贲门腺癌组织中TSP1高甲基化与TGF-β1及细胞免疫水平的关系

Correlation of hypermethylation of TSP1 gene with TGF-β1 level and T cell immunity in gastric cardia adenocarcinoma

背景与目的:凝血酶敏感蛋白1(thrombospondin-1,TSP1)是一种内源性的血管生成抑制因子,其在肿瘤中的高甲基化可导致其基因沉默。本研究探讨贲门腺癌中TSP1基因的甲基化状态及其与TGF-β1含量及细胞免疫之间的关系。方法:应用甲基化特异性PCR方法和免疫组织化学法分别检测贲门癌组织及相应癌旁组织的TSP1甲基化状况和TSP1蛋白表达情况,ELISA方法检测贲门癌患者血清中TGF-β1的含量,流式细胞术对贲门癌细胞免疫水平进行分析。结果:贲门癌组织TSP1甲基化率为35.4%(34/96),显著高于癌旁组织(3.1%,P<0.01)。Ⅲ期和Ⅳ期贲门癌患者中TSP1基因甲基化率显著高于Ⅰ期和Ⅱ期患者(P<0.05)。贲门癌组织中TSP1的蛋白表达显著低于癌旁组织(P<0.05),且与其甲基化状态之间有明显的相关性。贲门腺癌患者血清中总TGF-β1的含量高于正常对照组(P<0.05),Ⅲ期和Ⅳ期贲门癌患者血清中总TGF-β1的含量显著高于Ⅰ期和Ⅱ期患者(P<0.05),贲门腺癌患者细胞免疫功能低于正常对照,TSP1发生甲基化的贲门癌患者其细胞免疫功能低于未发生甲基化的贲门癌患者(P<0.05)。结论:TSP1基因启动子区的高甲基化可能参与了贲门腺癌的发生发展过程,其高甲基化有可能影响患者的细胞免疫功能。

Background and Objective： Thrombospondin-1 （TSP1） is an inhibitor of angiogenesis and its promoter hypermethylation has been found resulting in gene silencing in some primary human carcinomas. This study was to investigate the promoter methylation of TSP1 and its correlation with TGF-151 level and T cell immunity in gastric cardia adenocarcinoma （GCA）. Methods.. Methylation specific polymerase chain reaction （MSP） approach and immunohistochemistry method were used to examine the methylation status of the 5＇ CpG island and expression of TSP1 protein, respectively, The level of TGF-β1 was measured by ELISA and T cell immunity of GCA by flow cytometry analysis. Results： TSP1 methylation frequency was significantly higher in tumor specimens than in corresponding normal tissues （35.4% vs. 3.1%, P〈0.001） and significanty higher in Stages III and IV tumor tissues than in Stages Ⅰ and Ⅱ tumor tissues （P〈0.05）. TSP1 protein expression was significantly lower in the tumor tissues than in corresponding normal tissues （P〈0.05） and statistically correlated with its methylation status （P〈0.01）. The total level of TGF-β was significanty higher in the GCA patients than in healthy controls （P〈0.05） and significanty higher in Stages Ⅲ and Ⅳ GCA patients than in Stages Ⅰ and ⅡGCA patients （P〈0.05）. The level of active TGF-β1 was significanty higher in the GCA patients with hypermethyration of TSP1 than in the GCA patients without methylation of TSP1 （P〈0.05）, but there was no statistical difference （P〉0.05）. The function of T cell immunity was significantly different between the GCA patients with hypermethylation of TSP1 and those without methylation of TSP1 （P〈0.05）. Conclusion： Promoter hypermethylation of TSP1 may play an important role in the development of GCA and reflect the biological behaviours of GCA.