Shih Yu-Lueng, Shyu Rong-Yuan, Hsieh Chung-Bao, Lai Hung-Cheng, Liu Kuo-Yu, Chu Tang-Yuan, Lin Ya-Wen
Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei, Taiwan.
Cancer. 2006 Aug 1;107(3):579-90. doi: 10.1002/cncr.22023.
The secreted frizzled-related protein 1 gene (SFRP1) encodes a Wnt/beta-catenin signaling antagonist and frequently is inactivated by promoter methylation in many tumors. However, the role of SFRP1 in hepatocellular carcinoma (HCC) is not clear. Therefore, the authors investigated whether methylation of the SFRP1 promoter is common in HCC and whether it may influence SFRP1 expression.
Four HCC cell lines, 54 HCCs, 42 cirrhotic livers, 21 livers with chronic hepatitis, and 15 normal control tissues were analyzed for 1) SFRP1 promoter methylation by using methylation-specific polymerase chain reaction analysis and bisulfite sequencing, 2) SFRP1 messenger RNA expression by using quantitative reverse transcriptase-polymerase chain reaction analysis, and 3) loss of heterozygosity (LOH) by using microsatellite markers flanking the SFRP1 locus. HCC cells were treated with the demethylating agent 5-aza-2'-deoxycytidine to determine whether it could restore SFRP1 expression.
SFRP1 promoter methylation was observed in 75%, 48.2%, 21.4%, 14.3% and 0% in HCC cell lines, primary HCCs, cirrhotic livers, livers with chronic hepatitis, and normal control tissues, respectively. Methylation of the SFRP1 promoter region in HCCs increased significantly compared with control tissues. All samples with SFRP1 methylation showed down-regulation of SFRP1 expression. Demethylation treatment with 5-aza-2'-deoxycytidine in HCC cells restored SFRP1 expression. Moreover, LOH of markers D8S505 and D8S1722 was found in 25% and 27.6% of the informative samples, respectively.
The current results suggested that promoter hypermethylation of SFRP1 is a common event in HCC and plays an important role in the regulation of SFRP1 expression. In addition to methylation-mediated down-regulation of SFRP1, LOH also may play a role.
分泌型卷曲相关蛋白1基因（SFRP1）编码一种Wnt/β-连环蛋白信号拮抗剂，在许多肿瘤中常因启动子甲基化而失活。然而，SFRP1在肝细胞癌（HCC）中的作用尚不清楚。因此，作者研究了SFRP1启动子甲基化在HCC中是否常见，以及它是否可能影响SFRP1的表达。
对4种HCC细胞系、54例HCC、42例肝硬化肝脏、21例慢性肝炎肝脏和15例正常对照组织进行分析，以检测：1）使用甲基化特异性聚合酶链反应分析和亚硫酸氢盐测序检测SFRP1启动子甲基化；2）使用定量逆转录聚合酶链反应分析检测SFRP1信使核糖核酸表达；3）使用位于SFRP1基因座侧翼的微卫星标记检测杂合性缺失（LOH）。用去甲基化剂5-氮杂-2'-脱氧胞苷处理HCC细胞，以确定其是否能恢复SFRP1表达。
在HCC细胞系、原发性HCC、肝硬化肝脏、慢性肝炎肝脏和正常对照组织中，SFRP1启动子甲基化的发生率分别为75%、48.2%、21.4%、14.3%和0%。与对照组织相比，HCC中SFRP1启动子区域的甲基化显著增加。所有SFRP1甲基化的样本均显示SFRP1表达下调。用5-氮杂-2'-脱氧胞苷对HCC细胞进行去甲基化处理可恢复SFRP1表达。此外，在25%和27.6%的信息样本中分别发现标记D8S505和D8S1722的LOH。
目前的结果表明，SFRP1启动子高甲基化是HCC中的常见事件，在SFRP1表达调控中起重要作用。除了甲基化介导的SFRP1下调外，LOH也可能起作用。
