CN114717191A - 一株人肝内胆管癌细胞株icc-x3及其应用 - Google Patents
一株人肝内胆管癌细胞株icc-x3及其应用 Download PDFInfo
- Publication number
- CN114717191A CN114717191A CN202210448281.1A CN202210448281A CN114717191A CN 114717191 A CN114717191 A CN 114717191A CN 202210448281 A CN202210448281 A CN 202210448281A CN 114717191 A CN114717191 A CN 114717191A
- Authority
- CN
- China
- Prior art keywords
- bile duct
- duct cancer
- intrahepatic bile
- icc
- cell line
- 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
- 208000006990 cholangiocarcinoma Diseases 0.000 title claims abstract description 42
- 208000014899 intrahepatic bile duct cancer Diseases 0.000 title claims abstract description 40
- 201000007450 intrahepatic cholangiocarcinoma Diseases 0.000 title claims abstract description 32
- 206010028980 Neoplasm Diseases 0.000 claims abstract description 29
- 206010027476 Metastases Diseases 0.000 claims abstract description 9
- 230000007246 mechanism Effects 0.000 claims abstract description 9
- 230000009401 metastasis Effects 0.000 claims abstract description 9
- 208000005623 Carcinogenesis Diseases 0.000 claims abstract description 4
- 230000036952 cancer formation Effects 0.000 claims abstract description 4
- 231100000504 carcinogenesis Toxicity 0.000 claims abstract description 4
- 238000003745 diagnosis Methods 0.000 claims abstract description 4
- 230000005856 abnormality Effects 0.000 claims abstract description 3
- 238000010171 animal model Methods 0.000 claims abstract description 3
- 238000011161 development Methods 0.000 claims abstract description 3
- 230000018109 developmental process Effects 0.000 claims abstract description 3
- 230000004069 differentiation Effects 0.000 claims abstract description 3
- 230000008595 infiltration Effects 0.000 claims abstract description 3
- 238000001764 infiltration Methods 0.000 claims abstract description 3
- 238000004321 preservation Methods 0.000 claims abstract 2
- 229940079593 drug Drugs 0.000 claims description 2
- 239000003814 drug Substances 0.000 claims description 2
- 210000003228 intrahepatic bile duct Anatomy 0.000 claims description 2
- 238000012216 screening Methods 0.000 claims 1
- 210000004027 cell Anatomy 0.000 abstract description 87
- 238000011160 research Methods 0.000 abstract description 6
- 210000004102 animal cell Anatomy 0.000 abstract description 2
- 230000000813 microbial effect Effects 0.000 abstract description 2
- 238000011580 nude mouse model Methods 0.000 description 7
- 230000012010 growth Effects 0.000 description 6
- 210000001519 tissue Anatomy 0.000 description 5
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 4
- 241000699660 Mus musculus Species 0.000 description 4
- 238000004458 analytical method Methods 0.000 description 4
- 201000011510 cancer Diseases 0.000 description 4
- 210000000349 chromosome Anatomy 0.000 description 4
- 238000002474 experimental method Methods 0.000 description 4
- 230000001575 pathological effect Effects 0.000 description 4
- 210000004881 tumor cell Anatomy 0.000 description 4
- 241000699670 Mus sp. Species 0.000 description 3
- 201000010099 disease Diseases 0.000 description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 3
- 239000012634 fragment Substances 0.000 description 3
- 230000002068 genetic effect Effects 0.000 description 3
- 238000007490 hematoxylin and eosin (H&E) staining Methods 0.000 description 3
- 238000001727 in vivo Methods 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- IAKHMKGGTNLKSZ-INIZCTEOSA-N (S)-colchicine Chemical compound C1([C@@H](NC(C)=O)CC2)=CC(=O)C(OC)=CC=C1C1=C2C=C(OC)C(OC)=C1OC IAKHMKGGTNLKSZ-INIZCTEOSA-N 0.000 description 2
- 206010004593 Bile duct cancer Diseases 0.000 description 2
- 208000031639 Chromosome Deletion Diseases 0.000 description 2
- 206010059866 Drug resistance Diseases 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- 108091092878 Microsatellite Proteins 0.000 description 2
- 230000003698 anagen phase Effects 0.000 description 2
- 210000000013 bile duct Anatomy 0.000 description 2
- 210000003855 cell nucleus Anatomy 0.000 description 2
- 230000030944 contact inhibition Effects 0.000 description 2
- 238000012864 cross contamination Methods 0.000 description 2
- 210000004748 cultured cell Anatomy 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 230000029087 digestion Effects 0.000 description 2
- 229940088598 enzyme Drugs 0.000 description 2
- 230000003203 everyday effect Effects 0.000 description 2
- 206010073071 hepatocellular carcinoma Diseases 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 239000012188 paraffin wax Substances 0.000 description 2
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 description 2
- 230000004083 survival effect Effects 0.000 description 2
- 231100000588 tumorigenic Toxicity 0.000 description 2
- 230000000381 tumorigenic effect Effects 0.000 description 2
- NHBKXEKEPDILRR-UHFFFAOYSA-N 2,3-bis(butanoylsulfanyl)propyl butanoate Chemical compound CCCC(=O)OCC(SC(=O)CCC)CSC(=O)CCC NHBKXEKEPDILRR-UHFFFAOYSA-N 0.000 description 1
- 108700028369 Alleles Proteins 0.000 description 1
- 102000007325 Amelogenin Human genes 0.000 description 1
- 108010007570 Amelogenin Proteins 0.000 description 1
- 108090000145 Bacillolysin Proteins 0.000 description 1
- 102000008186 Collagen Human genes 0.000 description 1
- 108010035532 Collagen Proteins 0.000 description 1
- 108020004414 DNA Proteins 0.000 description 1
- 238000002738 Giemsa staining Methods 0.000 description 1
- 206010061598 Immunodeficiency Diseases 0.000 description 1
- 208000029462 Immunodeficiency disease Diseases 0.000 description 1
- 102000035092 Neutral proteases Human genes 0.000 description 1
- 108091005507 Neutral proteases Proteins 0.000 description 1
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 108010019160 Pancreatin Proteins 0.000 description 1
- 102100023361 SAP domain-containing ribonucleoprotein Human genes 0.000 description 1
- 101710139423 THO complex subunit 1 Proteins 0.000 description 1
- 208000035199 Tetraploidy Diseases 0.000 description 1
- 230000002159 abnormal effect Effects 0.000 description 1
- 229960000583 acetic acid Drugs 0.000 description 1
- 230000001464 adherent effect Effects 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 208000026900 bile duct neoplasm Diseases 0.000 description 1
- 210000003445 biliary tract Anatomy 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 230000002759 chromosomal effect Effects 0.000 description 1
- 229960001338 colchicine Drugs 0.000 description 1
- 229920001436 collagen Polymers 0.000 description 1
- 238000012790 confirmation Methods 0.000 description 1
- 239000006059 cover glass Substances 0.000 description 1
- 238000009795 derivation Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000008030 elimination Effects 0.000 description 1
- 238000003379 elimination reaction Methods 0.000 description 1
- 210000002919 epithelial cell Anatomy 0.000 description 1
- 201000008819 extrahepatic bile duct carcinoma Diseases 0.000 description 1
- 210000003194 forelimb Anatomy 0.000 description 1
- SDUQYLNIPVEERB-QPPQHZFASA-N gemcitabine Chemical compound O=C1N=C(N)C=CN1[C@H]1C(F)(F)[C@H](O)[C@@H](CO)O1 SDUQYLNIPVEERB-QPPQHZFASA-N 0.000 description 1
- 229960005277 gemcitabine Drugs 0.000 description 1
- 230000007614 genetic variation Effects 0.000 description 1
- 239000012362 glacial acetic acid Substances 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 230000007813 immunodeficiency Effects 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 201000007270 liver cancer Diseases 0.000 description 1
- 208000014018 liver neoplasm Diseases 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 230000003211 malignant effect Effects 0.000 description 1
- 238000009126 molecular therapy Methods 0.000 description 1
- 230000000877 morphologic effect Effects 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 229940055695 pancreatin Drugs 0.000 description 1
- 230000008506 pathogenesis Effects 0.000 description 1
- 229910052697 platinum Inorganic materials 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000004393 prognosis Methods 0.000 description 1
- 230000000644 propagated effect Effects 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 238000001959 radiotherapy Methods 0.000 description 1
- 239000012925 reference material Substances 0.000 description 1
- 230000008261 resistance mechanism Effects 0.000 description 1
- 230000000630 rising effect Effects 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 239000002356 single layer Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000010183 spectrum analysis Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000001356 surgical procedure Methods 0.000 description 1
- 238000011521 systemic chemotherapy Methods 0.000 description 1
- 230000005740 tumor formation Effects 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
- C12N5/0693—Tumour cells; Cancer cells
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New or modified breeds of animals
- A01K67/027—New or modified breeds of vertebrates
- A01K67/0271—Chimeric vertebrates, e.g. comprising exogenous cells
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/5005—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
- G01N33/5008—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
- G01N33/5011—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics for testing antineoplastic activity
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2207/00—Modified animals
- A01K2207/12—Animals modified by administration of exogenous cells
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2227/00—Animals characterised by species
- A01K2227/10—Mammal
- A01K2227/105—Murine
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2267/00—Animals characterised by purpose
- A01K2267/03—Animal model, e.g. for test or diseases
- A01K2267/0331—Animal model for proliferative diseases
-
- 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
- C12N2503/00—Use of cells in diagnostics
- C12N2503/02—Drug screening
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2500/00—Screening for compounds of potential therapeutic value
- G01N2500/10—Screening for compounds of potential therapeutic value involving cells
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Chemical & Material Sciences (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Zoology (AREA)
- Immunology (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Genetics & Genomics (AREA)
- General Health & Medical Sciences (AREA)
- Molecular Biology (AREA)
- Urology & Nephrology (AREA)
- Hematology (AREA)
- Environmental Sciences (AREA)
- Organic Chemistry (AREA)
- Wood Science & Technology (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Physics & Mathematics (AREA)
- General Engineering & Computer Science (AREA)
- Pathology (AREA)
- Oncology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Physics & Mathematics (AREA)
- Toxicology (AREA)
- Analytical Chemistry (AREA)
- Medicinal Chemistry (AREA)
- Animal Behavior & Ethology (AREA)
- Animal Husbandry (AREA)
- Biodiversity & Conservation Biology (AREA)
- Food Science & Technology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
一株人肝内胆管癌细胞株ICC‑X3及其应用，属于微生物动物细胞系领域。提供的肝内胆管癌细胞系为人肝内胆管癌细胞株ICC‑X3，已于2022年02月22日保藏于中国典型培养物保藏中心，保藏编号：CCTCC NO：C202260。人肝内胆管癌细胞株ICC‑X3可作为肝内胆管癌发生、发展或转移机制研究的细胞模型。人肝内胆管癌细胞株ICC‑X3也可以用于建立肝内胆管癌动物模型。所述人肝内胆管癌细胞株ICC‑X3可以作为研究肝内胆管癌的分化机制、细胞形态及功能异常、肿瘤浸润转移机制及指导临床综合诊治等的细胞模型。
Description
技术领域
本发明属于微生物动物细胞系领域，具体是涉及一株人肝内胆管癌细胞株ICC-X3及其应用。
背景技术
肝内胆管癌(Intrahepatic Cholangiocarcinoma，ICC)是第二常见的肝癌类型，在全球的发病率一直在上升。该肿瘤起源于胆道上皮细胞，其占所有胆管恶性肿瘤的10％～20％；手术只有在疾病早期才有可能的治愈性，而晚期不可切除的患者(70％～90％)预后较差，诊断后的生存时间小于12个月。以吉西他滨单独或联合铂类化疗药的全身化疗和放疗对改善患者的长期生存效果有限。
人类肿瘤细胞系，特别是那些具有完整数据和随访的细胞系，是肿瘤生物学研究的重要工具。20世纪80年代以来，科学家们建立并鉴定了多株胆管恶性肿瘤细胞系，但主要以肝外胆管癌为主，肝内胆管癌的细胞系报道很少。
来自胆道系统原发肿瘤的细胞系对于探索生物学和胆管肿瘤生成、检测药物敏感性、开发分子治疗靶点和耐药性研究耐药机制作用巨大。一个完善的肿瘤细胞系库应该反映出肿瘤表型的多样性并能够提供不同肿瘤异质性的细胞系；考虑到胆道系统肿瘤的不同病因及其与病因和种族相关的遗传变异，使用反映这些特征的适当的临床前模型是很重要的。
2002年，国内建成首例肝内胆管癌细胞系，为肝内胆管癌的基础和临床研究提供重要的细胞实验模型。但是，随着体外传代次数增多,细胞系一些特有的生物学特征会逐渐改变或消失，甚至会产生起源细胞本不具备的特征。另外，很多细胞系被检测到细胞交叉污染的现象，最常见的污染细胞源是HeLa,T-24和M14，使用这些细胞进行研究也会出现错误的结果。通过对从不同地区、不同实验室共收集到37株系的细胞共计46个样品，分别进行种属确认及STR谱分析，发现细胞交叉污染现象严重，错误率达62.6％。生物试剂和参考材料的成本约占总成本的三分之一。错误识别、污染、遗传漂变和克隆进化的细胞系是很多研究不可复制的重要原因，并影响到每个从事细胞研究的人。
新建立的细胞系可以最大程度上保持与原发肿瘤相似的特性，使用这种细胞系进行研究得出的结果最接近人体实际情况。因遗传背景不同，为更好地研究中国人的疾病，建立中国人特有的疾病模型非常必要。基于以上原因，不断建立新的细胞系并淘汰陈旧的细胞系，已经成为研究肝内胆管癌发病机理和治疗方法的重要环节。
发明内容
本发明的目的是提供一种人肝内胆管癌细胞系及其应用。
本发明提供的肝内胆管癌细胞系为人肝内胆管癌细胞株ICC-X3，已于2022年02月22日保藏于中国典型培养物保藏中心，地址：中国武汉武汉大学，邮编：430072，保藏中心保藏编号：CCTCC NO：C202260。
人肝内胆管癌细胞株ICC-X3可作为肝内胆管癌发生、发展或转移机制研究的细胞模型。人肝内胆管癌细胞株ICC-X3也可以用于建立肝内胆管癌动物模型。
本发明采用取自一62岁女性肝内胆管癌手术切除标本转移病灶，经Ⅱ型胶原酶/中性蛋白酶两者混合消化后作原代培养，经细胞培养技术建立了一个肝内胆管癌细胞系，命名为ICC-X3。
本细胞株具有以下生物学特性：
1、细胞贴壁生长，无接触抑制，可出现叠加生长现象。
2、细胞倍增时间接近36h。
3、细胞STR结果显示：ICC-X3细胞和同一患者的肿瘤组织的STR结果一致，细胞来源于同一肿瘤样品，并且在培养过程中没有被其它细胞污染。
4、染色体分析提示：本株细胞为稳定的亚四倍体细胞系，有较多的染色体缺失、异位及衍生现象。
5、裸鼠接种本细胞后全部成瘤，移植瘤组织学特性与原发瘤相似。
本发明所述的人肝内胆管癌细胞株ICC-X3可以作为研究肝内胆管癌的分化机制、细胞形态及功能异常、肿瘤浸润转移机制及指导临床综合诊治等的细胞模型。
附图说明
图1为ICC-X3细胞来源肿瘤组织的病理结果。
图2为ICC-X3细胞在显微镜下的形态学观察图(×100)。
图3为ICC-X3细胞STR结果。
图4为ICC-X3细胞的生长曲线。
图5为ICC-X3细胞染色体分析结果。
图6为ICC-X3细胞免疫缺陷小鼠体内成瘤性实验结果。
图7为裸鼠移植瘤的病理切片结果(×200)。
具体实施方式
为了使本发明的目的、技术方案及优点更加清楚明白，以下实施例将结合附图对本发明进行作进一步的说明。应当理解，此处所描述的具体实施例仅仅用于解释本发明，并不用于限定本发明。相反，本发明涵盖任何由权利要求定义的在本发明的精髓和范围上做的替代、修改、等效方法以及方案。下述实施例中所述实验方法，如无特殊说明，均为常规方法。
人肝内胆管癌细胞株ICC-X3的建立及鉴定。
一、人肝内胆管癌细胞株ICC-X3的建立
临床采集肝内胆管癌患者肿瘤转移灶，经混合酶消化后作原代培养，进而成功建立一株能够连续传代的肝内胆管癌细胞系，目前传至75代，细胞性状保持稳定。该患者病理结果为中-低分化肝内胆管癌，部分为未分化癌(图1)。
二、人肝内胆管癌细胞株ICC-X3的生物学特性检测
1、细胞形态学：细胞生长稳定并传化后，对培养细胞进行活细胞观察；另将生长于盖玻片上的单层细胞以体积分数为95％乙醇固定，行HE染色，光镜观察。结果显示：相差显微镜下见细胞呈上皮样细胞排列，细胞贴壁生长，有重叠生长现象(图2)。
2、短片段重复序列(STR)鉴定
短串联重复序列又称为微卫星DNA，是指染色体上，由数个碱基对作为核心单位(2-6个碱基对)，串联重复形成的一类DNA序列(重复次数为10～60多次，基因片段在400碱基对以下)；每个核心单位重复的次数会出现个体差异，从而形成片段长度不同的等位基因。因此，一组STR序列的重复次数在不同个体中几乎是唯一的，是个体的基因身份特征，也是细胞生物学对细胞身份和来源进行鉴定的主要方法。
收集新鲜培养的ICC-X3细胞和同一患者的肿瘤组织样品，抽提基因组DNA，由苏州鉴达生物科技有限公司提供STR检测服务，分别使用5’端荧光标记的引物进行PCR扩增，对所得产物进行测序，分析包括Amelogenin，THO1，TPOX，D13S317，vWA，D16S539，D5S818，CSF1PO以及D7S820等20个STR位点的序列重复数。上述序列和ATCC，DSMZ等细胞保存库的数据库进行查询对比，未返回相同的遗传图谱。ICC-X3细胞和同一患者的肿瘤组织的STR结果一致，说明细胞来源于同一肿瘤样品，并且在培养过程中没有被其它细胞污染(图3)。
3、细胞生长曲线测定：取处于对数生长期的第18代细胞制成1×105/ml的单细胞悬液，然后按照100ul/孔接种于96孔板，设置3个复孔。24h后开始加药测定490nm波长时的OD值，每天换液，共测9天。实验结束后绘制生长曲线，按公式Td＝t×lg2/lg(N1/N0)计算倍增时间。结果显示：本株细胞贴壁后第1天生长缓慢，第2天开始呈指数生长，在每天更换培养基营养充足的条件下细胞可出现叠加生长，无接触抑制现象；细胞平均倍增时间约36h(图4)。
4、染色体分析：取对数生长期的第30代细胞作核型分析。细胞经秋水仙素作用2h，经0.075mol/L KCl低渗，甲醇－冰醋酸固定处理，冰湿片滴片，室温老化后用胰酶处理，Giemsa染色显带分析。结果显示：本细胞系为稳定的亚三倍体细胞系。细胞染色体缺失、异位及衍生染色体等现象很常见，可见本细胞系具有恶性肿瘤细胞的特征(图5)。
5、免疫缺陷小鼠体内成瘤实验：取4周龄BALB/C雌性小鼠3只，右前肢皮下分别注射1×107个/100μl的培养细胞，4周后颈椎脱臼法处死裸鼠，取出移植瘤，福尔马林固定，石蜡包埋切片，HE染色，镜下观察，拍照记录实验数据。结果显示：裸鼠皮下接种细胞后1周，即可见移植瘤长出。3只裸鼠全部成瘤，移植瘤呈结节状，与皮肤有粘连(图6)。
6、肿瘤的病理切片：肿瘤组织福尔马林常规固定、石蜡包埋后切片及HE染色。结果显示：裸鼠体内成瘤标本镜下观察细胞生长旺盛，细胞核增大，深染，异型性明显，其组织学形态与临床标本相似(图7)。
上述实施例仅为本发明的较佳实施例，不能被认为用于限定本发明的实施范围。凡依本发明申请范围所作的均等变化与改进等，均应仍归属于本发明的专利涵盖范围之内。
Claims (5)
1.一株人肝内胆管癌细胞株ICC-X3，已于2022年02月22日保藏于中国典型培养物保藏中心，保藏中心保藏编号：CCTCC NO：C202260。
2.如权利要求1所述一株人肝内胆管癌细胞株ICC-X3在建立肝内胆管癌发生、发展或转移的细胞模型中的应用。
3.如权利要求1所述一株人肝内胆管癌细胞株ICC-X3在建立肝内胆管癌动物模型中的应用。
4.如权利要求1所述一株人肝内胆管癌细胞株ICC-X3在研究肝内胆管癌的分化机制、细胞形态及功能异常、肿瘤浸润转移机制及指导临床综合诊治的细胞模型中的应用。
5.如权利要求1所述一株人肝内胆管癌细胞株ICC-X3在研究肝内胆管癌的发生机理、筛选防治肝内胆管癌药物中的应用。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210448281.1A CN114717191B (zh) | 2022-04-26 | 2022-04-26 | 一株人肝内胆管癌细胞株icc-x3及其应用 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210448281.1A CN114717191B (zh) | 2022-04-26 | 2022-04-26 | 一株人肝内胆管癌细胞株icc-x3及其应用 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN114717191A true CN114717191A (zh) | 2022-07-08 |
| CN114717191B CN114717191B (zh) | 2023-06-16 |
Family
ID=82246622
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202210448281.1A Active CN114717191B (zh) | 2022-04-26 | 2022-04-26 | 一株人肝内胆管癌细胞株icc-x3及其应用 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN114717191B (zh) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115125213A (zh) * | 2022-07-14 | 2022-09-30 | 复旦大学附属中山医院 | 一种小鼠肝内胆管癌细胞系mIC-22及其应用 |
| CN115125214A (zh) * | 2022-07-14 | 2022-09-30 | 复旦大学附属中山医院 | 肺转移肝内胆管细胞癌(icc)原代细胞的建立方法 |
| CN115786262A (zh) * | 2022-11-29 | 2023-03-14 | 兰州大学第一医院 | 一种人肝门部胆管癌细胞系cbc3t-1及应用 |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104560878A (zh) * | 2015-01-04 | 2015-04-29 | 复旦大学附属中山医院 | 一种人肝内胆管癌细胞系及其用途 |
| CN107177551A (zh) * | 2016-03-11 | 2017-09-19 | 中国人民解放军第二军医大学东方肝胆外科医院 | 一种具有高成瘤能力的人肝内胆管癌细胞系及其应用 |
| US20190315832A1 (en) * | 2015-07-06 | 2019-10-17 | Immatics Biotechnologies Gmbh | Novel peptides and combination of peptides for use in immunotherapy against esophageal cancer and other cancers |
-
2022
- 2022-04-26 CN CN202210448281.1A patent/CN114717191B/zh active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104560878A (zh) * | 2015-01-04 | 2015-04-29 | 复旦大学附属中山医院 | 一种人肝内胆管癌细胞系及其用途 |
| US20190315832A1 (en) * | 2015-07-06 | 2019-10-17 | Immatics Biotechnologies Gmbh | Novel peptides and combination of peptides for use in immunotherapy against esophageal cancer and other cancers |
| CN107177551A (zh) * | 2016-03-11 | 2017-09-19 | 中国人民解放军第二军医大学东方肝胆外科医院 | 一种具有高成瘤能力的人肝内胆管癌细胞系及其应用 |
Non-Patent Citations (2)
| Title |
|---|
| GUILIANA CAVALLONI等: "Establishment and characterization of a human intrahepatic cholangiocarcinoma cell line derived from an Italian patient", TUMOR BIOL. * |
| 杨斌等: "安罗替尼对人肝内胆管细胞癌细胞系HCCC-9810作用研究", 中国医药导刊 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115125213A (zh) * | 2022-07-14 | 2022-09-30 | 复旦大学附属中山医院 | 一种小鼠肝内胆管癌细胞系mIC-22及其应用 |
| CN115125214A (zh) * | 2022-07-14 | 2022-09-30 | 复旦大学附属中山医院 | 肺转移肝内胆管细胞癌(icc)原代细胞的建立方法 |
| CN115786262A (zh) * | 2022-11-29 | 2023-03-14 | 兰州大学第一医院 | 一种人肝门部胆管癌细胞系cbc3t-1及应用 |
| CN115786262B (zh) * | 2022-11-29 | 2023-10-10 | 兰州大学第一医院 | 一种人肝门部胆管癌细胞系cbc3t-1及应用 |
Also Published As
| Publication number | Publication date |
|---|---|
| CN114717191B (zh) | 2023-06-16 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN114717191B (zh) | 一株人肝内胆管癌细胞株icc-x3及其应用 | |
| CN114606194B (zh) | 一株人肝内胆管癌细胞株icc-x1及其应用 | |
| Liu et al. | A high-risk retinoblastoma subtype with stemness features, dedifferentiated cone states and neuronal/ganglion cell gene expression | |
| Pearse et al. | Evolution in a transmissible cancer: a study of the chromosomal changes in devil facial tumor (DFT) as it spreads through the wild Tasmanian devil population | |
| Wistuba et al. | Comparison of features of human breast cancer cell lines and their corresponding tumors. | |
| CN105296430B (zh) | 一种人类结肠癌细胞系dxh-1及其应用 | |
| Hu et al. | Establishment, characterization, karyotyping, and comparative genomic hybridization analysis of HKESC-2 and HKESC-3: two newly established human esophageal squamous cell carcinoma cell lines | |
| CN107541494B (zh) | 一种人胆管癌细胞系及其应用 | |
| Dehner et al. | Malignant T cell activation by a bacillus species isolated from cutaneous T-cell lymphoma lesions | |
| CN102067828A (zh) | 人黑色素瘤高转移模型、细胞亚系以及建立方法和转移的动态检测 | |
| CN105255832A (zh) | 一种人胆管癌细胞系及其应用 | |
| CN115786262B (zh) | 一种人肝门部胆管癌细胞系cbc3t-1及应用 | |
| CN116426480A (zh) | 一种人胶质母细胞瘤细胞系y1203及其应用 | |
| CN107460171B (zh) | 人甲状腺未分化癌细胞系及其应用 | |
| KR20180000991A (ko) | 환자특이적 환자 유래 세포의 제조방법 및 이의 이용 | |
| US6316189B1 (en) | Evaluation of changes in breast cells using nipple aspirate fluid | |
| CN110317789A (zh) | 一种去除肿瘤细胞中支原体污染的方法及其应用 | |
| CN108048401A (zh) | 人胆道癌细胞系及应用 | |
| CN117625538A (zh) | 人肝内胆管癌细胞系icc-x2及其应用 | |
| CN117736993A (zh) | 一株人胰腺癌细胞系pdac-x3及其应用 | |
| Teixeira et al. | High frequency of clonal chromosome abnormalities in prostatic neoplasms sampled by prostatectomy or ultrasound‐guided needle biopsy | |
| CN105925530A (zh) | 高转移胰腺癌细胞株及其构建方法和应用 | |
| CN108467855A (zh) | 新的肺特异性转移肝癌细胞及其制备 | |
| CN117165534B (zh) | 一种永生化人胃癌成纤维细胞株及其构建和增殖强化方法 | |
| CN117431215A (zh) | 人下咽鳞癌细胞系及其应用 |
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 |