抑制野生型p53诱导的蛋白磷酸酶1提高肺腺癌细胞对砷剂的敏感性

发布时间：2018-07-04 11:05

本文选题：野生型p诱导的蛋白磷酸酶 + 三氧化二砷　；参考：《卫生研究》2017年03期

【摘要】：目的研究抑制野生型p53诱导的蛋白磷酸酶1(wild-type p53-induced phosphatase 1,WIP1)提高肺腺癌细胞对砷剂的敏感性及其分子机制。方法设计并合成WIP1编码基因的靶向siRNA,以Lipofectimient 2000转染入肺腺癌A549细胞,从而抑制WIP1蛋白的表达。运用Western-blot检测抑制WIP1对三氧化二砷(As_2O_3)诱导的P53磷酸化蛋白及其下游效应因子的影响;采用流式细胞术检测抑制WIP1后细胞凋亡情况;以噻唑蓝(MTT)法检测抑制WIP1后细胞的存活率。结果以siRNA技术成功将A549细胞中WIP1 mRNA和蛋白表达水平均抑制70%左右。相同浓度As_2O_3(5~40μmol/L)处理后,WIP1表达抑制组细胞中P53 ser15表达低于对照组,而P53 ser46表达显著高于对照组。此外,相同浓度As_2O_3(10~40μmol/L)处理后,WIP1表达抑制组中P53 ser15下游效应因子——P53上调凋亡诱导蛋白(P53 up-regulated modulator of apoptosis,PUMA)的表达水平显著低于对照组,而P53ser46下游效应因子——细胞周期调节蛋白P21的表达显著高于对照组。与之对应的是,相同剂量As_2O_3(5~40μmol/L)作用下,WIP1表达抑制组细胞的凋亡率和细胞死亡率均显著高于对照组。结论抑制肺腺癌细胞中WIP1的表达可通过促进P53磷酸化进程提高砷引发肺腺癌细胞凋亡的效率。
[Abstract]:Objective to study the molecular mechanism of inhibition of wild-type p53 induced protein phosphatase 1 (wild-type p53-induced phosphatase 1 WIP1) on the susceptibility of lung adenocarcinoma cells to arsenic. Methods targeted siRNAs encoding WIP1 gene were designed and synthesized and transfected into lung adenocarcinoma A549 cells with lipofectious 2000 to inhibit the expression of WIP1 protein. Western-blot was used to detect the effect of inhibition of WIP1 on p53 phosphorylated protein and its downstream effector factors induced by arsenic trioxide (as _ S _ 2O _ 3), flow cytometry was used to detect the cell apoptosis after inhibition of WIP1, and MTT method was used to detect the survival rate of cells after inhibition of WIP1. Results the expression of WIP1 mRNA and protein in A549 cells was inhibited by 70% by siRNA technique. The expression of p53 ser15 and p53 ser46 in the control group were significantly lower than those in the control group after treatment with the same concentration of as _ 2O _ 3 (5 ~ 40 渭 mol / L), but the expression of p53 ser46 was significantly higher than that in the control group. In addition, the expression level of p53 up-regulated modulator of apoptosis-inducing protein (Puma) was significantly lower in the group treated with the same concentration of as _ 2O _ 3 (10 ~ 40 渭 mol / L) than in the control group. The expression of P21, the downstream effector factor of P53ser46, was significantly higher than that of the control group. In contrast, the apoptosis rate and cell death rate of the WIP1 expression inhibition group were significantly higher than those of the control group under the same dose of As-Sh _ 2O _ 3 (5 ~ 40 渭 mol / L). Conclusion inhibiting the expression of WIP1 in lung adenocarcinoma cells can increase the efficiency of arsenic induced apoptosis of lung adenocarcinoma cells by promoting p53 phosphorylation.
【作者单位】：四川农业大学食品学院;四川大学华西公共卫生学院;
【基金】：国家自然科学基金面上项目(No.81372945)
【分类号】：R734.2

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