XAV939抑制人神经母细胞瘤细胞的增殖

Effect of XAV939 on proliferation inhibition of human neuroblastoma cell

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 study the effect of antiproliferative and apoptosis induction of small molecule inhibitor XAV939 on neuroblastoma （NB） cell line SH-SY5Y cells, and its possible mechanism. Methods The MTT assay was used to detect the inhibition effect of XAV939 on the viability of SH-SY5Y cells, and to determine the optimal concentration. Annexin V-FITC assay was used to detect the percentage of early apoptotic cells after XAV939 treatment. Hoechst33342 staining was used for observing the apoptotic nuclear morphology. Colony forming assay was applied for detecting the proliferation effect of XAV939 on SH-SY5Y cells. The expression of key proteins of Wnt/β-catenin pathway and anti-apoptotic protein Bcl-2 were detected by Western blotting. Results The MTT assay showed that the proliferation inhibition rate of SH-SYSY cells increased significantly by 1μmol/L XAV939 for 24hours. The percentages of apoptotic cells were significantly higher than those in control groups after treated with XAV939 for 48hours and 72hours, and the cells showed varying degrees of apoptotic morphology. In addition, XAV939 significantly reduced the colony formation rate of SH-SY5Y cells in vitro. The results of Western blotting showed that XAV939 reduced the expression of key proteins of Wnt/β-catenin signaling pathway as well as anti-apoptotic protein Bcl-2. Conclusion XAV939 may inhibit the proliferation of SH-SY5Y cells in part by inhibiting the Wnt/β-catenin signaling pathway.