IR-780联合阿霉素对肝癌干细胞样细胞的杀伤效应研究

IR-780 enhances cytotoxicity of adriamycin against cancer stem-like cells from hepatocellular carcinoma cells

目的探讨七甲川花菁类荧光小分子IR-780联合阿霉素对肝癌干细胞样细胞的杀伤作用。方法利用CCK-8试剂盒检测不同浓度IR-780(0、1.250、2.500、5.000、10.000、20.000μmol/L)及IR-780联合阿霉素(0、0.025、0.050、0.100、0.200、0.400μmol/L)对4种不同肝癌细胞(QGY-7703、SMMC-7721、Huh-7和Hep G2)的杀伤效应,并确定后续实验IR-780和阿霉素的浓度,采用细胞划痕实验检测IR-780联合阿霉素对肝癌细胞迁移能力的影响;采用无血清悬浮培养的方法从Huh-7细胞株中分离并鉴定出具有干细胞特性的肿瘤细胞,检测IR-780联合阿霉素对肝癌干细胞样细胞的杀伤作用及迁移能力的影响;在裸鼠皮下荷瘤模型中观察IR-780联合阿霉素对肝癌干细胞样细胞生长的抑制作用。结果低浓度的IR-780对肝癌细胞无明显杀伤作用;采用1.250μmol/L的IR-780联合阿霉素可对多种肝癌细胞产生较强的杀伤作用,其中半数致死浓度LC50较单药阿霉素降低25%;采用无血清悬浮培养的方法培养出Huh-7细胞肿瘤球,比较其与贴壁细胞的克隆形成能力、致瘤能力和细胞周期分布,证明悬浮培养的肿瘤细胞具有肿瘤干细胞特性;IR-780联合阿霉素对肝癌干细胞样细胞具有较强的杀伤作用,单用阿霉素和IR-780联合阿霉素对肝癌干细胞样细胞的LC50差异有统计学意义[(0.30±0.02)vs(0.22±0.04)μmol/L,P<0.05];IR-780联合阿霉素可明显抑制肝癌干细胞样细胞的迁移能力和裸鼠皮下肿瘤的生长。结论 IR-780可增强阿霉素的抗肿瘤作用,提高对肝癌干细胞样细胞的杀伤效应。

Objective To determine the cytotoxic effects of adriamycin （ADM） in combination with heptamethine cyanine dye IR-780 iodide on cancer stem-like cells from hepatocellular carcinoma （HCC） cell line. Methods CCK-8 assay was used to detect the cell viability and LCs0 of ADM in the HCC cell lines QGY-7703, SMMC-7721, Huh-7 and HepG2 treated with IR-780 （0, 1. 250, 2. 500, 5. 000, 10. 000 or 20. 000 μmol/L） alone or in combination with ADM （0, 0.025, 0.050, 0.1（30, 0.200 and 0.400 μmol/L）. The effect of IR-780 combined with ADM on cell migration was assessed with scratch wound healing assay. Cancer stem-like cells were isolated from Huh-7 cells cultured in serum-free medium under non-adherent conditions and identified for their stemness. The effects of IR-780 combined with ADM on the cell viability, migration, and tumorigenicity of the cancer stem-like cells were evaluated both in vitro and in a nude mouse model bearing the cell xenograft. Results Low-dose IR-780 （1. 250μmol/L） produced no significantcytotoxicity in the HCC cell lines. IR-780 at 1. 250 μmol/L combined with ADM resulted in a strong cytotoxic effect in the cell lines with a reduced LC50 of ADM by 25% compared to that of ADM used alone. Comparison of the colony formation and tumorigenic abilities and cell cycle distribution between Huh-7 tumorspheres cultured in serum-free medium and the adherent cells demonstrated the sternness of the isolated cells. IR-780 combined with ADM showed a stronger cytotoxic effect than ADM alone in the cancer stem-like cells with a significantly reduced LC50 of ADM （0.22 ± 0.04 vs 0. 30 ± 0.02 μmol/L, P 〈 0. 05 ）. The combination of IR- 780 and ADM significantly inhibited the migration and tumorigenicity of the cancer stem-like cells both in vitro and in the tumor-bearing nude mouse model. Conclusion IR-780 can enhances the cytotoxic effect of ADM against cancer stem-like cells isolated from HCC cells.