Hematopoietic neoplasm coexpressing CD4 and CD56 includes a subset of acute myeloid leukemia with myelomonocytic differentiation, plasmacytoid monocyte tumor, and other immature hematopoietic neoplasms of undefined origin. Herein, we report a CD4+CD56+CD68+ hematopoietic tumor that was thought to be a tumor of plasmacytoid monocytes. This case is unique in the absence of accompanying myelomonocytic leukemia and the faint expression of cCD3 on the tumor cells. The patient was a 22-yr old man presented with multiple lymphadenopathy and an involvement of the bone marrow. Tumor cells were large and monomorphic with an angulated eosinophilic cytoplasm of moderate amount. Nuclei of most tumor cells were eccentric and round with one or two prominent nucleoli. Rough endoplasmic reticulum was prominent in electron microscopic examination. Tumor cells expressed CD4, CD7, CD10, CD45RB, CD56, CD68, and HLA-DR and were negative for CD1a, CD2, sCD3, CD5, CD13, CD14, CD20, CD33, CD34, CD43, CD45RA, TIA-1, S-100, and TdT. cCD3 was not detected in the immunostaining using paraffin tissue, but was faintly expressed in flow cytometry and immunostaining using a touch imprint slide. T-cell receptor gene rearrangement analysis and EBV in situ hybridization showed negative results. Cytochemically, myeloperoxidase, Sudan black B, and alpha naphthyl butyrate esterase were all negative.
Adult ; Antigens, CD/*biosynthesis ; Antigens, CD3/*biosynthesis ; Antigens, CD4/*biosynthesis ; Antigens, CD45/biosynthesis ; Antigens, CD56/*biosynthesis ; Antigens, Differentiation, Myelomonocytic/*biosynthesis ; Bone Marrow Cells/pathology ; Cell Nucleus/pathology ; Eosinophils/metabolism ; Flow Cytometry ; Gene Rearrangement ; Humans ; Immunohistochemistry ; In Situ Hybridization ; Leukemia, Lymphocytic, Chronic, B-Cell/diagnosis/*metabolism ; Lymph Nodes/pathology ; Male ; Microscopy, Electron ; Monocytes/*metabolism ; Receptors, Antigen, T-Cell/metabolism

OBJECTIVESTo study the relationship between the mutation of Leu60Val in HBV core region and the cellular immunity in patients with chronic hepatitis B (CHB).

METHODSHBV DNA C gene mutation was confirmed by polymerase chain reaction (PCR) and sequencing the products directly. The cytokines (IFN-gamma, TNF-alpha and IL-2) levels in serum were measured by enzyme linked immunosorbent assay (ELISA). The distribution of T-lymphocyte subpopulations in peripheral blood was detected by flow cytometry (FCM).

RESULTSThe mutation of Leu60Val was found in 19 out of the 91 CHB patients. With the CHB severity, the mutation rate was getting higher, especially in the severe hepatitis group. The IFN-gamma and TNF-alpha levels were much higher in mutant strain group than those in wild strain group (t=2.584, 4.766, P<0.01), so was the ratio of CD4+/CD8+ (t=2.275, P<0.05).

CONCLUSIONThe mutant strain of 60Val may increase affinity to HLA-I molecule, or up-regulate the expression of HLA-I molecule, resulting in the activation of CTL to release the cytokines and cause immune response in liver.

Adult ; Aged ; CD4-CD8 Ratio ; Flow Cytometry ; Hepatitis B Core Antigens ; genetics ; Hepatitis B, Chronic ; immunology ; virology ; Histocompatibility Antigens Class I ; metabolism ; Humans ; Interferon-gamma ; biosynthesis ; Middle Aged ; Mutation ; T-Lymphocytes, Cytotoxic ; immunology ; Tumor Necrosis Factor-alpha ; biosynthesis

OBJECTIVETo investigate the effect of mitogen Phorbol 12-myristate 13-Acetate (PMA) on CD3, CD4 and CD8 expression of human T-lymphocytes.

METHODSPeripheral blood mononuclear cells from 37 blood samples stimulated in vitro with PMA at different concentrations (2,5,10,20 and 50 ng/ml for 4 hours) and time (10 ng/ml for 2,4 and 6 hours) were analyzed by 4-color flow cytometry (FCM).

RESULTSUnder different PMA stimulation protocols,significant CD4 down-regulation was observed,which was negatively correlated with intracellular cytokine secretion (r= 0.601,P<.001), except for PMA stimulation at 10 ng/ml for 2-hours which showed no significant intracellular cytokine secretion. The expressions of CD3 and CD8 molecules after PMA activation were not significantly affected as compared with pre-activation. Among CD3 positive T lymphocytes, CD4/CD8 double-negative cells only account for 5.52%.

CONCLUSIONPMA has a significant down-regulation effect on CD4 molecules of Th cells, without altering the CD3 and CD8 expression. For quantitative analysis of Th1/Th2 variation, indirect method such as CD3(+)CD8( ) T cells can be used to define CD4(+) Th cells stimulated by PMA in the future.

CD3 Complex ; blood ; CD4 Antigens ; blood ; CD8 Antigens ; blood ; Child ; Humans ; Interferon-gamma ; biosynthesis ; Interleukin-4 ; biosynthesis ; Lymphocyte Activation ; drug effects ; T-Lymphocytes ; chemistry ; drug effects ; Tetradecanoylphorbol Acetate ; pharmacology ; Th1 Cells ; immunology ; Th2 Cells ; immunology

OBJECTIVETo study the influence of percutaneous microwave ablation (PMA) and surgical resection for patients with small primary hepatocellular carcinoma (PHC) on dissemination of tumor cells in peripheral blood determined by AFP mRNA.

METHODSForty patients with small PHC (The maximal diameter < or = 5 cm) confirmed histologically were included in this study. All the patients had single tumor nodule only without metastasis. Of the 40 patients, 19 were treated by PMA and 21 by surgical resection. Blood samples were collected and tested immediately before treatment, 30 min after the mass ablated/resected, 1 d and 7 d later by RTD-Nested-RT-PCR for AFP mRNA. The CD3, CD4, CD8 and CD4/CD8 in blood, and hepatic function were tested at the same time points as well.

RESULTSAfter treatment, ALT and AST in peripheral blood increased in both groups, but more intensely in the surgical group. The CD3, CD4 and CD4/CD8 in peripheral blood decreased at 30 min, 1 day and 7 days after surgical resection, and the lowest value was at 30 min after surgery. The immune function was kept at the same level as pre-treatment in the PMA group. AFP mRNA copies in blood could be detected in 27 of 40 patients (67.5%) in two groups before treatment, and the copy number was increased after treatment. There was no significant difference between the two groups. The patients were followed up for 1 - 16 months. AFP mRNA copies in blood could be detected persistently in the 4 patients with extrahepatic metastasis or liver recurrence.

CONCLUSIONSurgical resection and microwave ablation may cause PHC cells dissemination into the blood circulation in patients with small PHC, and there was no difference between the two treatment groups. The cellular immune function in peripheral blood is decreased after surgical resection, but is maintained at the same level as pre-treatment in the PMA group. The impairment of liver function is less severe after PMA treatment than surgical resection. PMA may provide certain value for clinical management of small hepatocellular carcinoma.

Adult ; Aged ; CD3 Complex ; blood ; CD4 Antigens ; blood ; CD4-CD8 Ratio ; CD8 Antigens ; blood ; Carcinoma, Hepatocellular ; blood ; surgery ; therapy ; Catheter Ablation ; methods ; Female ; Follow-Up Studies ; Hepatectomy ; Humans ; Liver Neoplasms ; blood ; surgery ; therapy ; Male ; Microwaves ; therapeutic use ; Middle Aged ; Neoplasm Recurrence, Local ; RNA, Messenger ; biosynthesis ; genetics ; alpha-Fetoproteins ; biosynthesis ; genetics

Severe chronic active Epstein-Barr virus (EBV) infection is a rare and life-threatening illness. Although the criteria for diagnosis include chronic or recurrent infectious mononucleosis-like symptoms lasting more than 6 months and high titers of anti-EBV antibodies, clinical and laboratory findings may be heterogeneous and flexible application of those criteria is necessary in cases showing typical clinical and pathologic findings. We report a case of severe chronic active EBV infection in a 62-yr-old female patient who showed classical clinical findings with infiltration of EBV-infected T lymphocytes in the bone marrow, spleen, and lymph nodes, and died four months after presentation.
Antigens, CD3/biosynthesis ; Antigens, CD4/biosynthesis ; Antigens, CD8/biosynthesis ; Bone Marrow Cells/virology ; Epstein-Barr Virus Infections/*diagnosis/*mortality ; Female ; Herpesvirus 4, Human/genetics ; Human ; Immunohistochemistry ; Lymph Nodes/virology ; Lymphocytes/metabolism ; Middle Aged ; Organ Weight ; Spleen/pathology/virology ; Support, Non-U.S. Gov't ; T-Lymphocytes/virology

T cell anergy has been successfully induced under different conditions in cloned CD4(+) T cells, but induction of T cell anergy in vivo has been difficult and controversial. Due to the low frequency of naturally occurring T cell population with specificity to a defined antigen, it is very difficult to study anergy of naïve T cells without prior in vivo priming which complicates the interpretation of experimental data. To solve this problem, we adopted the HNT-TCR transgenic mice which have homogeneous antigen specific CD4(+) T cell population. In this study, we generated an influenza virus hemagglutinin (HA) peptide-specific CD4(+) T cell clone from the HNT-TCR transgenic mice and induced anergy using APCs which were treated with the crosslinker, ECDI (1-ethyl-3-3(3-dimethylaminopropyl) carbodiimide). The proliferative response of the cloned or freshly purified naïve CD4(+) transgenic T cells after treatment with ECDI-treated APCs and the HA peptide antigen was monitored as the index of anergy induction. The results showed that anergy was successfully induced in the cloned HNT-TCR transgenic CD4(+) T cells. It was determined that the induced anergy was antigen- and MHC-specific. By contrast, anergy was not observed in freshly purified naïve CD4(+) transgenic T cells under the same conditions. The results suggest that naïve CD4(+) T cells may have different anergy inducing requirements, or that cloned CD4(+) T cells may have certain priming or in vitro cloning artifact which makes them more susceptible to anergy induction. We propose that induction of T cell anergy may depend on the T cell growth, activation and differentiation state or cloning conditions. The results from the present study may have important implications for the study of the mechanism(s) underlying T cell anergy induction in vivo and for applications of immune tolerance based therapy.
Animals ; Antigen-Presenting Cells ; immunology ; metabolism ; Antigens, CD ; genetics ; immunology ; metabolism ; CD4 Antigens ; immunology ; CD4-Positive T-Lymphocytes ; cytology ; immunology ; Clonal Anergy ; genetics ; immunology ; Clone Cells ; immunology ; Epitopes, T-Lymphocyte ; biosynthesis ; Immune Tolerance ; genetics ; Major Histocompatibility Complex ; immunology ; Mice ; Mice, Transgenic ; Receptors, Antigen, T-Cell ; physiology

Human immunodeficiency virus (HIV) infects the host cells by the fusion of viral and cell membranes. Blocking the combining between HIV and the receptors can prevent HIV from entering the host cells. We designed an invasion-inhibitor for HIV-1 targeting dendritic cell (DC), including 2 important HIV-1 receptors CD4 and CCR5, and 2 molecules Flt3-L and Mip-3alpha. With the synthetic gene of the inhibitor, 2 eukaryotic expression vectors pABK-CKR5-CD4/Flt3L-Mip3alpha (pABK-HIV-MF) and pABK-CKR5-CD4 (pABK-HIV-MT) were constructed and transfected into HEK 293 cells for expression. Results from RT-PCR, immunofluorescent assay, ELISA and Western blot approved that the invasion-inhibitor for HIV-1 was successfully and exactly expressed in the eukaryotic cells. Current study formed a solid base for the further research on the function of inhibitors for HIV-1 and elimination targeting DC.
Artificial Gene Fusion ; CD4 Antigens ; biosynthesis ; genetics ; Chemokine CCL20 ; biosynthesis ; genetics ; Dendritic Cells ; immunology ; metabolism ; Genetic Vectors ; genetics ; HEK293 Cells ; HIV Envelope Protein gp120 ; genetics ; HIV-1 ; physiology ; Humans ; Receptors, CCR5 ; biosynthesis ; genetics ; Receptors, HIV ; antagonists & inhibitors ; Transfection ; fms-Like Tyrosine Kinase 3 ; biosynthesis ; genetics

Increasing importance is being given to the stimulation of Th1 response in cancer immunotherapy because its presence can shift the direction of adaptive immune responses toward protective immunity. Based on chemokine receptor expression, CXCR3+CCR4-CD4+ T cells as Th1-type cells were investigated its capacity in monocyte-derived dendritic cell (DC) maturation and polarization, and induction of antigen specific cytotoxic T lymphocytes (CTL) in vitro. The levels of IL-4, IL-5 and IL-10 were decreased to the basal level compared with high production of IFN-gamma, TNF-alpha, and IL-2 in CXCR3+CCR4-CD4+ T cells stimulated with anti-CD3 and anti-CD28 antibodies. Co-incubation of activated CD4+ or CXCR3+CCR4-CD4+ T cells with DC (CD4+/DC or CXCR3+CD4+/DC, respectively) particularly up-regulated IL-12 and CD80 expression compared with DC matured with TNF-alpha and LPS (mDC). Although there was no significant difference between the effects of the CXCR3+CCR4-CD4+ and CD4+ T cells on DC phenotype expression, CXCR3+CD4+/DC in CTL culture were able to expand number of CD8+ T cells and increased frequencies of IFN-gamma secreting cells and overall cytolytic activity against tumor antigen WT-1. These results demonstrated that the selective addition of CXCR3+CCR4-CD4+ T cells to CTL cultures could enhance the induction of CTLs by DC in vitro, and implicated on a novel strategy for adoptive T cell therapy.
Antigens, CD4/*immunology ; Cell Line ; Cells, Cultured ; Cytokines/immunology ; Cytotoxicity, Immunologic ; Dendritic Cells/cytology/*immunology ; Humans ; Interferon-gamma/biosynthesis ; Receptors, CCR4/*immunology ; Receptors, CXCR3/*immunology ; T-Lymphocytes, Cytotoxic/*cytology/immunology ; Th1 Cells/*immunology

The aim of this study was to separate and amplify CD4(+)CD25(+)Treg cells from splenocytes of sensitized mice. The percentage of CD4(+)CD25(+)Treg cells was detected by flow cytometry in sensitized and normal control mice. CD4(+)T, CD4(+)CD25(+)Treg and CD4(+)CD25(-) T cells were isolated from mouse splenocytes by MACS. CD4(+)CD25(+)Treg cells were expanded in vitro cultures in addition of CD3/CD28 MACSiBead and IL-2. The activity of cells was detected with 0.4 trypan blue staining. The purity of cells after sorting, the main surface marker and the level of Foxp3 were detected by flow cytometry. The results showed that CD4(+)CD25(+)Treg cell proportion was higher in sensitized mice than normal control mice (P < 0.05). The average purity of CD4(+)CD25(+)Treg cells was 87. The activity of these cells was more than 97, and the expression of Foxp3 in these cells was high. The amplification multiples achieved 42 times after 2 weeks in vitro. The percentage of CD4(+)CD25(+) regulatory T cells was 85.32, and the expression of Foxp3 decreased from (76.92 ± 1.72) to (75.33 ± 2.11) (P > 0.05). It is concluded that the sorting of CD4(+)CD25(+)Treg cells is isolated successfully by MACS without affecting the vitality of target cells. The amplification of CD4(+)CD25(+)Treg cells is successful in vitro. Expression of surface markers and Foxp3 gene does not obviously change after amplification, so that to establish a practical method to recover and enlarge the amount of CD4(+)CD25(+)Treg cells in good condition.
Animals ; CD4 Antigens ; biosynthesis ; Flow Cytometry ; Forkhead Transcription Factors ; metabolism ; Immunomagnetic Separation ; methods ; Interleukin-2 Receptor alpha Subunit ; metabolism ; Lymphocyte Count ; Mice ; Mice, Inbred C57BL ; T-Lymphocytes, Regulatory ; cytology

OBJECTIVETo establish a rapid assay for assessment of drug susceptibility of human immunodeficiency virus type 1 isolates (Recombinant virus assay).

METHODSThis procedure allows the generation of viable virus with SI phenotype by homologous recombination of a RT-PCR-derived pool of reverse transcriptase (RT) coding sequences into an RT-deleted, noninfectious proviral clone, pHIV delta RTBstE II. Then the drug susceptibility of recombinant virus to RT inhibitors can be assessed in the Hela CD4+ plaque reduction assays.

RESULTSAnalysis of 7 HIV strains with SI or NSI phenotype showed that recombinant viruses accurately exhibited the same genotype as that of the original HIV1 isolates. The results of drug susceptibilities of HIV1 isolate got by recombinant virus assay were the same as that by standardized peripheral blood mononuclear cell culture assay.

CONCLUSIONRecombinant virus assay is a rapid and accurate method to assess the drug sensitivity of HIV1 isolates with SI or NSI phenotype.

Antiviral Agents ; pharmacology ; CD4 Antigens ; analysis ; DNA, Viral ; biosynthesis ; HIV Infections ; virology ; HIV-1 ; drug effects ; genetics ; HeLa Cells ; Humans ; Microbial Sensitivity Tests ; methods ; Phenotype ; Recombination, Genetic ; Virus Replication ; drug effects