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目的研究异硫氰酸苄酯对人胶质瘤U87MG细胞体外侵袭和凋亡的影响,并初步探讨其作用机制。方法采用MTS实验考察异硫氰酸苄酯对肿瘤细胞体外增殖的抑制作用;2,5μmol·L-1异硫氰酸苄酯作用24 h后,采用Transwell实验、黏附实验和划痕实验观察异硫氰酸苄酯对人胶质瘤细胞侵袭、黏附和迁移能力的影响;应用Real-time-PCR和Western blot法检测相应浓度异硫氰酸酯处理后人胶质瘤U87MG细胞中MMP-2、MMP-9、CD44、Survivin、Bcl-2、Net1、Roh A、caspase-3、caspase-8和p-AKT的表达变化;应用报告基因技术检测NF-κB转录活性的变化;采用ELISA法测定胞内8-OH-d G含量;10,20μmol·L-1异硫氰酸酯作用24 h后,采用流式细胞术观察其对细胞凋亡的作用。结果异硫氰酸苄酯可显著抑制人胶质瘤U87MG细胞体外增殖;与0μmol·L-1组相比,2,5μmol·L-1异硫氰酸苄酯对人胶质瘤细胞U87MG侵袭、黏附和迁移能力有明显的抑制作用;不同浓度异硫氰酸苄酯处理后,肿瘤细胞MMP-2、MMP-9、CD44、Survivin、Bcl-2、NET1和Rho A的m RNA和蛋白表达、AKT磷酸化水平和NF-κB转录活性明显下调,caspase-3和caspase-8表达以及8-OH-d G含量显著上调;10,20μmol·L-1异硫氰酸苄酯可显著诱导细胞凋亡。结论异硫氰酸苄酯抑制人胶质瘤细胞U87MG的侵袭能力,诱导细胞凋亡,其机制可能与抑制AKT/NF-κB信号转导途径,进而调节侵袭和凋亡相关基因表达有关。
AIM To investigate the effects of benzyl isothiocyanate on invasion and apoptosis of human glioma U87MG cells in vitro and its possible mechanism. Methods MTS assay was used to investigate the inhibitory effect of benzyl isothiocyanate on the proliferation of tumor cells in vitro. Transwell experiments, adhesion experiments and scratches were used to observe the effect of 2,5 μmol·L -1 benzisothiocyanate for 24 h The effect of benzyl thiocyanate on the invasion, adhesion and migration of human glioma cells was evaluated by Real-time-PCR and Western blot. The expression of MMP-2 in human glioma U87MG cells treated with isothiocyanate The changes of NF-κB transcriptional activity were detected by reporter gene technology. The expression of caspase-3, caspase-8, caspase-8 and p-AKT were detected by ELISA The intracellular 8-OH-d G content; 10,20μmol·L-1 isothiocyanate role 24 h after the use of flow cytometry to observe the role of apoptosis. Results Benzyl isothiocyanate significantly inhibited the proliferation of human glioma U87MG cells in vitro. Compared with 0μmol·L-1 group, the invasion of U87MG cells by 2,5 μmol·L -1 benzyl isothiocyanate , Adhesion and migration ability obviously inhibited. The m RNA and protein expression of MMP-2, MMP-9, CD44, Survivin, Bcl-2, NET1 and RhoA in tumor cells treated with different concentrations of benzyl isothiocyanate , AKT phosphorylation and NF-κB transcriptional activity were significantly downregulated, caspase-3 and caspase-8 expression and 8-OH-d G levels were significantly increased; 10,20μmol·L-1 benzylisothiocyanate significantly induced cells Apoptosis. Conclusion Benzyl isothiocyanate inhibits the invasiveness of U87MG cells and induces apoptosis. The mechanism may be related to the inhibition of AKT / NF-κB signal transduction pathway and the regulation of invasion and apoptosis related gene expression.