Department of Surgery, Saitama Medical Center, Jichi Medical University, Saitama, Japan.
Int J Oncol. 2012 Mar;40(3):686-94. doi: 10.3892/ijo.2011.1231. Epub 2011 Oct 13.
The Pleckstrin and Sec7 domain-containing (PSD) gene, which regulates skeletal rearrangements, has been found to be more frequently methylated both in ulcerative colitis (UC)-associated colorectal cancer tissues (5 of 7; 71.4%) and matched normal epithelia (4 of 7; 57.1%) compared to non-neoplastic UC epithelia (6 of 22; 27.3%) and sporadic colorectal cancer tissues (6 of 32; 18.8%). The levels of PSD mRNA were positively correlated with the methylation status of PSD, as shown by both MSP and bisulfite sequencing. To determine the potential role of PSD silencing in the mechanisms underlying UC-associated carcinogenesis, the levels of senescence, proliferation and apoptosis were evaluated in a normal human fibroblast cell line (NHDF) in which 93% of PSD expression was knocked down by a small-interfering RNA (si-RNA). Although there were no significant differences in the levels of senescence and proliferation caused by PSD knockdown, the level of apoptosis was significantly decreased by PSD knockdown (5.3% in siControl-treated cells vs. 0.67% in siPSD-treated cells, p=0.0001). In addition, reactive oxygen species inducers accelerated apoptosis in NHDF and a neutrophil-like cell line, which was significantly reduced by PSD knockdown. To verify the effect of PSD methylation in tissue sections including 21 samples from UC patients with or without tumors, we elucidated PSD promoting accumulation of filamentous-actin (F-actin) and apoptosis by immunohistochemistry and TUNEL assay, respectively. Both levels of accumulation of F-actin and apoptosis were significantly decreased in specimens from UC patients with PSD methylation compared to those without PSD methylation (F-actin: 0.69±0.86 with vs. 1.57±0.51 without, p=0.0031, apoptotic index: 0.31±0.63 with vs. 1.0±0.88 without, p=0.0277). In conclusion, our results indicate that PSD methylation plays a significant role in the mechanisms underlying UC-associated carcinogenesis through its inhibitory effect on apoptosis in the interaction between colorectal mucosa and neutrophils.
Pleckstrin 和 Sec7 结构域包含（PSD）基因，可调节骨骼重排，与非肿瘤性 UC 上皮（22 例中的 6 例；27.3%）和散发性结直肠癌组织（32 例中的 6 例；相比，溃疡性结肠炎（UC）相关结直肠癌组织（7 例中的 5 例；71.4%）和匹配的正常上皮（7 例中的 4 例；57.1%）中 PSD 的甲基化更为频繁。PSD mRNA 的水平与 PSD 的甲基化状态呈正相关，MSP 和亚硫酸氢盐测序均显示如此。为了确定 PSD 沉默在 UC 相关致癌作用机制中的潜在作用，在通过小干扰 RNA（siRNA）将 93%的 PSD 表达敲低的正常人成纤维细胞系（NHDF）中评估衰老、增殖和凋亡的水平。尽管 PSD 敲低导致衰老和增殖的水平没有显着差异，但 PSD 敲低显着降低了凋亡水平（siControl 处理的细胞为 5.3%，而 siPSD 处理的细胞为 0.67%，p=0.0001）。此外，活性氧诱导剂加速了 NHDF 和中性粒细胞样细胞系的凋亡，而 PSD 敲低则显着降低了这种凋亡。为了验证 PSD 甲基化在包括 21 例 UC 患者肿瘤和非肿瘤组织样本中的作用，我们通过免疫组织化学和 TUNEL 分析分别阐明了 PSD 促进纤维状肌动蛋白（F-actin）积累和凋亡的作用。与没有 PSD 甲基化的标本相比，UC 患者有 PSD 甲基化的标本中 F-actin 积累和凋亡的水平显着降低（F-actin：0.69±0.86 与 1.57±0.51 无，p=0.0031，凋亡指数：0.31±0.63 与 1.0±0.88 无，p=0.0277）。总之，我们的结果表明，PSD 甲基化通过抑制结直肠黏膜与中性粒细胞之间的相互作用中的凋亡，在 UC 相关致癌作用机制中发挥重要作用。
