人结直肠癌裸鼠移植转移模型构建的实验研究

发布时间：2018-05-11 11:29

本文选题：人结直肠癌 + 淋巴道转移　；参考：《重庆医科大学》2008年硕士论文

【摘要】： 结直肠癌是消化道常见肿瘤,恶性程度较高,易发生淋巴性转移和消化道种植播散。肿瘤浸润和转移不仅影响着临床治疗效果,也是患者肿瘤复发、死亡的主要原因,因而对结直肠癌转移机制及其防治对策的研究是当前的重要任务。而人结直肠癌裸鼠移植转移模型的建立是当前国内外关注的课题,为了能更好地研究结直肠癌转移机制和防治对策,建立稳定、可靠、重复性好的适合动物模型是十分必要的。这可为进一步研究结直肠癌的浸润转移机制、药物干预等抗转移治疗提供有效的实验平台,利于改观结直肠癌的预后和结局。但目前尚未构建成功较为理想的结直肠癌单纯性淋巴道转移动物模型及结直肠癌粘膜原位移植转移模型。因此,我们从模型的角度利用人结肠癌细胞株HCT-116,研究构建人结直肠癌裸小鼠爪垫单纯性淋巴道移植转移动物模型,以及组织块法直肠粘膜原位移植转移动物模型的可行性具有重要的科研价值和临床意义。第一部分人结直肠癌裸小鼠淋巴道转移模型的建立目的建立一种简便、可行、稳定的人结直肠癌裸小鼠淋巴道移植转移动物模型。方法裸鼠爪垫皮下注射人结肠癌细胞系HCT-116细胞,分别于接种后第1、2、3、4、5、6周及第8周不同时间采集种植瘤,乆窝和腹股沟淋巴结(第一级)、髂血管旁淋巴结(第二级)、肾门淋巴结(第三级)以及肝、肺脏器标本,进行HE染色及免疫组化检测人癌胚抗原(CEA)。观察种植瘤的成瘤率、生长及淋巴结转移情况。结果爪垫种植1周后成瘤率100%。瘤体在3周后成指数生长。第一级、第二级、第三级淋巴结转移率第1、2周均为0%;第3周时分别为40%、0%、0%;第4周时转移率分别为60%、0%、0%;第5周时转移率分别为60%、20%、0%;第6周时转移率分别为100%、40%、0%。第8周时转移率分别为100%、60%、20%。但以上各时点均未见肝、肺转移。HE染色证实淋巴结有弥漫性癌细胞转移浸润。免疫组化染色显示转移灶内CEA阳性细胞。结论裸鼠爪垫皮下注射人结肠癌细胞系(HCT-116)可成功建立人大肠癌淋巴道转移模型。此模型快速简便、转移集中、转移率高,为研究人大肠癌淋巴道转移机制,药物干预等抗转移治疗提供理想的动物模型。第二部分人结直肠癌裸小鼠直肠粘膜原位种植癌及转移模型的建立目的建立人大肠癌组织块裸小鼠直肠粘膜原位种植癌及转移模型,为研究结直肠癌浸润转移机制和抗转移治疗提供动物模型。方法人结肠癌细胞系HCT-116细胞注射接种于裸鼠直肠粘膜下,获得直肠种植瘤。取该肿瘤4周瘤体制备成新鲜组织小块,原位种植于直肠粘膜面,分别于接种后第2、4、6、8、10、12周不同时间采集直肠瘤块,直肠上动脉、肠系膜下动脉、腹主动脉旁淋巴结以及肝、肺脏器标本,进行HE染色及人癌胚抗原(CEA)免疫组化检测。观察种植瘤的成瘤率,生长及转移情况。结果直肠粘膜组织块法原位种植2周时成瘤率91%。肿瘤在6~12周成指数生长。直肠上动脉淋巴结、肠系膜下动脉淋巴结、腹主动脉旁淋巴结转移率第2周时均为0%,第4周时分别为60%、20%、0%,第6周时分别为80%、60%、40%,第8周时分别为100%、80%、80%,第10、12周时转移率均为100%。于第10周、12周时见肝、肺同时均发生转移,第10周转移率均为20%;第12周时肝、肺转移率均为40%。HE染色证实淋巴结有弥漫性癌细胞转移浸润及肝、肺有转移癌结节。免疫组化染色显示转移灶内CEA阳性细胞。结论裸小鼠直肠粘膜植块法能成功构建人结直肠癌原位种植癌及转移模型。此模型操作简便、制模稳定、转移率高,且生物学行为更能模拟临床疾病粘膜原位发生、发展演变过程,可为研究大肠癌转移机制和抗转移治疗提供理想的动物模型。
[Abstract]:Colorectal cancer is a common tumor in the digestive tract, with high malignancy and easy occurrence of lymphatic metastasis and dissemination of digestive tract. Tumor invasion and metastasis not only affect the effect of clinical treatment, but also the main cause of the recurrence and death of the patients. Therefore, the study on the mechanism of metastasis and the prevention and treatment of colorectal cancer is an important task at present. It is necessary to establish a stable, reliable and reproducible animal model for colorectal cancer transplanting and metastasis in nude mice. It is necessary to establish a stable, reliable and reproducible animal model in order to further study the mechanism of invasion and metastasis of colorectal cancer, drug intervention and other anti metastasis. The treatment provides an effective experimental platform to improve the prognosis and outcome of colorectal cancer. But the ideal model of colorectal carcinoma simple lymphatic metastasis and the model of orthotopic transplantation of colorectal carcinoma have not been established. Therefore, we use the human colon cancer cell line HCT-116 from the model point of view to study the construction of human node. It is of great scientific value and clinical significance that the animal model of simple lymphatic transplanting in the nude mice of rectal cancer and the feasibility of transferring the animal model by the tissue block method of rectal mucosa transplantation in situ are of great importance.
Part one establishment of lymphatic metastasis model of human colorectal cancer in nude mice
Objective to establish a simple, feasible and stable animal model for human lymphatic metastasis of human colorectal cancer in nude mice.
Methods human colon cancer cell line HCT-116 cells were subcutaneously injected into the nude mouse's claw pads. The tumors were collected at the first 1,2,3,4,5,6 and 8 weeks after the inoculation, and the lymph nodes of the socket and groin (grade 1), the iliac para lymph nodes (grade second), the renal hilar lymph nodes (third), and the liver and lung organs were examined by HE staining and immunohistochemical detection. Human carcinoembryonic antigen (CEA). Tumor formation rate, growth and lymph node metastasis were observed.
Results after 1 weeks, the tumor formation rate of the claw pad was grown exponentially after 3 weeks. The first, second, and third lymph node metastasis rates were 0% and 40%, 0%, 0%, respectively, at the third week, and fourth weeks, respectively, 60%, 0%, 0%, respectively. The transfer rate at the week of fourth weeks was respectively, and the metastasis rate of the week was respectively 0%.. The rates were 100%, 60%, 20%., but no liver was found at all time points..HE staining showed that the lymph nodes had diffuse infiltrating infiltration in the lymph nodes. Immunohistochemical staining showed the CEA positive cells in the metastases.
Conclusion the nude mouse claw pad subcutaneous injection of human colon cancer cell line (HCT-116) can successfully establish the lymphatic metastasis model of human colorectal cancer. This model is rapid, simple, centralized and high metastasis rate. It provides an ideal animal model for the study of the mechanism of lymphatic metastasis of colorectal cancer and the antimetastasis treatment of drug intervention.
The second part: establishment of orthotopic implantation cancer and metastasis model of human colorectal cancer in nude mice
Objective to establish the carcinoma and metastasis model of rectal mucosa in the nude mice of colorectal cancer tissue, and to provide an animal model for the study of the mechanism of invasion and metastasis of colorectal cancer and the treatment of anti metastasis.
Methods human colon cancer cell line HCT-116 cells were injected under the rectal mucous membrane of nude mice to obtain rectal implants. The tumor was prepared for 4 weeks by the tumor tissue. The tumor was implanted in the rectal mucosa in situ. The rectal tumor blocks, the superior rectal artery, the inferior mesenteric artery and the abdominal aorta were collected at the different time of the inoculation at the 2,4,6,8,10,12 week. HE staining and human carcinoembryonic antigen (CEA) immunohistochemical staining were used to detect lymph node, liver and lung organ specimens. The tumorigenicity, growth and metastasis of the tumor were observed.
Results the tumor rate of the rectal mucosal tissue mass was in situ for 2 weeks, and the tumor rate 91%. tumor grew exponentially at 6~12 weeks. The upper rectal artery lymph node, the inferior mesenteric artery lymph node, the abdominal aorta lymph node metastasis rate were 0% at second weeks, and 60%, 20%, 0% at the fourth week, respectively, 80%, 60%, 40%, respectively, eighth weeks, respectively, 100%, 80%, 10,1, eighth, respectively. At the 2 week, the metastasis rate was 100%. at tenth weeks, the liver was seen at 12 weeks and the metastasis rate was 20% at the tenth week. The liver and lung metastasis rates were both 40%.HE staining and diffuse infiltration of the lymph nodes and the liver and the metastatic nodules in the lung at the twelfth week. The immunohistochemical staining showed the CEA positive cells in the metastases.
Conclusion the nude mouse rectal mucosal graft method can successfully construct the orthotopic cancer and metastasis model of human colorectal cancer. This model is easy to operate, is stable, and has high metastasis rate. The biological behavior can simulate the occurrence of the clinical disease in situ and the evolution process. It can provide ideal animal for the study of the metastasis mechanism of colorectal cancer and the antimetastasis treatment. Model.

【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R735.35;R-332

【引证文献】