抑制白血病K562细胞FoxM1表达可增强细胞对高三尖杉酯碱的敏感性*

陈谨1,周敏然2,孙婷2,秦雪梅2,陈忠敏3,陈春燕2，于媛2△

(1安徽医学高等专科学校，安徽 合肥 230601；2山东大学齐鲁医院血液科，山东 济南 250012；3重庆理工大学药学与生物工程学院， 重庆400054)

[摘要]目的： 探讨抑制白血病K562细胞叉头框蛋白M1(FoxM1)是否增强细胞对高三尖杉酯碱(HHT)的敏感性。方法: HHT以不同浓度(0、0.015、0.030和0.045 μmol/L)和最低起效浓度不同时间(0.015 μmol/L，0、24、48和72 h)作用于K562细胞，real-time PCR和Western blot检测FoxM1 mRNA和蛋白表达；以0.015 μmol/L HHT作用K562细胞后转染FoxM1 siRNA，观察沉默K562细胞FoxM1后细胞对HHT的敏感性、细胞增殖和凋亡效应以及FoxM1相关靶分子c-Myc和Sp1表达状况。结果: 随着HHT浓度增加和时间延长FoxM1表达逐渐降低，说明HHT抑制K562细胞FoxM1表达；HHT处理K562细胞后转染FoxM1 siRNA，细胞生长和克隆形成显著下降，细胞凋亡增加，因此抑制FoxM1可增加K562细胞对HHT的敏感性；FoxM1 siRNA组c-Myc和Sp1表达显著降低，表明FoxM1可正性调控c-Myc和Sp1表达。结论: HHT可以抑制白血病K562细胞FoxM1表达，干扰FoxM1可增强细胞对HHT的敏感性。

[关键词]三尖杉酯碱； 叉头框蛋白M1； K562细胞； 药物敏感性

[中图分类号]R363.2[文献标志码]A

doi:10.3969/j.issn.1000-4718.2015.11.002

[文章编号]1000-4718(2015)11-1933-10

[收稿日期]2014-12-29[修回日期] 2015-08-14

[基金项目]*广州市属高校重点学科建设经费资助项目(穗教高教[2011]34号)

通讯作者△Tel: 020-84271652; E-mail: xuxia503@126.com

Inhibition of FoxM1 sensitizes leukemia K562 cells to homoharringtonineCHEN Jin1, ZHOU Min-ran2, SUN Ting2, QIN Xue-mei2, CHEN Zhong-min3, CHEN Chun-yan2, YU Yuan2

(1AnhuiMedicalCollege,Hefei230601,China;2DepartmentofHematology,QiluHospital,ShandongUniversity,Jinan250012,China;3SchoolofPharmacyandBioengineering,ChongqingUniversityofTechnology,Chongqing400054,China.E-mail:yuyuandoctor@163.com)

ABSTRACT[]AIM: To study whether inhibition of forkhead box protein M1(FoxM1) sensitizes leukemia K562 cells to homoharringtonine (HHT). METHODS: K562 cells were incubated with HHT at different concentrations (0 μmol/L, 0.015 μmol/L, 0.030 μmol/L and 0.045 μmol/L) for different time (0 h, 24 h, 48 h and 72 h). The mRNA and protein levels of FoxM1 were detected by real-time PCR and Western blot. FoxM1 siRNA was transfected into K562 cells with 0.015 μmol/L HHT after 6 h. After 72 h incubation, the cell proliferation was detected by cell counting and soft agar assay, and the proportion of apoptotic K562 cells was determined by flow cytometry. The expression of c-Myc and Sp1 were detected by real-time PCR and Western blot. RESULTS: FoxM1 expression was reduced time-dependently and dose-dependently, suggesting that HHT mediated the downregulation of FoxM1 in K562 cells. In K562 cells, treatment with FoxM1 siRNA and HHT inhibited the cell proliferation and promoted the apoptosis significantly. Therefore, inhibition of FoxM1 sensitized leukemia K562 cells to HHT. The expression of c-Myc and Sp1 was positively regulated by FoxM1. CONCLUSION: HHT inhibits Forkhead box protein M1 expression in K562 cells. Inhibition of FoxM1 sensitizes K562 cells to HHT.

[KEY WORDS]Homoharringtonine; Forkhead box protein M1; K562 cells; Drug sensitivity

慢性粒细胞白血病(chronic myelogenous leukemia，CML)是起源于多能造血干细胞的恶性克隆增殖性疾病，包含恶性程度递增的慢性期、加速期和急变期。CML一旦进入急变期，细胞增殖加快、分化受阻，骨髓和外周血中不成熟细胞大量积聚，对靶向抑制BCR/ABL酪氨酸激酶活性的伊马替尼等治疗反应差，病情恶化，预后不佳。研究表明CML在急变演进过程中分子调控方式发生了改变，其中涉及不依赖BCR/ABL的复杂调控过程[1-2]。……