急性双表型白血病的临床及生物学特征研究

目的分析急性双表型白血病（BAL）的临床、生物学特征及治疗疗效。方法对40例急性双表型白血病患者的临床和实验室资料,应用流式细胞术进行免疫分析,采用直接培养法或24/48h短期培养法G显带技术检测染色体。应用兼顾髓系和淋系的V（M）DALP方案联合化疗,Ph阳性患者部分加用伊马替尼治疗。结果符合急性双表型白血病诊断的40例,其中男26例,女14例,发病时WBC≥30×10^9/L者20例（50.0%）,WBC≥100×10^9/L者6例（15.0%）,合并肝、脾肿大者16例（40.0%）,2例患者（5.0%）合并中枢神经系统白血病;以同时表达髓系和B系抗原者最常见,占72.5%,髓系和T系抗原共同表达者占25.0%,B系和T系抗原共同表达者占2.5%。急性双表型白血病患者中CD34阳性高表达占80.0%;对30例患者进行了染色体核型分析,正常核型者13例,占43.3%,异常核型者17例,占56.7%,其中9例（30.0%）Ph染色体阳性表达;40例患者完全缓解19例（47.5%）,9例Ph染色体阳性患者完全缓解3例（33.3%）。结论急性双表型白血病高白细胞者多见,以表达髓系和B系抗原共表达者最常见,异常核型发生率高,Ph染色体阳性者多见,治疗效果较差,预后不良。     

43例鼻咽,鼻腔恶性淋巴瘤的临床病理及免疫表型研究

本文对遵义地区４３例鼻咽、鼻腔恶淋巴瘤的临床病理及免疫学表型进行了研究，结果显示：全部病例均为弥漫型，无１例滤泡型，组织学类型以多表细胞性淋巴瘤为主，共３３例（包括小细胞型１３例，中多形１５例，大多形５例），占７６．７％。用ＵＣＨＬ－１、ＣＤ２０和Ｍａｃ３８７等多单克隆抗体进行免疫表型研究。显示Ｔ细胞淋巴瘤３７例（８６％），Ｂ细胞淋巴瘤４例（９．３％），无１例组织细胞性淋巴瘤。鼻咽部Ｔ．Ｂ淋巴瘤比     

Natural killer lymphocyte blast crisis of chronic myelogenous leukemia

We describe for the first time a case report documenting a chronic myelogenous leukemia (CML) patient who developed a blast crisis of natural killer (NK) lymphocytes. Many of the blasts exhibited large granular lymphocytic (LGL) morphology. Single parameter immunophenotyping results determined that the granulated as well as the agranulated blast cells were NK lymphocytes (CD45, NKH1, CD2, LEU 17, and CD16 positive; CD3, CD8, and LEU 7 negative). Dual parameter flow cytometric testing also determined that some of the blasts expressed the CD11b and CD11c markers as reported for some types of NK lymphocytes. Approximately 10% of the cells were in the S phase of the cell cycle as determined by a modified Vindelov DNA content analysis test and may theoretically reflect some of those cells expressing CD11b and CD11c. The cells did not express in vitro NK lymphocyte functional activity against a K562 target and therefore similar to other reported cases of presumably immature NK lymphocytic leukemias. The NK lymphocyte blast crisis was successfully treated with vincristine and prednisone. The patient's disease eventually relapsed and transformed to a progenitor stem cell before she died (CD45, 13, CD38, and CD34 positive). The flow cytometric immunophenotyping results contributed significantly as an important adjunct in determining the appropriate diagnosis, helping to select the type of therapy, and monitoring the patient with this unusual type of blast crisis.     

Lymphocyte subset reference ranges in healthy Saudi Arabian children

In an attempt to establish the reference ranges for lymphocyte subsets in children, the distribution of lymphocyte population-bearing surface markers such as CD3 (T cells), CD 19 (B cells), CD4 (T helper/inducer cells), CD8 (T suppressor/cytotoxic cells), and CD 16 and/or CD56 on CD3” cells (NK cells) has been studied among healthy Saudi Arabian infants and children. Normal adult blood donors were used for comparison. Anticoagulat-ed peripheral blood was stained with monoclonal antibodies and the lymphocytes were analyzed by flow cytometry for the expression of the above markers. Absolute and percentage values for most lymphocyte populations differed substantially not only between children and adults but also among children from different age groups. Absolute numbers of all the lymphocyte subsets decreased with age from 1 month to 13 years; the median value declined from 4.1 to 1.9 (T cells), 1.6 to 0.6 (B cells), 0.5 to 0.3 (NK cells), 2.7 to 1.0 (CD4⁺ T cells) and 1.5 to 0.8 times 10³ cells/mm³ (CD8⁺ T cells). HLA-DR⁺ T cell counts changed significantly from 0.3 to 0.2 times 10³cells/mm³ during the same age period. In contrast, the lymphocyte percentage increased in all the subsets except B cells and CD4⁺ T cells with time. The percentage values increased from 66 to 74 (T cells), 8 to 11 (NK cells), 23 to 39 (CD8⁺ T cells) and 4 to 9 (HLA-DR⁺ T cells). The values changed from 24 to 12 and 46 to 39 for B cells and CD4⁺ T cells, respectively, with age from 1 month to 13 years. The variations in CD4⁺ and CD8⁺ T cells resulted in a decrease in CD4⁺/CD8⁺ ratio from 2.0 to 1.1 with age. These data should be useful as reference values for lymphocyte subsets in various diseases of infants and children.     

Mesenchymal stem cells (MSCs) from scleroderma patients (SSc) preserve their immunomodulatory properties although senescent and normally induce T regulatory cells (Tregs) with a functional phenotype: implications for cellular‐based therapy

Common variable immunodeficiency (CVID) is the most common severe primary immunodeficiency, but the pathology of this condition is poorly understood. CVID involves a defect in the production of immunoglobulin from B cells, with a subsequent predisposition to infections. Approximately 10–20% of cases are inherited, but even in families with a genetic defect the penetrance is far from complete. A classification system for CVID has been suggested (EUROclass) based on B cell immunophenotyping, but it has not been shown that altered B cell immunophenotype is not a consequence of the complications and treatment of CVID. This study compares the EUROclass B cell immunophenotype of CVID patients (n = 30) with suitable disease controls with bronchiectasis (n = 11), granulomatous disease (Crohn's disease) (n = 9) and neurological patients on immunoglobulin treatment (n = 6). The results of this study correlate with previous literature, that alterations in B cell immunophenotype are associated strongly with CVID. Interestingly, three of the 11 bronchiectasis patients without known immunodeficiency had an altered B cell immunophenotype, suggesting the possibility of undiagnosed immunodeficiency, or that bronchiectasis may cause a secondary alteration in B cell immunophenotype. This study showed a significant difference in B cell immunophenotype between CVID patients compared to disease control groups of granulomatous disease and immunoglobulin treatment. This suggests that granulomatous disease (in Crohn's disease) and immunoglobulin treatment (for chronic neurological conditions) are not causal of an altered B cell immunophenotype in these control populations.     

人外周血淋巴细胞体外扩增培养前后CD25、CD29、HLA-DR系列表型研究

目的观察35例恶性肿瘤患者外周血淋巴细胞在体外经扩增培养后细胞CD25、CD29、HLA—DR系列表型的变化。方法体外诱导扩增恶性肿瘤患者外周血淋巴细胞及单个核细胞,应用流式细胞术测定扩增前后CD25^＋、CD29^＋、HLA—DR^＋、CD4^＋CD25^＋、CD4^＋CD29^＋、CD3^＋HLA—DR^＋CD3^＋HLA-DR^-、CD4^＋HLA—DR^＋、CD4^＋HLA—DR^-、CD8^＋HLA—DR^＋、CD8^＋HLA—DR^-细胞百分比的变化。结果经体外扩增培养后,CD25^＋,CD29^＋,CD3^＋HLA—DR^＋,CD8^＋HLA-DR^＋和CD8^＋HLA—DR^-细胞比例较培养前明显增加（P〈0．01）;而CD4^＋CD25^＋,CD4^＋CD29^＋CD4^＋HLA—DR^＋和CD4^＋HLA—DR^-细胞比例则明显降低（P〈0．01）;HLA—DR^＋细胞和CD3^＋HLA—DR^＋细胞培养前后细胞比例变化无统计学意义（P〉0．05）。结论经体外扩增培养后,CD4^＋CD25^＋调节性T细胞（Treg）细胞比例降低,活化性T细胞的比例与培养前无统计学差异。     

Cytometry by time of flight identifies distinct signatures in patients with systemic sclerosis,systemic lupus erythematosus and Sjögrens syndrome

Systemic sclerosis (SSc), systemic lupus erythematosus (SLE) and primary Sjögrens syndrome (pSS) are clinically distinct systemic autoimmune diseases (SADs) that share molecular pathways. We quantified the frequency of circulating immune-cells in 169 patients with these SADs and 44 healty controls (HC) using mass-cytometry and assessed the diagnostic value of these results. Alterations in the frequency of immune-cell subsets were present in all SADs compared to HC. Most alterations, including a decrease of CD56^hi NK-cells in SSc and IgM⁺ Bcells in pSS, were disease specific; only a reduced frequency of plasmacytoid dendritic cells was common between all SADs Strikingly, hierarchical clustering of SSc patients identified 4 clusters associated with different clinical phenotypes, and 9 of the 12 cell subset-alterations in SSc were also present during the preclinical-phase of the disease. Additionally, we found a strong association between the use of prednisone and alterations in B-cell subsets. Although differences in immune-cell frequencies between these SADs are apparent, the discriminative value thereof is too low for diagnostic purposes. Within each disease, mass cytometry analyses revealed distinct patterns between endophenotypes. Given the lack of tools enabling early diagnosis of SSc, our results justify further research into the value of cellular phenotyping as a diagnostic aid.     

Relevance of immunophenotypes to prognostic subgroups of age,WBC, platelet count,and cytogenetics in de novo acute myeloid leukemia

Li X, Li J, Du W, Zhang J, Liu W, Chen X, Li H, Huang S, Li X. Relevance of immunophenotypes to prognostic subgroups of age, WBC, platelet count, and cytogenetics in de novo acute myeloid leukemia. APMIS 2010. Immunophenotyping is one of the independent prognostic factors in acute myeloid leukemia (AML). Relevance of immunophenotypes to prognostic subgroups of age, white blood cells (WBC), platelet count, and cytogenetics in de novo AML was comprehensively investigated in this study for the first time. Human leukocyte antigen (HLA)‐DR and CD14 expression associated with the elderly, highest WBC count, and unfavorable‐risk cytogenetics; CD4, CD7, and CD11b expression correlated with highest WBC count and unfavorable‐risk cytogenetics; CD64 expression was associated with higher WBC count while that of CD13 was associated with lower platelet count; CD22, CD34, CD123, and terminal deoxynucleotidyl transferase (TdT) expression correlated with unfavorable‐risk cytogenetics; CD5 expression was associated with normal platelet count while that of CD19 was associated with children and favorable‐risk cytogenetics; CD117 expression was associated with low WBC and lower platelet counts; myeloperoxidase (MPO) expression correlated with lower platelet count; and MPO and glycophorin A (Gly‐A) expression was associated with lower WBC count and favorable‐risk cytogenetics. The results of the relevance analysis revealed the distribution characteristics of antigen expression in different AML prognostic subgroups. The majority of antigens associated with good or poor prognostic subgroups were in accordance with the previous reports of correlation of expression of these antigens with prognosis. Antigens associated with good (or poor) prognostic subgroups were defined as good (or poor)‐risk antigens.     

T、B细胞表型抗体免疫组化敏感性与特异性分析

目的：探讨Ｔ、Ｂ细胞非霍奇金淋巴瘤免疫表型染色的特异性和交叉反应性。方法：应用Ｓ－Ｐ免疫组织化学方法对６２例非霍奇金淋巴瘤进行ＣＤ２０、ＣＤ７４、ＣＤ４５ＲＯ、ＣＤ４３和ＣＤ３免疫表型测定。结果：发现Ｂ细胞表型抗体ＣＤ２０和ＣＤ７４对Ｂ细胞非霍奇金淋巴瘤染色阳性率达８８．２％～１００％,但ＣＤ７４的特异性不强,与Ｔ细胞有２６．７％（４／１５）～３５．７％（５／１４）的交叉反应,而ＣＤ２０表现出高度的特异性,交叉反应为０。Ｔ细胞表型抗体ＣＤ４５ＲＯ、ＣＤ４３和ＣＤ３对Ｔ细胞非霍奇金淋巴瘤染色阳性率达８５．７％～１００％,但ＣＤ４５ＲＯ和ＣＤ４３对Ｂ细胞的交叉反应高达４０％（６／１５）～４７．１％（８／１７）,特异性不强,而多克隆抗体ＣＤ３与Ｂ细胞仅有５．９％（１／１７）的交叉反应,表现出明显的特异性。结论：提示在进行Ｔ、Ｂ细胞免疫表型划分时,应使用一组包括ＣＤ２０、ＣＤ７４,ＣＤ４５ＲＯ、ＣＤ４３和ＣＤ３等抗体,ＣＤ３是一种具有高度敏感性和高度特异性的Ｔ细胞标志抗体,值得推荐。     

The role of flow cytometric immunophenotyping in improving the diagnostic accuracy in referred fine-needle aspiration specimens

Flow cytometric (FCM) immunophenotyping has an important role in the diagnostic work up of fine-needle aspiration (FNA) specimens obtained from lymphoid lesions. The objective of the present study was to evaluate the feasibility of this method with respect to referred FNA specimens. One hundred and two FNA specimens referred to our laboratory for FCM analysis during the last 3 years were studied. Specimens were sent, accompanied by cytological smears, from 11 distant hospitals by ordinary mail. The evaluation of potential B-cell monoclonality, the main diagnostic issue to be resolved using FCM, was possible in 86 of these 102 cases. The remaining 16 samples could not be analyzed or adequately interpreted because of sparse or necrotic material. A monoclonal B-cell population was found in 17/86 satisfactory cases, of which 16 were histologically confirmed. Eight cases contained cells positive for the epithelial marker Ber-EP4 and were diagnosed accordingly as carcinomas. FCM analysis of specimens obtained with a clinical question of Hodgkin lymphoma or T-cell lymphomas did not yield definitive data. The time lapse between sampling and analysis (12-84 hr) did not affect the results. This probably was due to the fact that all aspirates were taken in Roswell Park Memorial Institute (RPMI) cell medium, supplemented with 50% fetal calf serum. In conclusion, this retrospective study establishes that it is possible, in the majority of cases, to refer FNA material for FCM immunophenotyping by mail, and that results regarding B-cell clonality in the case of small-cell lymphomas are reliable also after a transportation period of 3-4 days. Carcinoma may be similarly diagnosed and a diagnosis of lymphoma may be excluded in reactive proliferations. Cases with only a few atypical cells or specimens from patients suspected of having Hodgkin lymphoma or T-cell lymphomas are not suitable for analysis by FCM.