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目的探讨小分子抑制剂XAV939对人神经母细胞瘤(NB)细胞系SH-SY5Y细胞的抗增殖和诱导凋亡的作用,及其可能机制。方法采用MTT法检测XAV939对SH-SY5Y细胞活力的抑制作用,并确定最佳作用浓度。然后采用Annexin V-FITC法检测XAV939处理后早期凋亡细胞百分比,Hoechst33342染色法观察凋亡细胞核形态,克隆形成实验检测细胞体外增殖能力的变化。Western blotting检测Wnt/β-连环蛋白(β-catenin信号通路的关键蛋白和抗凋亡蛋白Bcl-2的变化。结果 MTT结果显示,1μmol/L XAV939作用24h后,SH-SY5Y细胞增殖的抑制率有明显上升。XAV939处理后48h及72h,凋亡细胞百分比显著高于对照组,且细胞呈现出不同程度的凋亡形态。此外,XAV939可显著降低SH-SY5Y细胞的体外克隆形成率,降低Wnt/β-catenin信号通路关键蛋白β-catenin,Cyclin D1和c-Myc及抗凋亡蛋白Bcl-2的表达。结论 XAV939可能部分通过抑制Wnt/β-catenin信号通路而抑制SH-SY5Y细胞的增殖。
Objective To investigate the effect of small molecule inhibitor XAV939 on antiproliferation and apoptosis induction in human neuroblastoma (NB) cell line SH-SY5Y and its possible mechanism. Methods The inhibitory effect of XAV939 on the viability of SH-SY5Y cells was detected by MTT assay and the optimal concentration was determined. Then the percentage of apoptotic cells in early stage after XAV939 treatment was detected by Annexin V-FITC method. The morphology of apoptotic nuclei was observed by Hoechst33342 staining. The proliferation of cells was detected by colony formation assay. Western blotting was used to detect the changes of Wnt / β-catenin, a key protein in the β-catenin signaling pathway, and the anti-apoptotic protein Bcl-2.Results MTT results showed that the inhibitory rate of proliferation of SH-SY5Y cells treated with 1μmol / L XAV939 for 24h Significantly increased.The percentage of apoptotic cells at 48h and 72h after treatment with XAV939 was significantly higher than that of the control group, and the cells showed different degrees of apoptotic morphology.In addition, XAV939 significantly reduced the rate of clonogenic SH-SY5Y cells in vitro and decreased Wnt / β-catenin signaling pathway.Conclusion XAV939 may inhibit the proliferation of SH-SY5Y cells partly by inhibiting the Wnt / β-catenin signaling pathway .