口腔鳞癌组织和口腔拭子中Dickkopf-1基因甲基化状态与疾病进展的相关性研究

Relationship between Dickkopf-1 gene methylation status in oral squamous cell carcinoma and oral swabs and disease progression

目的研究口腔鳞状细胞癌(OSCC)组织和口腔拭子(OS)中Dickkopf-1(DKK-1)基因甲基化状态与疾病进展的关系及其临床意义。方法选取诊断为OSCC的67例患者作为OSCC组,收集病变同侧和对侧OS标本及手术组织标本。另选择30例阻生智齿患者的健康口腔黏膜组织作为健康对照组,30例口腔黏膜下纤维病变(OSF)患者作为癌前病变组。采用免疫组化法检测组织DKK-1蛋白表达;采用重亚硫酸盐转化和定量甲基化特异性聚合酶链反应(qMSP)法检测DKK-1基因甲基化水平。结果癌前病变组(66.67%,20/30)和OSCC组(20.90%,14/67)DKK-1蛋白阳性表达率低于健康对照组(93.33%,28/30),且OSCC组低于癌前病变组,差异有统计学意义(P<0.001)。OSCC组肿瘤组织和OS标本中DKK-1基因甲基化水平高于癌前病变组和健康对照组,差异有统计学意义(P<0.001)。Spearman相关系数分析显示,组织标本与OS病变同侧标本中DKK-1基因甲基化水平呈正相关性(r_(s)=0.672,P<0.001)。组织和OS病变同侧标本中DKK-1基因甲基化水平用于鉴别诊断OSCC和癌前病变的曲线下面积(AUC)分别为0.799(95%CI:0.713~0.884)、0.843(95%CI:0.762~0.924)。OS样本中DKK-1甲基化在早期(Z=2.354,P=0.021)和高分化肿瘤患者中更显著(Z=3.731,P<0.001)。结论从病变部位采集的OS标本中DKK-1基因高甲基化可用于OSF癌变监测以及OSCC的早期诊断。

Objective To investigate the relationship between Dickkopf-1(DKK-1)gene methylation status in oral squamous cell carcinoma(OSCC),oral swabs and disease progression as well as its clinical significance.Methods A total of 67 OSCC patients were selected as OSCC group,whose ipsilateral and contralateral OS specimens and surgical tissue specimens were collected.The healthy oral mucosa of 30 patients with impacted wisdom teeth and 30 patients with oral submucosal fibrosis(OSF)were selected as control group and pre-cancerous group.Immunohistochemical staining method was used to detect DKK-1 protein expression,while the bisulphite-converting method and quantitative methylation-specific polymerase chain reaction(qMSP)were used to detect the methylation level of DKK-1 gene.Results The positive expression rates of DKK-1 protein in the pre-cancerous group(66.67%,20/30)and OSCC group(20.90%,14/67)were significantly lower than that in control group(93.33%,28/30),and the OSCC group was significantly lower than the pre-cancerous group(P<0.001).The methylation levels of DKK-1 gene in tumor tissues and OS samples of the OSCC group were significantly higher than those in pre-cancerous group and control group(P<0.001).Spearman correlation coefficient analysis showed that the methylation level of DKK-1 gene in the ipsilateral OS speciments was positively correlated with that of DKK-1 in the tissues(r_(s)=0.672,P<0.001).The area under the curve(AUC)of DKK-1 gene methylation level in tissue and ipsilateral OS specimens for differential diagnosis of OSCC and precancerous lesions was 0.799(95%CI,0.713 to 0.884)and 0.843(95%CI,0.762 to 0.924),respectively.The methylation of DKK-1 in OS samples was more pronounced in early stage(Z=2.354,P=0.021)and in patients with well-differentiated tumors(Z=3.731,P<0.001).Conclusion Hypermethylation of DKK-1 gene in OS specimens collected from the lesion sites can be used for monitoring OSF carcinogenesis and early diagnosis of OSCC.