急性白血病p53基因P1启动子区域DNA甲基化研究

Aberrant DNA methylation in promoter region of p53 gene in acute leukemia

目的:通过检测急性白血病(AL)中p53基因P1启动子异常甲基化,探讨p53基因异常甲基化在急性白血病中的意义。方法:分别使用限制性内切酶MspⅠ、HpaⅡ、EcoRⅡ、BstNⅠ酶切提取基因组DNA,然后分别使用酶切后产物及基因组DNA为模板进行PCR扩增。产物电泳后在凝胶成像分析系统内观测电泳条带及摄像;部分标本的电泳条带经凝胶回收纯化后进行测序。结果:急性白血病患者p53基因第一启动子甲基化阳性率为38.7%,而急性淋巴细胞白血病(ALL)与急性非淋巴细胞白血病(ANLL)患者分别为45.5%、35.0%。正常对照标本中未检测到p53基因的异常甲基化。p53基因甲基化情况在急性白血病病人与正常人之间经过统计学检验,P<0.05;但急性淋巴细胞白血病与急性非淋巴细胞白血病患者之间无显著差异,P>0.05。分析急性白血病患者p53基因甲基化与患者临床资料之间的关系,其中,p53基因异常甲基化与肝脾淋巴结是否肿大之间的关系经统计学分析P<0.05。结论:①部分急性白血病患者存在p53基因第一启动子异常甲基化,正常对照中未检测到p53基因的甲基化;②p53基因第一启动子在急性淋巴细胞白血病与急性非淋巴细胞白血病均可发生异常甲基化,两者之间发生甲基化的概率无统计学差异;③p53基因异常甲基化与肝脾淋巴结肿大有显著差异,但p53基因异常甲基化与急性白血病治疗效果及预后的关系尚不能确定,须进一步研究确定。

AIM： To investigate the significance of aberrant p53 gene promoter methylation in acute leukemia by detecting the occurrence of p53 gene promoter methylation. METHODS： Genomic DNA was digested using restriction endonuclease MspⅠ , HpaⅡ , EcoR Ⅱ, BstN Ⅰ , respectively. PCR amplification was conducted and the products after digestion and genomic DNA were used as template. The PCR product was subjected to electrophoresis and the results were analyzed by gel imaging and analysis system. Parts of the separated DNA were sequenced after purification from gel. RESULTS： The prevalence of methylation in acute leukemia group was 38. 7% , of which ALL was 45.5% （5 of 11 ） and ANLL 35.0% （7 of 20）. No methylation was detected in normal control group. There was significant difference between the prevalence of methylation in acute leukemia group and the normal control group （ Fisher＇s exact test, P 〈 0. 05 ）. However, the prevalence between ALL and ANLL was not significantly different （ Fisher＇s exact test, P 〉 0. 05 ）. Compared the relationship between aberrant methylation of p53 gene and clinical data, statistical significance between aberrant methylation of p53 gene and enlargement of lymph nodes, liver or spleen（ P 〈 0. 05 ） was observed. CONCLUSION ： ①Aberrant DNA methylation in P1 promoter region of p53 gene exits in part of acute leukemic patients, but not in health people. ②The prevalence of aberrant DNA methylation between ALL and ANLL is not significantly different. ③The patients with aberrant methylation of p53 gene seem to show more frequently the manifestations of enlarged lymph nodes, liver or spleen than usual.