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目的探讨erbB家族受体在急性单核细胞白血病细胞系U937中的表达及其对细胞生长的影响。方法采用逆转录聚合酶链反应(RT-PCR)检测erbB家族成员在U937细胞中的表达情况,Western blot检测erbB2在蛋白水平的表达。以erbB受体抑制剂EKI-785处理细胞后,采用四甲基偶氮唑蓝(MTT)实验并绘制细胞生长曲线,观察细胞的生长变化情况；应用Annexin V/PI双染和流式细胞仪计数,观察细胞是否发生凋亡；Western blot检测细胞增殖生长信号的变化。结果在U937细胞中,除了erbB1外,其他erbB受体均有表达,且erbB2有蛋白水平的表达。经过1、2、4和8μmoL/L EKI-785处理48h后,U937细胞的生长受到不同程度的抑制,抑制率分别为(10．49±3．69)%、(13．79±2．13)%、(31．42±3．90)%和(44．05±1．56)%,并呈现出比较明显的浓度依赖关系。EKI-785诱导U937细胞发生了早期凋亡,4μmol/L EKI-785处理U937细胞48和72h,Annexin- V~+/PI~-标记的早期凋亡细胞所占的比例分别为相应对照组的2．03倍和6．64倍。经4μmol/L EKI- 785作用24、48、72h后,磷酸化的MEK和Akt蛋白水平随着时间延长不断下降,72h时磷酸化的MEK和Akt几乎消失。结论erbB家族受体很可能在某些类型的白血病发生中发挥重要作用,有望成为这些白血病的新治疗靶点,EKI-785具有治疗白血病的应用前景。
Objective To investigate the expression of erbB family receptor in acute monocytic leukemia cell line U937 and its effect on cell growth. Methods The expression of erbB family members in U937 cells was detected by reverse transcriptase-polymerase chain reaction (RT-PCR), and the protein expression of erbB2 was detected by Western blot. Cells treated with erbB receptor inhibitor EKI-785 were treated with MTT assay and cell growth curve was drawn to observe the changes of cell growth; Annexin V / PI double staining and flow cytometry Counting to observe the cell apoptosis; Western blot detection of cell proliferation and growth signal changes. Results In U937 cells, except for erbB1, all other erbB receptors were expressed, and erbB2 had protein expression. After treated with EKI-785 at 1, 2, 4 and 8μmoL / L for 48h, the growth of U937 cells was inhibited to a certain extent with the inhibition rates of (10.49 ± 3.69)%, (13.79 ± 2.13 )%, (31.42 ± 3.90)% and (44.05 ± 1.56)%, respectively, and showed a more obvious concentration dependence. EKI-785 induced early apoptosis in U937 cells. At 48 and 72 hours after U437 cells were treated with 4μmol / L EKI-785, the proportion of early apoptotic cells in Annexin-V ~ + / PI ~ 2.03 times and 6.64 times. The phosphorylated MEK and Akt protein levels decreased with time after treated with 4μmol / L EKI-785 for 24,48,72h, and phosphorylated MEK and Akt almost disappeared at 72h. Conclusion The erbB family of receptors is likely to play an important role in the pathogenesis of certain types of leukemia and is expected to become a new therapeutic target for these leukemias. EKI-785 has potential application in the treatment of leukemia.