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目的:在原位移植瘤模型上验证CD44~+胃癌细胞的肿瘤干细胞特性。方法:选取MKN-45胃癌细胞株,通过流式细胞仪分选为CD44~+和CD44-两组,以未分选的MKN-45胃癌细胞为对照。体外部分,通过MTT法和克隆形成实验验证CD44~+胃癌细胞体外增殖能力,通过Transwell实验验证其侵袭力。体内部分,将上述细胞通过原位移植法接种至裸鼠,同等条件下饲养8周后处死裸鼠,检验其成瘤率;分离肝脏,检测肝转移瘤数目;H-E染色观察胃肿瘤和肝转移瘤形态;通过免疫荧光法检测两处肿瘤CD44~+细胞数量。结果:CD44~+胃癌细胞相比CD44-胃癌细胞具有明显增强的细胞增殖力(P<0.01)和侵袭力(P<0.01),更容易成瘤且更容易发生转移(P<0.05),但和MKN-45未分选组相比差异无统计学意义(P>0.05)。不论是在原发肿瘤还是转移瘤中,CD44~+胃癌细胞均能产生CD44-细胞,但后者不能产生CD44~+细胞。结论:CD44~+MKN-45胃癌细胞相比CD44-胃癌细胞具备更强的增殖力和侵袭力、更容易成瘤和发生转移,符合肿瘤干细胞部分特征,值得进一步研究。
OBJECTIVE: To verify the characteristics of tumor stem cells in CD44 ~ + gastric cancer cells in situ xenograft models. Methods: MKN-45 gastric cancer cell lines were selected and sorted into CD44 + and CD44- by flow cytometry. The unsorted MKN-45 gastric cancer cells were used as control. In vitro, the in vitro proliferation of CD44 ~ + gastric cancer cells was verified by MTT assay and colony formation assay. The invasion ability of CD44 ~ + gastric cancer cells was verified by Transwell assay. In vivo, the above-mentioned cells were inoculated into nude mice by orthotopic transplantation, and the nude mice were sacrificed after 8 weeks of rearing under the same conditions to test the tumorigenicity rate. The liver was separated and the number of liver metastases was detected. HE staining was used to observe the gastric tumor and liver metastasis The morphology of tumor was detected by immunofluorescence. The number of CD44 ~ + cells in two tumors was detected by immunofluorescence. Results: CD44 ~ + gastric cancer cells showed significantly enhanced cell proliferation (P <0.01) and invasiveness (P <0.01) as compared with CD44- gastric cancer cells, and were more likely to metastasize (P <0.05) but more likely to metastasize There was no significant difference between untreated group and MKN-45 group (P> 0.05). Whether in primary or metastatic tumors, CD44 ~ + gastric cancer cells can produce CD44- cells, but the latter can not produce CD44 ~ + cells. CONCLUSION: CD44 ~ + MKN-45 gastric cancer cells have stronger proliferative and invasive ability than CD44- gastric cancer cells, and are easier to tumorigenesis and metastasis, which is in line with the characteristics of cancer stem cells and deserves further study.