Department of Surgery, Penn State Milton S. Hershey Medical Center and College of Medicine, Hershey, Pennsylvania, USA.
Dis Colon Rectum. 2010 Mar;53(3):257-63. doi: 10.1007/DCR.0b013e3181c522c7.
Inflammatory bowel disease (IBD)-associated colorectal carcinogenesis involves dysregulation of multiple cellular pathways, including p53 signaling and cytokine action. The purpose of the current study was to evaluate the effects of tumor necrosis factor alpha (TNF-alpha) on p53 and p53 up-regulated modulator of apoptosis (PUMA), a downstream effector of p53 in the apoptotic pathway in colorectal cancer cells.
The cell lines HT29 (which express mutant p53) and HCT116 (which express wild-type p53) were treated with TNF-alpha (0, 50, 100, or 500 ng/mL) for 1, 12, 24, or 48 hours. Protein expression and subcellular localization of p53 and PUMA were determined by immunoblot and immunofluorescence. Changes in p53 and PUMA mRNA expression were determined by quantitative real time polymerase chain reaction.
Nuclear p53 expression was increased in TNF-alpha-treated HT29 cells; in contrast, expression was decreased or minimally changed in TNF-alpha-treated HCT116 cells, as determined by immunoblot and immunofluorescence. At 24 hours, p53 mRNA transcript levels were minimally increased in HT29 cells, whereas PUMA increased 34-fold.
TNF-alpha increased nuclear p53 expression in HT29 cells, which express p53 mutation, but not in HCT116 cells, which are wild type for p53. In addition, TNF-alpha markedly up-regulated PUMA mRNA levels in HT29 cells. Our findings suggest that TNF-alpha may be a factor in carcinogenesis in IBD in cells carrying a p53 mutation.
炎症性肠病（IBD）相关结直肠癌的发生涉及多个细胞途径的失调，包括 p53 信号和细胞因子作用。本研究的目的是评估肿瘤坏死因子-α（TNF-α）对结直肠癌细胞中 p53 信号和凋亡途径下游效应物 p53 上调凋亡调节剂（PUMA）的影响。
用 TNF-α（0、50、100 或 500ng/ml）处理 HT29（表达突变型 p53）和 HCT116（表达野生型 p53）细胞系 1、12、24 或 48 小时。通过免疫印迹和免疫荧光法测定 p53 和 PUMA 的蛋白表达和亚细胞定位。通过定量实时聚合酶链反应测定 p53 和 PUMA mRNA 表达的变化。
TNF-α处理的 HT29 细胞中核 p53 表达增加；相反，TNF-α处理的 HCT116 细胞中表达减少或几乎不变，通过免疫印迹和免疫荧光法测定。在 24 小时时，HT29 细胞中 p53 mRNA 转录本水平仅略有增加，而 PUMA 增加了 34 倍。
TNF-α增加了 HT29 细胞中 p53 突变表达的核表达，但对 HCT116 细胞（野生型 p53）没有影响。此外，TNF-α显著上调 HT29 细胞中 PUMA mRNA 水平。我们的发现表明，TNF-α可能是 IBD 中携带 p53 突变的细胞发生癌变的一个因素。
