BACKGROUND: The aberrant, leukemia-associated antigen expression patterns allow us to discriminate leukemic blasts from normal precursor cells. Our major goal was to determine a guideline for the detection of minimal residual disease using CD20+/CD34+ and myeloid Ag+/CD19+ combination in the bone marrow of acute leukemia in complete remission (CR) after chemotherapy. METHODS: Bone marrow samples from 117 patients with acute leukemia in complete remission after chemotherapy and from 22 healthy controls were immunophenotyped by triple staining and measured by flow cytometry. RESULTS: The CD20+/CD34+ cells in the large lymphocyte gate (R1) ranged from 0% to 3.24% (0.8+/-0.82%, P=0.000) in CD20+/CD34+ B-lineage ALL CR (N=31), from 0.03% to 4.2% (0.7+/-0.83%, P=0.000) in CD20-/CD34- B-lineage ALL CR (N=66), from 0.1% to 0.96% (0.45+/-0.32%, P=0.016) in T-ALL CR (N=10), and from 0.02% to 0.48% (0.18+/-0.15%, P=0.776) in AML CR (N=10). The CD13,33+/CD19+ cells in R1 gate ranged from 0% to 2.69% (0.37+/-0.48%, P<0.001) in CD13,33+/CD19+ B-lineage ALL CR (N=31), from 0% to 1.8% (0.31+/-0.28%, P<0.001) in CD13,33-/CD19+B-lineage ALL CR (N=65), from 0.02% to 0.64% (0.29+/-0.22%, P=0.071) in T-ALL CR (N=9), and from 0% to 0.17% (0.07+/-0.09%, P=0.341) in AML CR (N=3). CONCLUSIONS: Using an immunophenotypic method for the detection of early relapse or minimal residual disease of B-lineage ALL bone marrow in CR after chemotherapy, different cutoff values should be applied according to antigen combination and gating. When the proportion of aberrant antigen combination was less than 5% in large lymphocyte gate, the results should be interpreted with caution.
Acute Disease ; Antigens, CD/*metabolism ; Antigens, CD19/metabolism ; Antigens, CD20/metabolism ; Antigens, CD34/metabolism ; Antigens, Differentiation, Myelomonocytic/analysis/metabolism ; Bone Marrow Cells/*classification/metabolism ; Flow Cytometry ; Hematopoietic Stem Cells/classification/metabolism ; Humans ; Immunophenotyping ; Leukemia/*diagnosis/drug therapy ; Leukemia, Myeloid, Acute/diagnosis/drug therapy ; Neoplasm, Residual ; Remission Induction ; Tumor Markers, Biological/immunology

OBJECTIVETo evaluate the therapeutic effect of a new recombinant immunotoxin mMIP-1alpha-DT390 on experimental autoimmune encephalomyelitis (EAE).

METHODSEAE was induced in the low-sensitive strain C57BL/6 mice with intraperitoneal injection of myelin basic protein (MBP) to simulate the human disease multiple sclerosis, followed by intramuscular injection of cationic liposome carrying the plasmid DNA SRalpha-mMIP-1alpha-DT390 in the leg muscle to elicit resistance to EAE development. The mice were then examined daily for clinical signs of EAE by an observer blind to the treatment protocol. For immunohistochemistry the mice were anesthetized and perfused with sterile PBS and paraformaldehyde, and the cerebrum, cerebellum, medulla and spinal cord were removed for preparation of serial sections. The mononuclear cells (MNCs) from the EAE mouse spleens were prepared for three-color flow cytometry analysis of the surface markers with appropriate antibodies following the BD Pharmingen cytokine staining protocol.

RESULTSEAE model was successfully established by active MBP immunization in C57BL/6 mice. Administration of the immunotoxin mMIP-1alpha-DT390 significantly delayed the disease onset and lowered the mean clinical score for EAE as compared with the control mice. Immunohistochemistry demonstrated much less CCR5(+) infiltrating cells in the central nervous system in mMIP-1alpha-DT390-treated mice than in the control. The treatment also eliminated reactive T cells in the periphery blood without affecting the number of B cells.

CONCLUSIONThe immunotoxin mMIP-1alpha-DT390 can attenuate the disease activity of EAE in mice, suggesting its potential use in the treatment of other autoimmune disorders.

Animals ; Antigens, CD19 ; analysis ; B-Lymphocytes ; cytology ; metabolism ; CD3 Complex ; analysis ; Chemokine CCL3 ; genetics ; metabolism ; Diphtheria Toxin ; genetics ; metabolism ; Disease Models, Animal ; Encephalomyelitis, Autoimmune, Experimental ; drug therapy ; Female ; Flow Cytometry ; Immunoglobulin Fragments ; genetics ; metabolism ; Immunohistochemistry ; Immunologic Factors ; therapeutic use ; Immunotoxins ; therapeutic use ; Meninges ; chemistry ; pathology ; Mice ; Mice, Inbred C57BL ; Multiple Sclerosis ; drug therapy ; NIH 3T3 Cells ; Receptors, CCR5 ; analysis ; Recombinant Fusion Proteins ; genetics ; metabolism ; therapeutic use ; T-Lymphocytes ; cytology ; metabolism

The differential diagnosis of acute lymphoblastic leukemia (ALL) from other small round blue cell tumors in children is very important for proper treatment, but sometimes difficult. CD45 is expressed on almost all-human leukocytes and not expressed on other small round blue cell tumors. Moreover, CD19 is expressed on all stages of B lineage cells and loss of this antigen is very rare in precursor B-cell ALL. We report a case of ALL with atypical morphology and immunophenotype. A 6-yr-old girl presented with fever and weight loss. Many abnormal cells with variable sized, high nuclearcytoplasmic ratio and distinct nucleoli were counted 23% in bone marrow. The results of immunophenotyping were negative for CD45, CD19, CD10, CD20, CD3, CD5, CD7, CD56/16, CD13, and CD33 and positive for CD22, TdT, and CD34. The immunohistochemical staining of bone marrow biopsies was positive for CD79a, CD10, TdT and CD99. The cytogenetic study showed normal karyotype but amplification of MLL (myeloid/lymphoid or mixed lineage leukemia) gene was suggestive in the fluorescent in situ hybridization. The patient received the standard chemotherapy for acute lymphoblastic leukemia and reached complete remission.
Acute Disease ; Antigens, CD19/*analysis ; Antigens, CD45/*analysis ; Bone Marrow/*pathology ; Child ; Female ; Humans ; In Situ Hybridization ; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/diagnosis/*pathology

This study was aimed to establish a cytokine-independent human myelodysplastic cells line from bone marrow of a patient with MDS-CMML. This cell line was incubated in mixed culture of RPMI 1640 and DMEM with 15% bovine serum, but without cytokines; its biological characteristics were identified by morphology, surface marker profiles, cell proliferation, differentiation and apoptosis. The results showed that the established cell line could not depend on cytokines for long-term survival and growth, and could differentiate into colony-forming unit-macrophage, colony-forming unit-megakaryocyte. In conclusion, a cytokine-independent human myelodysplastic syndrome cell line, named MDS-JSN04 (MDS Nanjing Jiansu 04), was established. Its partial biological characteristics were identified and clarified.
Antigens, CD19 ; analysis ; Antigens, CD20 ; analysis ; Apoptosis ; drug effects ; Bone Marrow Cells ; metabolism ; pathology ; ultrastructure ; CD79 Antigens ; analysis ; Cell Differentiation ; drug effects ; Cell Line ; Cell Proliferation ; drug effects ; Culture Media ; pharmacology ; Cytokines ; pharmacology ; Flow Cytometry ; HLA-DR Antigens ; analysis ; Humans ; Lewis X Antigen ; analysis ; Male ; Microscopy, Electron, Scanning ; Microscopy, Electron, Transmission ; Middle Aged ; Myelodysplastic Syndromes ; metabolism ; pathology ; Sialic Acid Binding Ig-like Lectin 2 ; analysis ; Time Factors

Primary Sjogren's syndrome (pSS) is a chronic autoimmune disease with welldocumented association of lymphoid malignancies during the progress of the disease. Although several types of malignancy and pseudomalignancy have been reported in pSS, low-grade non-Hodgkin's lymphomas are the most frequently observed. Reactive plasmacytosis mimicking myeloma is a very rare condition in association with pSS. We describe a 72-yr-old woman with pSS who presented with hypergammaglobulinemia, and extensive bone marrow and lymph node plasmacytosis, which mimicked multiple myeloma. In this patient, there was an abnormal differentiation of memory B cells to plasma cells in the peripheral blood suggesting underlying pathogenetic mechanism for this condition.
Aged ; Antigens, CD19/analysis ; Antigens, CD27/analysis ; Bone Marrow Examination ; Diagnosis, Differential ; Female ; Fluorescent Antibody Technique/methods ; Humans ; Multiple Myeloma/*pathology ; Plasma Cells/chemistry/*pathology ; Sjogren's Syndrome/*pathology

OBJECTIVETo study the effect of natural killer (NK)-cell on reconstitution of hematopoiesis and immunity in mouse allogeneic bone marrow transplantation (allo-BMT).

METHODSLethally irradiated BALB/c (H-2(d)) mice were transplanted with C57BL/6 (H-2(b)) bone marrow plus peripheral T cells and/or NK cells. Recipients CD34(+) cells and H-2K(b+), CD3(+) and CD19(+) cells were detected by flow cytometry, peripheral white blood cell (WBC) by auto-cytometry, and the survival rates, engraftment, hematopoietic and immune recovery were observed.

RESULTSIn the transplantation with NK cell infusion group, the survival rates, the WBC and CD34(+) cell counts, and the H-2(b+) and CD19(+) cells were significantly higher than that in without NK cell infusion group (P < 0.01). Twenty-eight days after transplantation, the CD3(+) cells in the NK cell infusion group [(33.69 +/- 3.36)%] were lower than that in without [(50.4 +/- 5.06)%] (P < 0.01), and there was no longer difference between these groups 60 days after transplantation (P > 0.05).

CONCLUSIONIn mouse allo-BMT, alloreactive NK cell enhances engraftment, promotes reconstitution of hematopoiesis and immunity and increases survival rates.

Animals ; Antigens, CD19 ; analysis ; Antigens, CD34 ; analysis ; Bone Marrow Transplantation ; immunology ; methods ; Cells, Cultured ; Female ; Flow Cytometry ; Graft Survival ; immunology ; Hematopoiesis ; immunology ; Killer Cells, Natural ; cytology ; immunology ; transplantation ; Lymphocyte Transfusion ; methods ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Transplantation, Homologous

OBJECTIVETo investigate the expression of CD19 on childhood acute leukemia (AL) and its significance, and to provide evidence for the diagnosis and differential diagnosis as well as monoclonal antibody-targeting treatment of leukemia.

METHODSThere were 210 cases of childhood AL, of which 130 cases were male and 80 were female with a mean age of 9 years old. Using a panel of 27 fluorochrome directly labeled monoclonal antibodies, 210 samples from the patients were analyzed with CD45/SSC double parameters and multi-color flow cytometry to determine the expression of CD19.

RESULTSIn the 93 cases of B lineage acute lymphoblastic leukemia (ALL), the positive rate (98.9%, 92/93) of CD19 was significantly higher than that of the other B cell related antigens, such as CD10 (88.2%, 82/93, P = 0.003), CD20 (24.7%, 23/93, P = 0.001) and CD22 (60.2%, 56/93, P = 0.001). CD19 was expressed on all 8 cases of B/myeloid (My) hybrid acute leukemia (HAL) and 1 case of B/T HAL, but was not expressed on all 24 cases of T lineage leukemia and 5 cases of T/My HAL. In the 79 cases of acute myeloid leukemia (AML), only 5 (6.3%) cases expressed CD19. The positive rate (6.3%) of CD19 on AML was significantly lower than that on B lineage ALL (98.9%, P = 0.001). The percentage of CD19 positive cells in B/My HAL (41.6% - 88.7% with a mean of 73.8%) was significant higher than that in CD19(+)-AML (21.4% - 50.4% with a mean of 24.2%; Run Sum test, P = 0.0084). Of the 210 cases, 102 were B lineage related AL including B lineage ALL, B/My HAL and B/T HAL. In B lineage related AL, the sensitivity and the specificity of CD19 was 99.0% (101/102) and 95.4% (13/108) while the positive predictive and the negative predictive values to B lineage were 95.3% (101/106) and 99.0% (103/104), respectively. Using CD19(+) as a single reagent to diagnose B lineage, the false positive rate was 4.6% (5/108) and the false negative rate was 1.0% (1/102) with a general diagnosis index (GDI) of 94.4% [GDI = 1-(false positive rate + false negative rate)].

CONCLUSIONCD19 is continuously and stably expressed on all stages of B lineage differentiation. It is a reliable cell membrane marker for diagnosing B lineage ALL and an ideal target for antibody-targeting treatment of leukemia as well; the expression degree of CD19 can be used to distinguish B/My HAL from CD19(+)-AML; CD19 didn't express on normal myeloid cells but did on some AML cells. Therefore it can be used to detect the minimal residual disease.

Adolescent ; Antigens, CD19 ; analysis ; Child ; Child, Preschool ; Female ; Flow Cytometry ; Humans ; Infant ; Male ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; classification ; immunology

To investigate the biological features of leukemic cells in bcr/abl fusion transcript-positive B-lineage acute lymphoblastic leukemia (B-ALL), 3- or 4-color flow cytometry with directly conjugated monoclonal antibodies was used to detect the immunophenotype of the cells in 26 patients with bcr/able-positive B-ALL and 32 patients with bcr/abl-negative B-ALL. bcr/abl fusion transcript was detected by RT-PCR. Immunoglobulin heavy chain (IgH) gene rearrangement was detected by PCR. The results showed that all of the B-ALL patients were positive for CD19. There was significant difference in expression of CD34 (96.2% vs 65.6%), CD10 (96.2% vs 71.8%) and CD38 (43.8% vs 95.4%) between bcr/abl-positive and -negative groups. In bcr/abl-positive B-ALL group, the co-expression rates of CD10(+)/CD19(+)/CD34(+), CD10(+)/CD34(+)/HLA-DR(+) and CD10(+)/CD34(+)/CD38(-) were 92.3% (24/26), 73.1% (19/26) and 56.2% (9/16), respectively. In bcr/abl-negative group, co-expression of CD10(+)/CD19(+)/CD34(+) and CD10(+)/CD34(+)/HLA-DR(+) were 43.8% (14/32) and 37.5% (12/32), respectively, there were significant differences (P < 0.05) between bcr/abl-positive and -negative groups, but none of the cases co-expressed CD10(+)/CD34(+)/CD38(-). The detection rate of monoclonal IgH gene rearrangement (58.8%, 10/17) was lower in bcr/abl-positive group than that (85.7%, 12/14) in bcr/able-negative group. It is concluded that the expression rates of CD34 and CD10 are higher, and CD38 and IgH gene rearrangement are lower in bcr/abl-positive B-ALL cases, CD10(+)/CD34(+)/CD38(-) is a unique feature of immunophenotype, and this phenotype of leukemia cells is closer to that of early B-lineage progenitor cells.
Adolescent ; Adult ; Aged ; Antigens, CD19 ; analysis ; Antigens, CD34 ; analysis ; Burkitt Lymphoma ; immunology ; Child ; Female ; Flow Cytometry ; Fusion Proteins, bcr-abl ; genetics ; Humans ; Immunophenotyping ; Leukocyte Common Antigens ; analysis ; Male ; Middle Aged ; Neprilysin ; analysis ; Polymerase Chain Reaction ; RNA, Messenger ; analysis

OBJECTIVETo study the effect on apoptotic of CD4+ T, CD19+ B in spleen of BXSB mice with systemic lupus erythematosus treated with Langchuangjing Granule and to probe into the mechanism of treatment.

METHODSThe apoptosis was examined by the flow cytometric analysis and immunofluorescence double-staining method.

RESULTSApoptosis of male BXSB mice speeds up. Langchuangjing Granule can restrain the excessive apoptosis of CD4+ T and CD19+ B cells in spleen.

CONCLUSIONLangchuangjing Granule treated systemic lupus erythematosus by restraining the excessive apoptotic of T, B lymphocytes, probably restraining the release of excessive amount of apoptotic DNA fragments, so decreasing abnormal proliferation of B cells and the produce of autoantibodies.

Animals ; Antigens, CD19 ; analysis ; Apoptosis ; drug effects ; B-Lymphocytes ; pathology ; CD4-Positive T-Lymphocytes ; pathology ; Drugs, Chinese Herbal ; pharmacology ; Female ; Lupus Erythematosus, Systemic ; immunology ; pathology ; Male ; Mice ; Mice, Inbred Strains ; Random Allocation ; Spleen ; pathology

Antigens, CD19 ; analysis ; Asthma ; blood ; Child ; Child, Preschool ; Female ; Flow Cytometry ; Humans ; Immunoglobulin E ; blood ; Lymphocytes ; immunology ; Male ; Receptors, IgE ; analysis