Zhuang Yong, Li Dong, Fu Jinqiu, Shi Qing, Lu Yuanyuan, Ju Xiuli
Department of Pediatrics, Qilu Hospital, Shandong University, Ji'nan, Shandong 250012, P.R. China.
Cryomedicine Laboratory, Qilu Hospital, Shandong University, Ji'nan, Shandong 250012, P.R. China.
Oncol Rep. 2014 Mar;31(3):1183-90. doi: 10.3892/or.2013.2964. Epub 2013 Dec 31.
The AIOLOS gene is important in the control of mature B-lymphocyte differentiation and proliferation. Previous research has shown that deregulated AIOLOS expression is associated with adult B-cell acute lymphoblastic leukemia (ALL) and chronic lymphocytic leukemia in human patients. However, the function of AIOLOS in childhood B-cell precursor (BCP)-ALL is not fully understood. In the present study, Nalm-6 cells were divided into three groups: the untransfected control (UT), the lentiviral vector control (Lenti-Mock) and the AIOLOS-overexpressing (Lenti-AIOLOS) group. Lenti-AIOLOS Nalm-6 cells were constructed by lentiviral transduction, followed by cell proliferation assay, cell-cycle analysis and apoptosis assay, to evaluate the effects of AIOLOS on proliferation, cell cycle distribution and apoptosis of Nalm-6 cells in vitro. Moreover, the expression levels of genes associated with apoptosis and the cell cycle, as well as the transcription factors IKZF1 and NF-κB, were investigated by quantitative reverse transcription-polymerase chain reaction and western blot analysis. The results showed that the proliferation of Nalm-6 cells in the Lenti-AIOLOS group was reduced by 16% on day 8 compared with cells in the UT group (P>0.05). The reduction peaked at 29% on day 10 (P<0.05). The percentage of Nalm-6 cells in the G0/G1 phase increased from 70.4 (UT) to 84.1% (Lenti-AIOLOS) (P<0.01), and the S-phase cells decreased from 20.3 (UT) to 11.7% (Lenti-AIOLOS) (P<0.01). Total apoptotic cells significantly decreased in AIOLOS-transfected Nalm-6 cells (10.75%) compared with those in the Lenti-Mock (17.00%) or UT group (19.05%) (P<0.01). In particular, the difference between the groups in the percentage of late apoptotic cells was significant (2.85 vs. 7.95%; P<0.01). In addition, overexpression of AIOLOS resulted in upregulation of BCL-2 and downregulation of CCND3, BAX, IKZF1 and NF-κB. No changes were detected on C-MYC and P27. Our findings indicate that lentivirus-mediated overexpression of AIOLOS in Nalm-6 cells could inhibit cell proliferation, suppress cell apoptosis and arrest the cell cycle at the G0/G1 phase in vitro.
AIOLOS基因在成熟B淋巴细胞的分化和增殖控制中起着重要作用。先前的研究表明，AIOLOS表达失调与人类患者的成人B细胞急性淋巴细胞白血病（ALL）和慢性淋巴细胞白血病有关。然而，AIOLOS在儿童B细胞前体（BCP）-ALL中的功能尚未完全了解。在本研究中，将Nalm-6细胞分为三组：未转染对照组（UT）、慢病毒载体对照组（Lenti-Mock）和AIOLOS过表达组（Lenti-AIOLOS）。通过慢病毒转导构建Lenti-AIOLOS Nalm-6细胞，随后进行细胞增殖测定、细胞周期分析和凋亡测定，以评估AIOLOS对Nalm-6细胞体外增殖、细胞周期分布和凋亡的影响。此外，通过定量逆转录-聚合酶链反应和蛋白质印迹分析，研究了与凋亡和细胞周期相关的基因以及转录因子IKZF1和NF-κB的表达水平。结果显示，与UT组细胞相比，Lenti-AIOLOS组Nalm-6细胞在第8天的增殖率降低了16%（P>0.05）。在第10天，降低幅度达到峰值，为29%（P<0.05）。Nalm-6细胞在G0/G1期的比例从70.4%（UT）增加到84.1%（Lenti-AIOLOS）（P<0.01），S期细胞从20.3%（UT）减少到11.7%（Lenti-AIOLOS）（P<0.01）。与Lenti-Mock组（17.00%）或UT组（19.05%）相比，AIOLOS转染的Nalm-6细胞中总凋亡细胞显著减少（10.75%）（P<0.01）。特别是，各组晚期凋亡细胞百分比之间的差异显著（2.85%对7.95%；P<0.01）。此外，AIOLOS的过表达导致BCL-2上调，CCND3、BAX、IKZF1和NF-κB下调。未检测到C-MYC和P27的变化。我们的研究结果表明，慢病毒介导的AIOLOS在Nalm-6细胞中的过表达可在体外抑制细胞增殖、抑制细胞凋亡并使细胞周期停滞在G0/G1期。
