As_2O_3和/或TGF-β1诱导NB4细胞凋亡中P27^(Kip1)、cyclin E、内源性TGF-β1的变化及其意义  

P27^(Kip1),cyclin E and Endogenous TGF-β1 Changes in Apoptosis of NB4 Cells Induced by As_2O_3 and/or TGF-β1 and Their Significance

作　　者：梁颖[1] 李艳[1] 王月[1] 李霞[1] 王萍萍[1] 王柏勋[1] 

机构地区：[1]中国医科大学第一附属医院血液科,辽宁沈阳110001

出　　处：《中国实验血液学杂志》2009年第1期74-79,共6页Journal of Experimental Hematology

基　　金：辽宁省攻关课题;编号2004225004-10

摘　　要：本研究探讨三氧化二砷(As2O3)和/或转化生长因子-β1(TGF-β1)作用于NB4细胞后的细胞凋亡情况、P27Kip1、cyclin E及内源性TGF-β1mRNA水平的变化及其意义。用MTT法检测As2O3对NB4细胞的细胞毒性及IC50,用瑞氏-姬姆萨染色观察凋亡细胞形态学的变化,流式细胞术检测细胞周期和凋亡,半定量RT-PCR检测P27Kip1、cyclin E以及内源性TGF-β1的mRNA水平。结果表明:As2O3、TGF-β1均能明显抑制NB4细胞的生长,促进NB4细胞的凋亡;As2O3对NB4细胞的抑制及促凋亡作用均呈剂量-时间依赖关系,24小时的IC50约为12μmol/L,48小时的IC50约为5μmol/L,72小时的IC50约为3μmol/L。As2O3和/或TGF-β1均可使NB4细胞出现细胞周期阻滞,5μmol/L As2O3使NB4细胞阻滞在G2/M期,5ng/ml TGF-β1使NB4细胞阻滞在G1期,两者联合处理组主要使S期阻滞。As2O3、TGF-β1分别作用于NB4细胞时均可见P27Kip1及内源性TGF-β1的mRNA表达上调,而cyclin E的mRNA表达下调;联合处理组结果显示TGF-β1可增强As2O3上述作用。结论:As2O3及TGF-β1均可诱导NB4细胞凋亡,引起细胞周期分布异常;As2O3及外源性TGF-β1可能通过上调内源性TGF-β1使P27Kip1高表达以诱导细胞凋亡;TGF-β1可能直接抑制了cyclin E的表达,或者通过上调P27Kip1的表达而反馈抑制cyclin E的活性,从而导致NB4细胞周期阻滞。This study was aimed to investigate the effects of arsenic trioxide （As2O3） and/or transfroming growth factor-β1 （ TGF-β1） on cell apoptosis and the changes of P27^Kip1, cyclin E and endogenous TGF-β1 mRNA levels in NB4 cells. As2O3 cytotoxicity to NB4 cells and the IC50 were assayed with MTT, the apoptotic morphological changes were observed by Wright-Giemsa staining; the cell cycle and apoptosis were detected with flow cytometry. Semiquantitative RT-PCR was used to examine P27^Kip1, cyclin E and endogenous TGF-β1 mRNA levels. The results showed that the As2O3 and TGF-β1 significantly suppressed the growth of NB4 cells, and promoted the apoptosis of these cells. The growth inhibition and apoptosis of NB4 cells treated with As2O3 were in dose-and time-dependent manners. IC50 were about 12 μmol/L for 24 hours, about 5 μmol/L for 48 hours, and about 3 μmol/L for 72 hours respectively. Cell cycle arrest in NB4 cells was induced by As2O3 and/or TGF-β1. The arrest of NB4 cells treated by 5 μmol/L AS2O3 was in G2/M phase, and 5 ng/ml TGF-β1 in G1 phase. However, the arrest of NB4 cells caused by combination of As2O3 and TGF-β1 was in S phase. After treating with As2O3, P27^Kip1 and endogenous TGF-β1 mRNA expressions of NB4 cells were up-regulated, and cyclin E mRNA expression was down-regulated. When NB4 cells were treated with TGF-β1 alone, P27^Kip1 and cyclin E mRNA expressions were the same as that treated by As2O3. Exogenous TGF-β1 enhanced the above effects of As2O3 in combination group. It is concluded that ms2O3 and TGF-β1 are able to induce apoptosis and cell cycle abnormal distribution in NB4 cells. As2O3 and exogenous TGF-β1 may up-regulate endogenous TGF-β1, which induce apoptosis of NB4 cells through consequently high expression of P27^Kip1. TGF-β1 may lead to cell cycle arrest by inhibiting the expression of cyclin E directly, or by the activity of cyclin E through the increased expression of P27^Kip1.

关 键 词：As2O3 TGF-Β1 NB4细胞 细胞凋亡 P27^KIP1 cyclin E 内源性 TGF-Β1 

分 类 号：R979.1[医药卫生—药品] R733.7[医药卫生—药学]