CN1490403A - Human inflammatory mammary cancer cell lines and its establishment - Google Patents
Human inflammatory mammary cancer cell lines and its establishment Download PDFInfo
- Publication number
- CN1490403A CN1490403A CNA031505384A CN03150538A CN1490403A CN 1490403 A CN1490403 A CN 1490403A CN A031505384 A CNA031505384 A CN A031505384A CN 03150538 A CN03150538 A CN 03150538A CN 1490403 A CN1490403 A CN 1490403A
- Authority
- CN
- China
- Prior art keywords
- breast cancer
- inflammatory breast
- clone
- posterity
- cell
- 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.)
- Pending
Links
Images
Abstract
A human inflammatory breast cancer's cell line is created by use of the transplanted tumor tissue from the naked mouse model of human inflammatory breast cancer as the tumor source and the external cutlure mode for subculturing to obtain said cell line with high transfer phynotype. Said cell line features that its cells appear as multipolar epthelial form, are collected to become flake or aggregate and adhered on wall firmly, and have the distorted number and structure of chromosomes. It can reproduce the clinic characteristics of inflammatory breast cancer.
Description
Technical field
The invention belongs to technical field of microbe cell line, be specifically related to a kind of foundation and establishment method thereof of people's inflammatory breast cancer clone.
Background technology
Mammary cancer is women's common malignancy, and the blood road shifts and lymphangitic spread is most important two routes of metastasis of mammary cancer.Distant metastasis is to influence tumor efficiency and prognosis, causes the principal element of death.Inflammatory breast cancer (inflammatory breast cancer, IBC) be a kind of rarer high malignancy, the specific type mammary cancer of prognosis extreme difference, its histopathology is characterized as the interior knurl bolt infiltration of lymphangitic spread-dermal lymphatics widely and causes " inflammatory " changes such as skin of mammary gland redness, occurs distant metastasis in a short time.Do not had the operation indication when most cases are gone to a doctor, the patient is lower than 2.5 years mean survival time (MST).
At present still not fully aware of to the molecular mechanism of the special biological property of inflammatory breast cancer, there are not ideal treatment plan and medicine yet.Up to now, the spontaneous animal model of inflammatory breast cancer is not successfully set up as yet, can't utilize early stage, responsive, the special index of exploring the prediction metastases at the biological study model of body metastases; Can't screen and verify original new drug; Can't provide technical support for the integrated control effect that further improves tumour.
The common method of setting up tumor cell line is to adopt tumour patient sample direct inoculation nude mice, breeds in a large number in nude mouse and passes through and set up stable transplanted tumor model in the body screening process, and capable again in-vitro cultivation obtains the stable clone that goes down to posterity.Another kind method be get patient tumors tissue block or cells in vitro directly carry out former be commissioned to train foster, screening and set up the stable clone that goes down to posterity.The factor that influences tumor cell line foundation is a lot, as the vigor state of tumor tissues, the size of tissue block, the kind selection of substratum etc.People's metastasis of cancer model of widespread usage is that people's cancer is inoculated in the immune deficiency animal at present, the human carcinoma cell line who mainly is meant the people's cancer nude mice metastasis model that forms and can takes place to shift in the immune deficiency animal body in the nude mouse body.Human breast carcinoma transplantation model success ratio only about 10%.
Summary of the invention
The purpose of this invention is to provide a kind of people's inflammatory breast cancer clone, for the research special biological characteristics of inflammatory breast cancer and the molecular mechanism of metastases provide cell model and research tool with high metastatic phenotype.
Another object of the present invention provides the establishment method of people's inflammatory breast cancer clone of high metastatic phenotype.
The present invention adopts human inflammatory breast cancer transplanted tumor in nude mice animal model (Shao Zhimin etc., Chinese surgical magazine), uses the in-vitro cultivation method, and screening is set up can stablize the inflammatory breast cancer clone that goes down to posterity, name IBC-1.
Inventor's inflammatory breast cancer clone IBC-1, cancer cells is multipole shape epithelium morphological specificity, assemble in blocks or cluster shape, adherent firm, indigestibility separates, karyomit(e) is the hypo-triploid caryogram, chromosomal structural aberration, 49 ~ 61 of modal number scopes, account for 71.6% of sum, cell generation cycle and apoptosis meet the tumour cell feature, S phase 42.90%, G2/G1:1.89, the tumor formation rate height, can reproduce the Clinical symptoms of inflammatory breast cancer, the exponential growth phase is at the 3rd ~ 6 day, the Western blot E-cadherin positive.Cell cryopreservation recovery back well-grown can continuous passage.
The present invention sets up inflammatory breast cancer clone by following method and step.
1, the former generation and the transplanting of going down to posterity:
1) by the aseptic tumor tissues that cuts of currently known methods, carries out subcutaneous in-situ inoculating of former generation.
2) the method continuation series of transplanting by former generation goes down to posterity.
2, oncocyte former be commissioned to train foster:
Get fresh inflammatory breast cancer transplanted tumor in nude mice tissue block, be cut into 1mm
3And evenly be attached at 50ml Corning
In the plastic culture bottle, add 5ml DMEM substratum, contain 20% foetal calf serum (FCS).CO
2Incubator, 37 ℃ of dry doubling walls are cultivated, treat around the tissue block free and grow cell, trysinization when propagation reaches certain area, M-199 substratum cultured continuously.
3, set up clone: go down to posterity 1 time every 5d, divided kind of rate 1: 2.Subculture in vitro separately to the carried out every biological indicator and detects after 16 generations, and note is made IBC-1.FBS reduces to 5% concentration and keeps and goes down to posterity in the substratum to 25 generations.
The subcutaneous in-situ inoculating tumor formation rate 100% of IBC-1 clone nude mice of the present invention has dermal lymphatics and vessel invasion.The spontaneous lung rate of transform is 100% after 4 weeks, and histopathological examination turns out to be adenocarcinoma metastatic.The tumor growth curve of above-mentioned transplanted tumor model meets the Gompertz function model, for being the exponential model growth after latent period.FCM analyzes showed cell, and to be distributed in people's inflammatory breast cancer clone IBC-1 proliferating cycle consistent with the transplanted tumor in nude mice cell, shows that the tumour cell division that goes down to posterity in the model is bred to tend towards stability, and can reproduce the biological property of people's inflammatory breast cancer growing multiplication preferably.
Description of drawings
Fig. 1 is an IBC-1 cellular form under the inverted microscope.
Fig. 2 a is an IBC-1 clone karyomit(e),
2b is IBC-1 chromosome karyotype analysis (hypo-triploid).
Fig. 3 is an IBC-1 nude mice in-situ inoculating transplanted tumor histopathology form (HE * 400).
Fig. 4 inflammatory breast cancer clone IBC-1 nude inoculation model of behaving.
Embodiment
Embodiment 1
Set up inflammatory breast cancer clone
1) the former generation and the transplanting of going down to posterity:
The aseptic tumor tissues that cuts carries out subcutaneous in-situ inoculating of former generation.
Orthotopic transplantation: remove non-tumor tissue and downright bad parts such as fiber, fat, the sterile saline rinsing, the knurl piece cuts into 0.5mm
3After the animal intraperitoneal anesthesia, the tincture of iodine, the female nude mouse mammary gland of alcohol disinfecting surface skin are inoculated in the subcutaneous lipids pad with tumor tissue.In former generation,, the transplanted tumor tumour grew to about 2cm
3The time, put to death nude mouse, cut former generation transplanted tumor;
The method continuation series of transplanting by former generation goes down to posterity.
2) oncocyte former be commissioned to train foster:
Get fresh inflammatory breast cancer transplanted tumor in nude mice tissue block, be cut into 1mm
3And evenly be attached at 50ml Cornin
In the plastic culture bottle, add 5ml DMEM substratum (20%FCS).Attaching is put into CO towards last with culturing bottle
2In the incubator, 37 ℃ of dry doubling wall 4h, after treating that tissue block is firmly adherent, the upset culturing bottle is dipped in tissue block and continues in the substratum to cultivate, and treats that tissue block is free on every side and grows cell, trysinization when propagation reaches at the bottom of the culturing bottle 60% area, M-199 substratum cultured continuously, the red corpuscle of sneaking in the cleavable cell mass, and be suppressed to the competitiveness growth of fibroblast.
3) set up clone: go down to posterity 1 time every 5d, divided kind of rate 1: 2.Subculture in vitro separately to the carries out the detection of every biological indicator after 16 generations, and note is made IBC-1.FBS reduces to 5% concentration and keeps and goes down to posterity in the substratum to 25 generations.
Experimentize and observe and checking, the IBC-1 of growth in vitro has following feature:
1) cellular form: cancer cells is multipole shape epithelium morphological specificity, assembles in flakes or the cluster shape, attaches firmly with the culturing bottle wall, separates than indigestibility.
2) chromosome analysis: the division phase cell of selecting karyomit(e) finely disseminated 100 mid-terms carries out chromosome analysis.Statistics shows that cell chromosome is the hypo-triploid caryogram, and chromosome number and structural aberration are arranged; 49 ~ 61 of modal number scopes account for 71.6% of sum.Table 1 is an inflammatory breast cancer clone IBC-1 chromosome karyotype analysis.
Table 1.
53，xx，der(1)t(1；7)，+add(1)(q21)，(q10；q10)，
+add(3)(p21)，
+add(3)(p13)，add(4)(q21)，+5，add(6)(p23)，add(7)
(q22)，add(8)(p21)，add(8)(p11.2)，add(9)(p12)，+add
(9)(q12)，add(11)(p11.2)，der(13)t(11；13)add(14)
(p11.2)，add(15)(q26)，+add(16)(q11.2)，？t(18；19)
(q11.2；q12)add(20)(q13.3)，add(21)(p12)，-22，+mar1，
+mar2[14]/
100~104，xxxx，idem，-12，-13，-14，-14，-17，-18，-18[CP2].
3) the genomic dna microsatellite sequence detects: people's inflammatory breast cancer clone IBC-1, former generation transplanted tumor, go down to posterity three of transplanted tumors at random the little satellite in site (D14S68, D18S69, D20S199) all amplify identical fragment, and nude mice normal galactophore tissue does not amplify corresponding banding pattern.
4) cell generation cycle and apoptosis: flow cytometry, detect the inflammatory breast cancer transplanted tumor cell S phase 39.38%, G2/G1:1.83; The IBC-1 clone S phase 42.90%, G2/G1:1.89, the two cell cycle all meets the tumour cell feature, has consistence.
5) tumor formation rate and growth pattern: tumor formation rate is 100%, and tumor growth meets the Gompertzian exponential Function Model.
6) inflammatory breast cancer clone IBC-1 growth curve, the exponential growth phase was at the 3rd ~ 6 day.
7) inflammatory breast cancer clone IBC-1 and orthotopic implantation model biological characteristics: Western blot detects and shows the IBC-1 and the transplanted tumor cell E-cadherin positive.
8) cell cryopreservation and recovery: cell cryopreservation recovery back well-grown, can continuous passage.Microbial contamination detects: cell does not have bacterium, fungi or mycoplasma contamination.
Claims (3)
1, a kind of people's inflammatory breast cancer clone, it is characterized in that can be external long term growth and stable going down to posterity, have following biological property and genetics feature: (1) cell is multipole shape epithelium morphological specificity, assemble in blocks or cluster shape, adherent firm, indigestibility separates, the forfeiture of contact growth-inhibiting; (2) cell is a hypo-triploid, chromosome number and structural aberration; (3) the in-situ inoculating tumor formation rate and the spontaneous lung rate of transform are 100%, can reproduce the Clinical symptoms of inflammatory breast cancer.
2, by the described people's inflammatory breast cancer of claim 1 clone, it is characterized in that 49 ~ 61 of described modal number scopes, account for 71.6% of sum.
3, a kind of by the described people's inflammatory breast cancer of claim 1 clone, it is characterized in that setting up by following method:
(1) the former generation and the transplanting of going down to posterity:
Asepticly carry out subcutaneous in-situ inoculating of former generation after cutting tumor tissues.
(2) oncocyte former be commissioned to train foster:
Get fresh inflammatory breast cancer transplanted tumor in nude mice tissue block, be attached in the culturing bottle, add 5mlDMEM substratum (20%FCS), CO
2Incubator, 37 ℃ of dry doubling wall 4h, adherent after, tissue block is dipped in the substratum and continues to cultivate propagation, trysinization, M-199 substratum cultured continuously.
(3) set up clone: go down to posterity 1 time every 5d, divided kind of rate 1: 2.Subculture in vitro separately to the is after 16 generations, and biological indicator detects, checking.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA031505384A CN1490403A (en) | 2003-08-25 | 2003-08-25 | Human inflammatory mammary cancer cell lines and its establishment |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA031505384A CN1490403A (en) | 2003-08-25 | 2003-08-25 | Human inflammatory mammary cancer cell lines and its establishment |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN1490403A true CN1490403A (en) | 2004-04-21 |
Family
ID=34156524
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNA031505384A Pending CN1490403A (en) | 2003-08-25 | 2003-08-25 | Human inflammatory mammary cancer cell lines and its establishment |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1490403A (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107988160A (en) * | 2018-01-11 | 2018-05-04 | 武汉大学深圳研究院 | Human breast cancer cell and its primary it is separately cultured and secondary culture method and purposes |
| CN108471732A (en) * | 2015-10-30 | 2018-08-31 | 杰克逊实验室 | With the relevant composition of tumor analysis and method |
| CN108504637A (en) * | 2017-07-10 | 2018-09-07 | 复旦大学附属肿瘤医院 | Chinese's clear cell carcinoma of ovary cell line |
| CN113667645A (en) * | 2021-08-19 | 2021-11-19 | 北京市农林科学院 | Canine breast cancer cell line, and establishment method and application thereof |
| CN113980904A (en) * | 2021-11-25 | 2022-01-28 | 华中农业大学 | Canine inflammatory breast cancer cell line and application thereof |
-
2003
- 2003-08-25 CN CNA031505384A patent/CN1490403A/en active Pending
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108471732A (en) * | 2015-10-30 | 2018-08-31 | 杰克逊实验室 | With the relevant composition of tumor analysis and method |
| CN108504637A (en) * | 2017-07-10 | 2018-09-07 | 复旦大学附属肿瘤医院 | Chinese's clear cell carcinoma of ovary cell line |
| CN107988160A (en) * | 2018-01-11 | 2018-05-04 | 武汉大学深圳研究院 | Human breast cancer cell and its primary it is separately cultured and secondary culture method and purposes |
| CN113667645A (en) * | 2021-08-19 | 2021-11-19 | 北京市农林科学院 | Canine breast cancer cell line, and establishment method and application thereof |
| CN113980904A (en) * | 2021-11-25 | 2022-01-28 | 华中农业大学 | Canine inflammatory breast cancer cell line and application thereof |
| CN113980904B (en) * | 2021-11-25 | 2023-11-28 | 华中农业大学 | Canine inflammatory breast cancer cell line and application thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102174469B (en) | Method for effectively culturing tumor infiltrating lymphocytes (TILs) | |
| CN101519646B (en) | CIK cell, as well as preparation method and cell preparation thereof | |
| CN102634482B (en) | Serum-free complete medium for mesenchymal stem cell | |
| CN102051344B (en) | Human osteosarcoma cell line group and mouse in-vivo transplantation model | |
| CN103422176B (en) | Construction method of human amniotic mesenchymal stem cell bank | |
| CN105296430B (en) | A kind of human colon cancer cells system DXH-1 and its application | |
| McAllister et al. | Colonial growth in agar of cells derived from adenovirus-induced hamster tumors | |
| CN1490403A (en) | Human inflammatory mammary cancer cell lines and its establishment | |
| CN1139655C (en) | Establishment of immortal human ovary carcinoma cell strain | |
| CN107653228A (en) | A kind of stomach cancer stem cell media and stomach cancer stem cell isolated culture method | |
| CN113667645A (en) | Canine breast cancer cell line, and establishment method and application thereof | |
| CN103131667B (en) | Pancreatic cancer cell line with lymphatic channel high migration activity | |
| CN103493735B (en) | The light regulate and control method that a kind of camplotheca acuminata callus is cultivated and bred | |
| CN105274058A (en) | Rat hepatoma cell strain with high metastatic potential and its preparation method and application | |
| CN101955911B (en) | Chinese lung adenocarcinoma cell line with high metastases potentiality of bone, lever and adrenal gland | |
| CN110295132A (en) | One plant has the active bacillus CCPM7645 of powerful anticancer and its application | |
| CN105602901A (en) | Bioreactor, stirring paddle thereof and method for culturing TIL cells by using bioreactor | |
| CN100408676C (en) | Establishing and application of scale cancer cell line of purify species New Zealand mice oral cavaty and palatal surface parts | |
| CN101429496A (en) | Culture medium for human peripheral blood source hemopoietic stem cell | |
| CN101429495A (en) | Cultivation method for human peripheral blood source hemopoietic stem cell | |
| CN103468746B (en) | Method for constructing tumor cell line | |
| Kwon et al. | Suspension culture of hematopoietic stem cells in stirred bioreactors | |
| CN111793605A (en) | Separation and extraction method of primary tumor cells of skin squamous carcinoma | |
| CN109042332B (en) | Tissue culture and rapid propagation method of tripterygium wilfordii | |
| CN101597591B (en) | Liver cancer cell line from liver cancer portal vein tumor thrombus of human and line establishing method thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C02 | Deemed withdrawal of patent application after publication (patent law 2001) | ||
| WD01 | Invention patent application deemed withdrawn after publication |