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目的研究不同分化程度的神经母细胞瘤(NB)中 E-cadherin、β—catenin 蛋白的表达情况,初步探讨 E-cadherin、β—catenin 异常表达的分子生物学机制及其与临床病理参数的关系。方法免疫组织化学 EnVision 法检测黏附分子 E-cadherin、β—catenin 在90例石蜡包埋组织中的表达;对7例NB 的新鲜样本和24例 NB 石蜡包埋组织采用甲基化特异性 PCR(MSP)方法检测 E-cadherin 基因启动子 CpG 岛甲基化状态;运用 PCR 扩增和测序方法,检测7例 NB 组织中β—catenin 基因第3外显子的突变情况,采用 SPSS 统计分析软件进行相关性分析。结果分化良好组织类型 NB β—catenin 的阳性表达率(47/70,67.1%)显著高于分化不良组织类型(8/20,40.0%);E-cadherin、β—catenin 在 NB 中普遍呈异常表达;有淋巴结转移组中 E-cadherin 和β—catenin 的阳性表达率较无淋巴结转移组显著下降;检测31例 NB E-cadherin 基因启动子的部分区域均为未甲基化状态;对β—catenin 第3外显子 PCR扩增产物进行 DNA 测序,7例 NB 中1例发生基因重排和点突变,2例发生点突变,其中2例在同一位点(27184)发生 T→A的突变。结论 NB 中黏附分子 E-cadherin 高异常表达率与实验中检测的启动子区域甲基化无关,是否与该启动子未检测区域 CpG 岛甲基化有关,还需进一步实验证实;β-catenin的高异常表达率可能与β—catenin 基因第3外显子的突变有关。
Objective To investigate the expression of E-cadherin and β-catenin in neuroblastoma (NB) with different degrees of differentiation and to investigate the molecular mechanisms of E-cadherin and β-catenin abnormalities and their relationship with clinicopathological parameters . Methods EnVision immunohistochemistry was used to detect the expression of E-cadherin and β-catenin in 90 paraffin-embedded tissues. Seven samples of NB and 24 specimens of NB paraffin-embedded tissue were detected by methylation-specific PCR MSP) was used to detect methylation status of CpG island of E-cadherin gene promoter. The mutation of exon 3 of β-catenin gene in 7 cases of NB was detected by PCR amplification and sequencing. SPSS statistical analysis software Correlation analysis. Results The positive expression rates of NB β-catenin in well-differentiated tissues (47/70, 67.1%) were significantly higher than those in poorly differentiated tissues (8/20, 40.0%). E-cadherin and β-catenin were generally abnormal in NB The positive expression rate of E-cadherin and β-catenin in lymph node metastasis group was significantly lower than that in non-lymph node metastasis group. Some of the 31 cases of NB E-cadherin gene promoter were unmethylated, catenin 3 exon PCR amplification products DNA sequencing, 1 case of 7 cases of NB gene rearrangement and point mutation occurred, 2 cases of point mutation occurred, of which 2 cases in the same locus (27184) T → A mutation . Conclusion The high abnormal expression rate of E-cadherin in NB is not related with the promoter methylation detected in the experiment, but whether it is related to the CpG island methylation in the non-detected region of the promoter. Further experiments confirm that the expression of β-catenin The high abnormal expression rate may be related to the mutation of exon 3 of β-catenin gene.