Lipopolysaccharide extracted from Porphyromonas gingivalis induces DNA hypermethylation of runt-related transcription factor 2 in human periodontal fibroblasts.
Lipopolysaccharide extracted from Porphyromonas gingivalis induces DNA hypermethylation of runt-related transcription factor 2 in human periodontal fibroblasts.
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DOI：
10.1016/j.jmii.2012.08.005
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发表时间：
2014-06
期刊：
影响因子：
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通讯作者：
O. Uehara;Y. Abiko;M. Saitoh;H. Miyakawa;F. Nakazawa
中科院分区：
文献类型：
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作者：
O. Uehara;Y. Abiko;M. Saitoh;H. Miyakawa;F. Nakazawa
背景和目的表观遗传学改变，如DNA甲基化和组蛋白乙酰化，是指不涉及DNA序列的基因表达模式的改变。牙龈卟啉单胞菌（Porphyromonas gingivalis）的脂多糖（LPS）可抑制成骨细胞的分化。我们研究了DNA超甲基化是否参与了LPS对人牙周膜成纤维细胞（HPDL）成骨分化的抑制作用。浓度为10 μg/ml，作用24 h。用DNA甲基转移酶抑制剂5-Aza-2 ′-脱氧胞苷（5Aza）处理细胞。未处理的细胞用作对照。使用细胞增殖试剂测定细胞活力。通过定量聚合酶链反应（RT-PCR）评估DNA甲基转移酶（DNMT 1）和runt相关转录因子2（RUNX 2）mRNA。采用定量甲基化特异性PCR分析RUNX 2 DNA甲基化。结果LPS刺激的细胞中RUNX 2的表达水平明显低于对照组。5Aza的存在增加了LPS刺激的细胞中RUNX 2的表达。LPS刺激后DNMT 1 mRNA表达水平明显高于对照组。5Aza的存在完全消除了LPS刺激的细胞中DNMT 1的上调表达。LPS刺激HPDL后，细胞内0.1 kb和-1.9 kb DNA甲基化水平显著高于对照组。结论LPS对HPDL成骨细胞分化的抑制作用可能与DNA甲基化有关。
Background/PurposeEpigenetic alterations such as DNA methylation and histone acetylation are described as changes in the pattern of gene expression not involving the DNA sequence. Lipopolysaccharide (LPS) derived fromPorphyromonas gingivalishas been shown to inhibit osteoblastic cell differentiation. We examined whether DNA hypermethylation was involved in the inhibitory effect of LPS on osteoblastic differentiation of fibroblasts derived from human periodontal ligament (HPDL).MethodsThe HPDL cells were incubated with LPS derived fromP. gingivalisat a concentration of 10 μg/ml for 24 h. The cells were treated with DNA methyltransferase inhibitor 5-Aza-2'-deoxycytidine (5Aza). Untreated cells were used as a control. Cell viability was determined using cell proliferation reagent. DNA methyltransferase (DNMT1) and runt-related transcription factor 2 (RUNX2) mRNAs were evaluated by quantitative polymerase chain reaction (RT-PCR). Analysis of RUNX2 DNA methylation was performed using quantitative methylation-specific PCR.ResultsThe expression level of RUNX2 was significantly lower in the cells stimulated with LPS than the controls. The presence of 5Aza increased the expression of RUNX2 in cells stimulated with LPS. The expression levels of DNMT1 mRNA in the cells stimulated with LPS were significantly higher than in the control. The presence of 5Aza completely abolished the upregulated expression of DNMT1 in cells stimulated with LPS. The methylation of DNA at 0.1 kb and −1.9 kb in the cells stimulated with LPS was significantly higher than the control.ConclusionThe results indicate that DNA hypermethylation may be involved in the inhibitory effect of LPS on osteoblastic differentiation in HPDL.