymal transition Effects 0.000 description 1
- 150000002148 esters Chemical class 0.000 description 1
- LVGKNOAMLMIIKO-QXMHVHEDSA-N ethyl oleate Chemical compound CCCCCCCC\C=C/CCCCCCCC(=O)OCC LVGKNOAMLMIIKO-QXMHVHEDSA-N 0.000 description 1
- 229940093471 ethyl oleate Drugs 0.000 description 1
- 230000005284 excitation Effects 0.000 description 1
- 230000029142 excretion Effects 0.000 description 1
- 239000000945 filler Substances 0.000 description 1
- 238000007667 floating Methods 0.000 description 1
- MHMNJMPURVTYEJ-UHFFFAOYSA-N fluorescein-5-isothiocyanate Chemical compound O1C(=O)C2=CC(N=C=S)=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 MHMNJMPURVTYEJ-UHFFFAOYSA-N 0.000 description 1
- 235000013355 food flavoring agent Nutrition 0.000 description 1
- 235000003599 food sweetener Nutrition 0.000 description 1
- 239000003205 fragrance Substances 0.000 description 1
- 239000001530 fumaric acid Substances 0.000 description 1
- 235000011087 fumaric acid Nutrition 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
- 238000012812 general test Methods 0.000 description 1
- 235000011187 glycerol Nutrition 0.000 description 1
- 229960002449 glycine Drugs 0.000 description 1
- 229930182470 glycoside Natural products 0.000 description 1
- 150000002338 glycosides Chemical class 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 229940093915 gynecological organic acid Drugs 0.000 description 1
- AIONOLUJZLIMTK-AWEZNQCLSA-N hesperetin Chemical compound C1=C(O)C(OC)=CC=C1[C@H]1OC2=CC(O)=CC(O)=C2C(=O)C1 AIONOLUJZLIMTK-AWEZNQCLSA-N 0.000 description 1
- AIONOLUJZLIMTK-UHFFFAOYSA-N hesperetin Natural products C1=C(O)C(OC)=CC=C1C1OC2=CC(O)=CC(O)=C2C(=O)C1 AIONOLUJZLIMTK-UHFFFAOYSA-N 0.000 description 1
- 235000010209 hesperetin Nutrition 0.000 description 1
- 229960001587 hesperetin Drugs 0.000 description 1
- FTODBIPDTXRIGS-UHFFFAOYSA-N homoeriodictyol Natural products C1=C(O)C(OC)=CC(C2OC3=CC(O)=CC(O)=C3C(=O)C2)=C1 FTODBIPDTXRIGS-UHFFFAOYSA-N 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 235000015243 ice cream Nutrition 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 229960000905 indomethacin Drugs 0.000 description 1
- 239000000411 inducer Substances 0.000 description 1
- 229910052500 inorganic mineral Inorganic materials 0.000 description 1
- 229940125396 insulin Drugs 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 230000003834 intracellular effect Effects 0.000 description 1
- 238000000185 intracerebroventricular administration Methods 0.000 description 1
- 238000007918 intramuscular administration Methods 0.000 description 1
- 238000001990 intravenous administration Methods 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 150000002576 ketones Chemical class 0.000 description 1
- 210000003127 knee Anatomy 0.000 description 1
- TYQCGQRIZGCHNB-JLAZNSOCSA-N l-ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(O)=C(O)C1=O TYQCGQRIZGCHNB-JLAZNSOCSA-N 0.000 description 1
- 239000004310 lactic acid Substances 0.000 description 1
- 235000014655 lactic acid Nutrition 0.000 description 1
- VMPHSYLJUKZBJJ-UHFFFAOYSA-N lauric acid triglyceride Natural products CCCCCCCCCCCC(=O)OCC(OC(=O)CCCCCCCCCCC)COC(=O)CCCCCCCCCCC VMPHSYLJUKZBJJ-UHFFFAOYSA-N 0.000 description 1
- 229940069445 licorice extract Drugs 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 229940057995 liquid paraffin Drugs 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 239000000314 lubricant Substances 0.000 description 1
- 229960003511 macrogol Drugs 0.000 description 1
- 235000019359 magnesium stearate Nutrition 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 230000035800 maturation Effects 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- WBYWAXJHAXSJNI-UHFFFAOYSA-N methyl p-hydroxycinnamate Natural products OC(=O)C=CC1=CC=CC=C1 WBYWAXJHAXSJNI-UHFFFAOYSA-N 0.000 description 1
- 239000011707 mineral Substances 0.000 description 1
- 235000010755 mineral Nutrition 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 235000013923 monosodium glutamate Nutrition 0.000 description 1
- PSZYNBSKGUBXEH-UHFFFAOYSA-N naphthalene-1-sulfonic acid Chemical compound C1=CC=C2C(S(=O)(=O)O)=CC=CC2=C1 PSZYNBSKGUBXEH-UHFFFAOYSA-N 0.000 description 1
- WGEYAGZBLYNDFV-UHFFFAOYSA-N naringenin Natural products C1(=O)C2=C(O)C=C(O)C=C2OC(C1)C1=CC=C(CC1)O WGEYAGZBLYNDFV-UHFFFAOYSA-N 0.000 description 1
- 235000007625 naringenin Nutrition 0.000 description 1
- 229940117954 naringenin Drugs 0.000 description 1
- 229930014626 natural product Natural products 0.000 description 1
- 229940053128 nerve growth factor Drugs 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- VVGIYYKRAMHVLU-UHFFFAOYSA-N newbouldiamide Natural products CCCCCCCCCCCCCCCCCCCC(O)C(O)C(O)C(CO)NC(=O)CCCCCCCCCCCCCCCCC VVGIYYKRAMHVLU-UHFFFAOYSA-N 0.000 description 1
- 239000011664 nicotinic acid Substances 0.000 description 1
- 229960003512 nicotinic acid Drugs 0.000 description 1
- 235000001968 nicotinic acid Nutrition 0.000 description 1
- 229910017604 nitric acid Inorganic materials 0.000 description 1
- 229920001220 nitrocellulos Polymers 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 239000003921 oil Substances 0.000 description 1
- 235000019198 oils Nutrition 0.000 description 1
- 239000002674 ointment Substances 0.000 description 1
- 229920001542 oligosaccharide Polymers 0.000 description 1
- 150000002482 oligosaccharides Chemical class 0.000 description 1
- 235000008390 olive oil Nutrition 0.000 description 1
- 239000004006 olive oil Substances 0.000 description 1
- 238000001543 one-way ANOVA Methods 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 235000005985 organic acids Nutrition 0.000 description 1
- 210000004409 osteocyte Anatomy 0.000 description 1
- 239000003002 pH adjusting agent Substances 0.000 description 1
- 239000012188 paraffin wax Substances 0.000 description 1
- 238000007911 parenteral administration Methods 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- LCLHHZYHLXDRQG-ZNKJPWOQSA-N pectic acid Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)O[C@H](C(O)=O)[C@@H]1OC1[C@H](O)[C@@H](O)[C@@H](OC2[C@@H]([C@@H](O)[C@@H](O)[C@H](O2)C(O)=O)O)[C@@H](C(O)=O)O1 LCLHHZYHLXDRQG-ZNKJPWOQSA-N 0.000 description 1
- 239000008188 pellet Substances 0.000 description 1
- 239000000825 pharmaceutical preparation Substances 0.000 description 1
- 239000006187 pill Substances 0.000 description 1
- 235000013550 pizza Nutrition 0.000 description 1
- 229920001223 polyethylene glycol Polymers 0.000 description 1
- 239000010318 polygalacturonic acid Substances 0.000 description 1
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 1
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 1
- 229920001155 polypropylene Polymers 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 239000001508 potassium citrate Substances 0.000 description 1
- 229960002635 potassium citrate Drugs 0.000 description 1
- QEEAPRPFLLJWCF-UHFFFAOYSA-K potassium citrate (anhydrous) Chemical compound [K+].[K+].[K+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O QEEAPRPFLLJWCF-UHFFFAOYSA-K 0.000 description 1
- 235000011082 potassium citrates Nutrition 0.000 description 1
- 230000002335 preservative effect Effects 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 230000001681 protective effect Effects 0.000 description 1
- 238000003753 real-time PCR Methods 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 238000007670 refining Methods 0.000 description 1
- 230000009703 regulation of cell differentiation Effects 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- BOLDJAUMGUJJKM-LSDHHAIUSA-N renifolin D Natural products CC(=C)[C@@H]1Cc2c(O)c(O)ccc2[C@H]1CC(=O)c3ccc(O)cc3O BOLDJAUMGUJJKM-LSDHHAIUSA-N 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 235000013580 sausages Nutrition 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 230000001953 sensory effect Effects 0.000 description 1
- 230000019491 signal transduction Effects 0.000 description 1
- 235000020183 skimmed milk Nutrition 0.000 description 1
- 235000011888 snacks Nutrition 0.000 description 1
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 1
- 229940073490 sodium glutamate Drugs 0.000 description 1
- 239000001488 sodium phosphate Substances 0.000 description 1
- 229910000162 sodium phosphate Inorganic materials 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 235000014347 soups Nutrition 0.000 description 1
- 241000894007 species Species 0.000 description 1
- UNFWWIHTNXNPBV-WXKVUWSESA-N spectinomycin Chemical compound O([C@@H]1[C@@H](NC)[C@@H](O)[C@H]([C@@H]([C@H]1O1)O)NC)[C@]2(O)[C@H]1O[C@H](C)CC2=O UNFWWIHTNXNPBV-WXKVUWSESA-N 0.000 description 1
- 239000003381 stabilizer Substances 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 238000007920 subcutaneous administration Methods 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 235000000346 sugar Nutrition 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 239000002511 suppository base Substances 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 239000003765 sweetening agent Substances 0.000 description 1
- 210000005222 synovial tissue Anatomy 0.000 description 1
- 239000000454 talc Substances 0.000 description 1
- 229910052623 talc Inorganic materials 0.000 description 1
- 239000011975 tartaric acid Substances 0.000 description 1
- 235000002906 tartaric acid Nutrition 0.000 description 1
- 235000013616 tea Nutrition 0.000 description 1
- 230000017423 tissue regeneration Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- VZCYOOQTPOCHFL-UHFFFAOYSA-N trans-butenedioic acid Natural products OC(=O)C=CC(O)=O VZCYOOQTPOCHFL-UHFFFAOYSA-N 0.000 description 1
- 239000003656 tris buffered saline Substances 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
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 1
- 230000003827 upregulation Effects 0.000 description 1
- VBEQCZHXXJYVRD-GACYYNSASA-N uroanthelone Chemical compound C([C@@H](C(=O)N[C@H](C(=O)N[C@@H](CS)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CS)C(=O)N[C@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)NCC(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H](CO)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CS)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O)C(C)C)[C@@H](C)O)NC(=O)[C@H](CO)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](NC(=O)[C@H](CC=1NC=NC=1)NC(=O)[C@H](CCSC)NC(=O)[C@H](CS)NC(=O)[C@@H](NC(=O)CNC(=O)CNC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CS)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)CNC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@H](CO)NC(=O)[C@H]1N(CCC1)C(=O)[C@H](CS)NC(=O)CNC(=O)[C@H]1N(CCC1)C(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@@H](N)CC(N)=O)C(C)C)[C@@H](C)CC)C1=CC=C(O)C=C1 VBEQCZHXXJYVRD-GACYYNSASA-N 0.000 description 1
- 235000015112 vegetable and seed oil Nutrition 0.000 description 1
- 239000008158 vegetable oil Substances 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
- 150000003722 vitamin derivatives Chemical class 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0652—Cells of skeletal and connective tissues; Mesenchyme
- C12N5/0655—Chondrocytes; Cartilage
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
-
- 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
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2200/00—Function of food ingredients
- A23V2200/30—Foods, ingredients or supplements having a functional effect on health
- A23V2200/306—Foods, ingredients or supplements having a functional effect on health having an effect on bone mass, e.g. osteoporosis prevention
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2500/00—Specific components of cell culture medium
- C12N2500/30—Organic components
- C12N2500/36—Lipids
-
- 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
- C12N2506/00—Differentiation of animal cells from one lineage to another; Differentiation of pluripotent cells
- C12N2506/13—Differentiation of animal cells from one lineage to another; Differentiation of pluripotent cells from connective tissue cells, from mesenchymal cells
- C12N2506/1346—Differentiation of animal cells from one lineage to another; Differentiation of pluripotent cells from connective tissue cells, from mesenchymal cells from mesenchymal stem cells
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Chemical & Material Sciences (AREA)
- Zoology (AREA)
- Biotechnology (AREA)
- General Health & Medical Sciences (AREA)
- Wood Science & Technology (AREA)
- Rheumatology (AREA)
- Organic Chemistry (AREA)
- Cell Biology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- Virology (AREA)
- Food Science & Technology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- Nutrition Science (AREA)
- Mycology (AREA)
- Developmental Biology & Embryology (AREA)
- Immunology (AREA)
- Polymers & Plastics (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- Epidemiology (AREA)
- Animal Behavior & Ethology (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The present invention relates to a composition for inducing differentiation of mesenchymal stem cells into chondrocytes, comprising ganglioside GM3 as an active ingredient, a method for inducing differentiation of mesenchymal stem cells into chondrocytes using the same, Or improving pharmaceutical composition and health functional food composition. The use of the composition containing ganglioside GM3 according to the present invention is excellent in the effect of inducing differentiation from mesenchymal stem cells into chondrocytes, and thus can be usefully used as a therapeutic, preventive or ameliorating agent for cartilage diseases by promoting cartilage regeneration .
Description
The present invention relates to a composition for inducing differentiation of mesenchymal stem cells into chondrocytes, comprising ganglioside GM3 as an active ingredient, a method for inducing differentiation of mesenchymal stem cells into chondrocytes using the same, Or improving pharmaceutical composition and health functional food composition.
Mesenchymal stem cells (MSCs), also referred to as multipotent stromal progenitor cells, have been implicated in the development of mesodermal systems such as chondrocytes, osteoblasts, and adipocytes And is a resource of regenerative medicine (
Gangliosides are complex glycosphingolipids containing sialic acids and are a major component of the cytoplasmic cell wall (non-patent document 8). Different cell types have various forms of gangliosides that accompany various biological processes such as cell death, cell proliferation, differentiation, surface interactions, and transmembrane signals (Non-Patent Documents 9-13). Many studies have confirmed that the type of ganglioside and its expression level are developmentally regulated and cell type specific (Non-Patent Literature 14, 15). In addition, recent research results of the present inventors have confirmed that the expression of ganglioside is closely related to various stem cells in vitro (Non-Patent Document 16-20). Interestingly, ganglioside GM3 regulates tyrosine phosphorylation of various growth factor receptors (Non-Patent Document 21-23). In particular, ganglioside GM3 induced by TGF-? 1 regulates the epithelial-mesenchymal transition (non-patent document 24).
Transforming growth factor-beta (TGF-β) superfamily (TGF-β1, 2, and 3) proteins play an important role in proliferation (Non-Patent Document 25). In addition, TGF-beta effectively upregulates various molecules involved in pre-cartilage cell formation and condensation and thus effectively induces chondrocyte differentiation (Non-Patent Document 26-28). Signaling associated with TGF- [beta] is initially internalized through serine / threonine phosphorylation of the TGF-beta receptor (TGF-betaR) after TGF-beta-R1 and TGF-beta-R2 are paired on the cell surface 29). Phosphorylated TGF-? R directly activates downstream mediator receptor-activated SMAD containing SMAD2 and SMAD3 (Non-Patent Documents 29-32).
Therefore, the present inventors confirmed that human SDMSC (hSDMSCs) can be differentiated into chondrocytes and continued the study on the effect of ganglioside GM3 on chondrocyte differentiation of the cells, and found that ganglioside GM3 inhibits the differentiation induction effect of hSDMSCs into chondrocytes The present invention has been completed.
It is an object of the present invention to provide a composition for inducing differentiation from mesenchymal stem cells into cartilage cells, which comprises ganglioside GM3 as an active ingredient.
Another object of the present invention is to provide a method for inducing differentiation from mesenchymal stem cells into chondrocytes, comprising culturing mesenchymal stem cells in vitro in the presence of ganglioside GM3.
It is still another object of the present invention to provide a pharmaceutical composition for treating or preventing cartilage diseases containing ganglioside GM3 as an active ingredient.
Another object of the present invention is to provide a health functional food composition for improving or preventing cartilage disease containing ganglioside GM3 as an active ingredient.
It is still another object of the present invention to provide a method for treating or preventing cartilage diseases, which comprises administering a composition containing glycoside GM3 as an active ingredient to a subject in need thereof.
One embodiment of the present invention provides a composition for inducing differentiation from mesenchymal stem cells into chondrocytes, comprising ganglioside GM3 as an active ingredient.
As used herein, the term "ganglioside " is a type of glycosphingolipid, which is an amphoteric substance composed of a hydrophilic sugar moiety and a hydrophobic ceramide, N-acetylneuraminic acid (sialic acid). Gangliosides of up to 100 kinds are now known as a combination of the number of sialic acids and their binding sites, with the diversity of neutral skeletal oligosaccharides except sialic acid. M, D, and T are mono, di and trisialo, and a, b, and c are bonded to a Gal at the reducing end of GM1, and G, G, And the number of sialic acids that are being used.
The ganglioside is located in an outer cell membrane that is in direct contact with the extracellular environment and regulates cell proliferation and differentiation. Regulation of cell differentiation depends on many extracellular and intracellular factor controls. Changes in the ganglioside expression pattern are observed during cell differentiation and in response to exposure to some cytokines and growth factors. Exogenous gangliosides have been shown to respond to exposure to several growth factors and to produce platelet-derived growth factor (PDGF) receptors, epidermal growth factor (EGF) receptors, insulin-like growth factor (IGF) nerve growth factor) regulates the function of the growth factor receptor such as the receptor.
As used herein, the term "ganglioside GM3" is named II3NeuAc-LacCer according to the IUPAC-IUB nomenclature and may be represented, for example,
[Chemical Formula 1]
The ganglioside GM3 can be obtained by a method generally known in the art or commercially available.
The present invention is characterized in that ganglioside GM3 has a function of promoting the induction of differentiation from mesenchymal stem cells into cartilage cells.
The ganglioside GM3 may be treated with a concentration of 0.1 to 1000 μM and may be treated with a concentration of 0.1 to 100 μM, more specifically a concentration of 0.1 to 10 μM, more specifically a concentration of 1 to 10 μM The mesenchymal stem cells can be treated with mesenchymal stem cells at a concentration of, for example, 2 to 5 [mu] M, but the present invention is not limited thereto. Concentrations having an inducing effect can be selected by an ordinary person skilled in the art.
As used herein, the term "active ingredient" refers to a component that exerts the effects of a drug or quasi-drug. In the present invention, it refers to a component that exerts a chondrocyte differentiation inducing effect.
Mesenchymal Stem Cells (MSCs) are one of stem cells collected from bone marrow and umbilical cord blood, and it is known that there are about 1 million stem cells in the body. Mesenchymal stem cells (MSCs) differentiate into mesenchymal cells such as chondrocytes, osteoblasts and adipocytes, thus providing cellular resources for cartilage regeneration. In addition, MSCs are easy to separate without significant donor-site morbidity and are easy to cultivate in vitro . MSCs are known to secrete a variety of growth factors, and some growth factors regulate immune function control activity.
The mesenchymal stem cells may be mesenchymal stem cells derived from a variety of adult tissues such as synovium-derived, bone marrow-derived, umbilical cord blood-derived, other placenta, muscle, fat and nerve tissue, Stem cells, but are not necessarily limited thereto. The term " synovium "refers to a membrane wrapping the inside of a joint pocket. When mesenchymal stem cells derived from synovium-derived mesenchymal stem cells (SDMSCs) are used as the mesenchymal stem cells, they have unique characteristics such as UDPDG activity and COMP and GAG expression as compared with other tissue-derived MSCs, The chondrocyte differentiation effect can be excellent.
The chondrocyte (cartilage cell) refers to a cell constituting cartilage tissue.
As used herein, the term "inducing differentiation from mesenchymal stem cells into chondrocytes" may mean inducing mesenchymal stem cells to differentiate into chondrocytes.
Accordingly, the composition for inducing differentiation of mesenchymal stem cells into chondrocytes according to the above embodiment can be used as a composition for regenerating cartilage tissue. The above-mentioned "cartilage tissue regeneration" may mean regenerating damaged cartilage tissue or inducing generation of insufficient cartilage tissue to regenerate cartilage tissue.
Another embodiment of the present invention provides a method for inducing differentiation from mesenchymal stem cells into chondrocytes, comprising culturing mesenchymal stem cells in vitro in the presence of ganglioside GM3.
The ganglioside GM3 may be treated with a concentration of 0.1 to 1000 μM and may be treated with a concentration of 0.1 to 100 μM, more specifically a concentration of 0.1 to 10 μM, more specifically a concentration of 1 to 10 μM The mesenchymal stem cells can be treated with mesenchymal stem cells at a concentration of, for example, 2 to 5 [mu] M, but the present invention is not limited thereto. Concentrations having an inducing effect can be selected by an ordinary person skilled in the art.
The mesenchymal stem cells may be mesenchymal stem cells derived from a variety of adult tissues such as synovium-derived, bone marrow-derived, umbilical cord blood-derived, other placenta, muscle, fat and nerve tissue, Stem cells, but are not necessarily limited thereto.
The term "in vitro" refers to cultivation in vitro , not in vivo , and may mean culturing cells isolated from a living body.
In addition to the ganglioside GM3, it may further include a chondrocyte differentiation inducer, for example, an insulin-transferrin-selenium (ITS), ascorbate-2-phosphate, dexamethasone, TGF-beta (Transforming Growth Factor), and the like, but the present invention is not limited thereto.
The chondrocyte induced differentiation from mesenchymal stem cells by the above method can be used as a cell therapy agent for treating cartilage diseases. Herein, cartilage diseases that can be treated using cartilage cells include, for example, osteoarthritis, degenerative arthropathy, cartilage dysplasia, degenerative arthritis, rheumatoid arthritis, osteomalacia, fibrous osteitis, and intractable bone disease And can be used for treatment of various diseases caused by abnormal annihilation of chondrocytes or damage of cartilage tissue.
Another embodiment of the present invention provides a pharmaceutical composition for treating or preventing cartilage disease comprising ganglioside GM3 as an active ingredient.
The cartilage disease may be cartilage disease caused by abnormal annihilation of cartilage cells or damage of cartilage tissue, and may be caused by, for example, osteoarthritis, degenerative arthropathy, cartilage dysgenesis, degenerative arthritis, rheumatoid arthritis, osteomalacia, Although bone disease is a representative example, it is not limited thereto.
As used herein, the term "treatment" refers to any action in which administration of a composition containing ganglioside GM3 improves or alleviates symptoms of cartilage disease. Thus, treatment of the cartilage disease may mean that the damage of cartilage tissue is treated by regenerating cartilage damaged by the composition.
As used herein, the term "prevention" means any action that inhibits or delays cartilage disease by administration of a composition containing the ganglioside GM3.
In the above pharmaceutical composition, naringenin and hesperetin are preferably used as such, but derivatives thereof may be prepared and used in consideration of solubility and stability characteristics.
The pharmaceutical composition according to the above embodiments may be various oral or parenteral formulations. In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules, and the like, which may contain one or more excipients such as starch, calcium carbonate, sucrose or lactose lactose, gelatin and the like. In addition to simple excipients, lubricants such as magnesium stearate, talc, and the like are also used. Liquid preparations for oral administration include suspensions, solutions, emulsions, syrups and the like. Various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included in addition to water and liquid paraffin, which are simple diluents commonly used. have. Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories.
Examples of the pharmaceutical composition according to the present invention include non-aqueous solutions, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, injectable esters such as ethyl oleate, and the like. Examples of the suppository base include witepsol, macrogol, tween 61, cacao paper, laurin, glycerogelatin and the like.
Dosage forms of the pharmaceutical compositions according to the above embodiments may be used in the form of their pharmaceutically acceptable salts, and may be used alone or in combination with other pharmaceutically active compounds as well as in suitable aggregates. The salt is not particularly limited as long as it is pharmaceutically acceptable so long as it is pharmaceutically acceptable and includes, for example, hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, hydrofluoric acid, hydrobromic acid, formic acid acetic acid, tartaric acid, lactic acid, citric acid, fumaric acid, , Benzenesulfonic acid, toluenesulfonic acid, and naphthalenesulfonic acid.
The pharmaceutical composition according to the above embodiments may be administered parenterally or orally according to the purpose, and may be administered in one to several divided doses in an amount of 0.1 to 500 mg and 1 to 100 mg per kg of body weight per day . The dosage for a particular patient may vary depending on the patient's body weight, age, sex, health condition, diet, time of administration, administration method, excretion rate, severity of disease, and the like.
The pharmaceutical compositions according to the above embodiments may be formulated into oral preparations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups and aerosols, external preparations such as ointments and creams, suppositories and sterilized injection solutions And may be formulated in any form suitable for pharmaceutical preparations.
The pharmaceutical compositions according to the above embodiments may be administered to mammals such as rats, mice, livestock, and humans in various routes such as parenteral, oral, etc., and all manner of administration may be expected, Or by intravenous, intramuscular, subcutaneous, intramural or intracerebroventricular injection.
Another embodiment of the present invention provides a health functional food composition for improving or preventing cartilage disease containing ganglioside GM3 as an active ingredient.
As used herein, the term "improvement" means any action that improves or alters the symptoms of cartilage disease by administering a composition containing the ganglioside GM3.
As used herein, the term "health functional food" refers to a food prepared by processing a specific ingredient as a raw material for the purpose of health assisting or by extracting, concentrating, refining, or mixing a specific ingredient contained in a food raw material , The health functional food composition can perform functions related to prevention of cartilage disease and recovery of cartilage disease.
The health functional food may contain flavoring agents such as various nutrients, vitamins, minerals (electrolytes), synthetic flavors and natural flavors, coloring agents and thickening agents (cheese, chocolate etc.), pectic acid and its salts, alginic acid and its salts, Organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonating agents used in carbonated drinks, and the like. It can also contain natural fruit juices and pulp for the production of fruit juices and vegetable drinks. These components may be used independently or in combination. The health functional food may be any one of meat, sausage, bread, chocolate, candy, snack, confectionery, pizza, ramen, gum, ice cream, soup, beverage, tea, functional water, drink, alcohol and vitamin .
In addition, the health functional food may further include food additives, and the suitability of the food functional food as a "food additive" is not limited to the corresponding items in general rules and general test methods approved by the Food and Drug Administration Shall be determined according to the relevant standards and standards.
Examples of the products that have been used in the above-mentioned "food additives" include natural products such as ketones, chemical products such as glycine, potassium citrate, nicotinic acid and cinnamic acid, sensory coloring matter, licorice extract, crystalline cellulose, high- - Mixed preparations such as a sodium glutamate preparation, a noodle-added alkaline agent, a preservative preparation, a tar coloring agent and the like.
At this time, the content of ganglioside GM3 added to the food during the production of the health functional food can be appropriately increased or decreased as needed. The ganglioside GM3 may be added in an amount of 0.1 wt% to 50 wt% to 100 wt% of the food.
Another embodiment of the present invention provides a method for treating or preventing cartilage disease comprising administering a composition containing ganglioside GM3 as an active ingredient to a subject in need thereof.
As used herein, the term "individual" refers to all animals including humans who have already developed or are capable of developing cartilage disease, and the composition of the present invention is effective for preventing and treating the disease by administration to a subject have.
The composition is preferably a pharmaceutical composition and can be administered in a therapeutically effective amount.
As used herein, the term "therapeutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment and the effective dosage level will vary depending on the species and severity, 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 the treatment, factors including co-administered drugs, and other factors well known in the medical arts. The composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents. And can be administered singly or multiply. It is important to take into account all of the above factors and to administer the amount in which the maximum effect can be obtained in a minimal amount without side effects, which can be easily determined by a person skilled in the art.
Previous studies have demonstrated the role of TGF-β in many cell fates such as maturation, cell proliferation and differentiation. Specifically, in chondrocyte differentiation, the TGF-β signal is transmitted to the nucleus through the SMAD pathway, which activates the Sox-9 transcription factor for cartilage gene expression, such as type II collagen and aggrecan. In the present invention, it was confirmed that treatment of ganglioside GM3 during chondrocyte cell differentiation activates Sox-9, type II collagen, agricans and COMP. Also, according to an embodiment of the present invention, ganglioside GM3 has been shown to modulate serine phosphorylation of TGF-s-R1 and -R2 and SMAD2 / 3 during chondrocyte cell differentiation.
In conclusion, it was confirmed that exogenous ganglioside GM3 did not affect cytotoxicity of hSDMSCs. However, exogenous ganglioside GM3 significantly increased the tissue weight of chondrocyte cell groups differentiated from the chondrocyte cell group differentiation, especially in the 5 [mu] M treated group, and the amount of toluidine blue was high, and aggrecan, Sox-9, 2 expression of cartilage cell specific marker genes such as collagen and COMP was increased and GAG accumulation was increased. In addition, exogenous ganglioside GM3 enhanced the TGF-beta signaling pathway through
The use of the composition containing ganglioside GM3 according to the present invention is excellent in the effect of inducing differentiation from mesenchymal stem cells into chondrocytes, and thus can be usefully used as a therapeutic, preventive or ameliorating agent for cartilage diseases by promoting cartilage regeneration .
Figure 1 shows the characteristics of human synovium-derived mesenchymal stem cells (hSDMSCs): (A) MSC negative surface proteins, CD4, CD34 and CD45 (upper), and positive surface proteins, CD44, CD73, CD90 and CD105 The results of FACS analysis of the expression of the expression of And (B) staining results of osteoblast differentiated from hSDMSCs (left: Alizarin red S staining) and adipocyte differentiated from hSDMSCs (right: Oil red O staining).
Figure 2 shows the effect of ganglioside GM3 on cell viability in hSDMSCs: hSDMSCs were treated with ganglioside GM3 (0, 1, 2, 5 and 10 [mu] M) for 7 days. Cell viability was calculated according to the formula [(OD value after treatment with ganglioside GM3) / OD value in the group treated with 0 [mu] M ganglioside GM3) x 100]. Values are expressed as mean ± SEM from 6 independent experiments.
Figure 3 shows the effect of ganglioside GM3 on cartilage cell population differentiation: hSDMSCs treated ganglioside GM3 (0, 1, 2, 5 and 10 [mu] M) for 21 days during chondrocyte cell population changes. (A) Analysis of the tissue weight of differentiated chondrocytes at day 21 after chondrocyte differentiation. Values are expressed as mean ± SEM from 6 independent experiments. ** p < 0.01 and *** p < 0.001 compared to GM3 treated group at 0 μM; And (B) Toluidine blue and H & E staining results to confirm the histological characteristics of the cell groups of all groups.
FIG. 4 is a graph showing the effect of ganglioside GM3 on the tissue weight of differentiation of chondrocytic cell group: (A) the result of analysis of the weight of the differentiated tissue of the cartilage cell group on the 7th day and (B) 14th day. Values are expressed as mean ± SEM from 6 independent experiments. * P < 0.05 and ** p < 0.01 compared to 0 μM GM3 treated group.
5 shows the effect of ganglioside GM3 on the histological analysis of differentiation of chondrocyte cell group. For histological analysis of differentiated chondrocytic cell groups on
FIG. 6 is a graph showing a comparison of chondrocyte-specific marker expression in chondrocyte cell group differentiation: (A) Aggrecan, (B) Sox-9, (C )
FIG. 7 shows a comparison of chondrocyte-specific marker expression in differentiated chondrocyte cell groups: (A) aggrecan, (B) Sox- 9, (C)
FIG. 8 is a graph showing the results of biochemical analysis for evaluating the differentiation of chondrocyte cell group. After 21 days of chondrocyte cell differentiation, the cell group was digested with papain and S-GAG (sulfated glycosaminoglycans) Respectively. GAG content was normalized to the total DNA content of each sample. Values are expressed as mean ± SEM from 3 independent experiments. * P < 0.05 and *** p < 0.001 as compared to 0 μM GM3 treated group.
Fig. 9 is a diagram showing the results of biological analysis for evaluation of chondrocyte cell differentiation. After 7 and 14 days of chondrocyte cell differentiation, the cells were digested with papain and S-GAG (sulfated glycosaminoglycans) And the content thereof was measured. GAG content was normalized to the total DNA content of each sample. Values are expressed as mean ± SEM from 3 independent experiments per week. * P < 0.05, and *** p < 0.001 compared to GM3 treated group at 0 μM.
Figure 10 shows the effect of ganglioside GM3 on chondrocyte cell differentiation: hSDMSCs that underwent chondrocyte cell differentiation were treated with various concentrations of GM3 (0, 1, 2, 5, and 10 [mu] M) for 21 days. After SDS-PGE and Western blotting, serine (TGF-β-R1 and -R2) and phosphorylation of
Figure 11 shows the effect of differentiating chondrocyte cell cultures of ganglioside GM3 on differentiation: hSDMSCs that underwent differentiation into chondrocyte cell groups were cultured at various concentrations of GM3 (0, 1, 2, 5, and 10 [mu] M) 14 days. After SDS-PGE and Western blotting, phosphorylation of serine (TGF-β-R1 and -R2),
Hereinafter, the present invention will be described in more detail in the following Examples. It should be noted, however, that the following examples are illustrative only and do not limit or limit the scope of the present invention. It will be understood by those of ordinary skill in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims.
Example One. hSDMSCs Culture and on hSDMSCs About Ganglioside Evaluation of GM3 cytotoxic effect
1-1. hSDMSCs Isolation and Culture
Primary cell lines were obtained from primary synovial membrane tissues isolated from human knees.
Specifically, human synovium-derived mesenchymal stem cells (hSDMSCs) derived from human synovium were obtained from the synovium of osteoarthritis patients according to the regulations of the Ethics Committee of Yonsei Sarang Hospital. Synovial tissue was chopped, digested with 0.25% trypsin EDTA (Hyclone), and incubated at 37 ° C for 30 minutes. After 0.25% trypsin EDTA was removed, the cells were treated with 0.3% collagenase (Worthington) in DMEM (high glucose Dulbecco's modified Eagle's medium; Hyclone) and cultured at 37 ° C for 90 minutes. Single cell suspensions were centrifuged and supernatant removed. Cells were resuspended in pre-warmed alpha-modified Eagle's medium (Hyclone) supplemented with 20% fetal bovine serum (HBS) and 1 x penicillin / streptomycin (Hyclone) I moved. When adherent cells proliferated and reached confluence, cells were trypsinized and transferred back to the dish for expansion, which was used as hSDMSCs.
1-2. Intermediate lobe Identification of stem cells
In order to identify the hSDMSCs cells obtained in Example 1-1, FACS analysis and the ability to differentiate (osteocyte and adipocyte differentiation) were performed.
FACS analysis was performed as follows. After trypinization, the hSDMSCs of Example 1-1 above were resuspended at 1.0 x 10 cells / ml in ice cold PBS containing 0.5% BSA (bovine serum albumin). The cells were then incubated in ice for 20 minutes with PerCP-conjugated anti-human CD4, PE-conjugated anti-human CD34, APC-conjugated anti-human CD45, PE- CD40, FITC-conjugated anti-human CD73, PerCP-conjugated anti-human CD90, or PerCP-conjugated anti-human CD105 (BD Biosciences). Analysis was performed using CellQuest software (BD Biosciences) by flow cell counting (BD Biosciences).
The ability to differentiate into osteoblasts was confirmed as follows. the hSDMSCs in 24-well tissue culture plates 5 x 10 4 cells / well were seeded and grown for one day in a standard growth medium. The medium was replaced with an osteoblast-producing medium composed of 1-penicillin / streptomycin, 10% FBS, 100 nM dexamethasone, 10 mM? -Glycerophosphate (Sigma-aldrich) and 50 μM ascorbic acid in high- Lt; / RTI > The medium was changed every 2 days. After differentiation, the cultured cells were fixed with 4% PFA (paraformaldehyde) and stained with 2% Alizarin red S pH 7.2 (Sigma-aldrich). Samples were observed through a light microscope (Leica, DMI 3000B).
The ability to differentiate into adipocytes was confirmed as follows. the hSDMSCs in 12-well tissue culture plates 5 x 10 3 cells / well were seeded and grown for one day in a standard growth medium. The medium was supplemented with 1 × penicillin / streptomycin, 10% FBS, 1 μM dexamethasone, 200 μM indomethacin (Sigma-aldrich), 0.5 mM IBMX (3-butyl-1-methylxanthine) mL insulin (GIBCO) and replaced with an adipocyte-derived medium consisting of complete high-glucose DMEM for 21 days. The medium was changed every 2 days. After differentiation, the cultured cells were fixed with 4% PFA (paraformaldehyde) and stained with 2% Oil O Red solution (Sigma-aldrich). Samples were observed through a light microscope (Leica, DMI 3000B).
As a result, it was confirmed that the cells obtained in Example 1-1 were mesenchymal stem cells (MSCs) (FIG. 1).
1-3. Ganglioside Cytotoxicity assay of GM3
To assess ganglioside GM3 toxicity, hSDMSCs cells were incubated with various concentrations of ganglioside GM3.
Specifically, the cell viability assay was performed as follows. HSDMSCs cells of Example 1-1 were inoculated on a 24-well plate at a density of 1 x 10 4 cells / well. Cells were treated with GM3 (1, 2, 5, and 10 μM) (Matreya) and then cell proliferation was assessed by MTT (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide) Based tests. Each well was incubated with MTT solution (Sigma-Aldrich) for 4 hours, and the absorbance of each well was measured at 590 nm using a spectrophotometer.
Experimental The statistical analysis was carried out in the following manner in all experiments related to the present invention. All data are expressed as mean ± S.E.M. Comparisons of multiple groups were performed by one-way ANOVA followed by Bonferroni post-test and compared in pairs. Differences were considered statistically significant at P <0.05. All data was analyzed using GraphPad Prism software, version 5.00 (Graph-Pad software).
As a result, the GM3 treatment group had no significant difference in the cell survival rate at the respective concentrations for 7 days from the GM3 treatment group of 0 μM. Therefore, it was found that ganglioside GM3 did not affect cytotoxicity of hSDMSCs (Fig. 2).
Example 2. Ganglioside The induction effect of GM3 on differentiation of chondrocytic cell group
2-1. Tissue weighing of chondrocytes
An aggregates culture system was used to confirm the effect of ganglioside GM3 on chondrocyte differentiation of hSDMSCs. During chondrocyte cell differentiation, cell populations of all groups were weighed.
Specifically, chondrocyte cell population differentiation was carried out by the following method. The hSDMSCs in Example 1-1 in polypropylene tubes were inoculated with 1.0 x 10 6 cells / ml. The cells were centrifuged at 1,500 rpm for 5 minutes for use in an aggregate culture system. Cells were prepared and transfected with 1xITS (Insulin-Transferrin-Selenium; GIBCO), 50 mM ascorbate-2-phosphate, 100 nM dexamethasone (Sigma), 5 ng / And cultured at 37 占 폚 in chondrocyte-producing medium consisting of growth factor-beta (Prospec) and GM3 (or without GM3). Cells were formed as free-floating masses in the first 24-hour culture. The medium was replaced every 2 days for 21 days.
As a result, the tissue weights did not differ in the respective groups during the 7 days of chondrocyte cell differentiation. However, especially in the GM3 treated group of 2, 5 and 10 μM, the tissue weight increased on the 14th day after the differentiation of chondrocyte cell group. On the 21st day after chondrocyte cell differentiation, the mean values of the pellet weights in the 2 μM and 5 μM GM3-treated groups were 10.13 ± 0.37 mg and 11.37 ± 0.59 mg, respectively, which were significantly increased compared with the 0 μM GM3-treated group (Figs. 3A and 4).
Therefore, the induction of chondrocyte differentiation was promoted by treatment with ganglioside GM3, and in particular, it was confirmed that the effect of inducing chondrocyte differentiation in the GM3 treatment group of 2 and 5 μM was excellent.
2-2. Histological analysis of chondrocytes
Histological analysis was performed by staining with toluidine blue and H & E for each of the experimental groups of Example 2-1.
Specifically, histological analysis was performed as follows. Cells were fixed with 10% formaldehyde for 10 min and washed twice with PBS (phosphate buffered saline). Cells were placed in paraffin and cut into 5-μm sections. The sections were stained according to conventional protocols with toluidine blue (for matrix proteoglycan staining), H & E (haematoxylin and eosin) staining. Samples were observed through a light microscope (Leica, DMI 3000BI).
As a result, the GM3-treated group of 5 [mu] M had a considerable amount of toluidine blue (Fig. 3B and Fig. 5). Since the content of glycosaminoglycan in the cartilage tissue can be visually confirmed according to the toluidine blue staining intensity, it was confirmed that the GM3-treated group, especially 5 μM of the ganglioside GM3-treated group, Respectively.
2-3. Cartilage cell specific Marker Gene expression measurement
Expression of chondrocyte-specific marker genes such as aggrecan, Sox-9,
Specifically, the analysis of mRNA expression was performed in the following manner. Total RNA was prepared from samples using the Trizol Kit according to the manufacturer's instructions (Invitrogen). 2 μg of total RNA was reverse transcribed (RT) according to the manufacturer's instructions (Qiagen) using Omniscript RT kit and oligo dT. Real-time PCR was performed using an ABI PRISM 7500 sequence detection system (Applied Biosystems). The reaction was carried out in three independent experiments. GAPDH (glyceraldehyde 3-phosphate dehydrogenase) was used as a control. The primer sequences are shown in Table 1 below. Expression data were normalized to the geometric mean of the expression gene GAPDH at all time to control for changes in expression levels and analyzed using the 2 - ΔΔCT method.
As a result, at day 21 after chondrocyte cell differentiation, chondrocyte-specific markers were highly expressed in 5 μM ganglioside GM3 (3.00 ± 0.02 fold compared to 0 μM GM3 treated group, 3.00 ± 0.02 fold, Sox-9, 2.85 占 .55 times;
2-4. Of GAGs Accumulation analysis
In Example 2-1, the accumulation of GAGs (glycosaminoglycans) normalized to whole DNA in all groups of cells during chondrocyte differentiation was analyzed.
Specifically, the measurement of the GAG (glycosaminoglycan) content was carried out in the following manner. After 21 days of chondrocyte induction, all cell populations were stimulated with 125 μg / mL papain dissolved in PBE buffer (10 mM EDTA, 100 mM sodium phosphate, pH 6.5) containing 5 mM L-cysteine- Gt; 65 C < / RTI > per sample. The chondrocyte productivity was determined by measuring the production of S-GAG (sulfated glycosaminoglycans) according to the manufacturer's recommended protocol (Blyscan ™ Glycosaminoglycan Assay Kit) using the 1,9-dimethylmethylene blue (DMMB) test. Absorbance was measured at 656 nm using a microplate immunoreader (Sunrise ™, TECAN). To quantify the cell density, the amount of DNA in the papain digest was analyzed using the Quant-iT PicoGreens dsDNA Assay kit (Invitrogen) according to the manufacturer's recommended protocol. (Excitation at 485 nm; emission at 538 nm) using a CytoFluors Series 4000 (Applied Biosystems). The GAG content was normalized to the total DNA content.
As a result, the GAG / DNA ratio was 16.48 ± 1.45 μg / μg in the group treated with 5 μM ganglioside GM3 (0 μM, 3.69 ± 0.12 μg / μg; 1 μM, 5.05 ± 1.08 μg / μg; 2 μM, 4.93 ± 0.21 μg / μg; 10 μM, 6.28 ± 0.41 μg / μg) (FIGS. 8 and 9).
Therefore, it was confirmed that GAG (glycosaminoglycan), which is important in articular cartilage formation, was highly effective in inducing chondrocyte differentiation because of its high concentration in GM3 treatment group of 5 μM, especially in ganglioside GM3 treatment group.
Example 3. Ganglioside In the differentiation of GM3 chondrocytes TGF -β Signal path Upward adjustment ( UpRegulation ) effect
Ganglioside GM3 involves the phosphorylation of SMAD2 / 3, which is serine, TGF-beta receptor-1 (TGF-beta-R1) and TGF- beta-R2 and downstream of the TGF- Lt; RTI ID = 0.0 > TGF-beta < / RTI > signaling.
Specifically, Western blot analysis was performed as follows. Cells were homogenized with RIPA buffer (Sigma-Aldrich) and centrifuged at 15,000 x g for 30 minutes. Protein concentrations were measured using the Bradford method. The same amount of protein (30 μg) was separated by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to a Hybond ECL nitrocellulose membrane (Amersham Pharmacia). Blot was stopped at room temperature for 2 hours with 5% (w / v) skim milk powder dissolved in a Tris-buffered saline containing 0.05
As a result, phosphorylation of TGF-βRs, particularly TGF-β-R2, and SMAD2 / 3 was increased in the ganglioside GM3 treated group at 5 μM (FIGS. 10 and 11). These results indicate that GM3 and TGF-β are closely related to the chondrocyte cell differentiation of hSDMSCs in TGF-β receptor activation.
<110> TJC Life <120> Composition containing ganglioside GM3 for inducing chondrogenic differentiation, and uses thereof <130> P16E30D0461 <160> 10 <170> KoPatentin 3.0 <210> 1 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Aggrecan forward primer <400> 1 ctgagtgaaa ccacctctgc att 23 <210> 2 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Aggrecan reverse primer <400> 2 gacgcctcgc cttcttga 18 <210> 3 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> SOX-9 forward primer <400> 3 ccccaacaga tcgcctacag 20 <210> 4 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> SOX-9 reverse primer <400> 4 tctggtggtc ggtgtagtcg ta 22 <210> 5 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Collagen type 2 forward primer <400> 5 ggcaatagca ggttcacgta ca 22 <210> 6 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Collagen type 2 reverse primer <400> 6 cgataacagt cttgccccac tt 22 <210> 7 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> COMP forward primer <400> 7 caagaagtcc tatcgttggt tcct 24 <210> 8 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> COMP reverse primer <400> 8 ctcagggccc tcatagaatc g 21 <210> 9 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> GAPHD forward primer <400> 9 ggcatcctgg gctacactga 20 <210> 10 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> GAPHD reverse primer <400> 10 gaggagtggg tgtcgctgtt 20
Claims (8)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1020160035243A KR101794503B1 (en) | 2016-03-24 | 2016-03-24 | Composition containing ganglioside GM3 for inducing chondrogenic differentiation, and uses thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1020160035243A KR101794503B1 (en) | 2016-03-24 | 2016-03-24 | Composition containing ganglioside GM3 for inducing chondrogenic differentiation, and uses thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| KR20170113733A true KR20170113733A (en) | 2017-10-13 |
| KR101794503B1 KR101794503B1 (en) | 2017-11-08 |
Family
ID=60139675
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| KR1020160035243A KR101794503B1 (en) | 2016-03-24 | 2016-03-24 | Composition containing ganglioside GM3 for inducing chondrogenic differentiation, and uses thereof |
Country Status (1)
| Country | Link |
|---|---|
| KR (1) | KR101794503B1 (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114206330A (en) * | 2019-08-02 | 2022-03-18 | 三得利控股株式会社 | Composition for promoting cartilage regeneration |
| WO2022114845A1 (en) * | 2020-11-27 | 2022-06-02 | 주식회사 강스템바이오텍 | Method for preparing injectable injection composition derived from animal cartilage, and use thereof |
-
2016
- 2016-03-24 KR KR1020160035243A patent/KR101794503B1/en active IP Right Grant
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114206330A (en) * | 2019-08-02 | 2022-03-18 | 三得利控股株式会社 | Composition for promoting cartilage regeneration |
| EP4008401A4 (en) * | 2019-08-02 | 2023-05-03 | Suntory Holdings Limited | Composition for cartilage regeneration promotion |
| WO2022114845A1 (en) * | 2020-11-27 | 2022-06-02 | 주식회사 강스템바이오텍 | Method for preparing injectable injection composition derived from animal cartilage, and use thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| KR101794503B1 (en) | 2017-11-08 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Nimura et al. | Increased proliferation of human synovial mesenchymal stem cells with autologous human serum: comparisons with bone marrow mesenchymal stem cells and with fetal bovine serum | |
| US20080233195A1 (en) | Synthetic triterpenoids and tricyclic-bis-enones for use in stimulating bone and cartilage growth | |
| KR102035273B1 (en) | Composition for preventing or treating osteoporosis comprising stem cell-derived exosomes | |
| US11213572B2 (en) | Cartilage regeneration composition containing HAPLN1 as active ingredient | |
| KR20180127355A (en) | Colony-forming medium and uses thereof | |
| KR101980453B1 (en) | Composition For Promoting Production of Stem Cell-derived Exosomes | |
| JP2022533277A (en) | Composition for promoting stem cell-derived exosome production and enhancing stem cell ability | |
| KR101794503B1 (en) | Composition containing ganglioside GM3 for inducing chondrogenic differentiation, and uses thereof | |
| KR20230109119A (en) | Compositions for enhancing stemness and uses thereof | |
| EP3822343A1 (en) | Production method for neuron-like cells | |
| Zhang et al. | Regulation of human mesenchymal stem cell differentiation by TREM-2 | |
| CA3188486A1 (en) | A composition comprising mesenchymal precursor or stem cells and their use | |
| WO2007136157A1 (en) | Compositions and methods for treating motor neuron diseases | |
| US20210147482A1 (en) | Peptide for inhibiting bone resorption | |
| Zhang et al. | Neuritin Promotes Bone Marrow-Derived Mesenchymal Stem Cell Migration to Treat Diabetic Peripheral Neuropathy | |
| US11466254B2 (en) | Composition containing, as active ingredient, culture of chicken bone marrow-derived osteochondral progenitor cells for promoting osteogenesis or inducing chondrogenic differentiation | |
| WO2019045451A1 (en) | Cartilage regeneration composition containing hapln1 as active ingredient | |
| KR20130085863A (en) | Composition for inducing migration of mesenchymal stem cells | |
| KR102580518B1 (en) | Method for selecting mesenchymal stem cell with improved self-maintenance and mesenchymal stem cells selected by thereof | |
| KR101998257B1 (en) | Composition for preventing or treating cardiofaciocutaneous syndrome | |
| JP5854492B2 (en) | Chondrocyte differentiation inducer | |
| KR102308505B1 (en) | COMPOSITION FOR TREATMENT OF SARCOPENIA OR OSTEOPOROSIS THROUGH THE MECHANISM OF PROMOTING MYOBLAST DIFFERENTIATION OR INHIBITING THE DIFFERENTIATION OF OSTEOCLAST COMPRISING Cyclo (L-phenylalanyl-L-proline) | |
| US20110223138A1 (en) | Mesenchymal stem cells that express increased amounts of anti-apoptotic proteins | |
| KR101805771B1 (en) | A pharmaceutical composition for preventing or treating blood dyscrasia comprising TMEM100 protein | |
| KR20240054468A (en) | Medium composition for inducing differentiation from pluripotent stem cells to macrophages and a differentiation method using the same |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| A201 | Request for examination | ||
| E902 | Notification of reason for refusal | ||
| E701 | Decision to grant or registration of patent right | ||
| GRNT | Written decision to grant |