筛选自人卵巢癌细胞系 HO8910的干细胞生物学特性分析

摘要

目的：探讨筛选自人卵巢癌细胞系HO8910干细胞的生物学特性。方法将前期筛选卵巢癌细胞系HO8910的干细胞在无血清培养基中进行传代培养，以HO8910细胞作为对照。体外观察两种细胞的球体形成能力；将两种细胞接种于含血清培养基，观察其分化能力，并检测干细胞相关标记物；采用药敏试验检测两种细胞对顺铂、多柔比星及米托蒽醌的敏感性；将无血清培养基中进行传代培养的两种细胞接种于NOD／SCID裸鼠，观察其体内成瘤情况。结果 HO8910细胞在无血清培养基中的球体形成能力明显低于卵巢癌细胞（P＜0．05）。卵巢癌干细胞分化前CD133、CD117阳性表达率明显高于分化后（P均＜0．05）。卵巢癌干细胞分化后ABCG2、Nanog、Oct4、BCRP、E-cadherin表达均低于分化前（P均＜0．05），与HO8910细胞分化前后上述指标比较差异均无统计学意义（P均＞0．05）。0．25、0．5μg／mL顺铂，0．5、1．5μmol／L多柔比星，0．05、0．25μg／mL米托蒽醌作用于卵巢癌干细胞、HO8910细胞48 h时，卵巢癌干细胞的相对活性均高于HO8910细胞（P均＜0．05）。裸鼠接种1×103个卵巢癌干细胞即可成瘤；随着干细胞接种数量的增加，成瘤比率逐渐增加，成瘤时间逐渐缩短（P均＜0．05）。结论筛选自人卵巢癌细胞系HO8910的卵巢癌干细胞具有自我更新和分化、体内成瘤、高表达干细胞基因以及对化疗耐药等特性，符合肿瘤干细胞的理论标准。%Objective To identify the biological characteristics of human ovarian cancer cell line HO8910-derived stem cells.Methods The pre-screening of ovarian cancer cell line HO8910-derived stem cells were subcultured ( HO8910 cells were used as the control group) in serum-free medium.The capacities of forming spheroids and self-renew were ob-served.Then ovarian cancer stem cells ( CSCs) were seeded in medium containing serum and cultured to observe the differ-entiation capacity, and the stem cell-specific markers were also tested.We tested the sensitivity of stem cells to cisplatin, doxorubicin and mitoxantrone by using drug susceptibility test.Finally, we inoculated the ovarian CSCs after passaging from culturing in serum-free media to NOD/SCID ( non-obese diabetic/severe combined immunodeficient mice) mice in order to observe the tumorigenicity in vivo.Results The forming spheroid capacity of HO8910 cells in the serum-free medium was significantly lower than that of the ovarian cancer cells (P<0.05).The positive expression rates of CD133 and CD117 in the ovarian CSCs before differentiation were higher than those after differentiation (all P<0.05).The expression of AB-CG2, Nanog, Oct4, BCRP and E-cadherin in the ovarian CSCs after differentiation was lower than that before differentia-tion (all P<0.05).No statistical significance was found in the above indexes before and after HO8910 cells differentiation (all P>0.05).The relative activities in the ovarian CSCs were higher than in HO8910 cells when they were respectively treated with 0.25 and 0.5μg/mL cisplatin, 0.5 and 1.5μmol/L doxorubicin and 0.05 and 0.25μg/mL mitoxantrone for 48 h (all P<0.05).The tumor formed when the nude mouse was inoculated with 1 ×103 ovarian CSCs.With the increase of inoculated stem cells, the tumor formation ratio was gradually increased ( all P<0.05 ) , and the time was gradually shortened (all P<0.05).Conclusion The ovarian CSCs derived from human ovarian cancer cell line HO8910 have the abilities of self-renew, differentiation, in vivo tumorigenicity, highly expressed stem cell genes and multidrug resistance, which meet the standard of tumor stem cell theory.