Inhibition of DNA methyltransferase by antisense oligodeoxynucleotide modifies cell characteristics in gastric cancer cell lines.
Inhibition of DNA methyltransferase by antisense oligodeoxynucleotide modifies cell characteristics in gastric cancer cell lines.
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DOI：
10.3892/or.12.3.527
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发表时间：
2004-09
期刊：
影响因子：
4.2
通讯作者：
Y. Saikawa;T. Kubota;Shingo Maeda;Y. Otani;K. Kumai;M. Kitajima
中科院分区：
文献类型：
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作者：
Y. Saikawa;T. Kubota;Shingo Maeda;Y. Otani;K. Kumai;M. Kitajima
DNA（cytosine-5-）-methyltransferase 1（DNMT 1）通过复杂的信号通路和转录因子网络在维持DNA甲基化模式中发挥重要作用，与细胞分化或癌变有关。本研究设计了DNMT 1的反义寡核苷酸（AS/MT：5 '-CGGTAC GCGCCGGCATCT-3'），并在体外胃癌细胞中成功地在蛋白水平上抑制DNMT 1的表达。AS/MT组E-cadherin蛋白表达增加，cyclin D1和PCNA表达减少。AS/MT还诱导抑制细胞生长，通过BrDU摄取掺入测定，以剂量依赖性方式，表明AS/MT的特异性。与此同时，2 μ M AS/MT孵育24 h后，在TMK-1和MKN-45细胞中观察到形态学改变。细胞从其原始形式改变形状为分散的成纤维细胞样细胞，具有神经突样突起，伴随着细胞粘附能力的增加。使用裸鼠系统的腹膜播散的体内模型显示AS/MT处理的TMK-1细胞的恶性潜能增加，如通过与NS/MT处理组相比，AS/MT中更多数量的腹膜肿瘤结节所证明的，分别为34.8+/-4.3对22.4+/-3.0个结节（p=0.0039）。AS/MT组的总湿肿瘤重量（350+/-47.4 g）显著大于NS/MT组（248+/-41.5 g）（p=0.0065）。总之，DNMT 1的反义寡核苷酸抑制DNA甲基化影响细胞形态和粘附，以及在体外胃癌细胞的细胞生长。此外，癌细胞特性的这些改变导致在体内裸鼠系统中附着到腹膜上的能力增加，这表明使用这种DNA甲基化抑制剂将需要严格的临床指南，因为它并不总是意味着治疗性抗肿瘤策略。
DNA (cytosine-5-)-methyltransferase 1 (DNMT1) plays an important role in the maintenance of DNA methylation patterns via complicated networks including signaling pathways and transcriptional factors, relating to cell differentiation or carcinogenesis. In the present study, we designed an antisense oligodeoxynucleotide of DNMT1 (AS/MT: 5'-CGGTAC GCGCCGGCATCT-3') and demonstrated successful inhibition of DNMT1 expression by AS/MT at the protein level, using gastric cancer cell lines in vitro. E-cadherin protein expression was increased, and both cyclin D1 and PCNA were decreased by AS/MT treatment. AS/MT also induced suppression of cell growth as determined by BrDU uptake incorporation, in a dose-dependent manner, suggesting specificity of AS/MT. Simultaneously, morphological alterations were observed in both TMK-1 and MKN-45 cells after 24 h incubation with 2 micro M of AS/MT. The cells changed shape from their original forms to dispersed, fibroblast-like cells with neurite-like processes, accompanied by an increased adhesive potential of the cells. An in vivo model of peritoneal dissemination using the nude mouse system showed an increased malignant potential of AS/MT treated TMK-1 cells as demonstrated by a greater number of peritoneal tumor nodules in the AS/MT as compared to the NS/MT treated group, 34.8+/-4.3 vs. 22.4+/-3.0 nodules, respectively (p=0.0039). The total wet tumor weight in the AS/MT group (350+/-47.4 g) was significantly greater than that in the NS/MT group (248+/-41.5 g) (p=0.0065). In conclusion, the inhibition of DNA methylation by DNMT1 by an antisense oligodeoxynucleotide influences cell morphology and adhesion, as well as cell growth in gastric cancer cells in vitro. Moreover, these alterations in the characteristics of cancer cells resulted in an increased ability to attach onto the peritoneum in the nude mouse system in vivo, suggesting that strict clinical guidelines will be necessary to utilize such a DNA methylation inhibitor, since it does not always mean a therapeutic antitumor strategy.