青蒿琥酯调节Shh/Gli1信号通路对脑胶质瘤细胞增殖、凋亡及血管生成拟态形成的影响

The effects of artesunate regulating the Shh/Gli1 signaling pathway on the proliferation, apoptosis and angiogenic mimicry formation of glioma cells

摘要:

目的：探究青蒿琥酯（ART）对脑胶质瘤细胞增殖、凋亡及血管生成拟态形成的影响，分析与音猬因子（Shh）/胶质瘤相关癌基因1（Gli1）通路的关系。方法：将脑胶质瘤细胞U251随机分为对照（CK）组、低、中、高剂量ART（L、M、H-ART）组、Shh激活剂PM（PM）组、H-ART+PM组。平板克隆实验、Hoechst33258染色、血管生成实验分别检测细胞增殖、细胞凋亡、细胞血管生成拟态形成；鬼笔环肽染色观察细胞骨架；Western blot检测蛋白表达；裸鼠移植瘤实验检测其质量及体积。结果：与CK组比，L、M、H-ART组细胞集落形成数、PCNA表达、血管生成拟态管状结构数、VEGF-A、VE-cadherin、Shh、Gli1、Snail表达降低，细胞凋亡率、Caspase-3表达、肌动蛋白荧光强度升高（P＜0.05）；与H-ART组比，H-ART+PM组细胞集落形成数、PCNA表达、血管生成拟态管状结构数、VEGF-A、VE-cadherin、Shh、Gli1、Snail表达升高，细胞凋亡率、Caspase-3表达、肌动蛋白荧光强度降低（P＜0.05）。与对照组比，ART组肿瘤质量、肿瘤体积降低（P＜0.05）；与ART组比，联合组肿瘤质量、肿瘤体积升高（P＜0.05）。结论：ART通过抑制Shh/Gli1通路，抑制脑胶质瘤细胞增殖、血管生成拟态形成，并促进其细胞凋亡。

Abstract:

Objective: To discuss the effects of artesunate (ART) on the proliferation, apoptosis and angiogenic mimicry formation of glioma cells, and to analyze its relationship with the hedgehog factor (Shh)/glioma-associated oncogene homologue 1 (Gli1) pathway. Methods: Glioma cells U251 were stochastically separated into the control (CK) group, low-, medium-, and high-dose ART (L, M, H-ART) groups, Shh activator PM (PM) group, and H-ART+PM groups. Cell proliferation, apoptosis and cell angiogenesis mimicry formation were respectively detected by plate cloning assay, Hoechst33258 staining and angiogenesis assay. Phalloidin staining was used to observe the cytoskeleton. The protein expression was measured by Western blot. Moreover, the nude mouse transplant tumor experiment was used to detect its mass and volume. Results Compared with the CK group, the number of cell colony formation, the expressions of PCNA, the number of angiogenic mimicry tubular structures, the expressions of VEGF-A, VE-cadherin, Shh, Gli1, and Snail in the L, M, and H-ART groups were lower, the apoptosis rate, the expressions of Caspase-3 and the fluorescence intensity of actin were higher (P < 0.05). Compared with the H-ART group, the number of cell colony formation, the expressions of PCNA, the number of angiogenic mimicry tubular structures, the expressions of VEGF-A, VE-cadherin, Shh, Gli1, and Snail in the H-ART+PM groups were higher, the apoptosis rate, the expressions of Caspase-3 and the fluorescence intensity of actin were lower (P < 0.05). Compared with the control group, the tumor mass and tumor volume in the ART group were smaller (P < 0.05). Compared with the ART group, the tumor mass and tumor volume in the combination group were increased (P < 0.05). Conclusion: ART inhibits the proliferation of glioma cells, the formation of angiogenic mimicry, and promotes their apoptosis by suppressing the Shh/Gli1 pathway.