髓系/NK细胞急性白血病一例

患者男，43岁。因“乏力、四肢皮肤淤点、淤斑3个月”于2005年9月7日收入我科。患者于3个月前无诱因出现乏力、皮肤淤点、淤斑，反复发作。入院体检：皮肤可见散在淤点、淤斑，浅表淋巴结未及，胸骨压痛阳性，肝、脾肋下均4cm；血象Hb114g／L，WBC59．8×10^9／L，PLT 9×10^9／L。骨髓象增生极度活跃，异常早幼粒细胞占0．82，该类细胞胞体大小不等，可见伪足，浆量多，浆内可见大量细小的紫红色颗粒，[第一段]     

急性髓细胞白血病CD34抗原与干细胞白血病基因表达及其意义

CD34抗原是造血干细胞的表面标志,随着造血干细胞分化、成熟而逐渐消失;干细胞白血病(SCL)基因也表达于造血干细胞中,它在维持造血干细胞增殖分化过程中起着重要作用,并随着造血干细胞的分化成熟而消失.CD34抗原与SCL基因在急性髓细胞白血病(AML)中有所表达,它们与AML的临床特征及预后存在一定关系[1,2],但也存在争论,国内报道较少.我们对167例AML进行免疫分型,应用逆转录-聚合酶链反应(RT-PCR)进行SCL基因检测,同时结合AML临床表现、血流学特点、治疗效果进行分析.     

化疗联合自体细胞因子诱导杀伤细胞治疗急性白血病的临床观察

目的评价化疗联合自体细胞因子诱导的杀伤细胞(CIK)治疗急性白血病(AL)的疗效。方法选择经化疗达完全缓解(CR)>6个月的AL患者41例。19例患者接受化疗联合自体CIK细胞治疗,22例接受同期单纯化疗作为对照组。CIK细胞治疗采用血细胞分离机大量采集患者的外周血单个核细胞,用抗CD3单抗、IL2、IFNγ培养10d,将细胞洗涤后经静脉于化疗后第一天回输给患者。结果CIK组19例患者化疗同时共接受52疗程CIK细胞治疗,每例患者平均接受2~3个疗程CIK细胞治疗。每疗程回输CIK细胞总数为(22~300)×109/L,平均(142±85)×109/L。4年持续CR(CCR)率CIK组为734%;单纯化疗组4年预期CCR率为273%。两者差异有统计学意义(P<0005)。接受≥3个疗程CIK治疗的10例患者至观察截止时均处于CCR,中位CCR期43个月(23~52个月);接受<3个疗程CIK治疗的患者4/9例复发。结论化疗联合自体CIK细胞治疗AL的CCR率明显优于单纯化疗;疗效与疗程有关,疗程≥3个患者的疗效优于疗程<3个的患者。     

脐血NK细胞的杀伤活性及其影响因素

目的探讨脐血NK细胞的杀伤活性及其影响因素。方法对成人脐血及外周血NK细胞的活化和抑制性受体、细胞毒性颗粒、胞内细胞因子的表达进行检测。结果脐血NK细胞与成人外周血NK细胞的大部分指标均无差异,但脐血NK细胞的NKG2A、CD94表达明显升高,而颗粒酶B表达则较低。结论脐血NK细胞抑制性受体NKG2A/CD94表达升高,颗粒酶B降低是脐血NK细胞活性低于外周血NK细胞的主要原因。     

体外诱导白血病源性树突状细胞及其对T细胞杀伤活性影响的研究

目的 :探讨体外培养白血病源性树突状细胞 (DC)及其对T细胞杀伤活性的影响。方法 :从慢性髓细胞性白血病 (CML)患者外周血中分离单个核细胞 ,加入 1× 10 6U/L粒 巨噬细胞集落刺激因子 (GM CSF)、1×10 6U/L白细胞介素 (IL) 4和 5 0× 10 3 U/L肿瘤坏死因子 (TNF α) ,每 3～ 4天换液 1次 ,连续培养 14天 ,获得DC。取外周血单个核细胞加入 5 0 0× 10 3 U/LIL 2 ,培养至第 7天 ,把细胞分成两组 ,其中一组加入培养的DC ,两组细胞继续培养 3～ 4天 ,然后测定T细胞杀伤活性。结果 :DC具有典型的树状突起 ,高表达CD1a,具有慢粒特异性染色体t(9;2 2 ) ,能增强T细胞对白血病细胞的杀伤活性。结论 :在体外利用细胞因子可以把CML细胞诱导分化成具有刺激T细胞产生细胞毒性反应的白血病源性DC。     

急性髓系白血病CD34和CD38抗原表达及其临床意义

目的:分析急性髓系白血病(AML)CD34、CD38抗原表达特点及临床意义。方法:采用单克隆抗体直接免疫荧光标记法的流式细胞术,对55例AML患者进行免疫表型检测。结果:AML患者CD34和CD38表达均达40．0％。AML CD34 患者完全缓解(CR)率(19．0％)明显低于CD34-患者(73．5％)(P<0．05);而CD38 患者CR率(52．4％)与CD38-患者CR率(52．9％)基本相同。结论:作为造血干细胞免疫标记的CD34对判断AML临床预后有一定的指导意义。     

急性髓系白血病患者预后分组方式的探讨

目的 评价原发、初治急性髓系白血病(AML)患者诱导治疗后不同时间骨髓幼稚细胞比例对预后的影响.将细胞遗传学与诱导治疗后不同时间骨髓幼稚细胞比例相结合,提出新的AML患者预后分组方法.方法 回顾性分析1999年1月1日至2008年2月1日于我院住院的原发、初治AML患者(非M3型)105例,所有患者在诱导化疗结束时(T1)和(或)骨髓抑制期(T2)进行骨髓穿刺检查.有细胞遗传学资料的患者97例.结果 (1)T1或T2时间点105例行骨髓穿刺检查的患者,骨髓幼稚细胞<0.05者和≥0.05者相比,T1时间点完全缓解(CR)率分别为86.0%、47.4%,3年无复发生存(RFS)率分别为46.2%、21.6%,3年总生存率分别为49.7%、25.6%.T2时间点二者CR率分别为86.3%、41.4%,3年RFS率分别为52.4%、18.9%,3年总生存率分别为61.1%、35.2%,差异均有统计学意义.且T1和,12时间点骨髓幼稚细胞比例具有相关性.(2)将染色体核型预后中等组患者根据T1或T2时间点骨髓幼稚细胞比例分为二组:骨髓幼稚细胞<0.05者和≥0.05者.前者预后与良好组相近,后者预后与不良组相近.(3)多因素分析表明T1或12时间点骨髓幼稚细胞比例是AML患者的独立预后因素.T1时间点骨髓幼稚细胞比例可能较T2时间点骨髓幼稚细胞比例意义更大.结论 以0.05为界,T1或T2时间点骨髓幼稚细胞比例是原发、初治AML患者(非M3型)CR率、RFS、总生存的独立预后因素.将染色体核型与T1和(或)T2时间点骨髓幼稚细胞比例相结合分组,可进一步区分中等组患者,有助于评估预后和选择治疗方案.     

急性白血病治疗进展：急性髓细胞白血病的治疗

近年来许多实验资料证明 ,化学药物剂量增加 1倍 ,对白血病细胞杀伤能力增加 10倍。因此 ,国内外不少学者主张对急性白血病患者采用强烈诱导化疗和早期大剂量强化治疗 ,但不等于无限制地盲目地增大剂量和延长用药时间。现将急性髓细胞白血病 (ANLL)的治疗原则总结如下 ,供同道参考。1　诱导缓解目前 ,国内外对急性白血病强烈诱导化疗的方案药物剂量标准不完全一致 ,但一般认为须达到骨髓明显抑制水平 ,外周血白细胞 <1× 10 9/ L,甚至 0 .5×10 9/ L以下。治疗急性 ANL L常用 DA及 HA方案。经过多年来的临床试验后认为 ,目前应用最广…     

枸杞多糖在白血病患者中的应用及对HL-60和NK细胞杀伤活性的影响

目的探讨枸杞多糖在白血病患者中的应用及对人前髓细胞HL-60表面靶细胞表面相应配体(MICA)蛋白表达水平和自然杀伤(NK)细胞杀伤活性的影响及机制。方法取白血病患者HL-60细胞,随机分为观察1组、观察2组及观察3组,分别放入浓度为6、8、10 mg/ml枸杞多糖共同培养,未经枸杞多糖作用的30例白血病患者HL-60细胞为对照组,采用流式细胞仪测定4组细胞表面MICA蛋白表达水平。将4组获得的溶液与NK细胞相互混合,20 h后采用四甲基偶氮唑蓝比色(MTT)法检测NK细胞对4组HL-60细胞的杀伤活性进行测定。结果观察1组MICA表达率显著低于观察2组、观察3组(P<0.05);观察1组、观察2组、观察3组MICA表达率高于对照组(P<0.05);随着靶细胞、效应细胞比的不断增加,4组NK细胞杀伤活性均出现上升趋势;观察1组、观察2组及观察3组NK细胞杀伤活性高于对照组(P<0.05);观察3组NK细胞杀伤活性,高于观察1组、观察2组(P<0.05)。结论白血病患者体内HL-60细胞在枸杞多糖的孵育下能提高细胞MICA表达率,且呈浓度正相关性。同时,枸杞多糖能提高机体内NK细胞的杀伤活性,能清除微小病灶。     

Resistance of some leukemic blasts to lysis by lymphokine activated killer (LAK) cells

Peripheral blood mononuclear cells (PBMC) from healthy donors and AML patients in remission were stimulated with phytohemagglutinin (PHA) and recombinant interleukin-2 (IL-2). These stimulated cells (lymphokine activated killer (LAK) cells) showed increased DNA synthesis as measured by 3H-Thymidine uptake. A synergistic effect of PHA and IL-2 was found. LAK cells' ability to kill acute myeloid leukemia (AML) blasts was investigated by the 51Cr release assay. LAK cells showed a cytotoxicity (over 10% specific 51Cr release) against 9/12 leukemic blasts, even at effector/target (E/T) ratios as low as 5:1. However, on average only 22.2% (SD 11.8) and 36.5% (SD 12.5) 51Cr release were obtained in 4- and 18-hour cytotoxicity assays, respectively, at an E/T ratio of 20:1. Leukemic blasts in 3/12 AML cases and normal PBMC were entirely resistant to lysis, even at an E/T ratio of 80:1. Susceptibility to lysis was not correlated to peanut-agglutinin receptor expression. LAK cells were more cytotoxic towards the K-562 cell line (natural killer activity) than unstimulated PBMC.     

Analysis of natural killer (NK) cell activity and adhesion molecules on NK cells from umbilical cord blood

The activity of natural killer (NK) cells in human umbilical cord blood (CB) has been reported to be low, compared with that in adult peripheral blood (PB) in vitro. To examine the cause of this, after dividing the CD56+/CD3- cells in CB and PB into CD56bright and CD56dim NK cells, the NK cell activities and the expression of various surface antigens were assayed for each fraction. The NK cell activity of CD56dim NK cells in CB was significantly lower than that in PB (P = 0.0003), whereas, there was no significant difference between the NK cell activity of CD56bright NK cells in PB and CB. The expression levels of adhesion molecules (CD2, CD11a, CD18, DNAX accessory molecule-1), CD16, and CD57 for CD56dim NK cells in CB were significantly lower than those in PB, and approximately one-third of CB CD56dim NK cells were capable of forming conjugates with K562 cells, compared with PB CD56dim NK cells. Furthermore, the inhibition of both the NK cell activities and binding of CD56dim NK cells in PB and CB by monoclonal antibody against each of these adhesion molecules suggests that they play an important role in NK cell activity. These findings show that the low NK cell activity in CB is caused by the low NK cell activity of CD56dim NK cells and that the low expression level of adhesion molecules on CB CD56dim NK cells may contribute to this low NK cell activity.     

Characterization of CD56-/CD16+ natural killer (NK) cells: a highly dysfunctional NK subset expanded in HIV-infected viremic individuals

Natural killer (NK) cells are an important component of the innate immune response against viral infections. NK cell-mediated cytolytic activity is defective in HIV-infected individuals with high levels of viral replication. In the present study, we examined the phenotypic and functional characteristics of an unusual CD56(-)/CD16(+) (CD56(-)) NK subset that is greatly expanded in HIV-viremic individuals. The higher level of expression of inhibitory NK receptors and the lower level of expression of natural cytotoxicity receptors observed in the CD56(-) NK fraction compared with that of CD56(+) NK cells was associated with extremely poor in vitro cytotoxic function of this subset. In addition, the secretion of certain cytokines known to be important in initiating antiviral immune responses was markedly reduced in the CD56(-), as compared with the CD56(+) NK cell subset. These data suggest that the expansion of this highly dysfunctional CD56(-) NK cell subset in HIV-viremic individuals largely accounts for the impaired function of the total NK cell population.     

An unusual case of CD4+ CD7+ CD56+ acute leukemia with overlapping features of type 2 dendritic cell (DC2) and myeloid/NK cell precursor acute leukemia

Type 2 dendritic cell (DC2) acute leukemia has been recently described. We report here an unusual case of a 17-yr-old adolescent with overlapping features of DC2 and myeloid/NK cell precursor acute leukemia as defined by Suzuki et al. The patient presented with lymphadenopathy and hepatosplenomegaly without extranodal manifestations in skin or elsewhere. The morphologic, cytochemical and immunophenotypic features were compatible with those described in DC2 acute leukemia, with co-expression of CD4, CD56 and CD123 antigens. The novel markers BDCA-4 and BDCA-2 considered specific for DC2s were co-expressed. However, bright CD7 positivity along with a dim expression of CD33 (57%) and CD117 (27%) were also noted. Additionally, there was bright expression of NG2 monoclonal antibody 7.1, a frequent finding in myeloid/NK cell precursor acute leukemia. The interpretation of the immunophenotypic profile leads to the hypothesis on the existence of borderline cases between DC2 and myeloid/NK cell precursor acute leukemia. Still, other hypotheses can not be overlooked, such as the possibility for a kind of variant monoblastic leukemia or of another rare entity of acute unclassified leukemia.     

妊娠早期孕妇CD56^brightCD16^-uNK、pNK细胞中CD44的表达变化及意义

目的观察妊娠早期妇女CD56^brightCD16^-子宫NK细胞（uNK细胞）和CD56^brightCD16^-外周血NK细胞（pNK细胞）表面cD。的表达变化,探讨CD56^brightCD16^-uNK细胞中CD44表达与母胎界面免疫耐受形成的关系。方法采集20例孕5—9周的正常妊娠妇女蜕膜和外周血以及20例正常非孕健康妇女外周血,用流式细胞仪检测其中CD56^brightCD16^-NK细胞的数量及其表面CD44的表达。结果早期妊娠妇女子宫蜕膜淋巴细胞中CD56^brightCD16^-NK细胞占71．86％±6．22％,CD56^brightCD16^-uNK细胞中CD44的阳性表达率为34．65％±9．96％,显著低于CD56^brightCD16^-pNK细胞的99．55％±0．35％,P〈0．05。非孕期妇女CD56^brightCD16^-pNK细胞CD44阳性表达率为99．66％±0．29％,与妊娠早期孕妇相近,但孕早期妇女外周血CD56^brightCD16^-NK细胞CD44平均荧光强度（MFI）为241．71±62．41,正常未孕妇女为354．62±41．28,两者相比P〈0．05。结论妊娠早期子宫蜕膜的淋巴细胞主要为CD56^brightCD16^-uNK细胞。CD56^brightCD16^-uNK细胞中CD44的表达明显低于pNK。这可能是妊娠早期维持母胎界面免疫耐受的重要因素。     

Defective natural killer (NK) cell-mediated cytotoxicity does not imply clonal involvement of NK cells in myelodysplastic syndromes

SUMMARY. The clonality of purified cells was examined in 10 myelodysplastic syndromes (MDS) patients by analysing the restriction fragment length polymorphism and methylation pattern of the phosphoglycerate-kinase gene. Natural killer (NK) cell-mediated cytotoxicity was also examined. The granulocytes and monocytes were monoclonal or oligoclonal in all cases, except for the monocytes in one case. Conversely, the NK and T cells had a polyclonal pattern in most cases, including all cases who had defective NK cellmediated cytotoxicty. The hypothesis that reduced NK cellmediated cytotoxicity in MDS is caused by a clonal involvement of NK cells was not supported by the present study.     

扩张型心肌病患者外周血CD4＋CD25＋T细胞变化及临床意义

目的探讨扩张型心肌病（DCM）患者外周血CD4＋CD2＋5T细胞变化及临床意义。方法采用流式细胞分析法检测30例DCM患者（DCM组）及20例健康者（对照组）的外周血CD4＋CD2＋5T细胞。结果外周血CD4＋CD2＋5T细胞占CD4＋T细胞的比例DCM组为（8.53±1.64）%,对照组为（11.4±2.17）%,两组比较有统计学差异（P〈0.01）;且DCM患者心功能越差,CD4＋CD2＋5T细胞占CD4＋T细胞的比例越低。结论DCM患者CD4＋CD2＋5T细胞比例减少,可能打破自身免疫耐受,发生针对心肌抗原的免疫反应,参与DCM发病。     

急性白血病患者调节性T细胞的检测及临床意义

目的:探讨急性白血病患者外周血中CD4+CD25+调节性T细胞(Treg)的变化及意义。方法:采用流式细胞技术及ELISA方法检测56例急性白血病患者外周血中CD4+CD25+ Treg、IL-10的变化,并与健康对照组进行比较。结果:急性白血病患者外周血中CD4+CD25+/CD4+CD25high Treg占(23.85±6.85)%及(5.21±1.20)%,明显高于健康对照组(P0.01);56例急性白血病患者血清中IL-10水平为(74.4±21.2)pg/ml,明显高于正常对照组(24.6±8.7)pg/ml(P0.01),与CD4+CD25+/CD4+CD25 high Treg存在正相关(r=0.253/r=0.511,P0.05)。结论:急性白血病患者外周血CD4+CD25+/CD4+CD25 high Treg水平明显升高,与急性白血病患者免疫逃逸密切相关。