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目的检测人胃癌KATO-Ⅲ、SGC7901、AGS及MKN-45细胞株中CD133基因及蛋白的表达,研究CD133阳性(CD133+)细胞与CD133阴性(CD133-)细胞的生物学特性的差异。方法采用半定量聚合酶链反应及免疫蛋白印迹法分别检测KATO-Ⅲ、SGC7901、AGS及MKN-45细胞株中CD133基因和蛋白的表达;免疫磁珠细胞技术将KATO-Ⅲ细胞株中CD133+和CD133-细胞分离纯化,对比两者生长形态特点、体外增殖能力及分化能力的差异;采用CCK-8法分析CD133+和CD133-细胞对不同浓度(0.05、0.10、0.20、0.50、1.00mg/ml)化疗药物5-氟尿嘧啶(5-FU)敏感性的差别。结果 KATO-Ⅲ细胞中CD133基因及蛋白表达量均明显高于SGC7901、AGS及MKN-45细胞(P<0.05)。CD133+细胞在无血清培养基中呈球形克隆悬浮生长,相比CD133-细胞具有更强的体外增殖能力及潜在分化能力。5-FU耐药性实验中,CD133+细胞在0.50mg/ml浓度下的抑制率较CD133-细胞明显降低(P<0.05),但其他浓度时二者比较差异无统计学意义(P>0.05)。结论人胃癌细胞株KATO-Ⅲ中的CD133+细胞具有一定增殖、分化及耐药潜能,CD133可作为胃肿瘤起始细胞亚群的表面标志之一。
Objective To detect the expression of CD133 gene and protein in human gastric cancer KATO-Ⅲ, SGC7901, AGS and MKN-45 cell lines and to study the biological characteristics of CD133 positive cells and CD133 negative cells. Methods The expression of CD133 gene and protein in KATO-Ⅲ, SGC7901, AGS and MKN-45 cell lines were detected by semi-quantitative polymerase chain reaction and immunoblotting. CD133 + and CD133 + CD133- cells were isolated and purified to compare their growth morphological characteristics, proliferation ability and differentiation ability in vitro; CCK-8 assay was used to analyze the effect of CD133 + and CD133- Chemotherapy drug 5-fluorouracil (5-FU) sensitivity difference. Results The expression of CD133 gene and protein in KATO-Ⅲ cells were significantly higher than those in SGC7901, AGS and MKN-45 cells (P <0.05). CD133 + cells grew in spherical suspension in serum-free medium with enhanced ability to proliferate and potentially differentiate in vitro compared to CD133- cells. In the 5-FU drug resistance experiment, the inhibitory rate of CD133 + cells at the concentration of 0.50mg / ml was significantly lower than that of the CD133- cells (P <0.05), but no significant difference was found at other concentrations (P> 0.05) . Conclusions CD133 + cells in human gastric cancer cell line KATO-Ⅲ have certain proliferation, differentiation and drug resistance potential. CD133 can be used as one of the surface markers of gastric cancer initiating cell subsets.