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目的通过研究髓母细胞瘤与幕上原始神经外胚叶肿瘤(SPNET)中 RASSF1A 基因的甲基化改变,探讨颅内原始神经外胚叶肿瘤(PNET)的不同亚型中该基因的表遗传学差异及其意义。方法收集25例原发髓母细胞瘤,9例原发 SPNET,3株髓母细胞瘤细胞系和2株 SPNET 细胞系。采用甲基化特异性聚合酶链反应(MSP)检测 RASSF1A 基因启动子区的甲基化状态。应用去甲基化试剂5-aza-2′deoxycytidine 处理存在基因表达缺失的细胞系,探讨基因表达与甲基化之间的关系。结果100%(25/25)的原发髓母细胞瘤、6/9的原发 SPNET 及全部 PNET 细胞系中均检测到RASSFIA 基因的甲基化。相反,该基因甲基化在全部正常组织(包括2例小脑,5例大脑)中均未检测到。并且,RASSF1A 在 SPNET 中的甲基化率明显低于髓母细胞瘤(Fisher 精确检验,P=0.014)。在经去甲基化试剂处理的 PNET 细胞中,该基因表达得以恢复,证明甲基化与该基因沉默相关。结论RASSF1A 甲基化是肿瘤特异性的,RASSF1A 甲基化与 PNET 的发生有一定关联,不同亚型的 PNET 之间 RASSF1A 基因的不同甲基化状态提示髓母细胞瘤和 SPNET 是表遗传学上存在差异的两类肿瘤。
Objective To investigate the methylation of RASSF1A gene in medulloblastoma and supratentorial primitive neuroectodermal tumor (SPNET) and to explore the epigenetic characteristics of this gene in different subtypes of intracranial primitive neuroectodermal tumor (PNET) Differences and Significance of Learning. Methods Twenty-five primary medulloblastomas, nine primary SPNETs, three medulloblastoma cell lines and two SPNET cell lines were collected. The methylation status of RASSF1A gene promoter region was detected by methylation-specific polymerase chain reaction (MSP). The demethylation reagent 5-aza-2’deoxycytidine was used to treat the cell lines with the absence of gene expression and to explore the relationship between gene expression and methylation. Results Methylation of the RASSFIA gene was detected in 100% (25/25) primary medulloblastoma, 6/9 primary SPNET and in all PNET cell lines. In contrast, methylation of this gene was not detected in all normal tissues (including 2 cerebellum and 5 cerebrum). Moreover, the methylation rate of RASSF1A in SPNET was significantly lower than that of medulloblastoma (Fisher’s exact test, P = 0.014). The gene expression was restored in demethylated PNET cells, demonstrating that methylation is associated with the gene silencing. Conclusion The methylation of RASSF1A is tumor-specific. The methylation of RASSF1A is related to the occurrence of PNET. The different methylation status of RASSF1A gene between different subtypes suggests that medulloblastoma and SPNET are epigenetically There are two types of tumors that differ.