CN105861441A - Hepatoma cell line STL-C1 derived from human hepatoma a-carcinoma tissue and establishment method thereof - Google Patents
Hepatoma cell line STL-C1 derived from human hepatoma a-carcinoma tissue and establishment method thereof Download PDFInfo
- Publication number
- CN105861441A CN105861441A CN201610202256.XA CN201610202256A CN105861441A CN 105861441 A CN105861441 A CN 105861441A CN 201610202256 A CN201610202256 A CN 201610202256A CN 105861441 A CN105861441 A CN 105861441A
- Authority
- CN
- China
- Prior art keywords
- cell
- cancer
- hepatoma
- stl
- beside organism
- 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.)
- Granted
Links
- 206010073071 hepatocellular carcinoma Diseases 0.000 title claims abstract description 45
- 238000000034 method Methods 0.000 title claims abstract description 24
- 201000009030 Carcinoma Diseases 0.000 title abstract 4
- 210000004027 cell Anatomy 0.000 claims abstract description 94
- 206010028980 Neoplasm Diseases 0.000 claims abstract description 62
- 201000011510 cancer Diseases 0.000 claims abstract description 45
- 230000015572 biosynthetic process Effects 0.000 claims abstract description 9
- 201000007270 liver cancer Diseases 0.000 claims description 48
- 208000014018 liver neoplasm Diseases 0.000 claims description 48
- 230000010412 perfusion Effects 0.000 claims description 23
- 239000000243 solution Substances 0.000 claims description 19
- 239000007788 liquid Substances 0.000 claims description 18
- 235000015097 nutrients Nutrition 0.000 claims description 17
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 claims description 16
- 102000029816 Collagenase Human genes 0.000 claims description 12
- 108060005980 Collagenase Proteins 0.000 claims description 12
- 229960002424 collagenase Drugs 0.000 claims description 12
- 239000007853 buffer solution Substances 0.000 claims description 10
- 238000011081 inoculation Methods 0.000 claims description 9
- 210000000130 stem cell Anatomy 0.000 claims description 9
- 238000010276 construction Methods 0.000 claims description 6
- 206010027476 Metastases Diseases 0.000 claims description 5
- 230000029087 digestion Effects 0.000 claims description 5
- 230000009401 metastasis Effects 0.000 claims description 5
- 238000010171 animal model Methods 0.000 claims description 4
- DEFVIWRASFVYLL-UHFFFAOYSA-N ethylene glycol bis(2-aminoethyl)tetraacetic acid Chemical compound OC(=O)CN(CC(O)=O)CCOCCOCCN(CC(O)=O)CC(O)=O DEFVIWRASFVYLL-UHFFFAOYSA-N 0.000 claims description 4
- 206010019695 Hepatic neoplasm Diseases 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
- 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
- 102000002070 Transferrins Human genes 0.000 claims description 2
- 108010015865 Transferrins Proteins 0.000 claims description 2
- 229940079593 drug Drugs 0.000 claims description 2
- 239000003814 drug Substances 0.000 claims description 2
- 239000004026 insulin derivative Substances 0.000 claims description 2
- 238000012216 screening Methods 0.000 claims description 2
- 238000011160 research Methods 0.000 abstract description 15
- 238000002474 experimental method Methods 0.000 abstract description 13
- 238000000338 in vitro Methods 0.000 abstract description 9
- 238000004321 preservation Methods 0.000 abstract description 5
- 238000012546 transfer Methods 0.000 abstract description 5
- 208000005189 Embolism Diseases 0.000 abstract description 3
- 230000007774 longterm Effects 0.000 abstract description 3
- 230000002440 hepatic effect Effects 0.000 abstract description 2
- 238000001727 in vivo Methods 0.000 abstract description 2
- 230000003902 lesion Effects 0.000 abstract description 2
- 210000004102 animal cell Anatomy 0.000 abstract 1
- 230000004791 biological behavior Effects 0.000 abstract 1
- 230000000813 microbial effect Effects 0.000 abstract 1
- 230000000381 tumorigenic effect Effects 0.000 abstract 1
- 210000001519 tissue Anatomy 0.000 description 19
- 230000012010 growth Effects 0.000 description 17
- 210000004185 liver Anatomy 0.000 description 16
- 230000001464 adherent effect Effects 0.000 description 9
- 238000011580 nude mouse model Methods 0.000 description 9
- 210000003240 portal vein Anatomy 0.000 description 9
- 230000007246 mechanism Effects 0.000 description 7
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 6
- 241000699660 Mus musculus Species 0.000 description 6
- 239000011248 coating agent Substances 0.000 description 6
- 238000000576 coating method Methods 0.000 description 6
- 230000006399 behavior Effects 0.000 description 5
- 210000000349 chromosome Anatomy 0.000 description 5
- 239000000463 material Substances 0.000 description 5
- 238000000926 separation method Methods 0.000 description 5
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical group CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 4
- 102000018651 Epithelial Cell Adhesion Molecule Human genes 0.000 description 4
- 108010066687 Epithelial Cell Adhesion Molecule Proteins 0.000 description 4
- 102100024785 Fibroblast growth factor 2 Human genes 0.000 description 4
- 108090000379 Fibroblast growth factor 2 Proteins 0.000 description 4
- 206010073069 Hepatic cancer Diseases 0.000 description 4
- 231100000844 hepatocellular carcinoma Toxicity 0.000 description 4
- 230000008569 process Effects 0.000 description 4
- 210000004881 tumor cell Anatomy 0.000 description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 4
- 206010027458 Metastases to lung Diseases 0.000 description 3
- 239000004677 Nylon Substances 0.000 description 3
- 108010019160 Pancreatin Proteins 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 3
- 230000008859 change Effects 0.000 description 3
- 238000004140 cleaning Methods 0.000 description 3
- 238000005520 cutting process Methods 0.000 description 3
- 238000001514 detection method Methods 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 239000012091 fetal bovine serum Substances 0.000 description 3
- 238000000684 flow cytometry Methods 0.000 description 3
- 239000001963 growth medium Substances 0.000 description 3
- 229920001778 nylon Polymers 0.000 description 3
- 229940055695 pancreatin Drugs 0.000 description 3
- 238000002271 resection Methods 0.000 description 3
- 229910052711 selenium Inorganic materials 0.000 description 3
- 239000011669 selenium Substances 0.000 description 3
- 229940091258 selenium supplement Drugs 0.000 description 3
- 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 2
- 241001465754 Metazoa Species 0.000 description 2
- 229930182555 Penicillin Natural products 0.000 description 2
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 2
- 108010087230 Sincalide Proteins 0.000 description 2
- 238000002835 absorbance Methods 0.000 description 2
- 239000000654 additive Substances 0.000 description 2
- 230000000996 additive effect Effects 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 238000010609 cell counting kit-8 assay Methods 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 208000019425 cirrhosis of liver Diseases 0.000 description 2
- 229960002897 heparin Drugs 0.000 description 2
- 229920000669 heparin Polymers 0.000 description 2
- 238000002347 injection Methods 0.000 description 2
- 239000007924 injection Substances 0.000 description 2
- 239000011159 matrix material Substances 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 230000009456 molecular mechanism Effects 0.000 description 2
- 230000000877 morphologic effect Effects 0.000 description 2
- 230000008520 organization Effects 0.000 description 2
- 230000001575 pathological effect Effects 0.000 description 2
- 229940049954 penicillin Drugs 0.000 description 2
- 239000008363 phosphate buffer Substances 0.000 description 2
- 230000003449 preventive effect Effects 0.000 description 2
- IZTQOLKUZKXIRV-YRVFCXMDSA-N sincalide Chemical compound C([C@@H](C(=O)N[C@@H](CCSC)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](N)CC(O)=O)C1=CC=C(OS(O)(=O)=O)C=C1 IZTQOLKUZKXIRV-YRVFCXMDSA-N 0.000 description 2
- 238000007920 subcutaneous administration Methods 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 238000001356 surgical procedure Methods 0.000 description 2
- 230000004083 survival effect Effects 0.000 description 2
- 230000005740 tumor formation Effects 0.000 description 2
- 238000011729 BALB/c nude mouse Methods 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- 208000005623 Carcinogenesis Diseases 0.000 description 1
- 206010008909 Chronic Hepatitis Diseases 0.000 description 1
- 208000000419 Chronic Hepatitis B Diseases 0.000 description 1
- 208000024678 Complex chromosomal rearrangement Diseases 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 206010016654 Fibrosis Diseases 0.000 description 1
- 239000012981 Hank's balanced salt solution Substances 0.000 description 1
- 241001582888 Lobus Species 0.000 description 1
- 206010050017 Lung cancer metastatic Diseases 0.000 description 1
- 206010058467 Lung neoplasm malignant Diseases 0.000 description 1
- 241000699666 Mus <mouse, genus> Species 0.000 description 1
- 241000610743 Psathyrotes ramosissima Species 0.000 description 1
- 208000007536 Thrombosis Diseases 0.000 description 1
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 230000006907 apoptotic process Effects 0.000 description 1
- 210000001367 artery Anatomy 0.000 description 1
- 239000012620 biological material Substances 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 230000036952 cancer formation Effects 0.000 description 1
- 231100000504 carcinogenesis Toxicity 0.000 description 1
- 239000012578 cell culture reagent Substances 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 230000004663 cell proliferation Effects 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 230000002759 chromosomal effect Effects 0.000 description 1
- 230000007882 cirrhosis Effects 0.000 description 1
- 238000010367 cloning Methods 0.000 description 1
- 230000001332 colony forming effect Effects 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 239000003636 conditioned culture medium Substances 0.000 description 1
- 210000002808 connective tissue Anatomy 0.000 description 1
- 239000013078 crystal Substances 0.000 description 1
- 230000034994 death Effects 0.000 description 1
- 238000000280 densification Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 230000009977 dual effect Effects 0.000 description 1
- 239000000975 dye Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 210000002950 fibroblast Anatomy 0.000 description 1
- 230000003328 fibroblastic effect Effects 0.000 description 1
- 239000000706 filtrate Substances 0.000 description 1
- 239000012737 fresh medium Substances 0.000 description 1
- 210000001652 frontal lobe Anatomy 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 239000003292 glue Substances 0.000 description 1
- 208000006454 hepatitis Diseases 0.000 description 1
- 208000002672 hepatitis B Diseases 0.000 description 1
- 210000003494 hepatocyte Anatomy 0.000 description 1
- 230000008676 import Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000011065 in-situ storage Methods 0.000 description 1
- 238000001802 infusion Methods 0.000 description 1
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Substances 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 1
- 229940125396 insulin Drugs 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 230000001788 irregular Effects 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 239000006193 liquid solution Substances 0.000 description 1
- 230000003908 liver function Effects 0.000 description 1
- 210000004072 lung Anatomy 0.000 description 1
- 201000005202 lung cancer Diseases 0.000 description 1
- 208000020816 lung neoplasm Diseases 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 239000012577 media supplement Substances 0.000 description 1
- 239000000155 melt Substances 0.000 description 1
- 230000001394 metastastic effect Effects 0.000 description 1
- 206010061289 metastatic neoplasm Diseases 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 210000001711 oxyntic cell Anatomy 0.000 description 1
- 230000007170 pathology Effects 0.000 description 1
- 230000002093 peripheral effect Effects 0.000 description 1
- 230000000258 photobiological effect Effects 0.000 description 1
- 238000010837 poor prognosis Methods 0.000 description 1
- 230000002980 postoperative effect Effects 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 238000012797 qualification Methods 0.000 description 1
- 238000004064 recycling Methods 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 229920006395 saturated elastomer Polymers 0.000 description 1
- 239000013049 sediment Substances 0.000 description 1
- 230000019491 signal transduction Effects 0.000 description 1
- 239000002356 single layer Substances 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 230000008961 swelling Effects 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 239000012588 trypsin Substances 0.000 description 1
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
- C12N5/0693—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
- C12N2500/05—Inorganic components
- C12N2500/10—Metals; Metal chelators
- C12N2500/20—Transition metals
- C12N2500/24—Iron; Fe chelators; Transferrin
- C12N2500/25—Insulin-transferrin; Insulin-transferrin-selenium
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/115—Basic fibroblast growth factor (bFGF, FGF-2)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2509/00—Methods for the dissociation of cells, e.g. specific use of enzymes
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)
- Oncology (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Cell Biology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The present invention relates to the field of microbial animal cells, and specifically a hepatoma cell line STL-C1 derived from human hepatoma para-carcinoma tissue. The cell line has a preservation number CCTCC No:C201630. The present invention also provides an establishment method and application of the cell line. The established hepatoma cell line STL-C1 is derived from human hepatocellular hepatoma para-carcinoma tissue, and can be cultured in vitro for a long term and has stable passage; and in vivo experiments show that the cell line still has good tumorigenic property, and is completely different from the currently existing established common hepatoma cell lines and even from the hepatoma cancer embolus cell line. The hepatoma cell line STL-C1 has important usage and reference value to the study of the formation of satellite lesions, and biological behaviors suahc as recurrence and transfer of primary hepatic carcinoma, is a novel cell model for basic and clinical research of hepatoma, and has broad application prospects.
Description
Technical field
The present invention relates to microorganism zooblast field, specifically, be a kind of isolated build the SMMC-7721 STL-C1 and its construction method and application being from human hepatocellular carcinoma cancer beside organism sample.
Background technology
Primary carcinoma of liver (Hepatocellular carcinoma, HCC) is one of modal malignant tumour of China, is only only second to lung cancer in tumour associated death, occupies second, patient's poor prognosis.Although surgery excision primary lesion is still current maximally effective liver cancer treatment measure, but has arrived middle and advanced stage during major part patient assessment and the inside and outside transfer of liver occurs, so Resection Rate is low, recurrence rate is high, clinical effectiveness is the most undesirable.Therefore, further investigation Primary Hepatic carcinogenesis, develop and the molecular mechanism that shift seem especially urgent and important for the liver cancer biological characteristics effective Results target spot of searching.
For the route of metastasis of liver cancer, being most commonly that tumour cell is directly diffused into other positions of liver by sinus hepaticus in liver and parasinoidal spaces, therefore MET presents the distribution of satellite shape, it is possible to away from primary tumor.And in clinical operation, surgeon also can retain residue liver on the premise of complete resection tumour (negative cutting edges) as far as possible as far as possible, it is to avoid patient's liver function is caused too much influence.Also having document to think, cancer beside organism refers mainly to the liver organization away from pathology 2cm, but cancer beside organism and cancerous tissue often have certain intersection in actual pathological tissue, can not distinguish with the clearest and the most definite boundary.So, even if operation can excise that primary tumor do not ensures that in primary tumor peripheral extent negative for tumor cells in the cancer beside organism of so-called " naked eyes are without obvious satellite stove " yet invade profit.Therefore, one of reason of still easily recurrence and transfer after this may also be operation of liver cancer.Make a general survey of current domestic and foreign literature, biological characteristics about the various aspects of HCC own has been made intensive research, equally, in the basic research of onset of liver cancer mechanism, cancer beside organism is also that the negative control as tumor tissues is widely used by vast researcher, but but rarely has report for the biological characteristics of HCC present in cancer beside organism.So, also the biological behaviour of the tumour cell that primary tumors cells and cancer beside organism are originated cannot be carried out comparative study, the most just be difficult to deeply open the molecular mechanism of hepatoma Metastasis, it is impossible to the effective clinical strategy finding prevention and treatment transfer.
In the research process of liver cancer, owing to primary carcinoma of liver has bigger heterogeneity, and the biological characteristics of different HCCs also has larger difference, so the external Bel7402 of foundation and Bel7402's animal model are the technological means that laboratory carries out that liver cancer research is indispensable.After Chen Rui inscription in 1962 establishes the first in the world strain SMMC-7721, domestic and international researcher in succession sets up and includes HepG2, a series of human hepatoma cell strain such as Hep3B, PLC/PRF/5.Because it continuous passage can be cultivated and can become knurl by Inoculation in vitro, preferably simulate generation development and its many-sided biological behaviour of liver cancer, have laid a good foundation for the research of onset of liver cancer mechanism and the exploration of clinical treatment, thus obtained relatively broad application.But along with going deep into liver cancer research, the research in terms of the biological behaviours such as the heterogeneity of tumour and Preventive is required more and more higher by researcher.Accordingly, it would be desirable to set up new hepatoma cell strain or SMMC-7721 animal model studies the biological characteristics of liver cancer different aspect further.
Chinese patent literature CN1338512A discloses the method for building up of people's liver cancer Lung metastases clone, i.e. utilizing human liver cancer tissue to inoculate in liver in situ makes nude mice become knurl and occur lung to shift, separating from metastatic lung cancer stove the most further and establish human hepatoma cell strain, mechanism and intervention target spot for research liver cancer Lung metastases provide preferable cell model.Chinese patent literature CN1232874A discloses the method for building up of the SMMC-7721 MHCC-97 with high metastatic potential.The nude mice model transplantable tumor that people's liver cancer height is shifted by researcher, as building the knurl source being, is alternately implemented by the way of inoculating in using in vitro culture associating nude mouse, thus is built up the human hepatoma cell strain with high Lung metastases rate.Chinese patent literature CN101597591A discloses and utilizes people's liver cancer and portal vein cancer MET is knurl source as building, by separating and the method for in-vitro cultivation establishes and derives from the SMMC-7721 of people's liver cancer and portal vein cancer, the biological behaviour of the research formation of liver cancer portal vein tumor thrombus, growth mechanism and cancer embolus derived cell system is provided important experiment material.And derive from the SMMC-7721 of people liver cancer cancer beside organism and construction method aspect has not yet to see report.
Summary of the invention
It is an object of the invention to provide the SMMC-7721 STL-C1 of a kind of people of deriving from liver cancer cancer beside organism and build system, method and application.
To achieve these goals, the present invention is by the following technical solutions:
A first aspect of the present invention, a kind of SMMC-7721 STL-C1 deriving from people liver cancer cancer beside organism is provided, its Classification And Nomenclature is the hepatoma cell strain STL-C1 deriving from cancer beside organism, it is preserved in China typical culture collection center (being called for short CCTCC), preservation date on February 26th, 2016, deposit number CCTCC No:C201630.
A second aspect of the present invention, it is provided that the construction method of the above-mentioned SMMC-7721 STL-C1 deriving from people liver cancer cancer beside organism, comprises the following steps:
A) perfusion of primary hepatoma cancer beside organism separates and obtains cell, carries out original cuiture；
B) formation and the expansion of cell island clone is cultivated；
C) the inoculation Secondary Culture of cell.
Wherein, the nutrient solution used in described original cuiture, expansion cultivation and Secondary Culture is containing 10%v/v hyclone (Fetal Bovine Serum, be called for short FBS) DMEM/F12 nutrient solution, possibly together with 10 μ g/ml rh-insulins, 5.5 μ g/ml transferrins, 5ng/ml sodium selenite and 20ng/ml people's recombinant bfgf stem cell factor in described nutrient solution.
Preferably, in described step a use tissue section multiple spot puncture combine under portal vein pipeline rout radioactivity coating puncture move under water perfusion method cancer beside organism is processed.
Preferably, cancer beside organism in described step a uses the method for buffer solution+collagenase perfusion liquid two step digestion to separate and obtain cancer beside organism's cell, described buffer solution is D-Hank ' the s buffer solution containing 0.02% (w/v) EGTA, and described collagenase perfusion liquid is D-Hank ' the s perfusion liquid containing 0.1% (w/v) IV Collagenase Type.
A third aspect of the present invention, it is provided that a kind of above-mentioned SMMC-7721 STL-C1 deriving from people liver cancer cancer beside organism application in preparing hepatoma model or animal model.
A fourth aspect of the present invention, it is provided that a kind of above-mentioned SMMC-7721 STL-C1 deriving from people liver cancer cancer beside organism is formed at research liver cancer satellite stove, the application in recurrence or metastasis inside and outside tumour liver.
A fifth aspect of the present invention, it is provided that a kind of above-mentioned SMMC-7721 STL-C1 deriving from people liver cancer cancer beside organism is in screening or the application of preparing in medicines resistant to liver cancer.
The invention has the advantages that:
1, STL-C1 SMMC-7721 has drawn from liver cancer patient cancer beside organism, has other current liver cancer or the incomparable advantage of cancer embolus clone to the mechanism of Preventive inside and outside research liver cancer satellite stove Forming Mechanism and tumour liver；In vitro can be with long-term cultivation and can stably pass on, experiment in vivo shows that this clone still has good oncogenicity, with the most existing and build the common SMMC-7721 being or even liver cancer opin cell system is entirely different, the satellite stove of research primary carcinoma of liver formed and the biological behaviour such as relapse and metastasis has important utilization and reference value；
2, Flow cytometry shows that this clone STL-C1 has the cell subsets of about about 4% can stably express surface marker CD133 and EpCAM that liver-cancer stem cell like cell subgroup is the most special, and prompting has a certain proportion of cell subsets to be likely to be of the characteristic of liver-cancer stem cell sample.Therefore, research liver cancer satellite stove is formed by the present invention and inside and outside liver, in transfer process, the effect of liver-cancer stem cell has certain reference significance and application prospect with mechanism；
3, cell of the present invention is in original cuiture period, owing to the separation method of external collagenase perfusion associating physical mechanical has considerable influence to cell state, simultaneously because the internal micro-environmental variation brought to external inoculated and cultured makes cell be in stress situation, survival is difficult to, and passes on the most difficult.By using appropriate Insulin-Transferrin-selenium additive (Insulin-Transferrin-Selenium in the present invention, and the DMEM/F12 nutrient solution (containing 10%v/v hyclone) of people's recombinant bfgf stem cell factor ITS), successfully maintain the survival that tumour cell is external, and gradually make it can stably expand growth and pass on, it is thus achieved that may being greatly increased of immortalization.At present this cell the most stably reaches about 20 generations, therefore, uses the conditioned medium of so combination can promote point cellifugal propagation and an amplification the most in vitro, and significantly improves and build the efficiency being；
4, the clone that the present invention stably sets up is examined under a microscope morphological feature mainly to show as size atypia obvious, in polygonal adherent growth, in vitro can long-term cultivation stably passing on, it has stronger Tumor formation to become knurl experiment display in nude mouse, illustrate that this clone still retains the biological characteristics of HCC, it is liver cancer basis and the new preferable cell model of clinical research, there is relatively broad application prospect.
5, primary hepatocyte is separated at present mainly by portal vein Seglen two step perfusion method, and the present invention is to be irrigated by the cancer beside organism that buffer solution+Post operation is taken off by the method for collagenase digesting liquid two step perfusion in vitro, and finally it is successfully separated required cell.Being different from situ liver has fixing portal vein and complete pipe-line system to can be used to the perfusion carrying out closing, and the outer cancer beside organism of human body is owing to operation reason of drawing materials is so existing multiple section, and pipeline is more, integrality is poor.According to such situation, the present invention has carried out a certain degree of improvement and adjustment to existing two step perfusions.Although it is little with perfusion liquid difference involved in seglen method in the selection of perfusion liquid and buffer solution, but the present invention is by separating a large amount of tissue samples summing up experience and gradually finding out tissue section multiple spot and puncture and combine under portal vein pipeline rout radioactivity coating the puncture method irrigated of moving under water and process cancer beside organism, and having preferable effect, cell separation effect is good, gained cytoactive is good.
The preservation information of biological material specimens:
Depositary institution: China typical culture collection center (is called for short CCTCC)
Address: Wuhan, China Wuhan University
Preservation date: on February 26th, 2016
Deposit number: CCTCC No:C201630
Classification And Nomenclature: human hepatoma cell strain STL-C1
Accompanying drawing explanation
The normal morphology figure (200x and 400x) that the STL-C1 HCC of Figure 1A-B: monolayer adherence growth is examined under a microscope.
Fig. 2: the growth curve chart of STL-C1 HCC in the DMEM/F12 nutrient solution (adding ITS and bFGF) containing 10% hyclone.
After Fig. 3: STL-C1 HCC inoculation, adherent rate changes over curve map.
Fig. 4: STL-C1 HCC plate clone forms lab diagram.
CD133 in Fig. 5: STL-C1 HCC (the 15th generation)+And EpCAM+Cell subsets streaming figure.
Fig. 6: STL-C1 HCC becomes knurl situation after being seeded to nude mice by subcutaneous.
Fig. 7: STL-C1 HCC chromosome karyotype analysis.
Detailed description of the invention
The detailed description of the invention provided the present invention below in conjunction with embodiment elaborates.
Embodiment 1
One, experiment reagent and material
1, experiment reagent
(1) cell culture reagent: DMEM/F12 (1:1) nutrient solution (Gibco, USA), 10% (v/v) hyclone (Biochrom AG, USA)；100U/L penicillin and the final working concentration of 0.1mg/L streptomysin (photo-biological company is won in Shanghai) are respectively 100U/mL and 0.1mg/mL；People's recombinant bfgf stem cell factor (Peprotech, USA)；Insulin-Transferrin-selenium additive (ITS Lipid Media Supplement, 100 ×) (Sigma, USA), it is made up of 1.0mg/ml rh-insulin, 0.55mg/ml HTrf and 0.5 μ g/ml sodium selenite, adds in the DMEM/F12 nutrient solution containing 10% (v/v) hyclone with the ratio of 1:100 during use；
(2) other cells are cultivated and cell separation material requested: D-Hank ' s balanced salt solution (pH value is 7.2-7.4), IV Collagenase Type (Sigma, USA), heparin injection, 50 μm aperture cell nylon mesh screen (BD Biosciences, USA), Percoll liquid, phosphate buffer (PBS) (pH=7.4), 0.25% (w/v) pancreatin (0.25g pancreatin melts in 100ml PBS) containing 0.5mM EDTA, cells frozen storing liquid (DMEM:FBS:DMSO forms with the volume ratio mixed preparing of 7:2:1) etc.；
(3) other Cell Biology Experiment materials: CCK-8 dyestuff (Dojindo, Japan), methanol solution, crystal violet, Anti-CD133-PE antibody and Anti-EpCAM-PE antibody (Miltenyi Biotec, USA), Matrix gel (BD Biosciences, USA) etc..
2, tissue-derived
Embodiment sample is taken at Dongfang Liver and Gall Surgery Hospital's 2012-11-01 specimens from pri (patient's admission number 129620), the male sex, 43 years old, because " B ultrasonic finds Liver masses 2 days " is admitted to hospital.Previously chronic hepatitis B, posthepatitic cirrhosis medical history.CT examination is pointed out: the right frontal lobe of liver is shown in nodositas low-density stove, diameter about 2cm, strengthens the strengthening of artery phase focus patch shape, portal vein phase and density period of delay and lowers, it is considered to primary carcinoma of liver.The preoperative sufferers themselves of obtaining agrees to and signs Informed Consent Form, detects and see in art: liver is that micronodular liver fibrosis changes.Tumour is positioned at lobus dexter V section, size about 1.5 × 1cm, is positioned at liver parenchyma, and quality is hard；Lump main body border is still clear, without obvious coating, periphery has no obvious sub-stove, and remaining liver does not touches lump.Excised cancerous swelling and part cancer beside organism sample, negative cutting edges.Postoperative pathological is diagnosed as hepatocellular carcinoma, thick beam type, II-III level；Chronic hepatitis C1S3.
3, animal used as test
BALB/c nude mice, mouse is 6-8 week age, purchased from Chinese Academy of Sciences's Shanghai Si Laike Experimental Animal Center, raises in liver and gall hospital signal transduction laboratory, east cleaning grade Animal House, ad lib, water inlet.Cell inoculation is all in purifying grade working table.
Two, experimental technique
We are from cancer beside organism's specimen sampling of this example liver cancer patient, the method using the digestion of " buffer solution+collagenase perfusion " two step separates and obtains cancer beside organism's cell, setting up, by steps such as original cuiture, the formation of cell island clone and the inoculation Secondary Culture expanding cultivation and cell, the clone that a strain can the most stably be passed on, concrete grammar is as follows:
1, sample obtains: take fresh cancer beside organism about 4g with scalpel from the sample of excision during above-mentioned patient's liver resection, ensures that section is neat when cutting as far as possible.Sample is placed in preprepared 50ml import centrifuge tube (the DMEM nutrient solution of prepackage 20ml serum-free in pipe), places on ice, and band is to laboratory；
2, the separation of liver cancer parietal cell and original cuiture:
With the clot of the other tissue surface of D-Hank ' the s buffer solution for cleaning liver cancer containing dual anti-(500U/mL penicillin and 500 μ g/mL streptomysins), and unnecessary connective tissue is wiped out, be placed in culture dish.First repeatedly irrigate D-Hank ' the s liquid containing 0.02% (w/v) EGTA and 50U/mL heparin injection of 38 DEG C of pre-temperature to go out organization internal clot with needle applicator along the tissue existing macroscopic entrance of section, then use tissue section multiple spot to puncture and combine the method puncturing perfusion of moving under water under portal vein pipeline rout radioactivity coating, give pre-temperature D-Hank containing 0.02% (w/v) EGTA ' s perfusion, until tissue block become from kermesinus the perfusion liquid of canescence and outflow become limpid till；Change D-Hank ' the s perfusion liquid containing 0.1% (w/v) IV Collagenase Type subsequently into continue to be irrigated according to the method described above, the recyclable recycling of collagenase perfusion liquid, until cancer beside organism's block gradually follows the string and perfusion liquid becomes cloudy, going out with the digested cell got off, naked eyes visible tissue surface presents turtleback crack, tissue inner structure becomes loose, easily dissociates.Tissue is carefully placed in new culture dish, adds 10ml DMEM nutrient solution, cut off tissue coating soft blunt separation tissue block, strip down having digested cell completely, and remove coating and the fibr tissue of last remaining.Being blown and beaten by digest subsequently is suspension, filters to 50mL centrifuge tube with the nylon screen (Nylon cell strainer) in 50 μm apertures.Removing after not digesting tissue, add 10mL DMEM culture medium suspendible in filtrate, 700rpm is centrifuged 1min, abandons supernatant, and the most resuspended sediment also adds 10ml DMEM nutrient solution and 5mL Percoll mixing, and under 4 DEG C of low temperature, 2500rpm is centrifuged 5min.After abandoning supernatant, again resuspended with culture medium and 700rpm is centrifuged 1min, gained cell precipitation is suspended in the DMEM/F12 culture medium containing 10% (v/v) hyclone, it is inoculated in 10cm culture dish after adding ITS and bFGF, puts into 37 DEG C of constant temperature, saturated humidity and 5%CO2Cell cultivate in incubator, and in the adherent situation of 12h observation of cell, and change liquid to remove the most adherent and dead cell, continue cultivation.
3, divide cellifugal clone to expand to cultivate
Fibroblast is removed by digestion-adherent method repeatedly.nullDuring adhere-wall culture，Seen from naked eyes, there is apoptosis in most cells，Still give aforementioned nutrient solution to cultivate，Fresh medium is changed in timing，After 2 weeks, culture dish local grows densification、Vigorous cell clone island，For obtaining the most purebred clone and avoiding the most fibroblastic pollution of other cells，Use cell clone digestion method，With marking pen, cell clone island bigger in culture dish is marked location the most under the microscope，Then phosphate buffer (PBS) rinses cell three times，0.25% pancreatin is dripped the digested 3min of incubator on cell clone，Then with fresh culture, postdigestive clone has been rushed sucking-off and has again been inoculated in 24 orifice plates，Add DMEM/F12 nutrient solution (containing 10%v/v hyclone) the continuation cultivation that 0.5mL contains 1 × ITS and 20ng/mL bFGF.
4, the passing on and cultivating of cell
Note observation of cell growing state and density, changed 2/3 fresh culture every 1-2 days, after treating that cell almost covers with 24 orifice plates, the most gradually digest and be passaged in 12 orifice plates, 6 orifice plates and 6cm dish.Routine observation according to the state of cell and the change of nutrient solution color, liquid is changed every 1-2 days, cell length passes on 0.25% trypsinization during 80-90% to bottom culture dish, and with the frozen stock solution containing 10% (v/v) DMSO, the cell of different algebraically is carried out frozen process.After in vitro culture 10 generation, cell proliferation is stable, detects its index of correlation, named STL-C1 SMMC-7721.Classification And Nomenclature is human hepatoma cell strain STL-C1, is preserved in China typical culture collection center (be called for short CCTCC), address: Wuhan, China Wuhan University, preservation date on February 26th, 2016, deposit number CCTCC No:C201630.
5, cellular morphology is observed
Living cells morphosis and the growth characteristic of cultivation is observed under inverted phase contrast microscope.
6, cell growth curve
Take the logarithm cell in good condition in growth period, resuspended with the DMEM/F12 nutrient solution (containing 1 × ITS and 20ng/mL bFGF) containing 10% (v/v) hyclone, with every hole 100 μ l (cell quantity 6 × 10 after Trypsin Induced3) be inoculated in 96 orifice plates, each time point intending measuring of each sample repeats 3 multiple holes.After choosing for 0 moment, 450nm wavelength absorbance value in 24 hours utilize CCK-8 method by ELIASA mensuration hole, continuous 6 days, with the time as transverse axis, draw growth curve with absorbance for drawing the longitudinal axis.
7, adherence rate
Ibid take the logarithm cell in good condition in growth period after digestion with 1.2 × 104Density is inoculated in 24 orifice plates, and each time point intending detection all prepares 3 multiple holes.Took one group of cell (3 secondary orifices) after inoculation every 3 hours and its attached cell is carried out trypsinization counting, till 24 hours.With the time point that measures as abscissa, attached cell number and total cell number ratio are that ordinate is drawn adherent rate and changed over figure.
8, plate clone forms experiment
Ibid processing, cell is with 1 × 103Density is inoculated in 6 orifice plates, the multiple hole of 3, each sample.Clone formation seen from naked eyes after cultivating 2 weeks, gives adding violet staining 15 minutes after methyl alcohol is fixed, carries out colony count after cleaning.Cloning efficiency=colony counts/inoculating cell number × 100%.
9, tumor stem cell ratio detection
Respectively choose cell in good condition about 2 × 105Respectively with 200 μ l precooling PBS resuspended after add Anti-CD133-PE and Anti-EpCAM-PE antibody 4 μ l, fully hatch 30 minutes on ice after mixing, it is centrifuged and washes twice with PBS, flow cytometry is carried out respectively, the ratio of whole cells shared by detection CD133 and EpCAM positive cell subgroup after resuspended.
10, subcutaneous one-tenth knurl experiment after cell inoculation
The STL-C1 HCC taken the logarithm growth period, with serum-free DMEM with Matrix gel glue 1:1 is resuspended is mixed into 5 × 10 after collected by trypsinisation is centrifugal7The cell suspension of cell/mL, according to 5 × 105With 1 × 107Two cell quantity levels are seeded in 6-8 week old male nude mouse dorsal sc respectively, observe into knurl situation after 2 weeks, and regular intervals of time is measured tumour major diameter and minor axis in 5 days, and weighed nude mice body weight, and gross tumor volume formula is: V=(minor axis2× major diameter)/2mm3.Take out tumour when the 45th day and take pictures.
11, chromosome karyotype analysis
Generation STL-C1 HCC in growth period the 15th of taking the logarithm is delivered to Beijing hundred Olympic Competition figure Genetic Biotechnologies Co., Ltd and is carried out karyotyping qualification.
Three, experimental result
1, morphological observation
Microscopic observation cell is polygonal adherent growth, not of uniform size, and form has different, irregular arrangement, and endochylema enriches, and core is big, substantially.See Fig. 1.
2, growth curve
Growth curve is as in figure 2 it is shown, cytotostatic growth in containing 10% (v/v) hyclone DMEM/F12 nutrient solution for cell, and growth rate is very fast.
3, adherence rate
Cell under above-mentioned condition of culture adherent well, it is adherent to plant 8 hours about 60-70%, and within 16 hours, basic 90% cell all can be with adherent growth.Adherence rate changes over sees Fig. 3.
4, plate clone experiment
Cell has certain clonality at above-mentioned condition of culture, and Colony forming is obvious, and formation rate is 4.5 ± 0.8%.Substantially Clone formation figure is shown in Fig. 4.
5, tumor stem cell ratio
CD133 positive cell subset proportions about 3.7% in flow cytometry results display STL-C1 HCC, EpCAM positive cell subset proportions is about 1.9%, as shown in Figure 5.
6, internal Tumor formation experiment
Take knurl to observe when cell infusion the 45th day, as shown in Figure 6,5 × 105With 1 × 107Three nude mices that two cell quantity levels are injected respectively all become knurl, and wherein 1 × 107The tumor mass that cell number magnitude inoculation nude mice is formed is noticeably greater than the tumor mass that low order of magnitude cell is formed.
7, chromosome karyotype analysis
Due to clone also exist complex chromosomal rearrangement, lack, the chromosomal variation (accounting for the 90% of total chromosome number) such as fracture, G shows band and cannot make and effectively match sequence, so experiment is only done chromosome counting and processed, result display clone is near diploid caryogram.
Below preferred embodiment to the invention is illustrated, but the invention is not limited to described embodiment, those of ordinary skill in the art it may also be made that modification or the replacement of all equivalents on the premise of the invention spirit, and modification or the replacement of these equivalents are all contained in the application claim limited range.
Claims (7)
1. deriving from a SMMC-7721 STL-C1 for people liver cancer cancer beside organism, its deposit number is
CCTCC No:C201630.
2. the SMMC-7721 STL-C1 deriving from people liver cancer cancer beside organism as claimed in claim 1
Construction method, comprise the following steps:
A) perfusion of primary hepatoma cancer beside organism separates and obtains cell, carries out original cuiture；
B) formation and the expansion of cell island clone is cultivated；
C) the inoculation Secondary Culture of cell.
The SMMC-7721 STL-C1's deriving from people liver cancer cancer beside organism the most according to claim 2
Construction method, it is characterised in that described original cuiture, expansion are cultivated and the cultivation of use in Secondary Culture
Liquid is the DMEM/F12 nutrient solution containing 10%v/v hyclone, in described nutrient solution possibly together with
10 μ g/ml rh-insulins, 5.5 μ g/ml transferrins, 5ng/ml sodium selenite and 20ng/ml people
Recombinant bfgf stem cell factor.
The SMMC-7721 STL-C1's deriving from people liver cancer cancer beside organism the most according to claim 2
Construction method, it is characterised in that the cancer beside organism in described step a uses buffer solution+collagenase perfusion
The method of liquid two step digestion separates and obtains cancer beside organism's cell, and described buffer solution is containing 0.02%w/v
D-Hank ' the s buffer solution of EGTA, described collagenase perfusion liquid is containing 0.1%w/v IV Collagenase Type
D-Hank ' s perfusion liquid.
5. the SMMC-7721 STL-C1 deriving from people liver cancer cancer beside organism as claimed in claim 1
Application in preparing hepatoma model or animal model.
6. the SMMC-7721 STL-C1 deriving from people liver cancer cancer beside organism as claimed in claim 1
Studying the application recurred inside and outside the formation of liver cancer satellite stove, tumour liver or in metastasis.
7. the SMMC-7721 STL-C1 deriving from people liver cancer cancer beside organism as claimed in claim 1
Screening or preparing the application in medicines resistant to liver cancer.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610202256.XA CN105861441B (en) | 2016-04-01 | 2016-04-01 | A kind of liver cancer cell lines STL-C1 from human hepatocellular carcinoma cancer beside organism and its build system, method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610202256.XA CN105861441B (en) | 2016-04-01 | 2016-04-01 | A kind of liver cancer cell lines STL-C1 from human hepatocellular carcinoma cancer beside organism and its build system, method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN105861441A true CN105861441A (en) | 2016-08-17 |
| CN105861441B CN105861441B (en) | 2018-06-12 |
Family
ID=56626724
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610202256.XA Expired - Fee Related CN105861441B (en) | 2016-04-01 | 2016-04-01 | A kind of liver cancer cell lines STL-C1 from human hepatocellular carcinoma cancer beside organism and its build system, method |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN105861441B (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108795851A (en) * | 2017-04-27 | 2018-11-13 | 大连医科大学 | The method for building up of people's salivary adenoid cystic carcinoma carcinoma-associated fibroblasts system CAF-SA a kind of and its application |
| CN108967417A (en) * | 2018-08-09 | 2018-12-11 | 爱克精医(北京)生物医药科技有限公司 | A kind of tumor tissues high activity storage transport liquid and preparation method thereof |
| CN109055302A (en) * | 2018-08-28 | 2018-12-21 | 重庆医科大学附属第医院 | The method of separation, culture and the identification of HBV transgenic mice primary hepatocyte |
| CN111321119A (en) * | 2020-03-18 | 2020-06-23 | 四川大学 | Liver cancer cell line suitable for large-scale serum-free adherent culture and establishment method and application thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101597591A (en) * | 2008-06-06 | 2009-12-09 | 中国人民解放军第二军医大学东方肝胆外科医院 | A kind of hepatoma cell line of the people's of deriving from liver cancer and portal vein cancer and build system, method |
| CN101693886A (en) * | 2009-08-10 | 2010-04-14 | 北京肿瘤医院 | Liver cancer cell line and application thereof |
| US20120295349A1 (en) * | 2009-02-20 | 2012-11-22 | Institute Of Nuclear Energy Research Atomic Energy Council, Executive Yuan | Mammal dedicated cell line from human hepatocellular carcinoma cell |
-
2016
- 2016-04-01 CN CN201610202256.XA patent/CN105861441B/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101597591A (en) * | 2008-06-06 | 2009-12-09 | 中国人民解放军第二军医大学东方肝胆外科医院 | A kind of hepatoma cell line of the people's of deriving from liver cancer and portal vein cancer and build system, method |
| US20120295349A1 (en) * | 2009-02-20 | 2012-11-22 | Institute Of Nuclear Energy Research Atomic Energy Council, Executive Yuan | Mammal dedicated cell line from human hepatocellular carcinoma cell |
| CN101693886A (en) * | 2009-08-10 | 2010-04-14 | 北京肿瘤医院 | Liver cancer cell line and application thereof |
Non-Patent Citations (1)
| Title |
|---|
| 江珊珊等: "肝癌肿瘤浸润淋巴细胞(TIL)体外扩增及特性研究", 《中国细胞生物学学报》 * |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108795851A (en) * | 2017-04-27 | 2018-11-13 | 大连医科大学 | The method for building up of people's salivary adenoid cystic carcinoma carcinoma-associated fibroblasts system CAF-SA a kind of and its application |
| CN108795851B (en) * | 2017-04-27 | 2022-03-18 | 大连医科大学 | Establishment method and application of human salivary adenoid cystic carcinoma related fibroblast cell line CAF-SA |
| CN108967417A (en) * | 2018-08-09 | 2018-12-11 | 爱克精医(北京)生物医药科技有限公司 | A kind of tumor tissues high activity storage transport liquid and preparation method thereof |
| CN108967417B (en) * | 2018-08-09 | 2019-05-10 | 爱克精医(北京)生物医药科技有限公司 | A kind of tumor tissues high activity storage transport liquid and preparation method thereof |
| CN109055302A (en) * | 2018-08-28 | 2018-12-21 | 重庆医科大学附属第医院 | The method of separation, culture and the identification of HBV transgenic mice primary hepatocyte |
| CN111321119A (en) * | 2020-03-18 | 2020-06-23 | 四川大学 | Liver cancer cell line suitable for large-scale serum-free adherent culture and establishment method and application thereof |
| CN111321119B (en) * | 2020-03-18 | 2022-09-20 | 四川大学 | Liver cancer cell line suitable for large-scale serum-free adherent culture and establishment method and application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN105861441B (en) | 2018-06-12 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Liang et al. | Aldehyde dehydrogenase-1 is a specific marker for stem cells in human lung adenocarcinoma | |
| JP2023521878A (en) | Culture medium, culture method, and use thereof for epithelial cells of esophageal squamous cell carcinoma | |
| Moutasim et al. | Cell migration and invasion assays | |
| CN105861441A (en) | Hepatoma cell line STL-C1 derived from human hepatoma a-carcinoma tissue and establishment method thereof | |
| CN102250840B (en) | Human pancreatic cancer cell line and its application | |
| CN103865876A (en) | Method for primary culture of tumor cells | |
| CN102465113B (en) | Human hepatoma carcinoma cell line and application thereof | |
| CN103451148B (en) | People's normal bronchial epithelial cell and primary separation and Culture thereof and Secondary Culture method and purposes | |
| CN105102614A (en) | Method for obtaining cell mass containing cancer stem cell | |
| CN102719403B (en) | A kind of human pancreas's adenosquamous carcinoma clone and establishment method thereof and application | |
| CN102690784B (en) | The foundation of hepatoma cell line HCC-LY10 and application | |
| CN103881975B (en) | Human Prostate Cancer Cells and primary separation and Culture thereof and Secondary Culture method and purposes | |
| CN104694476A (en) | Human non-small cell lung cancer cell line, and establishment method and application thereof | |
| CN102329775B (en) | A kind of human pancreatic cancer cell to gemcitabine resistance and application thereof | |
| CN110904026B (en) | Preparation method and application of hepatic precursor-like cells from different sources | |
| CN103966327B (en) | The application of a kind of miR-27a and diagnostic kit thereof | |
| CN107177551B (en) | Human intrahepatic bile duct cancer cell line with high tumor forming capability and application thereof | |
| CN104195109A (en) | Human lung adenocarcinoma cell line and application thereof | |
| CN101633922A (en) | Molecular marker hsa-miR-374a of breast carcinoma and application thereof | |
| CN102181399B (en) | Mouse liver tumor cell line for highly expressing CD133 and preparation method thereof | |
| CN104745530A (en) | Human hepatocellular carcinoma cell line, and establishing method and application thereof | |
| CN105062973B (en) | One plant carries HPV feminine gender penis squamous cell carcinomas system that TP53 is mutated and application thereof | |
| CN102168085A (en) | SiRNA for inhibiting expression of miR-130b gene and expression vector and application of siRNA or/and expression vector to preparation of medicament for improving treatment effect of liver cancer | |
| CN103911343B (en) | A kind of multiple organ shifts human bladder cancer cell | |
| CN108977410B (en) | Pancreatic cancer in-situ cancer cell line of Chinese people |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| CB03 | Change of inventor or designer information | ||
| CB03 | Change of inventor or designer information |
Inventor after: Wang Hongyang Inventor after: Yan Hexin Inventor after: Wang Mingda Inventor after: Wu Han Inventor after: Fu Gongbo Inventor after: Zhou Xu Inventor after: Zeng Min Inventor after: Huang Weijian Inventor before: Wang Hongyang Inventor before: Yan Hexin Inventor before: Wang Mingda Inventor before: Wu Han Inventor before: Fu Gongbo Inventor before: Zhou Xu |
|
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20180612 |