雷帕霉素对慢性髓性白血病细胞增殖的抑制作用及其机制

Inhibitory effects of rapamycin on proliferation of chronic myeiogenous leukemia cells and its mechanism

目的探讨雷帕霉素对慢性髓性白血病（CML）细胞增殖的抑制作用及可能机制。方法用不同浓度雷帕霉素处理CML细胞系K562细胞，用MTT法检测雷帕霉素对K562细胞的增殖抑制率；Westernblot法及RT-PCR法检测不同浓度雷帕霉素作用后的K562细胞中mTOR、4E．BP1、p70S6K蛋白及mRNA表达水平，Westernblot法检测CML慢性期患者骨髓原代细胞中mTOR、4E．BPl、p70S6K蛋白及磷酸化水平，并以健康人骨髓细胞为对照；数据采用）（2检验、Fisher确切概率计算、单因素方差分析（ANOVA）方法进行分析。结果CML慢性期患者骨髓中roTOR、4E—BP1、p70S6K蛋白磷酸化水平与正常对照相比明显增高（70．6％对30．0％，76．5％对40．0％，73．5％对20．0％，P值均〈0．05）；20nmol／L及更高浓度雷帕霉素对K562细胞增殖有明显抑制作用；雷帕霉素作用可降低K562细胞中mTOR磷酸化水平，及4E—BP1、p70S6K蛋白及mRNA表达水平（P值均〈0．05）。结论roTOR途径在CML的发病中发挥重要作用，其靶向抑制剂雷帕霉素可通过阻断roTOR途径抑制K562细胞增殖。

Objective To explore the inhibitory effects of rapamycin on proliferation of chronic my- elogenous leukemia（CML） cells and its possible mechanism. Methods The effects of rapamycin at various concentrations on cell proliferation of CML cell line K562 cells were analyzed by MTT. The expressions of mTOR, 4E-BP1 and p70S6K at protein and mRNA level in K562 cells with rapamycin treatment were detec- ted by Western blot and RT-PCR. The protein expressions and phosphorylation of roTOR, 4E-BP1 and pTOS6K in primary bone marrow cells from CML patients at chronic phase （CP） were also investigated by Western blot, bone marrow cells from healthy people were used as control. Data were analyzed by the X2 test, Fisher＇ s exact test and one-way analysis of variance （ ANOVA ）. Results The phosphorylation of roTOR, 4E-BPI and p70S6K were significantly increased in CML bone marrow cells compared with that of normal con- trol（70.6% vs 30.0% ,76.5% vs 40.0% ,73.5% vs 20.0% ,respectively, P 〈0.05 ）. The proliferation of I（562 cells was significantly inhibited with 20 nmol/L and more rapamycin treatment. The phosphorylation of mTOR was decreased after rapamycin treatment, as well as the expressions of 4E-BP1 and pTOS6K at protein and mRNA level （ P 〈 0.05 ）. Conclusion roTOR signaling played an important role in CML pathogenesis, and rapamycin could decrease CML cells proliferation by inhibiting the activity of mTOR signaling in vitro.