CN106834212B - 一种用于肺组织3d培养的培养基 - Google Patents
一种用于肺组织3d培养的培养基 Download PDFInfo
- Publication number
- CN106834212B CN106834212B CN201710183411.2A CN201710183411A CN106834212B CN 106834212 B CN106834212 B CN 106834212B CN 201710183411 A CN201710183411 A CN 201710183411A CN 106834212 B CN106834212 B CN 106834212B
- Authority
- CN
- China
- Prior art keywords
- culture
- lung
- tissue
- cells
- lung tissue
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 210000004072 lung Anatomy 0.000 title claims abstract description 59
- 239000001963 growth medium Substances 0.000 title claims abstract description 28
- 206010058467 Lung neoplasm malignant Diseases 0.000 claims abstract description 20
- 201000005202 lung cancer Diseases 0.000 claims abstract description 20
- 208000020816 lung neoplasm Diseases 0.000 claims abstract description 20
- 102100028412 Fibroblast growth factor 10 Human genes 0.000 claims abstract description 9
- 101000917237 Homo sapiens Fibroblast growth factor 10 Proteins 0.000 claims abstract description 9
- PWKSKIMOESPYIA-BYPYZUCNSA-N L-N-acetyl-Cysteine Chemical compound CC(=O)N[C@@H](CS)C(O)=O PWKSKIMOESPYIA-BYPYZUCNSA-N 0.000 claims abstract description 7
- 229960004308 acetylcysteine Drugs 0.000 claims abstract description 7
- 102000045246 noggin Human genes 0.000 claims abstract description 7
- 108700007229 noggin Proteins 0.000 claims abstract description 7
- HJCMDXDYPOUFDY-WHFBIAKZSA-N Ala-Gln Chemical compound C[C@H](N)C(=O)N[C@H](C(O)=O)CCC(N)=O HJCMDXDYPOUFDY-WHFBIAKZSA-N 0.000 claims abstract description 6
- 238000004113 cell culture Methods 0.000 claims description 13
- 238000010790 dilution Methods 0.000 claims description 12
- 239000012895 dilution Substances 0.000 claims description 12
- DFPAKSUCGFBDDF-UHFFFAOYSA-N Nicotinamide Chemical compound NC(=O)C1=CC=CN=C1 DFPAKSUCGFBDDF-UHFFFAOYSA-N 0.000 claims description 11
- 238000005516 engineering process Methods 0.000 claims description 10
- RJKFOVLPORLFTN-LEKSSAKUSA-N Progesterone Chemical compound C1CC2=CC(=O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H](C(=O)C)[C@@]1(C)CC2 RJKFOVLPORLFTN-LEKSSAKUSA-N 0.000 claims description 6
- 229960003966 nicotinamide Drugs 0.000 claims description 6
- 235000005152 nicotinamide Nutrition 0.000 claims description 6
- 239000011570 nicotinamide Substances 0.000 claims description 6
- KIDHWZJUCRJVML-UHFFFAOYSA-N putrescine Chemical compound NCCCCN KIDHWZJUCRJVML-UHFFFAOYSA-N 0.000 claims description 6
- 239000012583 B-27 Supplement Substances 0.000 claims description 3
- 108010052343 Gastrins Proteins 0.000 claims description 3
- 239000005700 Putrescine Substances 0.000 claims description 3
- AOXOCDRNSPFDPE-UKEONUMOSA-N chembl413654 Chemical compound C([C@H](C(=O)NCC(=O)N[C@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@H](CCSC)C(=O)N[C@H](CC(O)=O)C(=O)N[C@H](CC=1C=CC=CC=1)C(N)=O)NC(=O)[C@@H](C)NC(=O)[C@@H](CCC(O)=O)NC(=O)[C@@H](CCC(O)=O)NC(=O)[C@@H](CCC(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H]1N(CCC1)C(=O)CNC(=O)[C@@H](N)CCC(O)=O)C1=CC=C(O)C=C1 AOXOCDRNSPFDPE-UKEONUMOSA-N 0.000 claims description 3
- BVTBRVFYZUCAKH-UHFFFAOYSA-L disodium selenite Chemical compound [Na+].[Na+].[O-][Se]([O-])=O BVTBRVFYZUCAKH-UHFFFAOYSA-L 0.000 claims description 3
- 235000014666 liquid concentrate Nutrition 0.000 claims description 3
- 229960003387 progesterone Drugs 0.000 claims description 3
- 239000000186 progesterone Substances 0.000 claims description 3
- 229960001471 sodium selenite Drugs 0.000 claims description 3
- 235000015921 sodium selenite Nutrition 0.000 claims description 3
- 239000011781 sodium selenite Substances 0.000 claims description 3
- 101001011741 Bos taurus Insulin Proteins 0.000 claims description 2
- 101000825954 Homo sapiens R-spondin-1 Proteins 0.000 claims description 2
- 101000766306 Homo sapiens Serotransferrin Proteins 0.000 claims description 2
- 239000012580 N-2 Supplement Substances 0.000 claims description 2
- 102100022762 R-spondin-1 Human genes 0.000 claims description 2
- 239000000654 additive Substances 0.000 claims description 2
- 230000000996 additive effect Effects 0.000 claims description 2
- IXIBAKNTJSCKJM-BUBXBXGNSA-N bovine insulin Chemical compound C([C@@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@H]1CSSC[C@H]2C(=O)N[C@@H](C)C(=O)N[C@@H](CO)C(=O)N[C@H](C(=O)N[C@H](C(N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC=3C=CC(O)=CC=3)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC=3C=CC(O)=CC=3)C(=O)N[C@@H](CSSC[C@H](NC(=O)[C@H](C(C)C)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=3C=CC(O)=CC=3)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](C)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=3NC=NC=3)NC(=O)[C@H](CO)NC(=O)CNC1=O)C(=O)NCC(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)NCC(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C)C(O)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O)=O)CSSC[C@@H](C(N2)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](NC(=O)CN)[C@@H](C)CC)C(C)C)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CC(N)=O)NC(=O)[C@@H](NC(=O)[C@@H](N)CC=1C=CC=CC=1)C(C)C)C1=CN=CN1 IXIBAKNTJSCKJM-BUBXBXGNSA-N 0.000 claims description 2
- XNSAINXGIQZQOO-SRVKXCTJSA-N protirelin Chemical compound NC(=O)[C@@H]1CCCN1C(=O)[C@@H](NC(=O)[C@H]1NC(=O)CC1)CC1=CN=CN1 XNSAINXGIQZQOO-SRVKXCTJSA-N 0.000 claims description 2
- 102100021022 Gastrin Human genes 0.000 claims 1
- 210000001519 tissue Anatomy 0.000 abstract description 55
- 210000004027 cell Anatomy 0.000 abstract description 53
- 210000002220 organoid Anatomy 0.000 abstract description 35
- 238000011160 research Methods 0.000 abstract description 6
- -1 R-spondin1 Chemical compound 0.000 abstract description 5
- 235000015097 nutrients Nutrition 0.000 abstract description 4
- DFPAKSUCGFBDDF-ZQBYOMGUSA-N [14c]-nicotinamide Chemical compound N[14C](=O)C1=CC=CN=C1 DFPAKSUCGFBDDF-ZQBYOMGUSA-N 0.000 abstract description 3
- 230000002068 genetic effect Effects 0.000 abstract description 2
- 210000000130 stem cell Anatomy 0.000 abstract description 2
- 230000008827 biological function Effects 0.000 abstract 1
- 239000003636 conditioned culture medium Substances 0.000 description 14
- 238000012258 culturing Methods 0.000 description 13
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 12
- 239000002609 medium Substances 0.000 description 12
- 238000000338 in vitro Methods 0.000 description 9
- 238000000034 method Methods 0.000 description 9
- 108090000623 proteins and genes Proteins 0.000 description 9
- 239000006228 supernatant Substances 0.000 description 9
- 102000029816 Collagenase Human genes 0.000 description 8
- 108060005980 Collagenase Proteins 0.000 description 8
- 229960002424 collagenase Drugs 0.000 description 8
- 230000000052 comparative effect Effects 0.000 description 6
- 210000003743 erythrocyte Anatomy 0.000 description 6
- 102100033237 Pro-epidermal growth factor Human genes 0.000 description 5
- 230000008569 process Effects 0.000 description 5
- 102000004127 Cytokines Human genes 0.000 description 4
- 108090000695 Cytokines Proteins 0.000 description 4
- 230000029087 digestion Effects 0.000 description 4
- 230000014509 gene expression Effects 0.000 description 4
- 230000012010 growth Effects 0.000 description 4
- 238000003752 polymerase chain reaction Methods 0.000 description 4
- 238000002474 experimental method Methods 0.000 description 3
- 239000000706 filtrate Substances 0.000 description 3
- 210000005260 human cell Anatomy 0.000 description 3
- 238000001727 in vivo Methods 0.000 description 3
- 239000006166 lysate Substances 0.000 description 3
- 238000002826 magnetic-activated cell sorting Methods 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 210000000056 organ Anatomy 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 2
- 108091033409 CRISPR Proteins 0.000 description 2
- 102400000921 Gastrin Human genes 0.000 description 2
- 101150111110 NKX2-1 gene Proteins 0.000 description 2
- 206010028980 Neoplasm Diseases 0.000 description 2
- 230000001464 adherent effect Effects 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 2
- 238000005119 centrifugation Methods 0.000 description 2
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 2
- 238000010494 dissociation reaction Methods 0.000 description 2
- 230000005593 dissociations Effects 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 230000008014 freezing Effects 0.000 description 2
- 238000007710 freezing Methods 0.000 description 2
- 238000003125 immunofluorescent labeling Methods 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
- 210000004962 mammalian cell Anatomy 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 230000010076 replication Effects 0.000 description 2
- 238000010186 staining Methods 0.000 description 2
- OYHQOLUKZRVURQ-NTGFUMLPSA-N (9Z,12Z)-9,10,12,13-tetratritiooctadeca-9,12-dienoic acid Chemical compound C(CCCCCCC\C(=C(/C\C(=C(/CCCCC)\[3H])\[3H])\[3H])\[3H])(=O)O OYHQOLUKZRVURQ-NTGFUMLPSA-N 0.000 description 1
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 1
- GVJHHUAWPYXKBD-UHFFFAOYSA-N (±)-α-Tocopherol Chemical compound OC1=C(C)C(C)=C2OC(CCCC(C)CCCC(C)CCCC(C)C)(C)CCC2=C1C GVJHHUAWPYXKBD-UHFFFAOYSA-N 0.000 description 1
- 102000009027 Albumins Human genes 0.000 description 1
- 108010088751 Albumins Proteins 0.000 description 1
- 238000010354 CRISPR gene editing Methods 0.000 description 1
- 102000004266 Collagen Type IV Human genes 0.000 description 1
- 108010042086 Collagen Type IV Proteins 0.000 description 1
- 101800003838 Epidermal growth factor Proteins 0.000 description 1
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 1
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 1
- 102000018233 Fibroblast Growth Factor Human genes 0.000 description 1
- 108050007372 Fibroblast Growth Factor Proteins 0.000 description 1
- 206010064571 Gene mutation Diseases 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 229920002971 Heparan sulfate Polymers 0.000 description 1
- HTTJABKRGRZYRN-UHFFFAOYSA-N Heparin Chemical compound OC1C(NC(=O)C)C(O)OC(COS(O)(=O)=O)C1OC1C(OS(O)(=O)=O)C(O)C(OC2C(C(OS(O)(=O)=O)C(OC3C(C(O)C(O)C(O3)C(O)=O)OS(O)(=O)=O)C(CO)O2)NS(O)(=O)=O)C(C(O)=O)O1 HTTJABKRGRZYRN-UHFFFAOYSA-N 0.000 description 1
- 102000004877 Insulin Human genes 0.000 description 1
- 108090001061 Insulin Proteins 0.000 description 1
- 102000007547 Laminin Human genes 0.000 description 1
- 108010085895 Laminin Proteins 0.000 description 1
- 102000002274 Matrix Metalloproteinases Human genes 0.000 description 1
- 108010000684 Matrix Metalloproteinases Proteins 0.000 description 1
- 125000003047 N-acetyl group Chemical group 0.000 description 1
- FABQUVYDAXWUQP-UHFFFAOYSA-N N4-(1,3-benzodioxol-5-ylmethyl)-6-(3-methoxyphenyl)pyrimidine-2,4-diamine Chemical compound COC1=CC=CC(C=2N=C(N)N=C(NCC=3C=C4OCOC4=CC=3)C=2)=C1 FABQUVYDAXWUQP-UHFFFAOYSA-N 0.000 description 1
- 102100037369 Nidogen-1 Human genes 0.000 description 1
- AUYYCJSJGJYCDS-LBPRGKRZSA-N Thyrolar Chemical compound IC1=CC(C[C@H](N)C(O)=O)=CC(I)=C1OC1=CC=C(O)C(I)=C1 AUYYCJSJGJYCDS-LBPRGKRZSA-N 0.000 description 1
- 102000004338 Transferrin Human genes 0.000 description 1
- 108090000901 Transferrin Proteins 0.000 description 1
- DTOSIQBPPRVQHS-PDBXOOCHSA-N alpha-linolenic acid Chemical compound CC\C=C/C\C=C/C\C=C/CCCCCCCC(O)=O DTOSIQBPPRVQHS-PDBXOOCHSA-N 0.000 description 1
- 235000020661 alpha-linolenic acid Nutrition 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 235000020958 biotin Nutrition 0.000 description 1
- 239000011616 biotin Substances 0.000 description 1
- 229960002685 biotin Drugs 0.000 description 1
- 210000001124 body fluid Anatomy 0.000 description 1
- 239000010839 body fluid Substances 0.000 description 1
- 238000012832 cell culture technique Methods 0.000 description 1
- 230000030833 cell death Effects 0.000 description 1
- 230000024245 cell differentiation Effects 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 210000003850 cellular structure Anatomy 0.000 description 1
- 235000012000 cholesterol Nutrition 0.000 description 1
- 229940107161 cholesterol Drugs 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000012136 culture 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
- 230000004069 differentiation Effects 0.000 description 1
- 102000038379 digestive enzymes Human genes 0.000 description 1
- 108091007734 digestive enzymes Proteins 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 229940116977 epidermal growth factor Drugs 0.000 description 1
- 229940126864 fibroblast growth factor Drugs 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- 239000003102 growth factor Substances 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 239000003112 inhibitor Substances 0.000 description 1
- 229940125396 insulin Drugs 0.000 description 1
- 229960004488 linolenic acid Drugs 0.000 description 1
- KQQKGWQCNNTQJW-UHFFFAOYSA-N linolenic acid Natural products CC=CCCC=CCC=CCCCCCCCC(O)=O KQQKGWQCNNTQJW-UHFFFAOYSA-N 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 230000001404 mediated effect Effects 0.000 description 1
- 230000000955 neuroendocrine Effects 0.000 description 1
- 108010008217 nidogen Proteins 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 230000001766 physiological effect Effects 0.000 description 1
- 235000004252 protein component Nutrition 0.000 description 1
- 235000018102 proteins Nutrition 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 229960000342 retinol acetate Drugs 0.000 description 1
- QGNJRVVDBSJHIZ-QHLGVNSISA-N retinyl acetate Chemical compound CC(=O)OC\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C QGNJRVVDBSJHIZ-QHLGVNSISA-N 0.000 description 1
- 235000019173 retinyl acetate Nutrition 0.000 description 1
- 239000011770 retinyl acetate Substances 0.000 description 1
- 238000010839 reverse transcription Methods 0.000 description 1
- 239000011435 rock Substances 0.000 description 1
- 210000000813 small intestine Anatomy 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 238000013518 transcription Methods 0.000 description 1
- 230000035897 transcription Effects 0.000 description 1
- 239000012581 transferrin Substances 0.000 description 1
- 230000001052 transient effect 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
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
- C12N5/0688—Cells from the lungs or the respiratory tract
-
- 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
- C12N5/0693—Tumour cells; Cancer cells
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2500/00—Specific components of cell culture medium
- C12N2500/30—Organic components
- C12N2500/32—Amino acids
-
- 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
- C12N2500/38—Vitamins
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/11—Epidermal growth factor [EGF]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/119—Other fibroblast growth factors, e.g. FGF-4, FGF-8, FGF-10
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/15—Transforming growth factor beta (TGF-β)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/155—Bone morphogenic proteins [BMP]; Osteogenins; Osteogenic factor; Bone inducing factor
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/30—Hormones
- C12N2501/345—Gastrin; Cholecystokinins [CCK]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/40—Regulators of development
- C12N2501/415—Wnt; Frizzeled
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/70—Enzymes
- C12N2501/72—Transferases [EC 2.]
- C12N2501/727—Kinases (EC 2.7.)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/70—Enzymes
- C12N2501/73—Hydrolases (EC 3.)
- C12N2501/734—Proteases (EC 3.4.)
-
- 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
- C12N2513/00—3D culture
Landscapes
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Life Sciences & Earth Sciences (AREA)
- Biomedical Technology (AREA)
- Genetics & Genomics (AREA)
- Zoology (AREA)
- Organic Chemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- Chemical & Material Sciences (AREA)
- Wood Science & Technology (AREA)
- General Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Cell Biology (AREA)
- Pulmonology (AREA)
- Oncology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
本发明公开了一种3D培养人肺组织的培养基，所述述培养基含有以下成分：B27、N‑acetylcysteine、EGF、Noggin、R‑spondin 1、A83‑01、FGF10、Nicotinamide、Y‑27632*、WNT3a、Glutamax、N2、Gastin中的一种或多种。本发明的培养基营养成分适宜，肺组织及肺癌组织可以有效的维持组织细胞特异性、干细胞特性形成遗传及生物学功能稳定肺组织/肺癌组织类器官，满足科学研究需要。
Description
技术领域
本发明属于生物医药领域，特别是用于人肺组织类器官3D培养的培养基，属于细胞培养的培养基材料。
背景技术
细胞培养技术是由一个或少量细胞经过大量培养成为简单的单细胞或极少分化的多细胞的技术。细胞在体内环境中，由机体体液循环系统供给生存复制的能量和原料，当进行体外培养的时候，往往需要根据不同的细胞的生长特点配制具有针对性的营养基质。\
人源组织类器官(Organoids)培养技术作为近来肿瘤学研究的新手段，体外3D培养条件下使用成熟人体细胞培养人体组织类器官(organoid)，这种类器官几乎保留了来源人体组织相同的遗传学特征，可在体外表现出与体内细胞相似的遗传物质以及结构的稳定性。目前，科学家已成功在体外使用成熟肝脏、小肠、消化道组织细胞培养出肝脏类器官、小肠类器官及食管类器官等，并利用这一技术在相关领域开展进行了多种研究。相对于传统的细胞培养，这一技术可以为我们提供充足的遗传稳定、均一的体外培养的组织细胞，并将可能对直接研究基因在人体细胞中的作用提供巨大的帮助。
现有技术中对于肺组织细胞的主要是普通的培养技术，在二维培养过程中肺组织细胞难以或不能充分的表达出肺组织的特性，使得培养的肺组织细胞和活体的肺组织细胞有所差异，不利于研究的进行。而在3D培养中，缺少必要的适宜的培养基的情况下，肺组织细胞的培养分化同样不利，难以充分模拟出体内的肺组织结构的生理特性。
虽然多种人源组织在不同的培养条件下可在体外成功培养类器官，但目前暂无关于肺组织及肺癌组织类器官培养方法的研究及报道，尤其是具体的培养条件即培养基质配方尚无尝试及报道。
发明内容
本发明的目的在于克服现有技术中二维培养技术的肺组织细胞活性不佳，肺/肺癌组织细胞体外3D类器官培养缺少针对性培养基的问题，提供一种人肺/肺癌组织3D类器官培养的培养基。
为了实现上述发明目的，本发明提供了以下技术方案：
一种3D培养人肺/肺癌组织的培养基，所述述培养基含有以下成分：B27、N-acetylcysteine、EGF、Noggin、R-spondin 1、A83-01、FGF10、Nicotinamide、Y-27632*、WNT3a、Glutamax、N2、Gastin一种或几种。
进一步，所述培养基中至少含有FGF10，R-spondin1，Noggin，WNT3a，A83-01，B27/N2。
进一步，所述3D培养人肺组织的培养基，包括以下多种成分配制而成，
| B27 | 50X稀释 |
| N-acetylcysteine | 1mM |
| EGF | 50ng/ml |
| Noggin | 100ng/ml |
| R-spondin 1 | 250ng/ml(或30％条件培养基) |
| A83-01 | 200nM |
| FGF10 | 500ng/ml |
| Nicotinamide | 10mM |
| Y-27632<sup>*</sup> | 10uM |
| WNT3a | 25ng/ml(或10％条件培养基) |
| Glutamax | 100X稀释 |
| N2 | 100X稀释 |
| Gastrin | 1nM |
本发明的3D培养人肺/肺癌组织的类器官培养基包括了多种针对于人肺组织及肺癌组织细胞培养所需要的细胞因子，各种细胞因子相互之间密切影响，协调配合，使得肺组织细胞在培养的过程中能够更好的表现出其固有的活性特征，实现高度近似于活体肺组织的综合特性。
与现有技术相比，本发明的有益效果：
1.本发明的肺组织及肺癌组织类器官的培养基针对于肺组织来源细胞的培养生长特点，选用了多种细胞因子成分按照一定的比例进行调和，经过调和的培养基中细胞因子的含量适宜，肺组织细胞有效在3D环境中形成类器官。
2.本发明的培养基营养成分适宜，肺组织及肺癌组织可以有效的维持组织细胞特异性、干细胞特性、形成遗传及结构等生物学特点稳定肺/肺癌组织类器官，满足科学研究需要。同时采用本发明的培养基可以完成肺/肺癌组织类器官的传代培养，达到大规模复制培养肺组织类器官的需求，控制培养得到的类器官具有高度的一致性。
附图说明：
图1是原代肺及肺癌类器官培养流程示意图。
图2是新鲜肺组织通过胶原酶解离为单细胞后第2、6、14天(第3代)在Matrixgel中生长情况。
图3是新鲜肺癌组织通过胶原酶解离为单细胞后第10天在Matrixgel中生长情况。
具体实施方式
本发明中的部分英文缩写解释如下：
DMEM：是一种应用十分广泛的培养基，可用于许多哺乳动物细胞培养，购买自GIBCO公司。
DMEM/F12：是F12培养基和DMEM培养基按照1：1结合，称为DMEM/F12培养基。综合了F12含有较丰富的成分和DMEM含有较高浓度养分的优点。购买自GIBCO公司。
Martrigel，从富含胞外基质蛋白的EHS小鼠肿瘤中分离出，其主要成分由层粘连蛋白，Ⅳ型胶原，巢蛋白，硫酸肝素糖蛋白等组成，还包含生长因子和基质金属蛋白酶等。购买自BD公司。
B27，即B27补充剂，市售产品，可用于配制培养基。B27补充剂以50倍的液体浓缩液提供，除其它成分外其包含生物素、胆固醇、亚油酸、亚麻酸、黄体酮、腐胺、视黄醇、视黄醇乙酸酯、亚硒酸钠、三碘甲状腺原氨酸(T3)、DL-α-生育酚(维生素E)、白蛋白、胰岛素以及转铁蛋白。购自Life Technologies公司。
N-acetylcysteine：N-乙酰半胱氨酸，购买自Sigma公司。
EGF，表皮生长因子，市售产品，购买自R&D公司。
Noggin，细胞生长蛋白成分，市售产品，购买自Peprotech公司。
R-spondin 1，人细胞生长编码蛋白，市售产品，购买自Peprotech公司。
A83-01，TGF-β抑制剂，购买自Tocris Bioscience公司。
FGF10，成纤维细胞生长因子，购买自Peprotech公司。
Nicotinamide，烟酰胺，购买自Sigma公司。
Y-27632*，ROCK特异性通路阻断剂。购买自Abmole Bioscience公司。
WNT3a，WNT激动剂，细胞中激活TCF/LEF-介导的转录的因子，购买自PeproTech公司。
Glutamax，市售细胞培养添加剂，购自：GIBCO公司。
N2，N2补充剂以100倍的液体浓缩液提供，其包含500μg/ml人转铁蛋白，500μg/ml牛胰岛素，0.63μg/ml黄体酮，1611μg/ml腐胺和0.52μg/ml亚硒酸钠。购自LifeTechnologies公司。
Gastrin，胃泌素，购买自Sigma公司。
TrypLE，用于解离贴壁哺乳动物细胞的重组消化酶，购买自GIBCO公司。
下面结合试验例及具体实施方式对本发明作进一步的详细描述。但不应将此理解为本发明上述主题的范围仅限于以下的实施例，凡基于本发明内容所实现的技术均属于本发明的范围。
实施例1
条件培养基
肺/肺癌组织类器官3D培养的(条件)培养基，其配方如下：细胞因子B2750X稀释；N-acetylcysteine1mM；EGF50ng/ml；Noggin100ng/ml；R-spondin 1250ng/ml；A83-01200nM；FGF10500ng/ml；烟酰胺10mM；Y-27632*10uM；WNT3a25ng/ml；Glutamax100X稀释；N2100X稀释；Gastrin1nM。
实施例2
条件培养基
肺组织类器官3D培养的(条件)培养基，其配方如下：细胞因子B27 50X稀释；N-acetylcysteine 1mM；EGF 55ng/ml；Noggin 110ng/ml；R-spondin 1 30％条件培养基；A83-01200nM；FGF10 500ng/ml；烟酰胺(Nicotinamide)10mM；WNT3a或10％条件培养基；Glutamax 100X稀释；N2 100X稀释。
实施例3
肺正常组织类器官培养
按照如图1所示的工艺顺序进行3D培养肺组织/肺癌组织类器官。取肺组织冰上剪碎，加入10ml胶原酶重悬，在gentalMACS C tube胶原酶中运行Human Lung程序1。转移至37℃、220rpm摇床消化20min。使用gentalMACS运行Human Lung程序2。转移至37℃、220rpm摇床消化10min，用100μm细胞筛网过滤细胞，滤液加入10ml DMEM/F12终止消化，离心(4℃，200g，5min)，去除上清。
取5ml红细胞裂解液重悬，冰上裂解红细胞5min。在4℃离心机中，200g离心5min，去除上清液。加入10ml DMEM/F12重悬，4℃、200离心5min，去除上清液。
细胞计数，混合Martrigel，20,000细胞每40μL，滴于48孔板孔正中，放置培养皿至37℃(5％CO2)10min，凝固Martrigel。每孔加入150μL实施例1制备的条件培养基，37℃、5％CO2细胞培养箱培养。每间隔2-3天更换一次培养基。
显微镜观察，肺组织细胞在培养基中形成圆形实心或空心的细胞团，表明3D体外培养肺组织及肺癌组织成功。常规收取类器官，提取RNA，逆转录后PCR，检测肺组织中相关基因表达情况。结果显示培养的类器官表达人肺组织相关基因。
实施例4
肺癌组织类器官培养
实验过程中同实施例3，取肺癌组织冰上剪碎，加入10ml胶原酶重悬，在gentalMACS C tube胶原酶中运行Human Tumor程序1。转移至37℃、220rpm摇床消化20min。使用gentalMACS运行Human Tumor程序2。转移至37℃、220rpm摇床消化10min，用100μm细胞筛网过滤细胞，滤液加入10ml DMEM/F12终止消化，离心(4℃，200g，5min)，去除上清。
取5ml红细胞裂解液重悬，冰上裂解红细胞5min。在4℃离心机中，200g离心5min，去除上清液。加入10ml DMEM/F12重悬，4℃、200离心5min，去除上清液。
细胞计数，混合Martrigel，20,000细胞每40μL，滴于48孔板孔正中，放置培养皿至37℃(5％CO2)10min，凝固Martrigel。每孔加入150μL实施例5制备的条件培养基，37℃、5％CO2细胞培养箱培养。每间隔2-3天更换一次培养基，使用实施例1制备的条件培养基。
收集类器官，滴入事先准备好的OCT包埋剂中，-80℃冰冻后切片(染色常规)，鉴定细胞来源。免疫荧光染色NKX2.1及Ki67阳性，q-PCR检测组织中的细胞来源于人肺组织相关基因表达。
实施例5
小鼠肺组织类器官培养
实验过程中同实施例3，取小鼠肺组织冰上剪碎，加入10ml胶原酶重悬，在gentalMACS C tube胶原酶中运行mouse Lung程序1。转移至37℃、220rpm摇床消化20min。使用gentalMACS运行mouse Lung程序2。用100μm细胞筛网过滤细胞，滤液加入10ml DMEM/F12终止消化，离心(4℃，200g，5min)，去除上清。
取5ml红细胞裂解液重悬，冰上裂解红细胞5min。在4℃离心机中，200g离心5min，去除上清液。加入10ml DMEM/F12重悬，4℃、200离心5min，去除上清液。
细胞计数，混合Martrigel，20,000细胞每40μL，滴于48孔板孔正中，放置培养皿至37℃(5％CO2)10min，凝固Martrigel。每孔加入150μL实施例5制备的条件培养基，37℃、5％CO2细胞培养箱培养。每间隔3-4天更换一次培养基，使用实施例1制备的条件培养基。
收集类器官，滴入事先准备好的OCT包埋剂中，-80℃冰冻后切片(染色常规)，鉴定细胞来源。免疫荧光染色NKX2.1及Ki67阳性，q-PCR检测组织中的细胞来源肺组织相关基因表达。体外利用CRISPR/Cas9技术进行基因突变，研究相关基因在肺相关疾病中发生发展中的作用。
对比例1
对比其他的普通培养基
采用现有的普通培养基(DMEM+10％FBS)3D条件下培养的分散的肺组织细胞，37℃、5％CO2细胞培养箱培养。每间隔2-3天更换一次培养基，结果肺组织细胞在培养过程中细胞附着在培养皿底部，与一般细胞培养结果类似，无法形成有结构的、多细胞成分的类器官。
对比例2
对比低FGF10浓度(100ng/ml)条件培养基
采用低FGF10浓度(100ng/ml)条件培养基在3D条件下中培养肺组织细胞，37℃、5％CO2细胞培养箱培养。对本该发明中的全培养基，低浓度FGF10培养条件下肺组织类器官形成较缓慢，q-PCR检测肺组织相关基因表达发现其中神经内分泌相关基因水平低于来源的肺组织细胞，且限制肺组织类器官传代代数(短暂传代2-3代)。
对比例3
对比无WNT3a条件培养基
采用不添加WNT3a的条件培养基在3D条件下中培养肺组织细胞，37℃、5％CO2细胞培养箱培养。对比本发明中的全培养基，该培养条件下肺组织类器官形成较缓慢、形成率低，部分细胞分化或死亡，对肺组织类器官传代培养有限制。
对比例4
对比无R-spondin 1条件培养基
采用不添加R-spondin 1的条件培养基在3D条件下中培养肺组织细胞，37℃、5％CO2细胞培养箱培养。对比本发明中的全培养基，该培养条件下肺组织类器官较难形成，部分细胞分化贴壁生长或凋亡；即使部分细胞培养获得类器官，但仍无法进行传代实验。
Claims (1)
1.一种3D培养人肺组织/肺癌组织的培养基，包括以下多种成分配制而成，
其中，
B27，即B27补充剂，购自Life Technologies公司；
Glutamax，细胞培养添加剂，购自GIBCO公司；
N2，即N2补充剂以100倍的液体浓缩液提供，其包含500μg/ml人转铁蛋白，500μg/ml牛胰岛素，0.63μg/ml黄体酮，1611μg/ml腐胺和0.52μg/ml亚硒酸钠，购自Life Technologies公司。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710183411.2A CN106834212B (zh) | 2017-03-24 | 2017-03-24 | 一种用于肺组织3d培养的培养基 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710183411.2A CN106834212B (zh) | 2017-03-24 | 2017-03-24 | 一种用于肺组织3d培养的培养基 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106834212A CN106834212A (zh) | 2017-06-13 |
| CN106834212B true CN106834212B (zh) | 2021-02-09 |
Family
ID=59130472
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710183411.2A Active CN106834212B (zh) | 2017-03-24 | 2017-03-24 | 一种用于肺组织3d培养的培养基 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106834212B (zh) |
Families Citing this family (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110452876A (zh) * | 2018-05-07 | 2019-11-15 | 北京吉尚立德生物科技有限公司 | 一种用于肺癌实体瘤原代细胞培养的细胞消化液 |
| CN110452875A (zh) * | 2018-05-07 | 2019-11-15 | 北京吉尚立德生物科技有限公司 | 一种用于培养肺癌实体瘤原代细胞的培养基 |
| WO2020133436A1 (zh) * | 2018-12-29 | 2020-07-02 | 江秀秀 | 一种诱导胚胎干细胞分化形成子宫内膜腺上皮前体细胞的方法和培养基 |
| CN109652376B (zh) * | 2019-01-08 | 2021-10-15 | 创芯国际生物科技(广州)有限公司 | 一种用于卵巢癌组织3d培养的培养基 |
| AU2019460208A1 (en) * | 2019-08-05 | 2022-03-17 | Genex Health Co., Ltd | Method for culturing primary cells of lung cancer solid tumor and primary tumor cells of lung cancer pleural effusion, and supporting reagent |
| CN110734894B (zh) * | 2019-10-11 | 2021-09-24 | 纳肽得(青岛)生物医药有限公司 | 普适性癌症类器官体外培养培养基 |
| CN111057680A (zh) * | 2019-12-10 | 2020-04-24 | 重庆康克唯生物科技有限公司 | 用于肺部肿瘤细胞的培养基和三维培养方法 |
| CN111548997A (zh) * | 2020-06-03 | 2020-08-18 | 杭州联众医疗科技股份有限公司 | 一种肺癌组织体外培养的方法 |
| CN112080471B (zh) * | 2020-08-26 | 2021-07-09 | 北京科途医学科技有限公司 | 顺铂耐药肺癌类器官及其培养方法、培养用培养基和用途 |
| CN114480250B (zh) * | 2020-11-12 | 2023-09-08 | 四川大学华西医院 | 构建原位原发胃癌动物模型的方法 |
| CN112592896B (zh) * | 2020-11-22 | 2024-06-25 | 深圳市第二人民医院(深圳市转化医学研究院) | 一种肺腺癌类器官的培养液及其培养方法 |
| CN112501110B (zh) * | 2020-11-26 | 2023-08-25 | 海西纺织新材料工业技术晋江研究院 | 三维培养肺及肺癌组织类器官的标准化培养基及培养方法 |
| CN114763534B (zh) * | 2021-01-13 | 2024-04-26 | 合肥中科普瑞昇生物医药科技有限公司 | 原代胃肠间质瘤细胞培养基、培养方法及其应用 |
| CN113481162B (zh) * | 2021-07-01 | 2023-02-24 | 丹望医疗科技(上海)有限公司 | 用于快速培养肿瘤类器官的培养基、方法以及试剂盒 |
| CN114736869B (zh) * | 2022-04-20 | 2024-05-28 | 上海交通大学医学院附属第九人民医院 | 一种黏膜黑色素瘤3d类器官及其培养方法和应用 |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US8628964B2 (en) * | 2006-10-11 | 2014-01-14 | Drexel University | Fetal pulmonary cells and uses thereof |
| EP2412800A1 (en) * | 2010-07-29 | 2012-02-01 | Koninklijke Nederlandse Akademie van Wetenschappen | Liver organoid, uses thereof and culture method for obtaining them |
-
2017
- 2017-03-24 CN CN201710183411.2A patent/CN106834212B/zh active Active
Non-Patent Citations (2)
| Title |
|---|
| Foetal hepatic progenitor cells assume a cholangiocytic cell phenotype during two-dimensional pre-culture;Kazuya Anzai et al.;《Scientific Reports》;20160623;第6卷;第1-13页 * |
| Three-dimensional pancreas organogenesis models;A. Grapin-Botton;《Diabetes Obes Metab》;20160930;第18卷;第33-40页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN106834212A (zh) | 2017-06-13 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN106834212B (zh) | 一种用于肺组织3d培养的培养基 | |
| CN106967672B (zh) | 一种肺及肺癌组织培养方法以及用其构建肺癌小鼠动物模型方法 | |
| CN109609441B (zh) | 一种肾脏组织类器官3d培养的培养基及类器官培养方法 | |
| CN109679915B (zh) | 一种鼻咽癌类器官的培养与鉴定方法 | |
| KR101443478B1 (ko) | 간엽계 줄기 세포를 포함한 세포 제제 및 그의 제조 방법 | |
| CN111197030A (zh) | 一种体外培养膀胱癌类器官的方法 | |
| CN112852714B (zh) | 构建原位原发肺癌动物模型的方法 | |
| CN109652376B (zh) | 一种用于卵巢癌组织3d培养的培养基 | |
| CN112522201A (zh) | 一种膀胱癌类器官的培养基及培养方法 | |
| CN114075539B (zh) | 构建原位原发膀胱癌动物模型的方法 | |
| WO2008056963A1 (en) | Method for proliferating stem cells with leptin | |
| CN115094022B (zh) | 肺癌成纤维细胞与肺癌类器官共培养模型的构建方法 | |
| CN113801838A (zh) | 原发性肝细胞癌类器官的持续培养方法和培养基 | |
| CN116024159A (zh) | 构建鼠类皮肤类器官的方法 | |
| CN114752565A (zh) | 带有免疫细胞的视网膜类器官及其构建方法 | |
| CN118345040A (zh) | 用于培养结直肠癌类器官的新型培养基 | |
| CN113943755B (zh) | 构建原位原发食管癌动物模型的方法 | |
| CN114891749B (zh) | 一种胰腺癌类器官的培养基及胰腺癌类器官的培养方法 | |
| CN114807039B (zh) | 一种食管癌肿瘤类器官培养用的培养基及培养方法 | |
| CN107312745B (zh) | 一种无血清的上皮细胞培养液 | |
| CN115851577A (zh) | 一种基于单细胞的骨肉瘤类器官模型的构建方法 | |
| CN115975907A (zh) | 培养基及其应用以及培养心脏类器官的方法 | |
| CN114369573B (zh) | 构建原位原发鼻咽癌动物模型的方法 | |
| CN112592883B (zh) | 一种小鼠胰腺类器官培养基及其应用 | |
| CN115161261A (zh) | 一种人胆管胆囊细胞3d培养的培养基与培养方法 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |