三氧化二砷和肿瘤坏死因子相关凋亡诱导配体诱导的髓系恶性细胞系P15^(ink4b)表达

P15^(ink4b) expression in malignant myeloid cell lines after treatment of As_2O_3 and /or TRAIL

目的:观察骨髓增生异常综合征(MDS)髓系原始细胞系MDS-L及髓系白血病细胞系ML1经不同剂量和不同时间的三氧化二砷(As2O3)和肿瘤坏死因子相关凋亡诱导配体(TRAIL)处理后的抑癌基因P15ink4b变化。并研究DNA甲基化转移酶DNMT1在P15ink4b变化中的可能作用。方法:体外培养的MDS-L和ML1细胞经9种不同浓度的药物处理(As2O31 mmol/L;2 mmol/L;5 mmol/L;TRAIL 100μg/L;300μg/L;500μg/L;As2O31 mmol/L加Trail 100μg/L;As2O32 mmol/L加TRAIL 300μg/L;As2O35 mmol/L加TRAIL 500μg/L),在24 h、48 h和72 h后收获细胞。未经药物处理的细胞和药物处理后收获的细胞均提取总RNA,经半定量RT-PCR检测P15ink4bmRNA表达。对MDS-L细胞还同时检测DNMT1表达;正常人和5例MDS病例的P15ink4b和DNMT1检测作为对照。结果:未经处理的MDS-L和ML1细胞基本不表达P15ink4b,药物处理后P15ink4b表达增强;药物诱导MDS-L细胞表达P15ink4b的作用强于ML1细胞;未经处理的MDS-L和ML1细胞高表达DNMT1,药物处理24 h后DNMT1不同程度下降,但DNMT1表达状况与P15ink4b表达增强不显示相关性。结论:As2O3和(或)TRAIL处理能促进髓系恶性细胞抑癌基因P15ink4b表达,但并非主要通过抑制DNMT1功能而起作用。

Objective： To observe the P15^ink4b Expression and possible role of DNMT1 in it in MDS-L as well as ML1 cell lines after treatment of As2O3/TRAIL（TNF a related apoptosis inducing ligand）. Method： Cultured cells from MDS-L and ML1 cell lines were treated with 9 different concentration of drug （As2O3 1 mmol/L; 2 mmol/L; 5 mmol/L; TRAIl. 100 ug/L; 300 ug/L; 500 ug/Lt As203 plus Trail as 1 mmol＋100 ug/L; 2 mmol＋300 ug/ Lt 5 mmol/500 ug/L）, and then these treated cells were detected at 3 time points of 24 hours, 48 hours and 72 hours to observe P15ink4b and DNMT1 expression at mRNA level by semi-quantitive RT PCR. Untreated cells coming from these two cell line and cells from bone marrows of normal donors as well as 5 MDS patients were detected as controls. Result.. Untreated MDS-L and ML1 cells almost did not expressed P15^ink4b but showed enhanced expression after treatment of As2O3 or TRAIL. These effects appeared more obvious in MDS-L compared to ML1 cell line. On the other hand, untreated MDS-L and ML1 cells expressed higher DMNT1 than normal controls, and appeared a downregulation after 24 hours of treatment but with no evident corelation to P15^ink4b expression. Conclusion：As2O3 and TRAIL could seperately promote 15^ink4b expression in malignant myeloid cell lines but not mainly by DMNT1 pathway.