急性髓系白血病mRNA与非编码RNA差异表达谱及CeRNA调控网络分析

利用GEO数据库(gene expression omnibus database)通过生物信息学分析方法探讨急性髓系白血病(acute myelogenous leukemia,AML)的发病机制。检索GEO数据库中AML相关芯片数据集GSE142698、GSE142699和GSE96535。利用GEO2R分析得到差异mRNAs、miRNAs以及差异lncRNAs。利用在线生物信息学分析工具DAVID对差异mRNAs进行GO富集分析和KEGG通路分析。利用miRWalk数据库预测AML相关miRNAs的靶向mRNAs,利用Spongescan数据库预测AML相关miRNAs的靶向lncRNAs,构建lncRNA-miRNA-mRNA竞争性内源RNA （competing endogenous RNA,ceRNA）调控网络。共筛选出29个显著差异mRNAs、70个显著差异miRNAs和20 005个显著差异lncRNAs。GO富集分析和KEGG通路分析显示,差异表达基因主要涉及蛋白磷酸化、细胞分裂、细胞增殖的负调控、基因表达的正向调节、周期蛋白依赖的丝氨酸/苏氨酸激酶活性的调节等生物过程以及细胞周期、细胞衰老、癌症通路、PI3K-Akt通路等信号通路。将miRWalk数据库预测的靶向mRNAs与差异mRNAs取交集,Spongescan数据库预测的靶向lncRNAs与差异lncRNAs取交集,分别确定了25个mRNAs、6个lncRNAs参与AML相关ceRNA调控网络的构建。结果表明,lncRNAs可能作为关键的ceRNA,通过调控miRNA和相关靶基因参与AML的发生与发展,研究结果为AML诊断和治疗的分子生物学研究提供了新的依据。     

急性髓系白血病免疫表型特征及遗传学特征分析

目的:研究急性髓系白血病免疫表型特征以及遗传学特征。方法:选取2011年1月到2014年5月我院收治的急性髓系白血病患者169例,采用流式细胞术和相关的单克隆抗体来分析所有患者的骨髓免疫表型,采用染G染色体显带技术分析患者的核型,根据淋系抗原(lym Ag)的表达将患者分为lym Ag+组和lym Ag-组。结果:抗原CD13、CD33、CD117以及MPO等髓系抗原最常在急性髓系白血病患者中表达,其中CD117在M3型病例中表达为85.7%(24/28),而CD34、HLA-DR双阴性、较强的自发荧光、CD13、CD33和MPO对M3型的诊断也具有一定的价值;其中47.9%(81/169)的患者伴随着淋系抗原表达,以CD7和CD56为主;60.4%(102/169)的患者伴随着核型异常;而伴随着t(8:21)的M2患者中的CD15、CD19和CD56的表达显著增强,而t(15:17)均发生于M3型患者中;而lym Ag+组患者CD34的阳性患者为77.8%(63/81)显著高于lym Ag-组的47.7%(42/88),两组比较差异具有统计学意义(P0.05)。结论:免疫表型对急性髓系白血病的诊断具有重要的意义,且免疫表型和异常核型存在密切的联系。     

急性髓系白血病铁死亡相关基因筛选及预后模型的建立

基于急性髓系白血病(Acute Myeloid Leukemia,AML)临床大数据及多组学数据库探讨铁死亡相关基因在AML中的作用,并建立铁死亡基因表达相关预后模型。整合TCGA数据库中151例AML患者和GTEx数据库中337例正常人外周血的临床和转录组数据。将Wilcoxon检验和单因素Cox分析结果取交集,筛选出预后相关差异表达基因(Differential Expression Genes, DEGs),使用Lasso回归建立基因标志物预后模型,利用受试者工作特征曲线(Receiver Operating Characteristic Curve,ROC曲线)评价预测价值,Kaplan-Meier法进行生存分析,对AML患者临床数据进行单因素和多因素Cox回归分析,使用差异基因表达分析等方法比较高、低风险患者间的组学差异,最后,利用BeatAML数据库对基因标志物进行验证。将差异基因表达分析和单因素分析结果取交集,得到13个预后相关DEGs。构建了8个基因标志物的预后评分模型,并将患者分为高、低风险两组;ROC曲线分析证实了模型良好的预测性能;生存分析提示高、低风险组患者的生存率具有显著差异;单因素分析显示年龄和风险评分与患者整体生存显著相关,多因素分析显示,年龄和风险评分是独立预后指标。在2个风险组之间筛选出384个DEGs,GO富集分析结果显示,富集的基因大多与中性粒细胞和白细胞的趋化与迁移等免疫相关分子和通路显著相关,KEGG富集通路主要与TNF信号通路、细胞因子与细胞因子受体相互作用相关。BeatAML数据库验证结果显示,5个基因与预后显著相关。铁死亡相关基因在AML中显著表达,且高风险患者预后较差,该研究对AML铁死亡相关潜在生物标志物的发现和应用奠定了一定的基础。     

RNAi干预急性髓系白血病对p38信号通路的影响

目的 探讨RNAi技术治疗急性髓系白血病（acute myeloid leukemia,AML）对p38信号通路的影响。方法 以NC（HL-60细胞）和HK（含siRNA-FLT3的阴性对照的HL-60细胞）、F1（实验组,含siRNA-FLT3的HL-60细胞,本实验室已实验证明对FLT3有干扰作用）三种细胞株为研究对象,分为药物组（信号通道抑制作用组）和对照组（未加信号通道抑制剂）。通过RT-PCR和Western blot检测对照组和药物组p38 mRNA水平和蛋白表达的变化,MTT和FCM分别测定细胞活性和细胞凋亡率的变化。结果 药物组、对照组组内比较：以NC做参照,F1能诱导p38 mRNA水平和蛋白表达下降、从而抑制细胞的增殖,促进细胞凋亡,差异有统计学意义（均P 0.05）;与对照组相比,药物组p38 mRNA水平和蛋白表达下降,细胞活性下降,细胞凋亡率增加,差异有统计学意义（均P<0.05）;药物组的F1与药物组的NC、药物组的F1与对照组的F1分别比较,差异有统计学意义（均P s<0.05）。结论 p38信号通路是RNAi技术干扰HL-60细胞FLT3基因表达引起细胞凋亡的途径之一,SB203580（P38 MAPK抑制剂）对其有协同增效作用。     

急性髓系白血病发生机制研究进展

急性髓系白血病(acute myeloid leukemia,AML)是一种骨髓造血干细胞的克隆性扩增和分化缺失导致的恶性增生性疾病,发病率和死亡率高.阐明AML的发生机制,开发治疗药物将有效提高患者存活率,但目前其具体发生机制不明.研究报道,AML的发生与相关基因突变、信号通路异常、表观遗传调控、白血病微环境或免疫失...     

急性髓系白血病相关基因的DNA甲基化

急性髓系白血病(acute myeloid leukemia,AML)是异质性造血干细胞恶性克隆性疾病,主要表现为外周血、骨髓以及其他组织中的原始细胞克隆性扩增。近年来,AML发病率呈逐年增加的趋势,对人类健康产生巨大的威胁。在不同种族及不同性别中,AML发病率及死亡率存在显著差异。AML是一种复杂疾病,与遗传突变及异常表观遗传修饰密切相关。DNA甲基化是重要的表观遗传修饰,AML相关基因的异常DNA甲基化修饰对疾病的发生发展极其重要。该文对AML相关基因的异常甲基化修饰进行了系统的作用机制分析,并对重要基因进行了功能分类。该文总结的具有异常DNA甲基化修饰的基因,有望辅助预测AML的治疗及预后效果,并为设计个体化AML治疗方案提供全新的思路。     

探讨髓系白血病细胞株糖酵解表型特征

探讨髓系白血病细胞株的糖酵解表型特征及其潜在的调控机制。葡萄糖试剂盒和乳酸试剂盒分别检测5株白血病细胞培养上清液中的葡萄糖消耗（G）和乳酸生成含量（L）,计算L／G比值来评估糖酵解水平：定量PCR检测糖酵解相关基因GLUT、MCTlmRNA表达;CCK8法检测细胞体外增殖能力;Western blot检测NAKT蛋白磷酸化水平。结果显示,KG1和K562细胞体外培养24h后的L／G比值分别为1．78和1．71,接近糖酵解表型时L／G为2的比值,同时这两株细胞高表达糖酵解相关基因GLUTl和MCT1mRNA。低糖（0．5mmol／L）、中糖（5mmol／L）、高糖（10mmol／L）处理KGla和K562细胞40h后,两株细胞的增殖能力、葡萄糖消耗和乳酸生成随葡萄糖浓度增加而增强,高糖组增加更为显著（P〈0．05）。相反,若糖酵解抑制剂2-DG（0,5,10mmol／L）处理白血病细胞40h后,两株细胞的增殖能力及糖酵解代谢水平随2．DG浓度增加而降低,高浓度2．DG组（10mmol／L）降低更为显著（P〈0．05）。此外,AKT抑制剂低浓度（5gmol／L）短时间（12h）处理后能抑制白血病细胞AKT蛋白磷酸化水平,同时降低细胞的葡萄糖消耗和乳酸生成（P〈0．05）。该研究提示髓系白血病细胞具有高糖酵解表型,AKT可能参与调控白血病的糖代谢过程,这有助于阐明白血病的能量代谢特征以及为白血病的靶向抗代谢治疗奠定基础。     

急性髓系白血病ERG、MDR1及BAALC基因的表达水平及临床意义

目的:探讨急性髓系白血病(acutemyeloidleukemia,AML)患者骨髓单个核细胞的ETS相关基因(ETS related gene,ERG)、多药耐药基因1(Multidrug resistance gene 1,MDR1)、脑和急性白血病胞质(Brain and acute leukemia cytoplasmic,BAALC)基因的表达水平及临床意义。方法:选取90例成人AML患者为研究对象,均接受蒽环类药物诱导化疗联合阿糖胞苷等进行初步治疗,采用实时荧光定量检测PCR技术检测治疗后的骨髓单个核细胞ERG、MDR1、BAALC基因表达,并分析其与患者临床特征、危险度分层、治疗疗效、生存率的关系。结果:AML患者ERG、MDR1、BAALC基因均有强表达,三因子表达强弱同白细胞、血小板等临床指标无明显相关性(P0.05),不同危险度分层对应的ERG、MDR1、BAALC表达之间有明显差异,中危组、高危组患者表达强度均高于低危组(P0.05)。ERG、MDR1低表达组对应的疗效CR(93.1%%,89.6%)、OS(56.5%, 66.7%)较高表达组CR(61.4%, 81.3%)、OS(56.5%,66.7%)值更高(P0.05),不同BAALC表达者疗效CR及生存率OS比较差异无统计学意义(P0.05)。结论:AML患者单个核细胞的ERG、MDR1基因表达水平与其危险度分层、疗效和生存率呈负相关,以REG和MDR1同AML关系敏感,BAALC敏感度较低,联合检测REG、MDR1、BAALC基因表达可能提高AML患者危险度分层、疗效及预后判读的准确度。     

Nestin基因在髓系白血病的表达及意义

Nestin蛋白是一种神经干细胞标志物,参与组织修复,并且在一些肿瘤细胞中表达。最近研究报道称nestin可能参与肿瘤细胞恶性增殖及细胞侵袭。在急性淋巴细胞白血病(acute lymphoid leukemia,ALL)小鼠模型中,白血病增殖细胞(leukemia-propagating cells,LPCs)可形成一个抗药性niche,其中nestin阳性细胞参与该niche形成。为探索nestin基因在白血病患者的临床表达情况,通过RT-PCR和Western-blot实验方法 ,检测了50位髓系白血病患者的nestin基因的临床表达情况。结果发现,在50位髓系白血病患者(38位AML,12位CML)中,42位患者表达nestin基因mRNA,6位患者表达nestin蛋白,并且nestin基因表达水平和患者血红蛋白、血小板、细胞因子和T淋巴细胞数并不存在相关性,但是高白细胞白血病患者表现出nestin基因mRNA水平高表达。虽然nestin基因和患者完全缓解率(complete remission,CR)并不存在联系,但是nestin基因在髓系白血病异常表达,说明其可能作为一种诊断AML或CML的生物标记物。     

MicroRNAs and response to therapy in leukemia

A variety of epigenetic factors involved in leukemia pathogenesis. Among various epigenetic factors, microRNAs (miRNAs) have emerged as important players, which affect a sequence of cellular and molecular signaling pathways. Leukemia is known as progressive cancer, which is related to many health problems in the world. It has been shown that the destruction of the blood-forming organs could lead to abnormal effects on the proliferation and development of leukocytes and their precursors. Despite many attempts for approved effective and powerful therapies for patients with leukemia, finding and developing new therapeutic approaches are required. One of the important aspects of leukemia therapy, identification of underlying cellular and molecular mechanisms involved in the pathogenesis of leukemia. Several miRNAs (ie, miR-103, miR-101, mit-7, let-7i, miR-424, miR-27a, and miR-29c) and play major roles in response to therapy in patients with leukemia. miRNAs exert their effects by targeting a variety of targets, which are associated with response to therapy in patients with leukemia. It seems that more understanding about the roles of miRNAs in response to therapy in patients with leukemia could contribute to better treatment of patients with leukemia. Here, for the first time, we summarized various miRNAs, which are involved in response to therapy in the treatment patients with leukemia.     

Argonaute 2 sustains the gene expression program driving human monocytic differentiation of acute myeloid leukemia cells

MicroRNAs are key regulators of many biological processes, including cell differentiation. These small RNAs exert their function assembled in the RNA-induced silencing complexes (RISCs), where members of Argonaute (Ago) family of proteins provide a unique platform for target recognition and gene silencing. Here, by using myeloid cell lines and primary blasts, we show that Ago2 has a key role in human monocytic cell fate determination and in LPS-induced inflammatory response of 1,25-dihydroxyvitamin D₃ (D3)-treated myeloid cells. The silencing of Ago2 impairs the D3-dependent miR-17-5p/20a/106a, miR-125b and miR-155 downregulation, the accumulation of their translational targets AML1, VDR and C/EBPβ and monocytic cell differentiation. Moreover, we show that Ago2 is recruited on miR-155 host gene promoter and on the upstream region of an overlapping antisense lncRNA, determining their epigenetic silencing, and miR-155 downregulation. These findings highlight Ago2 as a new factor in myeloid cell fate determination in acute myeloid leukemia cells.     

Methylenetetrahydrofolate reductase gene polymorphisms contribute to acute myeloid leukemia and chronic myeloid leukemia susceptibilities: Evidence from meta-analyses

PurposeThe expression of methylenetetrahydrofolate reductase (MTHFR) is associated with acute myeloid leukemia (AML) and chronic myeloid leukemia (CML). Most studies have linked the common functional C677T and A1298C polymorphisms of the MTHFR gene and susceptibility to AML and CML, but the results were not consistent. The aim of the present study was to derive a more precise estimation of the relationship.MethodsMeta-analyses assessing the association of MTHFR C677T and A1298C variations with AML and CML were conducted. Eligible articles were identified from the PubMed and EMBASE databases. All statistical analyses were conducted using Review Manager Software.Results10 and 10 studies were included in the meta-analysis about the role of C677T polymorphism on the AML and CML risks, respectively; 6 and 4 studies were included about the role of A1298C polymorphism on the AML and CML risks, respectively. Overall, both the C677T and A1298C polymorphisms were significantly associated with CML risk under the recessive model (P = 0.04, OR = 1.35, 95% CI = 1.02–1.79 for C677T and P = 0.003, OR = 2.17, 95% CI = 1.29–3.63 for A1298C). In addition, the risk of CML was higher in 1298CC genotype carriers than in 1298AA genotype carriers (P = 0.004, OR = 2.17, 95% = 1.28–3.69). Conversely, the overall data failed to indicate a significant association of C677T or A1298C polymorphisms with AML risk under any model.ConclusionsThe findings provide evidence that C677T and A1298C polymorphisms are risk factors for CML risk.     

Interleukin-17 in acute myeloid leukemia

There are several reports that angiogenesis plays important roles in hematological malignancies including acute myeloid leukemia (AML). Human interleukin-17 (IL-17) is a proinflammatory cytokine produced by activated CD4 T cells. IL-17 plays a potential role in T cell mediated angiogenesis. The role of IL-17 in pathologic angiogenesis has not been evaluated yet. The aim of the study was to determine plasma level of IL-17 in patients with AML. IL-17 levels were measured by ELISA in plasma samples taken from 68 adult patients with AML before chemotherapy was administered. In addition 20 out of 68 patients were reanalysed after achieving complete remission (CR). Ten samples from healthy volunteers were evaluated as the control. In this study we have demonstrated that serum level of IL-17 is not elevated in AML patients. These results suggest that angiogenesis in AML is not mediated by CD4 T cells. To our knowledge this is the first report about IL-17 serum level in acute leukemias. We are currently evaluating IL-17 levels in others haematological malignancies.     

The role,mechanism and potentially therapeutic application of microRNA-29 family in acute myeloid leukemia

Abnormal proliferation, apoptosis repression and differentiation blockage of hematopoietic stem/progenitor cells have been characterized to be the main reasons leading to acute myeloid leukemia (AML). Previous studies showed that miR-29a and miR-29b could function as tumor suppressors in leukemogenesis. However, a comprehensive investigation of the function and mechanism of miR-29 family in AML development and their potentiality in AML therapy still need to be elucidated. Herein, we reported that the family members, miR-29a, -29b and -29c, were commonly downregulated in peripheral blood mononuclear cells and bone marrow (BM) CD34+ cells derived from AML patients as compared with the healthy donors. Overexpression of each miR-29 member in THP1 and NB4 cells markedly inhibited cell proliferation and promoted cell apoptosis. AKT2 and CCND2 mRNAs were demonstrated to be targets of the miR-29 members, and the role of miR-29 family was attributed to the decrease of Akt2 and CCND2, two key signaling molecules. Significantly increased Akt2, CCND2 and c-Myc levels in the AML cases were detected, which were correlated with the decreased miR-29 expression in AML blasts. Furthermore, a feed-back loop comprising of c-Myc, miR-29 family and Akt2 were found in myeloid leukemogenesis. Reintroduction of each miR-29 member partially corrected abnormal cell proliferation and apoptosis repression and myeloid differentiation arrest in AML BM blasts. An intravenous injection of miR-29a, -29b and -29c in the AML model mice relieved leukemic symptoms significantly. Taken together, our finding revealed a pivotal role of miR-29 family in AML development and rescue of miR-29 family expression in AML patients could provide a new therapeutic strategy.     

Karyotypic findings in chronic myeloid leukemia cases undergoing treatment

BACKGROUND:

Chronic myeloid leukemia (CML) is a clonal myeloproliferative expansion of primitive hematopoietic progenitor cells.

MATERIALS AND METHODS:

In the present study, CML samples were collected from various hospitals in Amritsar, Jalandhar and Ludhiana.

RESULTS:

Chromosomal alterations seen in peripheral blood lymphocytes of these treated and untreated cases of CML were satellite associations, double minutes, random loss, gain of C group chromosomes and presence of marker chromosome. No aberrations were observed in control samples. Karyotypic abnormalities have also been noted in the Ph-negative cells of some patients in disease remission.

CONCLUSION:

This is a novel phenomenon whose prognostic implications require thorough and systematic evaluation.     

The allometry of chronic myeloid leukemia

Chronic myeloid leukemia (CML) is an acquired neoplastic hematopoietic stem cell (HSC) disorder characterized by the expression of the BCR-ABL oncoprotein. This gene product is necessary and sufficient to explain the chronic phase of CML. The only known cause of CML is radiation exposure leading to a mutation of at least one HSC, although the vast majority of patients with CML do not have a history of radiation exposure. Nonetheless, in humans, significant radiation exposure (after exposure to atomic bomb fallout) leads to disease diagnosis in 3-5 years. In murine models, disease dynamics are much faster and CML is fatal over the span of a few months. Our objective is to develop a model that accounts for CML across all mammals. In the following, we combine a model of CML dynamics in humans with allometric scaling of hematopoiesis across mammals to illustrate the natural history of chronic phase CML in various mammals. We show how a single cell can lead to a fatal illness in mice and humans but a higher burden of CML stem cells is necessary to induce disease in larger mammals such as elephants. The different dynamics of the disease is rationalized in terms of mammalian mass. Our work illustrates the relevance of animal models to understand human disease and highlights the importance of considering the re-scaling of the dynamics that accrues to the same biological process when planning experiments involving different species.     

Risk of adult acute and chronic myeloid leukemia with cigarette smoking and cessation

BackgroundCigarette smoking is an established risk factor for adult myeloid leukemia, particularly acute myeloid leukemia (AML), but less is known about the nature of this association and effects of smoking cessation on risk.MethodsIn a large population-based case–control study of myeloid leukemia that included 414 AML and 185 chronic myeloid leukemia (CML) cases and 692 controls ages 20–79 years, we evaluated risk associated with cigarette smoking and smoking cessation using unconditional logistic regression methods and cubic spline modeling.ResultsAML and CML risk increased with increasing cigarette smoking intensity in men and women. A monotonic decrease in AML risk was observed with increasing time since quitting, whereas for CML, the risk reduction was more gradual. For both AML and CML, among long-term quitters (≥30 years), risk was comparable to non-smokers.ConclusionsOur study confirms the increased risk of myeloid leukemia with cigarette smoking and provides encouraging evidence of risk attenuation following cessation.     

Low expression of microRNA-340 confers adverse clinical outcome in patients with acute myeloid leukemia

MicroRNA-340 (miR-340) was considered as a tumor suppressor by affecting cancer cell proliferation, apoptosis, invasion, and migration, and was downregulated in diverse cancers. Moreover, dysregulation of miR-340 was also found to be associated with drug resistance and predicted patients’ survival in various cancers. Herein, we investigated miR-340 expression and its clinical significance in acute myeloid leukemia (AML). Real-time quantitative polymerase chain reaction was performed to detect miR-340 expression in bone marrow (BM) from 99 newly diagnosed AML patients except for acute promyelocytic leukemia (APL), 19 AML patients achieved complete remission (CR), and 29 healthy donors. BM miR-340 expression was significantly underexpressed in newly diagnosed AML patients as compared with controls (p = 0.031) and AML patients achieved CR (p = 0.025). No significant differences were observed between miR-340 expression and most of the clinicopathologic features (p > 0.05). However, low miR-340 expression was found to be associated with lower CR rate in both non-APL-AML and cytogenetically normal AML (CN-AML; p = 0.001 and 0.031, respectively), and acted as an independent risk factor for CR by logistic regression analysis (p = 0.001 and 0.021, respectively). More important, among both non-APL-AML and CN-AML, low expression of miR-340 was also associated with shorter overall survival (OS; p = 0.013 and 0.005, respectively), and was further validated by Cox regression (p = 0.031 and 0.039, respectively). Collectively, our study showed that BM miR-340 expression was downregulated in AML, and low expression of miR-340 correlated with adverse prognosis.