CN103131668B - A kind of Chinese's lung adenocarcinoma cell line without metastatic potential - Google Patents
A kind of Chinese's lung adenocarcinoma cell line without metastatic potential Download PDFInfo
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Abstract
The invention belongs to technical field of microbe cell line, relate to a kind of Chinese lung adenocarcinoma cell line Yang6 without metastatic potential and purposes.The present invention uses and takes from head and examine the hydrothorax of advanced pulmonary adenocarcinoma patient and make cell primary and cultivate, it is thus achieved that do not have the Chinese lung adenocarcinoma cell line Yang6 of metastatic potential, deposit number: CGMCC No.5404.The Chinese lung adenocarcinoma cell line Yang6 without metastatic potential of the present invention, adherent growth, there is not transitivity, cellular morphology is graceful rounded or polygonal segment, the speed of growth are slow, can be as the comparative study cell model of human lung adenocarcinoma, it has the molecular biological characteristic of uniqueness, heterogeneity, polymorphism, holds out broad prospects in the application of the metastasis and anti-metastasis drug screening that disclose human lung adenocarcinoma.
Description
Technical field
The invention belongs to technical field of microbe cell line, relate to Chinese's lung adenocarcinoma cell line, be specifically related to one and do not have
Have metastatic potential Chinese's lung adenocarcinoma cell line (The Chinese Human Lung Adenocarcinoma cell,
Yang6 especially parental cell tumorigenesis but Chinese's lung adenocarcinoma cell line (Yang6) having no transfer characteristic and application thereof.
Background technology
Pulmonary carcinoma is one of malignant disease of serious threat human survival and development, it has also become No.1 in tumor disease kills
Hands.Have statistics display, within 2006, pulmonary carcinoma new case about 174470 people makes a definite diagnosis in the U.S., account for whole tumor 13% (male) and
12% (women), sickness rate is only second to skin carcinoma；Die from pulmonary carcinoma person about 162460 people, account for whole tumor 31% (male) and
26% (women), mortality rate ranks first place [1～2]；The annual pulmonary carcinoma new cases of Japan more than 50000 people, mortality rate nearly 90%.
Shanghai City Incidence investigation in recent years shows, pulmonary carcinoma (21.56%) is in the Shanghai male malignancy sickness rate umber one,
Breast carcinoma then it is only second in women.
At present, in clinical treatment, lung cancer mortality for a long time high main reason is that be difficult to early diagnosis and its
High transitivity, many patient head are found to have multiple organ transfer when examining；Owing to the biological mechanism of lung cancer metastasis is the most not clear
Really, clinical lack effective Therapeutic Method, therefore, study of lung cancerometastasis pathogenesis to explore effective diagnosis and treatment method be domestic
Outer scholar striving direction for a long time.Currently, along with bases such as molecular biology, signal transduction path, protein function researchs
The development of subject, the mankind have had the development advanced by leaps and bounds to the research means of disease pathogenesis, but due to all objective restrictions,
The research in this field does not has important breakthrough yet.
Although host's microenvironment factor, right including osteoclast and some stromatins with somatomedin function etc.
The transfer tool of tumor has certain effect, but studies have found that, tumor cell own biological characteristic is undoubtedly Bone tumour and occurs
Main and direct reason.
Owing to most of advanced lung cancers all can shift, seldom have do not occur transfer clinical picture, therefore, if
Setting up the human lung adenocarcinoma cell model that transfer does not occurs while setting up effective human lung adenocarcinoma transfer cell model, this is not only
Can compare, screen, determine transfer mark, and can knock out, transfect disposal, for the generation of adenocarcinoma of lung, development, adenocarcinoma of lung mechanism
Research provides the good research/development platform of more preferable model, and this will assist in basic research level and the clinic diagnosis promoting adenocarcinoma of lung transfer
Level.
The list of references relevant with the present invention has:
[1] .American Cancer Society.Cancer facts and figures 2006.Atlanta, GA:
American Cancer Society；2006.pp.5～6
[2] .Alex G.Little a, E.Greer Gayb, Laurie E.Gasparc, Andrew
K.Stewartd.National survey of non-small cell lung cancer in the United
States:Epidemiology, pathology and patterns of care.Lung Cancer 2007,57 (2):
253-260
[3].Saburo Sone & Seiji Yano.Molecular pathogenesis and its
therapeutic modalities of lung cancer metastasis to bone.Cancer Metastasis
Rev.Springer Science+Business Media, LLC.2007
[4] .2003 Shanghai District Cancer Mortality. tumor, 2006,26 (7): 694.
[5] .J.S.Nam, A.M.Suchar, M.J.Kang, et al.Bone Sialoprotein Mediates the
Tumor Cell-Targeted Prometastatic Activity of Transforming Growth Factor beta
in a Mouse Model of Breast Cancer.Cancer Res.2006；66 (12): 6327-35.
[6] .Hayashi C, Rittling S, Hayata T, et al.Serum osteopontin, an enhancer
Of tumor metastasis to bone, promotes Bl6 melanoma cell migration.J Cell
Biochem.2007；101 (4): 979-86.
[7]. Yang Shunfang, Su Jianzhong, Cao Jie, Zhang Peiling, Lu Jianying, Xie Wenhui.Chinese Lungs gland cell system CPA-
The foundation of Yang1 and biological characteristics thereof.Lung cancer in China magazine 2009,12 (7): 753-759
[8]. Yang Shunfang, Su Jianzhong, Shi Meiping, Zhao Lanxiang, Zhang Peiling, Cao Jie, Lu Jianying, Xie Wenhui. Chinese Lungs gland
The foundation of cancerous cell line CPA-Yang2 and biological characteristics. lung cancer in China magazine, 2009,12 (10): 1055-1060.
[9]. Yang Shunfang, Shi Meiping, Cao Jie, Su Jianzhong, Zhao Lanxiang, thunder shellfish, Chang Cheng, Lu Jianying, Ye Jianding, Xie Wenhui.
Chinese Lungs gland cell system CPA-Yang3 and the foundation of Bone tumour cell thereof.Lung cancer in China magazine, 2011,14 (2): 79-
85。
Summary of the invention
It is an object of the invention to provide a kind of Chinese's lung adenocarcinoma cell line without metastatic potential；The present invention enters one
Step purpose is to provide the medical application of described lung adenocarcinoma cell line.
The present invention uses and takes from head and examine the hydrothorax of 65 years old female pulmonary adenocarcinoma patient and make cell primary culture materials, passes through
Cell is cultivated, it is thus achieved that does not has Chinese's lung adenocarcinoma cell line (Yang6) of metastatic potential, protects on October 28th, 2011
Hide, depositary institution: China Committee for Culture Collection of Microorganisms's common micro-organisms center (CGMCC), north, Chaoyang District, Beijing City
Occasion West Road 1 institute 3, deposit number: CGMCC No.5404, Classification And Nomenclature: Chinese lung adenocarcinoma cell line Yang6 (The
Chinese Human Lung Adenocarcinoma cell, Yang6).
The present invention is through experiment nude mice zoopery, as a result, it was confirmed that the described Chinese Lungs adenocarcinoma without metastatic potential
Cell strain (Yang6), adherent growth, there is not transitivity, the graceful rounded or polygonal segment of cellular morphology, the speed of growth is delayed
Slowly.
In the present invention, gathering advanced pulmonary adenocarcinoma head and examine the fresh hydrothorax 50 milliliters of patient, each Tissue Culture Flask inhales 8 milliliters
Hydrothorax and dual anti-after slightly shake up, put into cultivation temperature be 37 DEG C, CO2 concentration be 5% Germany congratulate Li Shi BB16 type carbon dioxide
Cultivate in incubating foster case；Light Microscopic observation after one week, finds sparse at the bottom of Tissue Culture Flask to grow cell；After 10 days, light Microscopic observation
Cell is grown the most at the bottom of Shi Faxian Tissue Culture Flask；Described cell growth speedup is unhappy, and cell is the least in adherent life
Long；Cellular morphology rule, polygon, uniformly；Initial stage of culture cell poor growth；Cellular morphology is unstable, reaches for the 9th generation, carefully
Born of the same parents' form finalizes the design almost, and speed of growth change is little；Nude mouse subcutaneous implantation cell 2 × 106/ only, often for equal tumorigenesis, but skin
Lower tumor poor growth, only grows 0.2-0.3cm in 3 months2Size tumor；After cryopreservation resuscitation, character is constant；
Above-mentioned cell growth status is observed: in attached cell, polygon and circle respectively account under difference inverted microscope
70% and 30%；Cell poor growth, it is seen that split coil method, multinuclear, it is thus achieved that the Chinese Lungs without metastatic potential of the present invention
Adenocarcinoma cell strain (Yang6).
In the present invention, in above-mentioned cell each generation, can make nude mice by subcutaneous tumorigenesis, parental cell and oncocyte plant 5 through left ventricle
Mice after 2 months row 99mTc-MDP micro pinhole Bone scans all do not detect Bone tumour stove；Each survival time of mice is all higher than
6 months.
Cell strain of the present invention has a following biological characteristics:
1, tumorigenesis rate is low, respectively for cell all can subcutaneous implantation tumorigenesis but tumor bulk-growth is slow；
2, shift without any Bone tumour and other organs.
The heretofore described Chinese lung adenocarcinoma cell line Yang6 without metastatic potential is special compatriots' lung gland
Cancerous cell line, its speed of growth is slow, and cellular morphology is abnormal, respectively for equal tumorigenesis but left ventricle modeling have no any Bone tumour and
Other organs shifts；Described cell strain Yang6 can be as the comparative study cell model of human lung adenocarcinoma, and it has dividing of uniqueness
Sub-biological characteristics, heterogeneity, polymorphism, have in the application of the metastasis and anti-metastasis drug screening that disclose human lung adenocarcinoma
Wide prospect.
Accompanying drawing explanation
Fig. 1 is p2 cell growth figure.
Fig. 2 shows p2 cell growth status.
Fig. 3 shows the form of 100 times of P3 cell.
Fig. 4 shows P6 cell form under 40 times of inverted microscopes, and wherein, after P6, cellular morphology is in stablizing, slowly giving birth to
Long.
Fig. 5 shows the forth generation censorship chromosome analysis situation of cell of the present invention.
Fig. 6 shows that cell of the present invention is hypo-triploid.
Fig. 7 shows the cultivation situation of cell of the present invention, wherein, turns out respectively from gathering fresh pleural effusion
For cell all can subcutaneous implantation tumorigenesis, subcutaneous implantation 1,000,000 cell but tumor bulk-growth is slow, after 3 experimental mouse being raised 5 months
Locate cell cultivation of after death drawing materials.
Fig. 8 shows Yang6 p6 cell H&E dyeing form.
Fig. 9 shows that 99mTc-MDP micro pinhole Bone scans left ventricle is injected million Yang6 cells and had no any bone
Transfer.
Figure 10 is fluorescent quantitative PCR result.
Detailed description of the invention
Embodiment 1
Take the faint yellow hydrothorax culture of first visit patients with lung adenocarcinoma, first for cell cultivation In vitro culture in he or she's hydrothorax
After, the lung adenocarcinoma cell that after 3 days, 5 bottles of Tissue Culture Flasks collect this patient reaches 300,000；With 2 × 106Cell concentration/0.1mL/ only, plants
Planting immunodeficient mouse BALB/c nu-nu nude mice oxter subcutaneous, SPF level raises that 1.5 months mices are subcutaneous grows 0.5cm3, 2
The moon only grows 0.5-0.8CM3, result shows, raises under nude mice SPF environment, and described cancerous cell can for cell from parental generation-p9
Subcutaneous tumorigenesis, but tumor poor growth；Subcutaneous tumors is taken out and does cell cultivation, test nude mice left ventricle after amplification and inject, 3 months
After to have not yet to see mice abnormal.
Yang6 cell growth status is observed: in attached cell, polygon and circle respectively account under difference inverted microscope
70% and 30%, cell poor growth, it is seen that split coil method, multinuclear；Under phase contrast microscope, p2 cell growth such as Fig. 1, shown in 2,
P3 cell 100 is the most as shown in Figure 3；40 times of inverted microscope P6 cells are as shown in Figure 4；After P6, cellular morphology is in stablizing, slowly giving birth to
Long；Under phase contrast microscope, living cells is the growth of atypical epithelium class cell, and sharpness of border, form be little and uniform, adherent growth；Carefully
Born of the same parents' form is with breast carcinoma MDA-MB-231, it is thus achieved that Chinese's lung adenocarcinoma cell line without metastatic potential of the present invention
(Yang6)。
The condition of culture of described culture, detection viable conditions be: culture medium for containing 10% hyclone RPMI1640 or
DMEM culture medium, pH=7, cultivation temperature is 37 DEG C, and 5%CO2 is incubated foster case and cultivated；Within 5-7 days, change culture fluid；
Described human lung adenocarcinoma cell (Yang6) 1 × 106, inject immunodeficient mouse blood system, claim every other day after two weeks
Weight, rises and carries out weekly Radionuclide Bone Imaging detection on the 3rd week, and Radionuclide Bone Imaging confirms this cell bone free transfer ability.This human lung adenocarcinoma
Cell (Yang6) can be as one rare, sluggish Chinese Lungs adenocarcinoma cell compares research.
Embodiment 2 gene expression dose comparative test
Use RT-PCR method that Chinese lung adenocarcinoma cell line Yang6 with A549 without metastatic potential obtained is carried out
The comparison of the expressions such as SERPINA1, IGFBP2, CDH1, KRT19, ARG2 and TMEM100 gene；Result shows, the present invention
Cell strain Yang6 be a special state Lu-csf-1, its speed of growth is slow, and cellular morphology is abnormal, and each generation all causes
Tumor but left ventricle modeling have no any Bone tumour and other organs transfer；
Further after utilizing this cell to do the sensitivity tests of 8 kinds of chemotherapeutics, clinical according to result of the test use promise
The patients with lung adenocarcinoma of Wei Ben+cisplatin clinical treatment embodiment 1, shows certain curative effect.Its tool of described cell strain Yang6
There are the molecular biological characteristic of uniqueness, heterogeneity, polymorphism, can disclose as the comparative study cell model of human lung adenocarcinoma
The application of the metastasis of human lung adenocarcinoma and anti-metastasis drug screening will there is wide prospect.
Claims (2)
1. Chinese's lung adenocarcinoma cell line without metastatic potential, it is characterised in that the Classification And Nomenclature of described cell strain:
Human lung adenocarcinoma cell line Yang6, deposit number: CGMCC No.5404, preservation date on October 28th, 2011.
2. the Chinese's lung adenocarcinoma cell line without metastatic potential described in claim 1 is screening anti-human adenocarcinoma of lung transfer medicine
Purposes in thing.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201110383668.5A CN103131668B (en) | 2011-11-26 | A kind of Chinese's lung adenocarcinoma cell line without metastatic potential |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201110383668.5A CN103131668B (en) | 2011-11-26 | A kind of Chinese's lung adenocarcinoma cell line without metastatic potential |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103131668A CN103131668A (en) | 2013-06-05 |
| CN103131668B true CN103131668B (en) | 2016-12-14 |
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1712521A (en) * | 2004-12-15 | 2005-12-28 | 上海市胸科医院 | Lung cancer strain with high potential power of bone transference and its generation |
| CN101717753A (en) * | 2008-10-09 | 2010-06-02 | 上海市胸科医院 | Chinese lung adenocarcinoma cell strain having metastatic potential of multiple organs |
| CN101717752A (en) * | 2008-10-09 | 2010-06-02 | 上海市胸科医院 | Parental human lung adenocarcinoma cell line with high potential of bone metastasis |
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1712521A (en) * | 2004-12-15 | 2005-12-28 | 上海市胸科医院 | Lung cancer strain with high potential power of bone transference and its generation |
| CN101717753A (en) * | 2008-10-09 | 2010-06-02 | 上海市胸科医院 | Chinese lung adenocarcinoma cell strain having metastatic potential of multiple organs |
| CN101717752A (en) * | 2008-10-09 | 2010-06-02 | 上海市胸科医院 | Parental human lung adenocarcinoma cell line with high potential of bone metastasis |
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