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目的通过观察含生长因子的无血清培养基(serum-free medium,SFM)培养的Colo205细胞,研究其对CD133与ALDH1表达的影响。方法用含生长因子的无血清培养基对Colo205细胞进行培养,以含血清培养基(serum-supplimented medium,SSM)组作为对照,流式细胞术检测两组细胞表面标志物CD133表达的阳性率;流式分选SFM组CD133+细胞和CD133-细胞,倒置显微镜观察CD133+细胞和CD133-细胞在无血清培养基中的生长特性;利用细胞免疫荧光检测CD133+细胞和CD133-细胞CD133和ALDH1的表达;分别将CD133+细胞和CD133-细胞在NOD/SCID小鼠皮下进行成瘤实验,采用免疫组织化学方法对肿瘤组织检测ALDH1表达。结果 SFM组CD133+细胞比例明显高于SSM组(P<0.05);CD133+细胞在SFM中可以形成肿瘤干细胞球,而CD133-则不能形成;CD133+细胞高表达CD133和ALDH1,且两者共表达,而CD133-细胞则是阴性表达;CD133+细胞和CD133-细胞成瘤后,CD133+细胞组肿瘤组织ALDH1阳性表达,而CD133-细胞组阴性表达。结论含生长因子的无血清培养基培养CD133+Colo205大肠癌细胞能够形成肿瘤干细胞球及高表达ALDH1,ALDH1可能是大肠癌干细胞候选的标志物之一。
Objective To investigate the effect of CD133 and ALDH1 expression on Colo205 cells cultured in serum-free medium (SFM) containing growth factors. Methods Colo205 cells were cultured with serum - free medium containing growth factors and serum - supplimented medium (SSM) group was used as a control. The positive rate of CD133 expression on the two groups was detected by flow cytometry. The CD133 + cells and CD133- cells were sorted by flow cytometry. The growth characteristics of CD133 + cells and CD133- cells in serum-free medium were observed under inverted microscope. The expressions of CD133 + and CD133- CD133 + ALDH1 were detected by immunofluorescence. The tumorigenicity experiments of CD133 + cells and CD133- cells in NOD / SCID mice were carried out. The expression of ALDH1 in tumor tissues was detected by immunohistochemistry. Results The percentage of CD133 + cells in SFM group was significantly higher than that in SSM group (P <0.05). CD133 + cells could form tumor stem cell spheres in SFM, while CD133- cells could not form. CD133 + cells overexpressed CD133 and ALDH1, CD133- cells were negative expression; CD133 + cells and CD133- cells after tumor formation, CD133 + cell group ALDH1 positive tumor tissue expression, while the negative group of CD133- cells. Conclusion CD133 + Colo205 colorectal cancer cells cultured with serum-free medium containing growth factors can form tumor stem cell spheres and ALDH1 overexpression, ALDH1 may be one of the markers of colorectal cancer stem cell candidates.