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目的研究苯是否通过引起DNA甲基化使抑癌基因p16转录失活。方法应用病例-对照研究对11名苯中毒患者和8名与苯中毒患者年龄(±5岁)、性别匹配,并且同工种同工龄的接苯但无苯中毒的对照进行检测。分别用实时荧光定量PCR及焦磷酸测序检测基因表达及甲基化水平。结果 p16的表达水平在苯中毒组(0.53)低于对照组(2.06)(P=0.064)。p16启动子区CpG位点甲基化平均水平苯中毒组(12.4%)高于对照组(11.3%)(P>0.05)。甲基化与基因表达水平相关分析发现,p16 CpG位点平均甲基化水平与基因表达水平负相关(pearson相关系数r=-0.64,P>0.05),其中p16启动子区第4个CpG位点位于嗅神经元转录因子共有结合序列内,其甲基化水平与基因表达水平显著负相关(pearson相关系数r=-0.88,P<0.05)。结论在苯中毒患者中p16的表达水平显著降低,基因启动子区CpG岛的高甲基化可能与该基因的低表达有关。需要进一步扩大样本量深入研究苯相关疾病抑癌基因表达异常的分子机制。
Objective To investigate whether benzene can inactivate the tumor suppressor gene p16 by causing DNA methylation. Methods A case-control study of 11 benzene poisoning patients and 8 patients with benzene poisoning age (± 5 years), gender-matched, and the same type of work with the same length of benzene-free benzene-free controls were detected. Real-time fluorescence quantitative PCR and pyrosequencing were used to detect the gene expression and methylation level. Results The expression level of p16 in benzene poisoning group (0.53) was lower than that in control group (2.06) (P = 0.064). The average level of methylation in p16 promoter CpG sites was higher in benzene poisoning group (12.4%) than that in control group (11.3%) (P> 0.05). Correlation analysis between methylation and gene expression level showed that the average methylation level of p16 CpG was negatively correlated with the gene expression (r = -0.64, P> 0.05), and the fourth CpG There was a significant negative correlation between the methylation level and the gene expression level (pearson correlation coefficient r = -0.88, P <0.05) in the co-binding sequence of olfactory neuronal transcription factor. Conclusions The expression level of p16 in benzene poisoning patients is significantly decreased. Hypermethylation of CpG island in gene promoter may be related to the low expression of this gene. Need to further expand the sample size in-depth study of benzene-related diseases, tumor suppressor gene expression abnormalities in molecular mechanisms.