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目的:研究Akt活性抑制剂LY294002抑制源自人小细胞肺癌NCI-H446细胞系肺癌球形成细胞即肺癌干细胞样细胞(LCSLCs)自我更新作用。方法:体外培养NCI-H446细胞系细胞。以干细胞条件培养基用超低粘附6孔细胞培养板悬浮培养富集和扩增LCSLCs。裸鼠皮下成瘤实验鉴别LCSLCs高致瘤特性。Western blot分析LCSLCs中Akt蛋白磷酸化水平。肿瘤球形成试验检测LY294002对LCSLCs自我更新的影响。结果:干细胞条件培养基悬浮培养6d,NCI-H446细胞系细胞呈三维非粘附性球体生长。LCSLCs具有高致瘤特性。与NCI-H446细胞系细胞比较,LCSLCs信号分子Akt组成性活化。LY294002有效降低LCSLCs中Akt磷酸化水平,并以剂量依赖方式抑制LCSLCs肺癌球形成(P<0.05)。结论:靶向干预小细胞肺癌LCSLCs信号分子Akt组成性活化可能成为抑制肺癌干细胞特性治疗小细胞肺癌的新策略。
AIM: To investigate the effect of LY294002, an inhibitor of Akt activity, on self-renewal of lung cancer stem cell-like cells (LCSLCs) derived from human small cell lung cancer NCI-H446 cell line. Methods: NCI-H446 cells were cultured in vitro. LCSLCs were enriched and expanded in stem cell-conditioned medium by suspension culture in ultra-low adhesion 6-well cell culture plates. Subcutaneous tumor formation experiment in nude mice to identify the high tumorigenicity of LCSLCs. Western blot analysis of Akt protein phosphorylation in LCSLCs. Tumor formation assay examines the effect of LY294002 on self-renewal of LCSLCs. RESULTS: Six days after suspension culture of stem cells, NCI-H446 cells grew in three-dimensional non-adherent spheres. LCSLCs have high tumorigenic properties. The LCSLCs signaling molecule Akt is constitutively activated compared to NCI-H446 cell line cells. LY294002 effectively reduced the phosphorylation of Akt in LCSLCs and inhibited the formation of lung cancer in LCSLCs in a dose-dependent manner (P <0.05). CONCLUSION: The constitutive activation of Akt signaling targeting LCSLCs in SCLC may be a new strategy to suppress the characteristics of lung cancer stem cells in the treatment of SCLC.