Zhang Hong-Mei, Sang Xiao-Guang, Wang Yan-Ze, Cui Can, Zhang Li, Ji Wan-Sheng
Hong-Mei Zhang, Li Zhang, Wan-Sheng Ji, Department of Gastroenterology, Affiliated Hospital of Weifang Medical University, Weifang 261031, Shandong Province, China.
World J Gastroenterol. 2017 Apr 21;23(15):2716-2722. doi: 10.3748/wjg.v23.i15.2716.
To investigate the role of Δ133p53 isoform in nuclear factor-κB (NF-κB) inhibitor pyrrolidine dithiocarbamate (PDTC)-mediated growth inhibition of MKN45 gastric cancer cells.
The growth rate of MKN45 cells after treatment with different concentrations of only PDTC or PTDC in combination with cisplatin was detected by the CCK-8 assay. mRNA expression levels of Δ133p53, p53β, and the NF-κB p65 subunit and p65 protein levels were detected by reverse transcription-polymerase chain reaction (RT-PCR) and immunofluorescence, respectively. Growth of MKN45 cells was significantly inhibited by PDTC alone in a dose-dependent manner ( < 0.01). Moreover, the inhibitory effect of cisplatin was remarkably enhanced in a dose-dependent manner by co-treatment with PDTC ( < 0.01).
RT-PCR analysis revealed that mRNA expression of p65 was curbed significantly in a dose-dependent manner by treatment with only PDTC ( < 0.01), and this suppressive effect was further enhanced when co-treated with cisplatin ( < 0.01). With respect to the other p53 isoforms, mRNA level of Δ133p53 was significantly reduced in a dose-dependent manner by treatment with only PDTC or PTDC in combination with cisplatin ( < 0.01), whereas p53β mRNA expression was not altered by PDTC treatment ( > 0.05). A similar tendency of change in p65 protein expression, as observed for the corresponding mRNA, was detected by immunofluorescence analysis ( < 0.01). Pearson correlation analysis demonstrated that Δ133p53 and p65 mRNA expression levels were positively related, while no significant relationship was observed between those of p65 and p53β ( = 0.076, > 0.01).
Δ133p53 isoform (not p53β) is required in PDTC-induced inhibition of MKN45 gastric cancer cells, indicating that disturbance in the cross-talk between p53 and NF-κB pathways is a promising target in pharmaceutical research for the development of treatment strategies for gastric cancer.
探讨Δ133p53亚型在核因子-κB（NF-κB）抑制剂吡咯烷二硫代氨基甲酸盐（PDTC）介导的MKN45胃癌细胞生长抑制中的作用。
采用CCK-8法检测不同浓度的单纯PDTC或PDTC联合顺铂处理后MKN45细胞的生长速率。分别通过逆转录-聚合酶链反应（RT-PCR）和免疫荧光法检测Δ133p53、p53β以及NF-κB p65亚基的mRNA表达水平和p65蛋白水平。单纯PDTC可显著抑制MKN45细胞生长，呈剂量依赖性（P<0.01）。此外，PDTC与顺铂联合处理可显著增强顺铂的抑制作用，呈剂量依赖性（P<0.01）。
RT-PCR分析显示，单纯PDTC处理可显著抑制p65的mRNA表达，呈剂量依赖性（P<0.01），与顺铂联合处理时这种抑制作用进一步增强（P<0.01）。对于其他p53亚型，单纯PDTC或PDTC联合顺铂处理可使Δ133p53的mRNA水平显著降低，呈剂量依赖性（P<0.01），而PDTC处理未改变p53β的mRNA表达（P>0.05）。免疫荧光分析检测到p65蛋白表达的变化趋势与相应mRNA相似（P<0.01）。Pearson相关性分析表明，Δ133p53与p65的mRNA表达水平呈正相关，而p65与p53β之间未观察到显著相关性（r=0.076，P>0.01）。
PDTC诱导的MKN45胃癌细胞抑制作用需要Δ133p53亚型（而非p53β）参与，表明p53与NF-κB信号通路之间的相互作用紊乱是胃癌治疗策略药物研发的一个有前景的靶点。
