HOX11＋急性T淋巴细胞白血病中MEIS1基因启动子区甲基化状态及其表达水平

Hypermethylation of CpG island in promoter region of MEIS1 gene and its expression in HOX11＋ acute T lymphoblastic leukemia

目的检测骨髓嗜病毒整合位点1（MEIS1）基因在同源异形盒11（HOX11）＋急性T淋巴细胞白血病（T—ALL）细胞株和T—ALL患者骨髓细胞中的甲基化状态和表达水平，探讨其表达水平与其启动子甲基化的关系。方法收集2009年1月至2011年12月北京大学人民医院北京大学血液病研究所38例T—ALL患者治疗前及其中29例治疗后完全缓解的T—ALL患者和8名健康供者的骨髓样本，及3株T—ALL细胞株（si1—ALL、DND41和RPMI）；采用反转录（RT）PCR检测MEIS1、HOX11mRNA表达水平，采用重硫酸盐测序法、甲基化DNA免疫沉淀技术和启动子寡核苷酸微阵列芯片分析MEIS1基因启动子区CpG岛甲基化状态和基因功能。结果根据HOX11表达的高低，将38例T—ALL患者分为HOX11高表达（HOX11＋）组（n=14）和HOX11低表达（HOX11-）组（n=24）。HOX11＋组患者骨髓中出现MEIS1基因启动子区域的甲基化。HOX11＋组患者MEIS1基因启动子区域甲基化比率明显高于HOX11一组患者（76．8％比29．5％，P〈0．01）。MEIS1mRNA在治疗后完全缓解患者和健康供者骨髓中的水平低于患者治疗前骨髓中的水平（均P〈0．05）；MEISImRNA在HOX11＋T—ALL细胞株（si1—ALL、DNIM1）中的表达明显低于HOX11-T—ALL细胞株（RPMI）（0．392±0．226、0．378±0．317比1．059±0．533，均P〈0．05），在HOX11＋组患者中的表达明显低于HOX11-组患者（0．462±0．281比0．916±0．437，P〈0．01）；且MEIS1mRNA的相对表达程度与HOX11mRNA的相对表达程度呈负相关（rs=-0．416，P〈0．01）。MEIS1基因在T．ALL中的表达水平与其甲基化呈负相关（rs=-0．383，P〈0．01）。结论MEIS1基因启动子甲基化是导致其在T．ALL中表达下调或基因沉默的原因，HOX11可能负调控MEIS1基因的表达。

Objective To detect the status of methylation in promoter region of MEIS1 gene and its expression in cell lines and patients with HOX11＋ T-cell acute lymphoblastic leukemia （T-ALL） and explore the relationship between the expression level of MEIS1 gene and methylation of CpG island in promoter region. Methods The methylation pattern in MEIS1 gene was detected with bisulfite sequencing PCR, DNA methylation immunoprecipitation and promoter oligonucleotide microarray. And the expression level of MEIS1 mRNA was detected with reverse transcription PCR in 3 T-ALL cell lines： sil-ALL, DNIM1 and RPMI as well as in 75 clinical bone marrow samples including 38 de novo T-ALL patients, 29 complete remission T-ALL patients and 8 normal samples from January 2009 to December 2011. Results The expressions of HOX11 mRNA in the patients were classified into high expression （HOX11 ~ ）groups （n = 14） and low expression （HOX11 - ）groups （n = 24 ）. There was hypermethylafion of CpG island in promoter region of MEIS1 gene in patients with HOX11＋ T-ALL. The methylation rate of promoter CpG islands of MEIS1 gene in HOX11＋ T-ALL （76. 8% ） was significantly higher than that of HOXll - T-ALL （29. 5% ） （P 〈 0. 01 ）. The expression MEIS1 in complete remission T-ALL patients and normal samples was significantly lower than that in de novo T-ALL patients （ both P 〈 0. 05 ）. The expression MEIS1 in HOX11 ＋ cell lines was significantly lower than that in HOXll - cell lines （0. 392 ±0. 226,0. 378 ±0. 317 vs 1. 059 ±0. 533, both P 〈 0.05） and the expression level of MEIS1 in HOX11 ＋ T-ALL patients （0.462 ± 0.281 ） significantly lower than that in HOX11 - T-ALL patients （0. 916± 0. 437 ） （ P 〈 0. 01 ）. The relationship between MEIS1 and HOXll mRNA expression level had a negative correlation（ rs= -0. 416, P 〈 0. 01 ）. The expression level of MEIS1 gene was negatively correlated with the methylation of CpG island in promoter region in T-ALL （ rs = - 0. 383, P 〈 0.01 ）. Conclusions The expression of MEIS1 becomes down- regulated or even lost by promoter methylation in T-ALL. HOXll maybe a negative regulator of MEIS1 gene.