姜黄素对UMI-77诱导急性T淋巴细胞白血病细胞凋亡的影响及其相关机制研究

Effect of Curcumin on Apoptosis of Acute T-Lymphoblastic Leukemia Cells induced by UMI-77 and Its Related Mechanism

目的:探究姜黄素对Mcl-1小分子抑制剂UMI-77诱导的人急性T淋巴细胞白血病(T-ALL)细胞凋亡的影响及其相关机制。方法:培养T-ALL细胞系Molt-4,用不同浓度的姜黄素和Mcl-1小分子抑制剂UMI-77分别处理细胞24 h, MTT法检测经不同浓度姜黄素和UMI-77分别处理后的细胞存活率;根据姜黄素和UMI-77的作用浓度,实验设置对照、姜黄素(20μmol/L姜黄素处理细胞)、UMI-77组(15μmol/L Mcl-1小分子抑制剂UMI-77处理细胞)和姜黄素+UMI-77(20μmol/L姜黄素加15μmol/L Mcl-1小分子抑制剂UMI-77处理细胞)共4组,MTT法检测细胞增殖抑制率,Annexin V-FITC/PI双染法和TUNEL染色检测细胞的凋亡情况,DCFH-DA探针检测细胞活性氧,JC-1荧光探针检测线粒体膜电位,Western blot检测细胞凋亡相关蛋白和Notch1信号通路相关蛋白的表达水平。结果:经过不同浓度的姜黄素和Mcl-1小分子抑制剂UMI-77处理Molt-4细胞后,细胞存活率降低(P<0.05);与对照组比较,姜黄素组和UMI-77组细胞增殖抑制率增加,细胞凋亡率升高,ROS水平提高,细胞内Bax、Caspase-3和Caspase-9蛋白表达均增加,Bcl-2蛋白表达降低(P<0.05);与姜黄素组和UMI-77组比较,姜黄素+UMI-77组细胞增殖抑制率和凋亡率进一步增加,ROS水平提高,同时,细胞内Bax、Caspase-3和Caspase-9蛋白表达均增加,而Bcl-2蛋白表达降低(P<0.05);此外,姜黄素处理后细胞的线粒体膜电位下降,Notch1和HES1蛋白表达均降低(P<0.05)。结论:姜黄素通过降低线粒体膜电位来增强Mcl-1小分子抑制剂UMI-77诱导的T-ALL细胞凋亡,其机制可能与抑制Notch1信号通路相关。

Objective: To explore the effect and mechanism of curcumin on human T-cell acute lymphoblastic leukemia(T-ALL) cell apoptosis induced by Mcl-1 small molecule inhibitors UMI-77. Methods: T-ALL cell line Molt-4 was cultured, and the cells were treated with different concentrations of curcumin and Mcl-1 small molecule inhibitor UMI-77 for 24 h. The MTT method was used to detect the cell survival rate after different treatment;According to the results of curcumin and UMI-77, the experimental settings were divided into control group, curcumin group(20 μmol/L curcumin treated cells), UMI-77 group(15 μmol/L Mcl-1 small molecule inhibitor UMI-77 treated cells) and curcumin+ UMI-77 group(20 μmol/L curcumin and 15 μmol/L Mcl-1 small molecule inhibitor UMI-77 treated cells), MTT method was used to detect cell proliferation inhibition rate, Annexin V-FITC/PI double staining method and TUNEL staining were used to detect cell apoptosis, DCFH-DA probe was used to detect cell reactive oxygen species, JC-1 fluorescent probe was used to detect mitochondrial membrane potential, Western blot was used to detect the expression levels of apoptosis-related proteins and Notch1 signaling pathway-related proteins. Results: After the treatment of Molt-4 cells with different concentrations of curcumin and Mcl-1 small molecule inhibitor UMI-77, the cell survival rate was decreased(P<0.05);Compared with the control group, the cell proliferation inhibition rate of the curcumin group and the UMI-77 group were increased, the apoptosis rate of cell was increased, the level of ROS was increased, the protein expression of Bax, Caspase-3 and Caspase-9 in the cells were all increased, and the protein expression of Bcl-2 was reduced(P<0.05);Compared with the curcumin group or UMI-77 group, the cell proliferation inhibition rate and apoptosis rate of the curcumin + UMI-77 group were further increased,and the level of ROS was increased. At the same time,the protein expression of Bax,Caspase-3 and Caspase-9 in the cells were all increased,the protein expression of Bcl-2 was reduced(P<0.05);In addition,the mitochondrial membrane potential of the cells after curcumin treatment was decreased,and the proteins expression of Notch1 and HES1 were reduced(P<0.05).Conclusion: Curcumin can enhance the apoptosis of T-ALL cells induced by Mcl-1 small molecule inhibitor UMI-77 by reducing the mitochondrial membrane potential,the mechanism may be related to the inhibition of Notch1 signaling pathway.