慢性髓细胞性白血病患者与健康人骨髓单个核细胞基因表达的差异

慢性髓细胞性白血病患者与健康人骨髓单个核细胞基因表达的
差异
周珏宇;马文丽;丁大鹏;石嵘;郑文岭
【期刊名称】《中国组织工程研究》
【年(卷),期】2006(010)009
【摘要】背景:慢性髓细胞性白血病是以造血干细胞的恶性克隆性增殖为特征.慢性期的慢性髓细胞性白血病患者如果不予以有效的治疗,必然演进为急变期,其预后往往非常差.因而,从全基因组水平上阐明慢性髓细胞性白血病的发生机制显得尤为重要,目的:应用Applied Biosystems表达芯片系统对慢性髓细胞性白血病患者骨髓单个核细胞的基因表达谱进行观察.设计:观察对比分析.单位:南方医科大学基因工程研究所.对象:两例骨髓样品(1例慢性髓细胞性白血病患者,1例健康者)来自于广州军区广州总医院血液科. 方法:实验于2004-10/2005-09在南方医科大学基因工程研究所完成.分别从一例慢性髓细胞性白血病患者与一例健康人的骨髓样品中分离单个核细胞,提取总RNA,纯化mRNA.通过逆转录体外线性扩增的方法对mRNA 样品进行标记,将标记好的cRNA样品与芯片杂交.利用ABI 1700化学发光芯片分析仪对骨髓单个核细胞的基因表达谱差异情况进行分析.通过比较同一样品不同芯片间的数据信息,对芯片结果的可重复性进行评估.主要观察指标:①总RNA及标记后的cRNA质量的评定.②芯片可重复性验证.③芯片杂交结果.④半定量反转录-聚合酶链反应结果.结果:①利用统计学数据分析工具,通过比较慢性髓细胞性白血病患者与健康人单个核细胞的基因表达差异,总共发现了6 706个差异基因.其中,与慢性髓细胞性白血病密切相关的差异基因为68个,上调的有17个,下调的有51个.②位于C/EBPalpha信号通路和CD40受体信号通路中的大部分差异表达基因,其表达
水平降低.③通过对重复实验结果的相关性及检测一致性分析,证实了芯片结果的可重复性较好.而两组重复实验间的相关系数分别是,慢性髓细胞性白血病组为0.991,健康组为0.988.④半定量反转录-聚合酶链反应验证了芯片分析结果的可靠性.结论:通过比较慢性髓细胞性白血病患者与健康人单个核细胞的基因表达谱的差异,发现了大量的差异表达基因.这些数据将为寻找可用于慢性髓细胞性白血病患者疾病治疗的分子靶标提供有用的信息.%BACKGROUND: Chronic myeloid leukemia (CML) is characterized by the clonal expansion of hematopoietic stem cells. Without effective treat ment, individuals in the indolent, chronic phase (CP) of CML will undergo blast crisis (BC), the prognosis for which is poor. Therefore, it is important to clarify the mechanism underlying CML from a whole-genome perspec tive. OBJECTIVE: To investigate the gene expression profile of bone marrow mononuclear cells from CML with Applied Biosystems Expression Array System.DESIGN: Observation and controlled analysis.SETTING: Institute of Gene Engineering, Southern Medical University PARTICIPANTS: Samples of two cases of bone marrow (a chronic myeloid leukemia patient and a healthy person).METHODS: This experiment was conducted at the Institute of Gene Engineering, Southern Medical University from October 2004 to September 2005.The total RNAs were extracted and purified from bone marrow mononuclear cells derived from a CML patient and a healthy person. mRNAs were purified using an oligo (dT)-cellulose mRNA purification kits and labeled using reverse transcription, in vitro transcription (RT-IVT), then hybridized with microarray. Gene expression differentiation of the bone marrow mononuclear cells were examined by ABI 1700 Chemiluminescent
Microarray Analyzer. Reproducibility of microarray results was assessed by comparing data sets obtained from the same sample and analyzed by two different arrays.MAIN OUTCOME MEASURES: ①Assessment of quality of total RNA and labled cRNA. ②Reproducibility of microarray. ③ Hybridization of array.④Results of semi-quantitative reverse transcription-polymerase chain reaction RESULTS: ①Using statistical data analysis tools, we identified 6 706 genes that were up- or down-regulated in CML patient compared with the healthy person. In these genes, we found that 17 genes were up-regulated while 51 genes were down-regulated among 68 genes closely related to CML. ②most differentially expressed genes in
C/EBPalpha mediated path way and CD40L signaling pathway had reduced expression. ③Go od repro ducibility of microarray was confirmed by analysis of correlation and detection concordance in technical replicates. The correlation coefficient of the detectable probe in technical replicates was 0.991 for the CML patient and 0.988 for the health y person. ④The results of semi-quantitative RT-PCR experiments supported the reliability of our microarray analysis.CONCLUSION: By comparing expression patterns of CML with those of the healthy person, we identified a large number of genes that, were up- or down-regulated in CML patients. These data should provide useful information for finding candidate genes whose products might serve as molecular targets for treatment of CML patients.【总页数】4页(P179-182)
【作者】周珏宇;马文丽;丁大鹏;石嵘;郑文岭
【作者单位】南方医科大学基因工程研究所,广东省广州市,510515;南方医科大学基因工程研究所,广东省广州市,510515;南方医科大学基因工程研究所,广东省广州市,510515;南方医科大学基因工程研究所,广东省广州市,510515;华南基因组研究中心,广东省广州市,510860
【正文语种】中文
【中图分类】R733.7
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