CN103194429A - Culturing and construction of human gastric cancer tumor stem cell line GAM-016S - Google Patents
Culturing and construction of human gastric cancer tumor stem cell line GAM-016S Download PDFInfo
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- CN103194429A CN103194429A CN2012100012635A CN201210001263A CN103194429A CN 103194429 A CN103194429 A CN 103194429A CN 2012100012635 A CN2012100012635 A CN 2012100012635A CN 201210001263 A CN201210001263 A CN 201210001263A CN 103194429 A CN103194429 A CN 103194429A
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Abstract
The invention relates to culturing and construction of a human gastric cancer tumor stem cell line GAM-016S. The invention is suitable for researches of relevant fields such as cytology, oncology, immunology, pharmacology, clinical medicine, and the like, and belongs to the technical area of cell culture. According to the invention, through in-vitro primary cell culture serum-free cell culture of patient gastric cancer tissue heterologously transplanted mice tumor mass, the tumor stem cell line GAM-016S is obtained. The cell line can form clone spheres under the serum-free cell culture condition. When 100 clone sphere cells are used in nude mice intraperitoneal injection, tumor mass can be formed. According to results of flow cytometry (FACS) and immunohistochemistry (IHC) analysis, the GAM-016S cell line expresses cancer stem cell marker molecules CD24 and CD44, and does not express CD133. The construction of GAM-016 cell line has important significances and values in the respects such as gastric cancer research, drug screening, clinical treatment, and the like.
Description
Technical field
The cultivation that the present invention relates to the strain of a kind of people's cancer of the stomach tumor stem cell is set up, and belongs to cytobiology cell culture technology field.
Background technology
The cancer cells that has the quantity rareness in the tumor tissues serves as the role of stem cell in neoplastic process, have the potential of self, propagation and differentiation, and its numerous character are similar to stem cell, therefore is called as tumor stem cell (TIC).Because the vital role in generation, development, recurrence and the transfer of tumour, TIC has become the focus of present tumor research.But in tumour cell group, TIC number rareness is cultivated unstablely relatively, and therefore developing efficiently vitro culture system cell amplification technology and keeping stem cell undifferentiated state technology has become the vital task of research now.
The research material of TIC is mainly from 3 aspects: patient's tumor specimen, tumor cell line and tumour transplatation knurl.Because tumor cell line obtains easily and cultivates, present many investigators use cell strain as the important materials of TIC research, regulate path in order to study TIC, for clinical treatment provides target spot.But can tumor cell line keep the hierarchical organization of former generation tumour to be queried always.The TIC surface markers that obtains by experiment in vitro or regulate adjusting molecule or gene on the path finally all will be verified by in vivo test again.Then solved this conforming problem with the tumour transplatation knurl to a great extent as the method for the research material of TIC.The heteroplastic transplantation tumour is to insert with coming from clinical tumor specimen that nude mice is subcutaneous to make it grow solid tumor, again the solid tumor that grows is continued the inoculation nude mice at the nude mice interior generation.Solid tumor passage number in nude mouse is more many, the more easy tumor cell line of setting up at short notice.The knurl block organization that obtains through heteroplastic transplantation is in the external preparation of carrying out single cell suspension, thereby the unicellular further separation and Culture that will prepare again obtains TIC.Though the heteroplastic transplantation meeting of tumour immunity deficient mice of former generation changes the living environment of stem cell, make the TIC that obtains in this way be still TIC near time of day in the body.
Summary of the invention
The research purpose of patent: cancer of the stomach is one of human modal malignant tumour, and its mortality ratio is number two in whole world cancer mortality.Though constantly there is the bibliographical information tumor stem cell extensively to be present in the various solid tumors, the research to the cancer of the stomach stem cell is very few for number at present.Because the significance of tumor stem cell in tumor research and treatment, separation and purifying that we use the solid tumor piece of allograft nude mice to carry out the cancer of the stomach stem cell are cultivated.
The result of study of patent: the present invention is by having obtained tumor stem cell strain GAM-016S to patient's stomach organization allograft mouse tumor piece in external primary cell culture, tumour clone formation experiment and serum-free cell cultures.This cell strain can form clone ball under the serum-free culture condition; Fluidic cell detects (FACS) and immunohistochemistry (IHC) analytical results shows GAM-016S expression of cell lines tumor stem cell marker molecule CD24, CD44, but does not express CD133; 100 clone ball cells of nude mice abdominal cavity injection can form the knurl piece.
The Research Significance of patent: the separation of people's cancer of the stomach tumor stem cell strain GAM-016S is built and is, can realize the large scale culturing to tumor stem cell, and then sets up corresponding perfect medicine sorting platform.Because the important biological action of tumor stem cell, the foundation of GAM-016S cell strain all has important theoretical meaning and practical value to the molecular mechanism of exploring the TIC biological behaviour, the clinical tumor research treatment that curing gastric cancer new drug and cancer of the stomach are researched and developed in screening.
Description of drawings
Fig. 1: GAM-016S clone forms and analyzes A. serum free medium B. nutrient agar
Fig. 2: the FACS of GAM-016S stem cell marker molecule and IHC analyze
Fig. 3: tumorigenesis capability analysis in the GAM-016S body
Embodiment
To come from clinical cancer of the stomach sample subcutaneous vaccination in B/C male nude mouse body, wait to grow the follow-up continued access kind of solid tumor nude mice at the nude mice interior generation.
To carry out trysinization from the heteroplastic transplantation tumor tissues that nude mice obtains and disperse to make single cell suspension, and separate with serum-free stem cell culture technique by primary cell culture, tumour clone formation experiment again and set up the cancer of the stomach stem cell.
1. tumor stem cell is cultivated: shear and use trysinization to obtain single cell suspension the knurl piece of heteroplastic transplantation; The single cell suspension that obtains is cultivated in the serum free medium that contains EGF, FGF-2; Use the culture dish of low-adhesion to cultivate to reduce the adhesive power of cell and support the growth that PD tumour cell is rolled into a ball.Change week about in the cell cultivation process and once contain the nutrient solution of cultivating the factor, till cell begins to form the cell mass of suspension.
2. the tumour clone forms experiment: the sepharose substratum (1: 1 agarose/nutrient solution) of preparation 0.5% is tiled in the culture dish bottom equably and prepares culture medium layer.The tumour cell group that serum-free culture is formed is separated into single cell suspension, and counting is also resuspended with 1: 1 agarose/nutrient solution, is laid on culture medium layer.It places 5%CO with culture dish
2, cultivate under 37 ℃ the wet condition.
3. fluidic cell detects (FACS) and immunohistochemistry detects (IHC): group is prepared into single cell suspension with the tumour clone cell, cleans and hatches with corresponding antibody (CD24, CD44, CD133) and carry out the FACS detection.IHC detection fixedly paraffin embedding film-making flake prepares sample, sample specimen is carried out antibody incubation again and detects.
4. tumorigenesis experiment in the body: group is prepared into single cell suspension with the tumour clone cell, counts and be resuspended in 1: among the 1PBS/Matrigel, be inoculated in the BALB/C nude mice under the cell skin with different quantities to observe tumorigenicity.GAM-016S is preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center (address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica); Preservation date is on December 9th, 2011; Deposit number: CGMCC No.5572; Classification name: people's cancer of the stomach stem cell.
Claims (4)
1. the present invention cultivates the cancer of the stomach tumor stem cell of having set up strain GAM-016s by name.
2. the present invention has used tumour heteroplastic transplantation technology, cultivates through primary cell culture, tumour clone formation experiment and serum-free stem cell and has obtained cancer of the stomach tumor stem cell strain GAM-016S.
3. the present invention builds the cancer of the stomach tumor stem cell strain GAM-016S that is, can form clone ball under the serum-free culture condition; 100 clone ball cells of nude mice abdominal cavity injection can form the knurl piece; Fluidic cell detects (FACS) and immunohistochemistry (IHC) analysis shows GAM-016S cell expressing tumor stem cell marker molecule CD24, CD44, but does not express CD133.
4. the present invention can be used at CD44, the development of CD24 targeting anti-tumor medicine.
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| CN2012100012635A CN103194429A (en) | 2012-01-05 | 2012-01-05 | Culturing and construction of human gastric cancer tumor stem cell line GAM-016S |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104593328A (en) * | 2014-12-30 | 2015-05-06 | 中国人民解放军第三军医大学第一附属医院 | Application and method for reversing Matrigel-induced tumor cells into tumor stem cells |
| CN110527666A (en) * | 2018-05-25 | 2019-12-03 | 香港中文大学 | A kind of sub-sieve enrichment method of cancer stem cell |
| WO2020206999A1 (en) * | 2019-04-11 | 2020-10-15 | 北京基石生命科技有限公司 | Method for culturing primary cells of gastric cancer and gallbladder and bile duct cancer, and supporting reagents |
| CN114533872A (en) * | 2020-11-19 | 2022-05-27 | 上海交通大学 | Method for treating gastric cancer targeting CD24 |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20110121410A (en) * | 2010-04-30 | 2011-11-07 | 부산대학교 산학협력단 | Novel cancer stem cell established from human gastric cancer tissues |
-
2012
- 2012-01-05 CN CN2012100012635A patent/CN103194429A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20110121410A (en) * | 2010-04-30 | 2011-11-07 | 부산대학교 산학협력단 | Novel cancer stem cell established from human gastric cancer tissues |
Non-Patent Citations (6)
| Title |
|---|
| ZHANG C ET AL: "Identification of CD44+ CD24+ gastric cancer stem cell", 《J CANCER RES CLIN ONCOL》 * |
| 凃琦 等: "CD44在胃癌中研究新进展", 《江西医药》 * |
| 张朝军 等: "肿瘤干细胞与胃癌", 《中国生物化学与分子生物学学报》 * |
| 袁亮 等: "胃癌干细胞的研究进展", 《福建医科大学学报》 * |
| 赵平: "《中国肿瘤临床年鉴》", 31 August 2008 * |
| 赵春华: "《干细胞原理技术与临床》", 31 May 2006 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104593328A (en) * | 2014-12-30 | 2015-05-06 | 中国人民解放军第三军医大学第一附属医院 | Application and method for reversing Matrigel-induced tumor cells into tumor stem cells |
| CN110527666A (en) * | 2018-05-25 | 2019-12-03 | 香港中文大学 | A kind of sub-sieve enrichment method of cancer stem cell |
| WO2020206999A1 (en) * | 2019-04-11 | 2020-10-15 | 北京基石生命科技有限公司 | Method for culturing primary cells of gastric cancer and gallbladder and bile duct cancer, and supporting reagents |
| CN114533872A (en) * | 2020-11-19 | 2022-05-27 | 上海交通大学 | Method for treating gastric cancer targeting CD24 |
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Application publication date: 20130710 |