中华血液学杂志
中華血液學雜誌
중화혈액학잡지
Chinese Journal of Hematology
2015年
1期
53-57
,共5页
廖旺%刘晓丹%彭红霞%徐令
廖旺%劉曉丹%彭紅霞%徐令
료왕%류효단%팽홍하%서령
白血病,淋巴样%儿童%微RNAs%抗药性,肿瘤
白血病,淋巴樣%兒童%微RNAs%抗藥性,腫瘤
백혈병,림파양%인동%미RNAs%항약성,종류
Leukemia,lymphocytic%Child%MicroRNAs%Drug resistance,neoplasm
目的 检测microRNA-181a(miR-181a)在急性淋巴细胞白血病(ALL)患儿中的表达水平,并研究其在ALL细胞株CCRF-CEM细胞及耐药株CEM-C1细胞中的功能.方法 采用实时荧光定量PCR方法检测ALL患儿骨髓样本、ALL细胞株CCRF-CEM细胞及其耐药株CEM-C1细胞中miR-181a的表达水平.采用电穿孔转染的方法抑制耐药株CEM-C1细胞并上调非耐药株CCRF-CEM细胞中miR-181a的表达,予不同浓度梯度(终浓度分别为0.01、0.1、1、10、100、1 000 ng/ml)喜树碱处理后,采用CCK-8法观察各浓度梯度喜树碱处理后细胞的存活情况,绘制细胞增殖抑制曲线并计算半数抑制浓度(IC50).结果 初诊-复发组患儿初诊及复发骨髓样本miR-181a相对表达水平(4.84±2.71及6.53±2.20)均高于对照组(1.41±0.53)(P=0.017、0.001),初诊-完全缓解组患儿初诊骨髓样本miR-181a水平(7.58±2.50)较对照组明显升高(P=0.000),而完全缓解后miR-181a水平下降至1.35±0.35,与对照组比较差异无统计学意义(P=0.863).CEM-C1细胞miR-181a相对表达水平(-4.39±0.08)较CCRF-CEM细胞(-2.32±0.03)明显升高(P=0.000).转染miR-181a抑制剂CEM-C1细胞较转染阴性对照组增殖抑制率明显升高(P＜0.05),IC50分别为30.61、2 255.00 ng/ml,耐药指数(RI) =73.67.miR-181a过表达CCRF-CEM细胞较阴性对照组增殖抑制率明显降低(P＜0.05),IC50分别为126.60、1.34 ng/ml,RI=94.26.结论 ALL患儿骨髓及CEM-C1细胞中miR-181a异常高表达,抑制CEM-C1细胞中miR-181a的表达可明显增加CEM-C1细胞的药物敏感性,在CCRF-CEM细胞中上凋miR-181a的表达能明显增加CCRF-CEM细胞的耐药性.
目的 檢測microRNA-181a(miR-181a)在急性淋巴細胞白血病(ALL)患兒中的錶達水平,併研究其在ALL細胞株CCRF-CEM細胞及耐藥株CEM-C1細胞中的功能.方法 採用實時熒光定量PCR方法檢測ALL患兒骨髓樣本、ALL細胞株CCRF-CEM細胞及其耐藥株CEM-C1細胞中miR-181a的錶達水平.採用電穿孔轉染的方法抑製耐藥株CEM-C1細胞併上調非耐藥株CCRF-CEM細胞中miR-181a的錶達,予不同濃度梯度(終濃度分彆為0.01、0.1、1、10、100、1 000 ng/ml)喜樹堿處理後,採用CCK-8法觀察各濃度梯度喜樹堿處理後細胞的存活情況,繪製細胞增殖抑製麯線併計算半數抑製濃度(IC50).結果 初診-複髮組患兒初診及複髮骨髓樣本miR-181a相對錶達水平(4.84±2.71及6.53±2.20)均高于對照組(1.41±0.53)(P=0.017、0.001),初診-完全緩解組患兒初診骨髓樣本miR-181a水平(7.58±2.50)較對照組明顯升高(P=0.000),而完全緩解後miR-181a水平下降至1.35±0.35,與對照組比較差異無統計學意義(P=0.863).CEM-C1細胞miR-181a相對錶達水平(-4.39±0.08)較CCRF-CEM細胞(-2.32±0.03)明顯升高(P=0.000).轉染miR-181a抑製劑CEM-C1細胞較轉染陰性對照組增殖抑製率明顯升高(P＜0.05),IC50分彆為30.61、2 255.00 ng/ml,耐藥指數(RI) =73.67.miR-181a過錶達CCRF-CEM細胞較陰性對照組增殖抑製率明顯降低(P＜0.05),IC50分彆為126.60、1.34 ng/ml,RI=94.26.結論 ALL患兒骨髓及CEM-C1細胞中miR-181a異常高錶達,抑製CEM-C1細胞中miR-181a的錶達可明顯增加CEM-C1細胞的藥物敏感性,在CCRF-CEM細胞中上凋miR-181a的錶達能明顯增加CCRF-CEM細胞的耐藥性.
목적 검측microRNA-181a(miR-181a)재급성림파세포백혈병(ALL)환인중적표체수평,병연구기재ALL세포주CCRF-CEM세포급내약주CEM-C1세포중적공능.방법 채용실시형광정량PCR방법검측ALL환인골수양본、ALL세포주CCRF-CEM세포급기내약주CEM-C1세포중miR-181a적표체수평.채용전천공전염적방법억제내약주CEM-C1세포병상조비내약주CCRF-CEM세포중miR-181a적표체,여불동농도제도(종농도분별위0.01、0.1、1、10、100、1 000 ng/ml)희수감처리후,채용CCK-8법관찰각농도제도희수감처리후세포적존활정황,회제세포증식억제곡선병계산반수억제농도(IC50).결과 초진-복발조환인초진급복발골수양본miR-181a상대표체수평(4.84±2.71급6.53±2.20)균고우대조조(1.41±0.53)(P=0.017、0.001),초진-완전완해조환인초진골수양본miR-181a수평(7.58±2.50)교대조조명현승고(P=0.000),이완전완해후miR-181a수평하강지1.35±0.35,여대조조비교차이무통계학의의(P=0.863).CEM-C1세포miR-181a상대표체수평(-4.39±0.08)교CCRF-CEM세포(-2.32±0.03)명현승고(P=0.000).전염miR-181a억제제CEM-C1세포교전염음성대조조증식억제솔명현승고(P＜0.05),IC50분별위30.61、2 255.00 ng/ml,내약지수(RI) =73.67.miR-181a과표체CCRF-CEM세포교음성대조조증식억제솔명현강저(P＜0.05),IC50분별위126.60、1.34 ng/ml,RI=94.26.결론 ALL환인골수급CEM-C1세포중miR-181a이상고표체,억제CEM-C1세포중miR-181a적표체가명현증가CEM-C1세포적약물민감성,재CCRF-CEM세포중상조miR-181a적표체능명현증가CCRF-CEM세포적내약성.
Objective To investigate the expression of miR-181 a in bone marrow (BM) samples of pediatric acute lymphoblastic leukemia (ALL) and explore the mechanism of miR-181a on ALL cell line CCRF-CEM and drug resistance cell line CEM-C1.Methods BM samples were obtained from 18 patients where matched samples at initial diagnosis and first BM relapse or complete remission were available.BM samples and cord blood samples (normal controls) were used to confirm the differential expression of miRNA-181a by quantitative real-time polymerase chain reaction (qRT-PCR).The expressions of miR-181a in both CCRF-CEM and its mutidrug-resistant counterpart CEM-C1 cells were also detected.Then,CCK-8 assay was performed to quantify the effects of miR-18 l a on CEM-C1 and CCRF-CEM cells growth and viability.Results Up-regulated miR-181a with higher fold changes in both initial diagnosis (4.84±2.71,7.58±2.50) and relapsed samples (6.53±2.20) compared to normal controls (1.41±0.53) (P=0.017,0.000,0.001,respectively) were observed,whereas the miR-181a expression in the samples of CR (1.35±0.35) compared to normal control showed no significant difference (P=0.863).The miR-181a expression level was higher in CEM-C1 cells (-4.39±0.08) than of in CCRF-CEM cells (-2.32±0.03) (P=0.000).CCK-8 assay revealed that suppression of miR-181a in CEM-C1 cells by transfecting the specific inhibitor of miR-18 1 a led to significantly higher cellular proliferation inhibition rate than negative control cells (P＜0.05),IC50 were 30.61 ng/ml and 2 255.00 ng/ml with RI as 73.67.While increased miR-181a in CCRF-CEM cells led to significantly lower CPIR than negative control cells (P＜0.01),IC50 were 126.60 ng/ml and 1.34 ng/ml with RI as 94.26.Conclusion Upregulation of miR-181a might play an important role in the development of drug resistance in CEM-C1 cells,and knockdown of miR-18 1 a could sensitize CEM-C1 cells to camptothecin; Meanwhile increased expression of miR-181a could promote CCRF-CEM drug resistance.These results suggested that suppression of miR-181a expression might provide a promising therapeutic in drug resistance of leukaemia.
