国际肿瘤学杂志

国际肿瘤学杂志 ›› 2021, Vol. 48 ›› Issue (12): 711-715.doi: 10.3760/cma.j.cn371439-20210514-00141

• 论著 • 上一篇    下一篇

IWR-1-endo通过抑制Wnt通路影响肝癌细胞的迁移和增殖

熊琳1, 张修云1, 张小余2, 黎越2, 徐细明2   

  1. 1武汉大学人民医院病理科 430060
    2武汉大学人民医院肿瘤中心 430060
  • 收稿日期:2021-05-14 修回日期:2021-06-17 出版日期:2021-12-08 发布日期:2022-01-12
  • 基金资助:
    国家自然科学基金(31971166)

IWR-1-endo affects the migration and proliferation of hepatocarcinoma cells by inhibiting the Wnt pathway

Xiong Lin1, Zhang Xiuyun1, Zhang Xiaoyu2, Li Yue2, Xu Ximing2   

  1. 1Department of Pathology, Renmin Hospital of Wuhan University, Wuhan 430060, China
    2Cancer Center, Renmin Hospital of Wuhan University, Wuhan 430060, China
  • Received:2021-05-14 Revised:2021-06-17 Online:2021-12-08 Published:2022-01-12
  • Supported by:
    National Natural Science Foundation of China(31971166)

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摘要： 目的 探讨Wnt通路抑制剂IWR-1-endo对人肝癌细胞Huh7生物学行为的影响及作用机制。 方法 体外培养人肝癌细胞Huh7,采用不同浓度(0、20、40、80、160、320 μmol/L)IWR-1-endo作用于Huh7细胞。采用划痕实验检测不同药物浓度下细胞迁移能力的变化;平板克隆实验检测细胞的增殖变化;蛋白质印迹法检测Wnt通路相关蛋白β-catenin表达的改变;免疫荧光染色检测β-catenin在细胞质和细胞核的表达情况。 结果 划痕实验结果显示,0、20、40、80、160、320 μmol/L浓度的IWR-1-endo处理Huh7细胞24 h划痕愈合率分别为(20.55±0.05)%、(12.10±0.08)%、(9.36±0.10)%、(3.62±0.09)%、(0.62±0.04)%、(0.23±0.02)%,差异有统计学意义(F=230.87,P<0.001);进一步两两比较,各浓度间24 h划痕愈合率差异均具有统计学意义(均P<0.001)。48 h划痕愈合率分别为(34.77±0.08)%、(17.69±0.05)%、(11.60±0.04)%、(5.68±0.07)%、(2.66±0.04)%、(1.75±0.02)%,差异有统计学意义(F=589.68,P<0.001);进一步两两比较,各浓度间48 h划痕愈合率差异均具有统计学意义(均P<0.001)。经0、20、40、80、160、320 μmol/L浓度的IWR-1-endo处理肝癌Huh7细胞后,克隆形成率分别为(61.67±0.21)%、(57.33±0.11)%、(50.00±0.25)%、(36.67±0.28)%、(23.33±0.12)%、(15.00±0.08)%,差异有统计学意义(F=403.56,P<0.001);进一步两两比较,各浓度间的克隆形成率差异均具有统计学意义(均P<0.001)。经0、20、40、80、160 μmol/L浓度的IWR-1-endo处理Huh7细胞24 h后,β-catenin的相对表达量分别为0.30±0.08、0.25±0.07、0.22±0.05、0.15±0.01、0.06±0.02,差异有统计学意义(F=247.00,P<0.001);与0 μmol/L相比,80、160 μmol/L浓度处理后的β-catenin的相对表达量差异均有统计学意义(P=0.014;P=0.008)。免疫荧光染色结果显示,与0 μmol/L相比,80 μmol/L IWR-1-endo处理后,Huh7细胞质和细胞核中的β-catenin表达均减少。结论Wnt通路抑制剂IWR-1-endo可通过抑制Wnt通路的活性抑制肝癌细胞Huh7的迁移和增殖,抑制作用均呈现剂量依赖性。

关键词: 肝肿瘤, Wnt通路, IWR-1-endo, 细胞增殖, 细胞运动

Abstract: Objective To investigate the effects and mechanisms of Wnt pathway inhibitor IWR-1-endo on the biological behaviors of human hepatocarcinoma cell Huh7. Methods Human hepatocellular carcinoma cell Huh7 was cultured in vitro, and Huh7 cells were treated with IWR-1-endo at different concentrations (0, 20, 40, 80, 160, 320 μmol/L). Scratch test was used to detect changes in cell migration ability at diffe-rent drug concentrations, plate cloning was used to detect changes in cell proliferation, Western blotting was used to detect changes in the expression of Wnt pathway related protein β-catenin, and immunofluorescence staining was used to detect the expression of β-catenin in cytoplasm and nucleus. Results The results of the scratch test showed that the 24 h scratch healing rates of Huh7 cells treated with 0, 20, 40, 80, 160, 320 μmol/L IWR-1-endo were (20.55±0.05)%, (12.10±0.08)%, (9.36±0.10)%, (3.62±0.09)%, (0.62±0.04)% and (0.23±0.02)%, respectively, and there was a statistically significant difference (F=230.87, P<0.001). Further pair comparison showed that there were statistically significant differences in 24 h scratch healing rates among different concentrations (all P<0.001). The 48 h scratch healing rates were (34.77±0.08)%, (17.69±0.05)%, (11.60±0.04)%, (5.68±0.07)%, (2.66±0.04)% and (1.75±0.02)%, respectively, and there was a statistically significant difference (F=589.68, P<0.001). Further pair comparison showed that there were statistically significant differences in 48 h scratch healing rates among different concentrations (all P<0.001). After treatment with IWR-1-endo at the concentration of 0, 20, 40, 80, 160, 320 μmol/L, the clone formation rates of Huh7 cells were (61.67±0.21)%, (57.33±0.11)%, (50.00±0.25)%, (36.67±0.28)%, (23.33±0.12)% and (15.00±0.08)%, respectively, and there was a statistically significant difference (F=403.56, P<0.001). Further pair comparison showed that there were statistically significant differences in clone formation rates among different concentrations (all P<0.001). After treatment with 0, 20, 40, 80, and 160 μmol/L IWR-1-endo for 24 h, the relative expression levels of β-catenin in Huh7 cells were 0.30±0.08, 0.25±0.07, 0.22±0.05, 0.15±0.01 and 0.06±0.02, respectively, and there was a statistically significant difference (F=247.00, P<0.001). Compared with 0 μmol/L, the relative expression levels of β-catenin treated with 80 and 160 μmol/L had statistical significance (P=0.014; P=0.008). Compared with 0 mol/L, immunofluorescence showed that the expressions of β-catenin in cytoplasm and nucleus were reduced after 80 μmol/L IWR-1-endo treatment. Conclusion Wnt pathway inhibitor IWR-1-endo can inhibit the migration and proliferation of hepatocarcinoma cells Huh7 by inhibiting the activity of Wnt pathway. The above inhibitory effects are dose-dependent.

Key words: Liver neoplasms, Wnt pathway, IWR-1-endo, Cell proliferation, Cell movement