Ma Li, Dong Shangwen, Zhang Peng, Xu Ning, Yan Huiqin, Liu Hongyu, Li Yongwen, Zhou Qinghua
Medical Department of Medical Oncology, Tianjin Medical University General Hospital, Anshan Road No. 154, Heping District, Tianjin 300052, China.
Clin Lab. 2010;56(9-10):407-16.
To study the expression of the Syk (Spleen Tyrosine Kinase) gene and methylation in its promoter region in lung cancer. To investigate the relationship between silencing of the Syk gene and DNA methylation of the Syk promoter region.
RT-PCR (Semi-quantitative reverse transcription PCR), Real-time PCR (Real-time quantitative PCR) and immunohistochemistry technique, the expression of Syk in specimens from 3 lung cancer cell lines and 16 lung cancer patients (tumor tissues and adjacent normal tissues). MSP (Methylation-specific PCR) was used to analyze the methylation status of the Syk promoter region. Then we also investigated the role of restoring Syk expression by using a DNA methyltransferase inhibitor, 5-aza-CdR (5-aza-2'-deoxycytidine), in suppressing invasion of lung cell lines.
No expression of the Syk gene was detected in the 3 lung cancer cell lines. In the 16 lung patient samples, Syk expression was significantly lower in the tumor tissues than in the adjacent normal tissues (P < 0.05). Consistently, immunohistochemistry analyses of Syk protein expression showed that in the cancer tissues Syk protein expression accounted for 5% (1/20), and in the adjacent normal tissues the rate of expression was 100% (20/20). The correlation was highly significant (chi2 = 36.19, P < 0.005). In the only case that showed a positive expression of cancer textus, the level was inferior to the adjacent tissue. In the two lung cancer cell lines (L9981, A549) that lack the endogenous Syk epression, 4uM demethylation agent 5-aza-CdR treatment was able to reactivate the Syk gene expression.
Hypermethylation leads to silencing of the Syk gene in human lung carcinoma. Methylation of the Syk promoter and loss of Syk expression in lung cancer are independent biomarkers, a determination which may offer guidance for selecting appropriate diagnoses and treatments. Syk may be a potential tumor suppressor in human lung cancer.
研究肺癌中Syk（脾酪氨酸激酶）基因的表达及其启动子区域的甲基化情况。探讨Syk基因沉默与Syk启动子区域DNA甲基化之间的关系。
采用RT-PCR（半定量逆转录PCR）、实时荧光定量PCR及免疫组化技术，检测3种肺癌细胞系及16例肺癌患者（肿瘤组织及癌旁正常组织）标本中Syk的表达。采用甲基化特异性PCR（MSP）分析Syk启动子区域的甲基化状态。此外，我们还研究了使用DNA甲基转移酶抑制剂5-氮杂-2'-脱氧胞苷（5-aza-CdR）恢复Syk表达对抑制肺癌细胞系侵袭的作用。
在3种肺癌细胞系中均未检测到Syk基因表达。在16例肺癌患者样本中，肿瘤组织中Syk表达明显低于癌旁正常组织（P < 0.05）。同样，Syk蛋白表达的免疫组化分析显示，癌组织中Syk蛋白表达率为5%（1/20），而癌旁正常组织中表达率为100%（20/20）。两者相关性高度显著（χ2 = 36.19，P < 0.005）。在唯一显示癌组织阳性表达的病例中，其表达水平低于癌旁组织。在两种缺乏内源性Syk表达的肺癌细胞系（L9981、A549）中，4μM去甲基化剂5-aza-CdR处理能够重新激活Syk基因表达。
高甲基化导致人肺癌中Syk基因沉默。Syk启动子甲基化及肺癌中Syk表达缺失是独立的生物标志物，这一判定可为选择合适的诊断和治疗提供指导。Syk可能是人类肺癌中的一种潜在肿瘤抑制因子。
