Comprehensive Epigenetic Analysis Using Oral Rinse Samples: A Pilot Study
Comprehensive Epigenetic Analysis Using Oral Rinse Samples: A Pilot Study
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DOI：
10.1016/j.joms.2011.04.021
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发表时间：
2012-06-01
影响因子：
1.9
通讯作者：
Sugihara, Kazumasa
中科院分区：
文献类型：
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作者：
Kusumoto, Takanobu;Hamada, Tomofumi;Sugihara, Kazumasa
目的：为了证明，染色质免疫沉淀法可以进行与口腔冲洗样品和开发一个协议，用于全面分析DNA甲基化，组蛋白修饰和基因表达之间的功能相互作用，使用这样的samples.Materials和方法：11个癌细胞系和口腔冲洗样品10例口腔鳞状细胞癌和3名健康受试者进行了检查。采用逆转录聚合酶链反应和免疫组化方法检测肿瘤抑制基因CDKN 2A的表达。通过甲基化特异性聚合酶链反应评估启动子DNA甲基化。染色质免疫沉淀法检测组蛋白H3赖氨酸9的二甲基化和乙酰化修饰。结果：CDK 2NA在11个肿瘤细胞系中均存在表观遗传调控。使用本方案，可以成功地对口腔冲洗液样品进行全面的表观遗传分析。所有患者均舒适地使用规定量（16 mL）的生理盐水漱口。9名患者（90%）和1名健康受试者（33%）显示组蛋白H3的赖氨酸9二甲基化。此外，8例患者（80%）表现出hypoacetylation的组蛋白H3，这是没有观察到在健康subjects.Conclusions：本研究表明，第一次染色质修饰可以使用口腔冲洗样品染色质免疫沉淀分析。为了评价组蛋白修饰在口腔鳞状细胞癌发生中的作用，未来应进行包括更多受试者的研究。(C)2012年美国口腔颌面外科医师协会J Oral Maxillofac Surg 70：1486-1494，2012
Purpose: To prove that chromatin immunoprecipitation assay can be performed with oral rinse samples and to develop a protocol for comprehensive analysis of functional interactions among DNA methylation, histone modification, and gene expression using such samples.Materials and Methods: Eleven cancer cell lines and oral rinse samples from 10 patients with oral squamous cell carcinoma and 3 healthy subjects were examined. The expression of CDKN2A, a tumor suppressor gene, was determined by reverse transcription/polymerase chain reaction and immunohistochemistry. Promoter DNA methylation was assessed by methylation-specific polymerase chain reaction. Chromatin modifications were analyzed by a chromatin immunoprecipitation assay using antibodies for dimethylation and acetylation of lysine 9 of histone H3.Results: Epigenetic control of CDK2NA was observed in vitro in 11 cancer cell lines. Using the present protocol, comprehensive epigenetic analysis could be successfully performed with oral rinse samples. All patients were comfortable using the prescribed amount (16 mL) of normal saline to rinse their mouths. Nine patients (90%) and 1 healthy subject (33%) showed dimethylation of lysine 9 of histone H3. Moreover, 8 patients (80%) showed hypoacetylation of lysine 9 of histone H3, which was not observed in healthy subjects.Conclusions: The present study showed for the first time that chromatin modifications can be analyzed using oral rinse samples by chromatin immunoprecipitation analysis. To evaluate the contribution of histone modifications for carcinogenesis of oral squamous cell carcinoma, studies including a larger number of subjects should be conducted in the future. (C) 2012 American Association of Oral and Maxillofacial Surgeons J Oral Maxillofac Surg 70: 1486-1494, 2012