中华医学杂志
中華醫學雜誌
중화의학잡지
National Medical Journal of China
2014年
36期
2816-2821
,共6页
多发性骨髓瘤%甲基化%DLC-1基因%三氧化二砷%U266细胞系
多髮性骨髓瘤%甲基化%DLC-1基因%三氧化二砷%U266細胞繫
다발성골수류%갑기화%DLC-1기인%삼양화이신%U266세포계
Multiple myeloma%Methylation%DLC-1 gene%Arsenic trioxide%U266 cell line
目的:探讨人多发性骨髓瘤(MM)肝癌缺失基因1(DLC-1)启动子区CpG岛甲基化情况,以及三氧化二砷( As2 O3)诱导DLC-1基因的去甲基化作用。方法采用甲基特异性PCR 法( MSP)定性检测2008至2012年来自福建医科大学附属协和医院血液内科的52例MM患者DIC基因甲基化状态,反转录( RT)-PCR检测MM患者和人骨髓瘤U266细胞系DLC-1基因的表达情况;采用重亚硫酸盐测序PCR( BSP)法定量检测As2 O3作用前、后人骨髓瘤细胞株U266其DLC-1基因的甲基化状态;荧光定量 PCR 检测U266细胞用药前、后DLC-1基因、DNA甲基转移酶基因( DNMT1、DNMT3a、DNMT3b) mRNA 的表达变化。结果 MM 患者 DLC-1基因的甲基化比例为71.15%(37/52)。 U266细胞DLC-1基因呈甲基化,DLC-1基因不表达;经0.5、1.0、2.0μmol/L As2 O3作用后72 h U266细胞DLC-1基因甲基化率由95.38%下降至63.07%、30.00%及7.69%;荧光定量PCR检测经0.5、1.0、2.0μmol/L As2 O3作用后72 h U266细胞DLC-1基因mRNA表达与未加药组相比分别为其(1.60±0.09)、(3.66±0.17)、(5.29±0.15)倍,而DNMT1、DNMT3a、DNMT3b基因mRNA表达下调(均P<0.05)。结论 DLC-1基因甲基化在MM患者中较为常见,这可能为MM的诊断和治疗提供借鉴;As2 O3可诱导DLC-1基因去甲基化,使DLC-1基因表达上调,恢复其活性,为MM去甲基化治疗提供新思路。
目的:探討人多髮性骨髓瘤(MM)肝癌缺失基因1(DLC-1)啟動子區CpG島甲基化情況,以及三氧化二砷( As2 O3)誘導DLC-1基因的去甲基化作用。方法採用甲基特異性PCR 法( MSP)定性檢測2008至2012年來自福建醫科大學附屬協和醫院血液內科的52例MM患者DIC基因甲基化狀態,反轉錄( RT)-PCR檢測MM患者和人骨髓瘤U266細胞繫DLC-1基因的錶達情況;採用重亞硫痠鹽測序PCR( BSP)法定量檢測As2 O3作用前、後人骨髓瘤細胞株U266其DLC-1基因的甲基化狀態;熒光定量 PCR 檢測U266細胞用藥前、後DLC-1基因、DNA甲基轉移酶基因( DNMT1、DNMT3a、DNMT3b) mRNA 的錶達變化。結果 MM 患者 DLC-1基因的甲基化比例為71.15%(37/52)。 U266細胞DLC-1基因呈甲基化,DLC-1基因不錶達;經0.5、1.0、2.0μmol/L As2 O3作用後72 h U266細胞DLC-1基因甲基化率由95.38%下降至63.07%、30.00%及7.69%;熒光定量PCR檢測經0.5、1.0、2.0μmol/L As2 O3作用後72 h U266細胞DLC-1基因mRNA錶達與未加藥組相比分彆為其(1.60±0.09)、(3.66±0.17)、(5.29±0.15)倍,而DNMT1、DNMT3a、DNMT3b基因mRNA錶達下調(均P<0.05)。結論 DLC-1基因甲基化在MM患者中較為常見,這可能為MM的診斷和治療提供藉鑒;As2 O3可誘導DLC-1基因去甲基化,使DLC-1基因錶達上調,恢複其活性,為MM去甲基化治療提供新思路。
목적:탐토인다발성골수류(MM)간암결실기인1(DLC-1)계동자구CpG도갑기화정황,이급삼양화이신( As2 O3)유도DLC-1기인적거갑기화작용。방법채용갑기특이성PCR 법( MSP)정성검측2008지2012년래자복건의과대학부속협화의원혈액내과적52례MM환자DIC기인갑기화상태,반전록( RT)-PCR검측MM환자화인골수류U266세포계DLC-1기인적표체정황;채용중아류산염측서PCR( BSP)법정량검측As2 O3작용전、후인골수류세포주U266기DLC-1기인적갑기화상태;형광정량 PCR 검측U266세포용약전、후DLC-1기인、DNA갑기전이매기인( DNMT1、DNMT3a、DNMT3b) mRNA 적표체변화。결과 MM 환자 DLC-1기인적갑기화비례위71.15%(37/52)。 U266세포DLC-1기인정갑기화,DLC-1기인불표체;경0.5、1.0、2.0μmol/L As2 O3작용후72 h U266세포DLC-1기인갑기화솔유95.38%하강지63.07%、30.00%급7.69%;형광정량PCR검측경0.5、1.0、2.0μmol/L As2 O3작용후72 h U266세포DLC-1기인mRNA표체여미가약조상비분별위기(1.60±0.09)、(3.66±0.17)、(5.29±0.15)배,이DNMT1、DNMT3a、DNMT3b기인mRNA표체하조(균P<0.05)。결론 DLC-1기인갑기화재MM환자중교위상견,저가능위MM적진단화치료제공차감;As2 O3가유도DLC-1기인거갑기화,사DLC-1기인표체상조,회복기활성,위MM거갑기화치료제공신사로。
Objective To explore the role of hypemethylation of DLC-1 gene in the pathogenesis of multiple myeloma ( MM ) and examine the effects of arsenic trioxide ( As2 O3 )-induced demethylation of DLC-1 gene in U266 cell line.Methods The methylation status of DLC-1 gene was detected by methylation specific PCR ( MSP ) in MM patients from 2008 to 2012.And the expression of DLC-1 gene mRNA was determined by reverse transcription-polymerase chain reaction ( RT-PCR ).Methylation statuses of DLC-1 gene exposed to As 2 O3 were detected by bisulfite sequencing PCR ( BSP ).And the mRNA expressions of DLC-1 and DNA methyltransferase ( DNMT1, T3a and 3b) were determined by real-time fluorescence quantitative PCR ( RTFQ-PCR).Results Hypermethylation of CpG island of DLC-1 gene was observed in 37/52 ( 71.15%) MM patients.DLC-1 gene was not expressed after methylation.As2 O3 could induce DLC-1 gene demethylation.After 72-houe treatments of 0.5, 1.0 and 2.0 μmol/L As2 O3 , the methylation rate of DLC-1 gene dropped from 95.38% to 63.07%, 30.00% and 7.69%.As compared with the untreated group , the expression of DLC-1 gene mRNA increased to ( 1.60 ±0.09 ) , ( 3.66 ±0.17 ) and (5.29 ±0.15) folds after exposures(all P<0.05).And As2O3 could induce the expression of DNMT1, DNMT3a,DNMT3b gene mRNA(all P<0.05).Conclusions Methylation of DLC-1 gene is essential in the pathogenesis of MM and may provide a new diagnostic technique and drug target for the treatment of MM .And As2 O3 may activate the expression of DLC-1 gene through demethylation.