Notch1信号通路对人肺癌A549细胞的影响。

Effects of the Notch1 signaling pathway on human lung cancer A549 cells.

作者信息

Zeng Yun, Yin Bijian, Wang Xinwei, Xia Guohao, Shen Zhengjie, Gu Wenzhe, Wu Mianhua

机构信息

a Department of Medical Oncology , Jiangsu Cancer Hospital , Nanjing , Jiangsu Province , China.

b First Clinical College , Nanjing University of Chinese Medicine , Nanjing , Jiangsu Province , China.

出版信息

Exp Lung Res. 2017 May-Jun;43(4-5):208-216. doi: 10.1080/01902148.2017.1341008. Epub 2017 Jul 18.

DOI:10.1080/01902148.2017.1341008

PMID:28718726

Abstract

PURPOSE

To evaluate the effects of the Notch1 signaling pathway on human lung cancer A549 cells.

MATERIALS AND METHODS

A549 cells were transfected with recombinant plasmids. Cell proliferation was detected by MTT assay. A tumor-bearing mouse model was established for intratumoral gene injection. Apoptosis-related factors were detected by immunohistochemical assay. Caspase-8, caspase-3, caspase-9, PI3K, pAkt and pSTAT3 expressions were detected by Western blotting.

RESULTS

Compared with A549-GFP and A549 cells, A549-ICN cell growth in mice decelerated, tumor volume significantly reduced (p < 0.01), and survival time significantly increased (p < 0.05). Cyclin E and phosphorylated Rb protein expressions were significantly down-regulated. Compared with control, apoptosis-related protein Bcl-2 expression in tumors injected with Notch1 gene was significantly inhibited. After Deltex1 transfection, A549 cell proliferation decelerated, growth was significantly inhibited (p < 0.05), and survival time was significantly extended (p < 0.05). Cyclin E and mutant p53 protein expressions in tumors were down-regulated, phosphorylated Rb expression was almost completely inhibited, and Bcl-2 expression was significantly inhibited. TNF-α-related apoptosis-inducing ligand (TRAIL) inhibited A549-ICN cell growth time- and dose-dependently. After treatment for 24 h or longer, TRAIL induced apoptosis of more A549-ICN cells. Cleaved caspase-3 and cleaved caspase-9 were detected only in A549-ICN cells after 6 h of 40 ng/mL TRAIL treatment, but cleaved caspase-8 was not detected. Combining Notch1 signal with TRAIL inhibited PI3K, phosphorylated Akt and phosphorylated STAT3 expressions.

CONCLUSION

The Notch1 signaling pathway may inhibit A549 cell growth in vitro and in vivo by regulating cell cycle-related and anti-apoptotic protein expressions. Notch1 activation also suppressed A549 cell apoptosis by inhibiting the PI3K/pAkt pathway and activating the caspase-3 pathway in cooperation with TRAIL.

摘要

目的

评估Notch1信号通路对人肺癌A549细胞的影响。

材料与方法

用重组质粒转染A549细胞。通过MTT法检测细胞增殖。建立荷瘤小鼠模型用于瘤内基因注射。通过免疫组化法检测凋亡相关因子。通过蛋白质印迹法检测Caspase-8、Caspase-3、Caspase-9、PI3K、pAkt和pSTAT3的表达。

结果

与A549-GFP和A549细胞相比，A549-ICN细胞在小鼠体内的生长减速，肿瘤体积显著减小（p < 0.01），生存时间显著延长（p < 0.05）。细胞周期蛋白E和磷酸化Rb蛋白表达显著下调。与对照组相比，注射Notch1基因的肿瘤中凋亡相关蛋白Bcl-2的表达受到显著抑制。转染Deltex1后，A549细胞增殖减速，生长受到显著抑制（p < 0.05），生存时间显著延长（p < 0.05）。肿瘤中的细胞周期蛋白E和突变型p53蛋白表达下调，磷酸化Rb表达几乎完全被抑制，Bcl-2表达显著被抑制。肿瘤坏死因子-α相关凋亡诱导配体（TRAIL）对A549-ICN细胞生长的抑制具有时间和剂量依赖性。处理24小时或更长时间后，TRAIL诱导更多A549-ICN细胞凋亡。仅在40 ng/mL TRAIL处理6小时后的A549-ICN细胞中检测到裂解的Caspase-3和裂解的Caspase-9，但未检测到裂解的Caspase-8。将Notch1信号与TRAIL联合使用可抑制PI3K、磷酸化Akt和磷酸化STAT3的表达。