人结肠癌移植瘤小鼠模型的建立及初步研究

目的利用中等分化的人结肠癌细胞建立小鼠皮下和原位移植瘤模型，以研究中国人种结肠癌特有的生物学特性，并为结肠癌的防治研究提供有用的实验工具。方法人新鲜结肠癌标本接种于BNX小鼠皮下，体内反复传代，待模型稳定后再转入BALB／c-nu／nu裸小鼠体内继续传代，使之生物学性状保持稳定，从而建立人结肠癌小鼠皮下移植瘤模型。比较人结肠癌标本在两种不同免疫缺陷动物体内的生长情况，并绘制生长曲线。采用组织块法建立人结肠癌原位移植瘤动物模型，观察原位成瘤、生长及转移等情况。组织病理学检测皮下移植瘤与人结肠癌标本的组织学差异，免疫组织化学检测皮下移植瘤中ESA、CEA、C-erbB-2、P53的表达情况，流式细胞技术检测移植瘤的细胞周期，并进行DNA倍体分析。结果通过体内反复传代筛选成功建立了中等分化的人结肠癌皮下移植瘤模型，利用皮下瘤成功建立了人结肠癌原位移植瘤模型，成瘤率达到5／8（62．5％），但肝脏、腹腔、淋巴结等未见明显转移。移植瘤组织切片观察显示肿瘤为腺癌Ⅰ-Ⅱ级，与人结肠癌组织标本相似。免疫组化显示ESA和CEA呈阳性至强阳性表达；C-erbB-2和P53呈阳性表达。流式细胞术检查发现肿瘤细胞处于G0-G1期的比例为67．50％±5．59％，G2-M期为4．47％±2．31％，S期为28．02％±7．13％，分析DNA倍体为1．74±0．02。结论本实验利用恶性程度较低的中等分化人结肠癌组织成功建立了结肠癌皮下及原位移植瘤模型，该模型生物学特性稳定，保持了人结肠癌标本的生物学特点。

Establishment of Colon Cancer Mouse Model and its Biological Characteristics

Objective To establish subcutaneous and orthotopic transplant mice models by using human colon cancer clinic sample, in order to provide a useful tool for studying the characteristics of colon cancer and preventing and curing the colon cancer. Method Fresh colon cancer sample was transplanted subcutaneously into BNX mice and nude mice, then the tumor xenografts were maintained by serial passages in BNX mice and nude mice in order to establish subcutaneous transplantable tumor animal model, tumor size was monitored regularly every week. The tumor tissues were collected and orthotopically implanted into the cecum sub-serosa. The rates of tumor formation and metastasis were observed after animal sacrifice. The expression of ESA, CEA, C-erbB-2, P53 proteins were detected by immunohistochemistry. DNA aneuploid and cell cycle distributions of tumor cells were estimated by flow cytometry. Result The subcutaneous transplant model of colon cancer was established, the tumor formation rate of orthotopic transplant model was 62.5% without metastasis. Biopsy observa- tion indicated that the xenograft tumor was pathologic stage II adenocarcinoma as same as the original human carcinoma. The expressions of ESA and CEA were strongly positive, C-erbB-2 and P53 were positive by immunohistochemistry. Flow cytometry was used to detect the change of cell cycle, the results indicated that the ratio in Go and Gl-phase was 67.50%±5.59%, 4.47%±2.31% in G2 and M-phase and 28.02%±7.13% in S phase. DNA heteroploid detection showed that the heteroploid ratio of xenograft tumor was 1.74±0.02. Conclusion The subcutaneous and orthotopic transplant colon cancer model in mice was successfully established, which had the same biological characteristic as original human colon cancer.