慢病毒介导的WWOX表达对急性淋巴细胞白血病细胞增殖活性的影响
| 作者: |
1罗剑锋,
1罗丹,
1文丹宁
1 武汉市金银潭医院感染科，武汉 430000 |
| 通讯: |
罗剑锋
Email: 189044065@qq.com |
| DOI: | 10.3978/j.issn.2095-6959.2018.11.006 |
摘要
目的：研究慢病毒介导的含WW域的氧化还原酶(WW Domain-containing Oxidoreductase，WWOX)表达对急性淋巴细胞白血病细胞增殖活性的影响。方法：用过表达WWOX重组慢病毒和阴性对照重组慢病毒感染急性淋巴细胞白血病细胞Jurkat，分别记为Lenti WWOX，Lenti NC；用Realtime PCR和Western印迹法检测细胞中WWOX表达水平。用含有Wnt信号通路抑制剂FH535的培养液培养感染过表达WWOX重组慢病毒和感染阴性对照重组慢病毒的Jurkat细胞，分别记为Lenti WWOX+FH535，Lenti NC+FH535；用Realtime PCR和Western印迹法检测细胞中β-catenin，c-myc表达水平。MTT测定细胞增殖活性，PI单染和Annexin V-FITC/PI双染法分别检测细胞周期和凋亡变化，Western印迹法检测细胞中周期相关蛋白Cyclin-D1，p27和凋亡蛋白C-Caspase-3表达水平。结果：Lenti WWOX细胞中WWOX表达水平均明显高于Lenti NC(P<0.05)。Lenti WWOX，Lenti NC+FH535，Lenti WWOX+FH535细胞中β-catenin，c-myc蛋白水平均明显低于Lenti NC，并且Lenti WWOX + FH535细胞中β-catenin，c-myc蛋白水平减少最多。Lenti WWOX，Lenti NC+FH535，Lenti WWOX+FH535细胞增殖能力降低，细胞G0/G1比例升高，细胞凋亡率升高，细胞中Cyclin-D1蛋白水平降低，p27，C-Caspase-3蛋白水平升高，与Lenti NC比较，差异均有统计学意义(P<0.05)。Lenti WWOX+FH535细胞增殖能力降低，细胞G0/G1比例升高，细胞凋亡率升高，细胞中Cyclin-D1蛋白水平降低，p27，C-Caspase-3蛋白水平升高，与Lenti WWOX，Lenti NC+FH535比较，差异均有统计学意义(P<0.05)。结论：慢病毒介导的WWOX表达能够抑制急性淋巴细胞白血病细胞增殖，阻滞细胞周期，诱导细胞凋亡，作用机制与抑制Wnt信号通路有关。
关键词：
急性淋巴细胞白血病；细胞周期；WWOX；Wnt信号通路
Effect of lentivirus mediated WWOX expression on proliferation of acute lymphoblastic leukemia cells
CorrespondingAuthor: LUO Jianfeng Email: 189044065@qq.com
DOI: 10.3978/j.issn.2095-6959.2018.11.006
Abstract
Objective: To study the effect of lentivirus mediated WW Domain-containing Oxidoreductase (WWOX) expression on the proliferation of acute lymphoblastic leukemia. Methods: WWOX recombinant lentivirus and infection negative control recombinant lentivirus infected acute lymphoblastic leukemia cells Jurkat were recorded as Lenti WWOX and Lenti NC, and WWOX expression levels were detected by Realtime PCR and Western blot methods. The Jurkat cells infected with WWOX recombinant lentivirus and negative control recombinant lentivirus were cultured with the culture solution containing the Wnt signaling pathway inhibitor FH535, and the Jurkat cells of the recombinant lentivirus and the negative control recombinant lentivirus were recorded as Lenti WWOX + FH535 and Lenti NC + FH535. The expression levels of β-catenin and c-myc were detected by Realtime PCR and Western blot. MTT assay cell proliferation activity, PI single staining and Annexin V-FITC/PI double staining were used to detect cell cycle and apoptosis respectively. The expression levels of Cyclin-D1, p27 and apoptotic protein C-Caspase-3 were detected by Western blot. Results: The expression level of WWOX in Lenti WWOX cells was significantly higher than that in Lenti NC (P<0.05). The levels of β-catenin and c-myc protein in Lenti WWOX, Lenti WWOX + FH535 and Lenti NC + FH535 were significantly lower than those in Lenti NC. Moreover, the level of β-catenin and c-myc protein in Lenti WWOX + FH535 cells decreased most. The proliferation of Lenti WWOX, Lenti WWOX + FH535 and Lenti NC + FH535 were increased, the rate of apoptosis was increased, the level of Cyclin-D1 protein in the cells decreased, the levels of p27 and C-Caspase-3 protein were elevated, compared with Lenti NC, the difference was statistically significant (P<0.05). The proliferation ability of Lenti WWOX + FH535 cells was reduced, the proportion of G0/G1 in cells increased, the rate of apoptosis was increased, the level of Cyclin-D1 protein in the cells decreased, the levels of p27 and C-Caspase-3 protein were elevated, compared with Lenti WWOX and Lenti NC + FH535, the difference was statistically significant (P<0.05). Conclusion: The expression of lentivirus mediated WWOX can inhibit the proliferation of acute lymphoblastic leukemia cells, block cell cycle, and induce apoptosis, the mechanism of action is related to the inhibition of Wnt signaling pathway.
Keywords：
acute lymphoblastic leukemia; cell cycle; WWOX; Wnt signaling pathway