肝素酶RNAi对HepG2肝癌细胞生物学行为的影响

Effect of heparanase RNA interference on biological behaviour of HepG2 liver cancer cells

目的利用筛选出的肝素酶有效干扰细胞株HepG2/RNAi/1和HepG2/RNAi/3研究肝素酶RNAi对HepG2肝癌细胞生物学行为的影响。方法通过MTT实验、流式细胞技术、平皿克隆形成实验、Transwell侵袭实验、裸鼠成瘤实验分别检测HepG2肝癌细胞肝素酶RNAi后生长速度、单个细胞形成克隆能力、侵袭能力及成瘤能力的变化。结果MTT实验表明HepG2/RNAi/1与HepG2/RNAi/3组细胞增殖能力明显低于HepG2组和HepG2/RNAi/N组;流式细胞生长周期实验表明,与HepG2和HepG2/RNAi/N细胞相比,HepG2/RNAi/1与HepG2/RNAi/3 G0/G1期细胞比例增加,而增殖指数下降;平皿克隆形成实验表明HepG2/RNAi/1与HepG2/RNAi/3组单个细胞形成克隆的能力与HepG2组和HepG2/RNAi/N组相比显著降低[分别为(34±4)、(26±5)和(138±7)、(123±22),P<0.05)];Transwell体外侵袭实验表明,HepG2/RNAi/1与HepG2/RNAi/3细胞穿膜数较HepG2和HepG2/RNAi/N细胞穿膜数明显减少[分别为(85.1±9.1)、(78.1±10.4)和(182.2±9.7)(、183.5±9.3),P<0.05)];裸鼠成瘤实验显示,HepG2、HepG2/RNAi/N细胞成瘤率为100%,虽然HepG2/RNAi/1细胞成瘤亦为100%,但裸鼠皮下肿瘤体积较HepG2、HepG2/RNAi/N细胞明显缩小[分别为(0.099±0.030)和(0.585±0.135)(、0.690±0.099),P<0.01)],而HepG2/RNAi/3细胞裸鼠皮下未见肿瘤形成。结论肝素酶RNA干扰可以明显抑制HepG2肝癌细胞增殖速度、单个细胞克隆形成能力、体外侵袭能力以及裸鼠皮下成瘤能力。

Objective To Utilize the screened heparanase RNA interference cell lines HepG2/RNAi/1 and HepG2/RNAi/3 to study the effect of heparanase RNA interference on biological behaviour of HepG2 liver cancer cells. Method Through MTT experiment, FCM, plate clone formation experiment Transwell invasion experiment and tumorigenesis in nude mice experiment we detected biological behaviour changes of HepG2 after heparanase RNA interference. Re- sult MTT result showed that cell proliferation ability of HepG2/RNAi/1 and HepG2/RNAi/3 cells was significantly lower than that of HepG2 and HepG2/RNAiAN cells. Plate clone formation experiment demonstrated that single cell clone formation capacity of HepG2/RNAi/1 and HepG2/ RNAi/3 cells was obviously lower than that of HepG2 and HepG2/RNAi/N cells （34± 4, 26 ±5 vs 138 ± 7, 123 ± 22, P 〈 0.05 ）. Flow cytometry demonstrated that HepG2/RNAi/1 and HepG2/RNAi/3 cells arresting in G0/G1 and decrease proliferative index （PI） of HepG2/RNAi/1 and HepG2/RNAi/3 cells. Transwell in vitro invasion experiment showed that penetrated cell number in HepG2/RNAi/1 and HepG2/RNAi/3 cells was much lower than in HepG2 and HepG2/ RNAi/N cells （85.1 ±9.1,78.1 ±10.4 vs 182.2± 9.7,183.5 ± 9.3, P 〈 0.05）. Tumorigenesis in nude mice experiment demonstrated that HepG 2 and HepG 2 / RNAi / N cells had a tumor formation ratio of 100 %. Although HepG2/RNAi/1 cells also had a tumor formation ratio of 100%, but its subcutaneous tumor volume is much smaller than that of HepG2 and HepG2/ RNAi/N cells （0. 099± 0. 030 vs 0. 585 ± 0. 135, 0. 690 ± 0. 099, P〈0.01）, while no subcutaneous tumor formed in group of HepG2/RNAi/3 cells. Conclusion Heparanase RNAi can effec- tively inhibit HepG2 liver cancer cells in proliferation, invasion, clone formation, and tumorigenesis in nude mice.