急性前髓细胞性白血病细胞诱导分化实验的研究进展

急性前髓细胞性白血病或急性早幼粒细胞白血病（APL）是一种特殊类型的血液系统恶性克隆性疾病,其特点是异常早幼粒细胞无限增殖伴分化受阻,是白血病中最危重的一种类型。95%以上的APL患者具有（t15;17）染色体异常,形成PML/RAR融合基因,几乎存在于所有的APL细胞中,成为APL细胞的一个特异性标志,是APL发病重要分子基础。自从全反式维甲酸（ATRA）成功用于临床诱导APL分化以来,对诱导分化剂的作用机制的研究已取得很大的进展。本文主要对APL细胞遗传学和分子生物学特征、发病机制、诱导分化机制、分化后细胞表型变化等方面对APL细胞诱导分化实验的研究进展进行综述。     

成人急性髓性白血病SOCS-1基因及其甲基化的研究

目的：通过检测成人急性髓性白血病中SOCS．1基因表达水平及其甲基化水平,研究其在白血病发病中的作用。方法：运用甲基化特异性PCR（Methylation specificPCR,MSP）方法,对24例急性髓性白血病患者和4株白血病细胞株（Jurkat、Raji、U937、NALM17）,进行SOCS-1基因甲基化水平的研究;同时运用Real—timePCR法定量分析SOCS—1基因表达水平。以10例健康人为正常对照组。结果：24例成人急性髓性白血病患者中,15例有SOCS-1基因甲基化（62．5％）,而正常对照组无SOCS-1基因甲基化（0％）,二者有显著差异（P〈0．05）;SOCS-1基因甲基化组与无SOCS-1基因甲基化组相比较,其SOCS—1基因相对表达量明显减少（P口0．05）;与患者临床病理特征相结合比较,发现SOCS-1基因的甲基化与患者年龄、性别和病程阶段无相关。4株白血病细胞株中,Jurkat和U937表现有SOCS—1甲基化（50％）,Raji和NALM17无SOCS—1甲基化,前者SOCS-1基因表达量较后者也明显降低（P〈0．05）。结论：SOCS—1基因在成人急性髓性白血病中甲基化水平明显增高,且SOCS-1基因甲基化后表达水平受到抑制,提示SOCS-1基因及其甲基化在急性髓性白血病的发生发展中可能具有一定作用。     

成人急性髓性白血病SOCS-1 基因及其甲基化的研究

目的:通过检测成人急性髓性白血病中SOCS-1基因表达水平及其甲基化水平,研究其在白血病发病中的作用。方法:运用甲基化特异性PCR(Methylation specific PCR,MSP)方法,对24例急性髓性白血病患者和4株白血病细胞株(Jurkat、Raji、U 937、NALM 17),进行SOCS-1基因甲基化水平的研究;同时运用Real-time PCR法定量分析SOCS-1基因表达水平。以10例健康人为正常对照组。结果:24例成人急性髓性白血病患者中,15例有SOCS-1基因甲基化(62.5%),而正常对照组无SOCS-1基因甲基化(0%),二者有显著差异(P<0.05);SOCS-1基因甲基化组与无SOCS-1基因甲基化组相比较,其SOCS-1基因相对表达量明显减少(P﹤0.05);与患者临床病理特征相结合比较,发现SOCS-1基因的甲基化与患者年龄、性别和病程阶段无相关。4株白血病细胞株中,Jurkat和U 937表现有SOCS-1甲基化(50%),Raji和NALM 17无SOCS-1甲基化,前者SOCS-1基因表达量较后者也明显降低(P<0.05)。结论:SOCS-1基因在成人急性髓性白血病中甲基化水平明显增高,且SOCS-1基因甲基化后表达水平受到抑制,提示SOCS-1基因及其甲基化在急性髓性白血病的发生发展中可能具有一定作用。     

SHIP1在急性髓细胞白血病患者中的表达及对人白血病细胞凋亡的影响研究

目的:探讨含SH2结构域的肌醇1(SHIP1)在急性髓细胞白血病患者中的表达及对人白血病细胞凋亡的影响。方法:采用Western blot检测收集的急性髓细胞白血病患者骨髓中SHIP1的表达。人白血病细胞U937转染SHIP1过表达载体(pEGFP-SHIP1组)及对照空载体(pEGFP组),同时设置对照组,对照组细胞不转染载体,其他步骤同pEGFP-SHIP1组和pEGFP组。流式细胞仪检测48 h的细胞凋亡情况,Western blot检测48 h细胞中SHIP1、Bcl-2、Bax、Akt、p-Akt的表达。结果:急性髓细胞白血病患者骨髓中SHIP1表达明显低于正常人(P0.05)。pEGFP-SHIP1组细胞中SHIP1、Bax表达和凋亡率均明显高于pEGFP组及对照组(P0.01),Bcl-2、p-Akt表达均明显低于对照组(P0.01)。结论:SHIP1在急性髓细胞白血病患者骨髓中表达下调,其可能通过Akt信号促进人白血病细胞凋亡。     

转录因子与急性髓性白血病分子生物学研究进展

转录因子是一类识别和综合特定ＤＮＡ调节顺序的核蛋白,能刺激或抑制转录。与人类白血病有关的ＴＦ基因是目前研究的热门课题,本对ＴＦ基序及髓性白血病ＴＦ特征等进行了综述。     

急性髓性白血病SCID小鼠模型的建立及鉴定

目的:探讨建立急性髓性白血病小鼠模型,并对模型建立过程进行观察、分析和鉴定.方法:重症联合免疫缺陷小鼠(SCID)建立HL-60细胞白血病动物模型,观察SCID小鼠一般情况,监测小鼠外周血HL-60细胞、白细胞、红细胞、血小板的变化,用流式细胞术、骨髓染色体检查等检测小鼠骨髓,濒死或死亡小鼠行组织病理学检查.结果:接种后3-5周,外周血白细胞上升,外周HL-60细胞占非红系有核细胞比例为3%-11%.骨髓HL-60细胞占非红系有核细胞中比例为40%-58%,骨髓细胞中CD33抗原阳性率8.18%-24.45%.小鼠骨髓细胞中有中、亚中部着丝点染色体.结论:HL-60细胞静脉接种SCID小鼠,白血病发病率为100%.其发病症状及组织浸润与人类白血病相类似,可用于研究白血病增殖、分化等机制,是探索新的白血病治疗手段及判断患者预后的重要载体.     

277例急性髓细胞白血病的细胞遗传学分析

目的：探讨急性髓细胞白血病（AML）染色体异常核型的分布情况、特点及其与预后的关系。方法：选择2009年1月-2012年12月在本院门诊和住院治疗的AML患者277例,采取新鲜骨髓,采用直接法和短期培养制备标本,以R显带技术进行染色体核型分析。结果：除47例无分裂相共检出异常核型122例,异常率为53．04％。结构异常（75例）以t（15;17）（56例）和t（8;21）（19例）常见,1．78％的M2、90．77％的M3、26．79％的M2和1．85％的M5分别有t（7;11）、t（15;17）、t（8;21）和t（11q23）异常,而100％的t（7;11）、100％的t（15;17）、55．56％的t（8;21）、100％的t（8;15）和66．7％的t（11q23）分别见于M2、M3、M2、M5和M4亚型。数I／1异常（28例）以＋8（12例）为常见,100％的＋22见于M4E0。结构和数目同时异常（19例）以t（8;21）伴性染色体丢失（8例）为最常见,100％为M2。核型异常与临床缓解率高度相关,伴t（15;17）易位组、伴t（8;21）易位组预后佳,正常核型组次之,其它异常核型预后较差,复杂核型预后最差。结论：特定的染色体异常与亚型及预后密切相关,染色体核型分析在急性髓细胞白血病诊断、分型、治疗及预后判断中具有十分重要的地位和作用。     

DLK1基因在急性白血病细胞系中的表达及其意义

目的:研究急性白血病细胞系DLK1基因的表达水平在红系分化中的作用.方法:采用RT-PCR、Western bitting时白血病细胞系K562、HL-60进行DLK1水平的检测.培养K562细胞,用氯化高铁血红素(hemin)诱导其分化,观察DLK1在红系分化中的变化.结果:K562细胞DLK1mRNA、蛋白水平存在明显表达,HL-60细胞DLK1则不表达.通过RT-PCR检测了hemin诱导K562细胞向红系分化过程中各时间点DLK1mRNA的变化,显示随着K562向红系分化,DLK1mRNA的水平逐渐下降.结论:K562细胞表达DLK1,HL-60不表达DLK1.DLK1基因可能参与K562细胞向红系分化的过程,可能抑制其分化.     

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。     

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。     

A new astrocytic cell line which is able to induce a blood-brain barrier property in cultured brain capillary endothelial cells

Astrocytes, a member of the glial cell family in the central nervous system, are assumed to play a crucial role in the formation of the blood-brain barrier (BBB) in vertebrates. It was shown that astrocytes induce BBB-properties in brain capillary endothelial cells (BCEC) in vitro. We now established an astroglial cell line of non-tumoral origin. The cloned cell line (A7) shows a highly increased proliferation rate and expresses the astrocytic marker glial fibrillary acidic protein. Furthermore, the clone A7 expresses S-100-protein and vimentin, which are also expressed by primary cultured astrocytes. This cell line therefore shows general astrocytic features. In addition, we were able to show that A7 cells re-induce the BBB-related marker enzyme alkaline phosphatase in BCEC, when these two cell types are co-cultured. Thus we have a cell line which can be readily cultured in large quantities, shows common astrocyte properties and is able to influence BCEC with respect to a BBB-related feature. This revised version was published online in July 2006 with corrections to the Cover Date.     

Low IL-23 levels in peripheral blood and bone marrow at diagnosis of acute leukemia in children increased with the elimination of leukemic burden

IL-23 is an IL-12 cytokine family member with pleiotropic functions that regulates tumour growth in various cancer types, exhibiting both anti-tumorigenic and pro-tumorigenic properties. Preclinical studies have shown a potential anti-leukemic action on childhood B-ALL cells. The study involved 65 children with acute leukemia [59 patients with acute lymphoblastic leukemia (ALL) and 6 patients with acute myeloid leukemia (AML)] and 27 healthy controls. Using an enzyme-linked immunosorbent assay, we aimed to determine the IL-23 levels in the peripheral blood (PB) and bone marrow (BM) of patients at diagnosis and at the end of the induction therapy (EIT). PB IL-23 levels were lower in leukemia patients compared to the healthy controls. In all acute leukemia patients, IL-23 levels were significantly lower at diagnosis both in PB (P = .015) and in BM (P = .037) compared to the PB and BM concentrations at the EIT. The same pattern was present in both subgroups of ALL and AML patients. The high leukemic burden at diagnosis was related with lower IL-23 levels, which were increased with the disease remission. Considering the anti-leukemic potential of this cytokine, the elevation of the IL-23 concentration at the disease remission indicates a beneficial role of IL-23 in paediatric acute leukemia.     

Differentiation of acute and chronic myeloid leukemic blasts into the dendritic cell lineage: analysis of various differentiation-inducing signals

Purpose: Ex vivo differentiation of myeloid leukemic blasts into dendritic cells (DCs) holds significant promise for use as cellular vaccines, as they may present a constellation of endogenously expressed known and unknown leukemia antigens to the immune system. Although variety of stimuli can drive leukemiaDC differentiation in vitro, these blast-derived DCs typically have aberrant characteristics compared with DCs generated from normal progenitors by the same stimuli. It is not clear whether this is due to underlying leukemogenic mechanisms (e.g., specific oncogenes), genetic defects, stage of maturation arrest, defects in cytokine receptor expression or signal transduction pathways, or whether different stimuli themselves induce qualitatively dissimilar DC differentiation. Methods: To assess what factors may contribute to aberrant leukemic blastDC differentiation, we have examined how the same leukemic blasts (AML and CML) respond to different DC differentiation signals—including extracellular (the cytokine combination GM-CSF+TNF-+IL-4) and intracellular (the protein kinase C agonist PMA, the calcium ionophore A23187, and the combination of PMA plus A23187) stimuli. Results: We have found that the same leukemic blasts will develop qualitatively different sets of DC characteristics in response to differing stimuli, although no stimuli consistently induced all of the characteristic DC features. There were no clear differences in the responses relative to specific oncogene expression or stage of maturation arrest (AML vs CML). Signal transduction agonists that bypassed membrane receptors/proximal signaling (in particular, the combination of PMA and A23187) consistently induced the greatest capability to activate T cells. Interestingly, this ability did not clearly correlate with expression of MHC/costimulatory ligands. Conclusions: Our findings suggest that signal transduction may play an important role in the aberrant DC differentiation of leukemic blasts, and demonstrate that direct activation of PKC together with intracellular calcium signaling may be an effective method for generating immunostimulatory leukemia-derived DCs.This work was supported by NIH CA85208, CA95829, and ASCO Young Investigator Award (M.A.K-D)     

The evolving landscape in the therapy of acute myeloid leukemia

Acute myeloid leukemia (AML) is a heterogeneous clonal disorder of myeloid precursors arrested in their maturation, creating a diverse disease entity with a wide range of responses to historically standard treatment approaches. While signifi cant progress has been made in characterizing and individualizing the disease at diagnosis to optimally inform those affected, progress in treatment to reduce relapse and induce remission has been limited thus far. In addition to a brief summary of the factors that shape prognostication at diagnosis, this review attempts to expand on the current therapies under investigation that have shown promise in treating AML, including hypomethylating agents, gemtuzumab ozogamicin, FLT3 tyrosine kinase inhibitors, antisense oligonucleotides, and other novel therapies, including aurora kinases, mTOR and PI3 kinase inhibitors, PIM kinase inhibitors, HDAC inhibitors, and IDH targeted therapies. With these, and undoubtedly many others in the future, it is the hope that by combining more accurate prognostication with more effective therapies, patients will begin to have a different, and more complete, outlook on their disease that allows for safer and more successful treatment strategies.     

miR-345-5p regulates proliferation,cell cycle,and apoptosis of acute myeloid leukemia cells by targeting AKT2

Acute myeloid leukemia (AML) is a malignant clonal hematopoietic disease, which is caused by hematopoietic stem cell abnormalities. Epigenetic regulation, especially of microRNAs (miRNAs), mostly results from external or environmental effects and is critical to AML. In this study, for the first time, we report that decreased expression of miR-345-5p facilitates the proliferation of leukemia cells in AML. Further study demonstrated that AKT1/2 was the target of miR-345-5p and was responsible for the dysregulation of leukemia cell proliferation and apoptosis. Inhibition of AKT1/2 ameliorated this malignant effect, which provides new insight into AML diagnosis, treatment, prognosis, and next-step translational investigations.     

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.     

High expression of miR-338 is associated with poor prognosis in acute myeloid leukemia undergoing chemotherapy

Acute myeloid leukemia (AML) is a heterogeneous disease with unfavorable outcomes. MicroRNAs (miRNAs) are important regulators and prognostic factors involved in AML. To determine the clinical role of miR-338 in AML, a total of 164 adults with de novo AML were collected. These patients were classified into a chemotherapy group and an allogeneic hematopoietic stem cell transplantation (allo-HSCT) group according to the clinical treatment, and then each group was divided into two subgroups based on the median miR-338 expression values. We found that upregulated miR-338 positively correlates with higher frequencies of complex karyotype, RUNX1 mutation, and poor risk status. In the chemotherapy group, high expression of miR-338 was independently associated with shorter EFS and OS. However, no significant differences were observed between the two subgroups within the allo-HSCT group. We also divided all patients into two groups according to the median miR-338 expression values of the whole cohort. In the miR-338 high expression group, patients receiving allo-HSCT had longer OS and EFS than those receiving chemotherapy only. In contrast, patients receiving different therapies had similar OS and EFS in the miR-338 low expression group. Our study suggests that high expression of miR-338 is an adverse prognostic biomarker in patients with AML undergoing chemotherapy and may guide treatment decisions for AML. Furthermore, allo-HSCT could significantly overcome the negative effect of high miR-338 expression, but it seemed to be unbeneficial and unnecessary for low miR-338 expressions.