Yes相关蛋白在脂多糖诱导肺癌A549细胞增殖中的作用研究

发布时间：2018-07-12 14:46

本文选题：脂多糖 + Yes相关蛋白　；参考：《医学研究生学报》2017年11期

【摘要】：目的慢性炎症与肿瘤的发生发展密切相关,但具体机制不详。文章旨在探讨脂多糖对肺腺癌A549细胞增殖的影响,并阐明YAP在其中所发挥的作用。方法培养人肺腺癌A549细胞,应用CCK-8法检测不同浓度LPS对A549增殖的影响,用Western blot、免疫荧光技术检测LPS处理后A549中YAP定量及定位的变化;实验设空白组、NCsiRNA组、YAP siRNA1组、YAP siRNA2组、YAP siRNA3组细胞,分别提取RNA及蛋白,通过siRNA抑制YAP表达,Western blot、RT-PCR检测siRNA抑制率;实验设对照组、LPS组、YAP siRNA组、YAP siRNA+LPS组,用CCK-8再次检测抑制YAP表达后LPS对A549细胞增殖影响。结果 LPS处理72 h后,与0μg/m L LPS处理后A值比较,0.01、0.1、1μg/m L处理后A值明显增大(P0.05),0.1μg/m L LPS处理后细胞增殖能力仍最强。Western blot检测发现,与0μg/m L浓度比较,其他浓度的LPS处理A549细胞48 h后,均能上调YAP总蛋白的表达(P0.05),其中以0.1μg/m L浓度最明显。RT-PCR检测结果显示,与空白组比较,YAP siRNA1组、YAP siRNA2组、YAP siRNA3组YAP mRNA的表达明显降低(P0.05);与NC siRNA组比较,YAP siRNA1组、YAP siRNA2组、YAP siRNA3组YAP mRNA的表达明显降低(P0.05)。Western blot检测结果与RT-PCR一致,转染YAP siRNA后能够显著抑制A549细胞中YAP蛋白的表达,其中以YAP siRNA1的抑制率最高(P0.05)。与对照组比较,LPS组24、48、72 h A549细胞均明显增殖(1.17±0.08 vs 1.47±0.08、1.95±0.16 vs 2.35±0.07、2.46±0.07 vs 3.01±0.08,P0.05);与LPS组比较,YAP siRNA+LPS组48、72 h A549细胞增殖能力(1.92±0.15、2.38±0.07)明显减弱(P0.05)。结论低浓度的LPS能够通过上调核内YAP的表达来促进A549细胞的增殖,这为LPS诱导的肺癌细胞增殖机制提供了新的理论依据。
[Abstract]:Objective chronic inflammation is closely related to the occurrence and development of tumor, but the specific mechanism is unknown. The aim of this study was to investigate the effect of lipopolysaccharide on the proliferation of lung adenocarcinoma cell line A549 and to elucidate the role of YAP in it. Methods Human lung adenocarcinoma A549 cells were cultured. The effects of LPS on the proliferation of A549 cells were detected by CCK-8 method. The quantitative and localization changes of YAP in A549 cells treated with LPS were detected by Western blot and immunofluorescence technique. RNA and protein were extracted from YAP siRNA2 cells, and the inhibition rate of siRNA was detected by Western blotRT-PCR in the control group (LPS group), YAP siRNA group (LPS group) and YAP siRNA group (LPS group). The effect of LPS on the proliferation of A549 cells was detected by CCK-8. Results after lipopolysaccharide (LPS) treatment for 72 h, the A value of A549 cells was significantly higher than that of 0 渭 g / mL LPS treatment (P0.05). Western blot analysis showed that compared with 0 渭 g / mL LPS treatment, the proliferation ability of A549 cells treated with other concentrations of LPS was higher than that of 0 渭 g / mL LPS for 48 h. All of them could up-regulate the expression of YAP total protein (P0.05), and the concentration of 0.1 渭 g / mL was the most obvious. RT-PCR showed that, The expression of YAP mRNA in YAP siRNA1 group was significantly lower than that in control group (P0.05), and the expression of YAP mRNA in YAP siRNA1 group was significantly lower than that in NC siRNA group (P0.05). The results of Western blot detection were consistent with that of RT-PCR, and the expression of YAP mRNA in YAP siRNA1 group was significantly lower than that in NC siRNA group (P0.05), and the expression of YAP mRNA in YAP siRNA2 group was significantly lower than that in NC siRNA group (P0.05). YAP siRNA transfection could significantly inhibit the expression of YAP protein in A549 cells, in which YAP siRNA1 had the highest inhibition rate (P0.05). Compared with the control group, the proliferation of A549 cells in LPS group was significantly increased (1.17 卤0.08 vs 1.47 卤0.08 卤1.95 卤0.16 vs 2.35 卤0.07 卤2.46 卤0.07 vs 3.01 卤0.08 P 0.05), and the proliferation ability of A549 cells in LPS group was significantly decreased (1.92 卤0.152.38 卤0.07) compared with LPS group. Conclusion low concentration of LPS can promote the proliferation of A549 cells by upregulating the expression of YAP in nucleus, which provides a new theoretical basis for the mechanism of LPS-induced proliferation of lung cancer cells.
【作者单位】：苏北人民医院/扬州大学临床医学院呼吸内科;
【基金】：江苏省自然科学基金(BK20160451)
【分类号】：R734.2

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