无血清培养人结肠癌球囊样细胞生物学特性研究

Biological properties of colon cancer spheroid cells cultured in serum-free medium

目的采用无血清培养方法从人结肠癌细胞株中分选出富含肿瘤干细胞的球囊样细胞，并分析其增殖和迁移特性。方法人结肠癌细胞株HCTll6、HT29细胞在特殊配制的无血清培养基（SFM）中悬浮培养，观察球囊样细胞的生成。流式细胞仪检测结肠癌干细胞分子标记CDl33表达，利用CCK一8比色法以及Transwell小室法研究结肠癌球囊样细胞的增殖情况和迁移能力。采用成组设计资料的t检验分析检测数据。结果HCTl16、HT29细胞在SFM培养条件下可以获得可稳定传代的球囊样细胞，SFM培养下HCTll6球囊样细胞CDl33阳性细胞占75．44％±11．41％，HT29球囊样细胞CDl33阳性细胞占76．22％±14．23％。与常规含血清培养基（SSM）培养的同系细胞比较，HCTll6及HT29球囊样细胞中结肠癌干细胞分子标记CDl33阳性细胞数明显增多（t：11．43，9．17，P〈0．05），其持续增殖能力和运动迁移能力均增强。结论无血清培养获得的结肠癌球囊样细胞高表达结肠癌干细胞分子标记CDl33，且具有更强的增殖和迁移能力。它可作为进一步研究结肠癌干细胞的良好模型。

Objective To obtain colon cancer spheroid cells from human colon cancer cell lines cultured in serum-free medium （SFM）, and investigate the proliferative and migratory properties of colon cancer spheroid cells. Methods Human colon cancer cell lines HCTll6 and HT29 were cultured in SFM, and then the generation of spheroid ceils was observed. The expression of stem cell surface marker CDl33 was detected by flow cytometry, and the proliferative and migratory properties of colon cancer spheroid cells were detected by cell counting kit-8 and Transwell migration assay, respectively. All data were analyzed by using the t test. Results Spheroid cells were obtained from colon cancer cell lines HCTll6 and HT29 in SFM. The ratios of spheroid cells with positive expression of CD133 generated by HCTII6 and HT29 were 75.44% ±11.41% and 76.22% ± 14.23% , respec- tively. Compared with original colon cancer cells cultured in serum supplemented medium, the number of HCT116 and HT29 spheroid cells with positive expression of CD133 was significantly greater （t = 11. 43, 9. 17, P 〈 0.05 ） , and the proliferative and migratory abilities were much stronger also. Conclusion Colon cancer spheroid cells cultured in SFM have higher positive expression of CD133 and stronger proliferative and migratory abilities, and it can be utilized as a feasible model for further studies of colonic stem cells.