Ⅰ型γ分泌酶抑制剂抗恶性胶质瘤的体外/体内实验研究

in vitro / in vivo Studies of Anti-tumor Effects of γ-secretase Inhibitor-Ⅰ on Glioblastoma Cell Lines

目的 明确γ-分泌酶抑制剂(γ-secretase inhibtor,GSI)对恶性胶质瘤细胞体外、体内的抗瘤作用,并初步探讨其机制。方法 通过MTT法观察GSI-Ⅰ对胶质瘤细胞株U87及U251生长曲线的抑制作用;以DAPI染色法观察GSI-Ⅰ作用后是否导致细胞凋亡;用荧光定量RT-PCR法检测GSI-Ⅰ对胶质瘤细胞中Notch信号的阻断作用,并通过Western-blot检测GSI-Ⅰ对胶质瘤细胞内抗凋亡蛋白Akt的表达及磷酸化水平的影响;最后,采用裸鼠成瘤实验验证GSI-Ⅰ对胶质瘤细胞体内增殖的抑制作用,以及GSI-Ⅰ对瘤组织的毒性作用。结果 GSI-Ⅰ可显著抑制U87及U251细胞的生长曲线,并直接诱导细胞凋亡。GSI-Ⅰ可有效阻断细胞内Notch信号通路转导,并下调抗凋亡蛋白Akt的活性。动物实验结果显示,GSI-Ⅰ预处理可降低U251细胞的体内成瘤能力,而瘤周注射GSI-Ⅰ可导致瘤组织大面积坏死。结论 GSI-Ⅰ有明确的体外、体内抗恶性胶质瘤效应,其可能机制为下调细胞内Notch信号通路及抗凋亡通路,但具体分子机制仍需进一步研究。

Objective To elucidate the anti-tumor effects of a γ-secretase inhibitor（GSI）on human glioblastoma cells and explore the underlying mechanisms.Methods MTT assays were performed to examine the inhibitory effects of GSI-Ⅰ on the growth curves of two glioblastoma cell lines U87 and U251.Cellular apoptosis induced by GSI-Ⅰ treatment was determined by DAPI staining.Quantitative real-time RT-PCR（qRT-PCR）was performed to evaluate the inhibitory effects of GSI-Ⅰ on Notch signaling of the cells.Additionally,the expression and phosphorylation level of the anti-apoptotic protein Akt was examined by Western-blot.Finally,subcutaneous xenografts experiments in athymia nude mice were carried out to test the anti-tumor effects of GSI-Ⅰ in vivo.Results GSI-Ⅰ significantly inhibited the growth curves of U87 and U251 cells and induced apoptosis.Notch signaling was abrogated by GSI-Ⅰ and activation of Akt was also downregulated.The in vivo anti-tumor effects of GSI-Ⅰ on glioblastoma cells were confirmed by subcutaneous xenografts experiments.Conclusions GSI-Ⅰ exerted anti-tumor effects on glioblastoma cells in vitro and in vivo.Down regulation of Notch signaling and the anti-apoptotic Akt signaling might be involved in the anti-tumor effects,however further studies are needed to elucidate the underlying mechanisms.