OBJECTIVETo study the expression and localization of co-stimulators in the mucosa of patients with ulcerative colitis (UC), and to explore its role in the pathogenesis of UC.

METHODSExpression of co-stimulators CD86 and inducible co-stimulator (ICOS) was studied by immunohistochemistry on paraffin-embedded mucosal tissue from patients with active UC (64 cases), inactive UC (51 cases) and normal controls (20 cases). Immunostaining for CD28 was also carried out on frozen fresh mucosal tissue sampled from patients with active UC (7 cases), inactive UC (2 cases) and normal controls (5 cases). In addition, expression of CD4, CD8 and CD20 were also examined.

RESULTSIn active UC, increased expression of CD86 was not only observed in lamina propria mononuclear cells but also in the intestinal epithelial cells, as compared with inactive UC and the normal controls (P < 0.01). Increased ICOS expression in lamina propria mononuclear cells was detected in active UC, as compared with inactive UC and the normal controls (P < 0.01). Increased ICOS expression in intestinal epithelial cells was also seen in active UC, as compared with that of inactive UC (P < 0.01). The expression of CD86 was higher in inactive UC than in the normal controls (P < 0.05 or P < 0.01). However, the expression of ICOS showed no statistically significant difference between inactive UC and normal controls. Increased expression of CD28 in active UC, compared with that in inactive UC and normal controls, was also noticed (P < 0.05 or P < 0.01). The number of CD4 or CD8-positive intraepithelial lymphocytes and lymphocytes infiltrating in the lamina propria and small vessel walls was much higher in active UC than in inactive UC and normal controls (P < 0.01). Moreover, the ratio of CD4/CD8 was highest in active UC (P < 0.01). The number of CD20-positive B lymphocytes in lamina propria was also higher in active UC than in inactive UC and normal controls (P < 0.01).

CONCLUSIONSIn active UC, CD86 and ICOS were over-expressed in the intestinal epithelial cells and lamina propria mononuclear cells. The phenomenon suggests that abnormal expression of co-stimulators may contribute to the deregulation of acquired immune responses in UC.

Adult ; Aged ; Antigens, Differentiation, T-Lymphocyte ; metabolism ; B7-2 Antigen ; metabolism ; CD28 Antigens ; metabolism ; CD4-CD8 Ratio ; Case-Control Studies ; Colitis, Ulcerative ; metabolism ; pathology ; Epithelial Cells ; metabolism ; pathology ; Female ; Humans ; Immunohistochemistry ; Inducible T-Cell Co-Stimulator Protein ; Intestinal Mucosa ; metabolism ; pathology ; Leukocytes, Mononuclear ; metabolism ; pathology ; Male ; Middle Aged ; Mucous Membrane ; metabolism ; pathology ; Young Adult

Since it is difficult to study human thymocyte maturation in vitro, we have developed an in vitro thymocyte culture system which has allowed us to select the optimal growth conditions for thymocyte subpopulations. Three thymocyte subpopulations (CD3-CD1-, CD1+CD3-, and CD3+CD1-) were isolated by a single step percoll density gradient centrifugation and indirect panning procedure using anti-CD1 and anti-CD3 monoclonal antibodies, and their purity was checked by flow cytometry. The combination of concanavalin A (Con A), tetradecanoylphorbol acetate (TPA), and IL-2 was shown to be the most reliable stimulus for the proliferation of CD3-CD1- thymocytes for up to 15 days in a culture system in vitro. Flow cytometric analysis for the phenotypic change of CD3-CD1- thymocytes revealed a steady increase of CD3 antigen after a 3-day cultivation, whereas there was no change in CD1 antigen intensity. A combination of Con A and IL-2 was both sufficient and necessary to induce growth of CD3+CD1- thymocytes. The major population of immature cortical thymocytes (CD3-CD1+ or CD3+CD1+), which are considered to be the most unresponsive dead-end cells, could not be maintained or stimulated with any combination used in this experiment, even in the presence of thymic accessory cells.
Antigens, CD ; Antigens, CD1 ; Antigens, CD3 ; Antigens, Differentiation, T-Lymphocyte ; Cell Cycle ; Cell Division/drug effects ; Cells, Cultured ; Humans ; Infant ; Infant, Newborn ; Ionophores/pharmacology ; *Lymphocyte Activation/drug effects ; Receptors, Antigen, T-Cell ; T-Lymphocytes/*cytology/drug effects/immunology ; Tetradecanoylphorbol Acetate/pharmacology

OBJECTIVE: To explore the role of DNAM-1 in the activation, proliferation and function of type Ⅰ regulatory T cells (Tr1 cells). METHODS: Anti-CD3/CD28 antibodies were used to stimulate mouse T cells derived from the spleen of wild-type (WT) mice, and the expression level of DNAM-1 in resting and activated Tr1 cells was evaluated with flow cytometry. Na?ve CD4⁺ T cells isolated by magnetic cell sorting from the spleens of WT mice and DNAM-1 knockout (KO) mice were cultured in Tr1 polarizing conditions for 3 days, after which CD25 and CD69 expressions were measured using flow cytometry. The induced Tr1 cells were labelled with CFSE and cultured in the presence of anti-CD/CD28 antibodies for 3 days, and their proliferative activity was analyzed. The expressions of IL-10 and p-STAT5 in DNAM-1-deficient Tr1 cells were detected before and after IL-2 stimulation. RESULTS: The expression level of DNAM-1 was significantly upregulated in CD4⁺ T cells and Tr1 cells after stimulation with anti-CD3/CD28 antibodies (P < 0.05). DNAM-1 knockout did not cause significant changes in the number or proportion of Tr1 cells, but but significantly increased the expression levels of the activation markers CD69 and CD25 (P < 0.05). Compared with WT Tr1 cells, DNAM-1-deficient Tr1 cells exhibited reduced proliferative activity in vitro (P < 0.05) with downregulated IL-10 production (P < 0.05) and decreased expressions of Il-10 and Gzmb mRNA (P < 0.05). In DNAM-1-deficient Tr1 cells, IL-2 stimulation significantly reduced IL-10 secretion level and the expression of p-STAT5 as compared with WT Tr1 cells. CONCLUSION DNAM-1 participate in the activation and proliferation of Tr1 cells and affect the biological functions of Tr1 cells through the IL-2/STAT5 pathway.
Animals ; Antigens, Differentiation, T-Lymphocyte ; CD28 Antigens/metabolism* ; Cell Proliferation ; Cells, Cultured ; Interleukin-10 ; Interleukin-2/metabolism* ; Mice ; RNA, Messenger ; STAT5 Transcription Factor/metabolism* ; T-Lymphocytes, Regulatory

Antigens, CD ; metabolism ; Antigens, Differentiation, T-Lymphocyte ; metabolism ; CD3 Complex ; metabolism ; Cells, Cultured ; Humans ; Kinetics ; Lectins, C-Type ; metabolism ; Lymphocyte Activation ; T-Lymphocyte Subsets ; drug effects ; immunology ; metabolism

Idiopathic thrombocytopenia purpura (ITP) is an autoimmune disease which is characterized by destruction of platelets by macrophages in the reticuloendothelial system. Recent studies suggest that ITP is related to the abnormal activation and apoptosis of T/B cells which lead to failure of immune tolerance. Now it is becoming clear that costimulatory signals are required for full T/B cell activation and assumed to modulate T/B cells responses as well as other aspects of the immune system. This review focuses on the role and state-of-the-art advancements of costimulatory signals in ITP.
Antigens, CD ; immunology ; Antigens, Differentiation ; immunology ; Antigens, Differentiation, T-Lymphocyte ; immunology ; B-Lymphocytes ; immunology ; CD28 Antigens ; immunology ; CTLA-4 Antigen ; Humans ; Immune Tolerance ; Inducible T-Cell Co-Stimulator Protein ; Purpura, Thrombocytopenic, Idiopathic ; immunology ; Receptors, Tumor Necrosis Factor ; immunology ; Signal Transduction ; physiology ; T-Lymphocytes ; immunology ; physiology

Cytomegalovirus (CMV) infection is a dangerous complication in patients with chronic graft versus host disease (cGVHD). CMV-specific immunity depends on the activity of T cells. This study was aimed to investigate the effect of CMV pp65 gene modified dendritic cells (DCs) on activation of autologous T cells. Lentivirus system was utilized to introduce the CMV full-length pp65 gene into mouse DCs; CpG-DNA was used to induce mature DCs; flow cytometry and immunofluorescence were used to determine the expression of antigen and IFNgamma in T lymphocytes. The results showed that the DCs were infected with lentivirus at a multiplicity of infection (MOI) of 50 with optimal infectious efficiency of 30%-40%; mature DCs expressing pp65 gene could stimulate autologous naive T cells to express CD69 specifically; mature DCs expressing PP65 could stimulate autologous CD4+ or CD8+ T cells to produce IFNgamma. It is concluded that CMV pp65-modified and CpG-DNA-induced mature DCs can activate CMV-specific T lymphocytes in vitro.
Animals ; Antigens, CD ; genetics ; metabolism ; Antigens, Differentiation, T-Lymphocyte ; genetics ; metabolism ; Antigens, Viral ; immunology ; CD4-Positive T-Lymphocytes ; immunology ; CD8-Positive T-Lymphocytes ; immunology ; CpG Islands ; genetics ; Cytomegalovirus ; immunology ; DNA ; genetics ; Dendritic Cells ; cytology ; immunology ; metabolism ; Humans ; Interferon-gamma ; genetics ; metabolism ; Lectins, C-Type ; Lentivirus ; genetics ; metabolism ; Mice ; Phosphoproteins ; genetics ; metabolism ; Viral Matrix Proteins ; genetics ; metabolism

OBJECTIVETo study the activating effect of protein transduction domain (PTD) mediated BCR/ABL protein on T cells from CML patients.

METHODSThe plasmid containing PTD and b3a2 bcr/abl of CML was constructed by genetic engineering and expressed in E. coli. The peripheral blood mononuclear cells from CML patients were stimulated in vitro with purified PTD-BCR/ABL protein and the expression of the early activation antigen CD(69) on CD(8)(+) and CD(4)(+) T cells was detected by flow cytometry (FCM).

RESULTSThe optimal concentration of PTD-BCR/ABL protein for activating CD(8)(+) T cells in vitro was 100 micro g/ml, CD(69) expression peaked in three days stimulation. CD(8)(+) T cells were activated in 10 of 15 CML patients, the expression rate of CD(69) was (15.01 +/- 3.75)%. CD(4)(+) T cells were activated in 4 of 15 patients, the expression rate of CD(69) was (10.32 +/- 3.08)%. Both CD(8)(+) and CD(4)(+) T cells were activated simultaneously in 3 of them. However, neither CD(4)(+) nor CD(8)(+) T cells was activated by stimulation with BCR/ABL protein in all 15 specimens, the expression rate of CD(69) on CD(8)(+) and CD(4)(+) T cells was (1.36 +/- 0.31)% and (1.41 +/- 0.43)%, respectively. There was no difference compared with that of PBS control group (P > 0.05).

CONCLUSIONBy using a PTD-mediated antigen delivering system, exogenous BCR/ABL protein can be delivered into APC, processed and presented onto surface of APC to activate Ag-specific CD(8)(+) and CD(4)(+) T cells in vitro.

Adolescent ; Adult ; Aged ; Amino Acid Sequence ; Antigens, CD ; analysis ; Antigens, Differentiation, T-Lymphocyte ; analysis ; CD4 Antigens ; analysis ; CD8 Antigens ; analysis ; Dose-Response Relationship, Drug ; Female ; Flow Cytometry ; Fusion Proteins, bcr-abl ; genetics ; metabolism ; Gene Products, tat ; genetics ; metabolism ; Humans ; Lectins, C-Type ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; blood ; Male ; Middle Aged ; Recombinant Fusion Proteins ; immunology ; metabolism ; pharmacology ; T-Lymphocytes ; drug effects ; immunology

OBJECTIVETo investigate the expression of T cell early activation marker (CD(69)) on peripheral CD(4)(+) and CD(8)(+) lymphocytes and serum levels of soluble tumor necrosis factor receptor 1 and 2 (sTNF-R1 and sTNF-R2) in serum and bone marrow in patients with aplastic anemia (AA) and their pathophysiological significance.

METHODSIn vitro activation of T lymphocytes was carried out by whole blood cell culture containing PHA (20 micro g/ml). The CD(69) expressions on CD(4)(+) and CD(8)(+) lymphocytes at 0 h and 4 h after PHA exposure were analyzed by two-color flow cytometry. The levels of sTNF-R1 and sTNF-R2 in serum and bone marrow were measured by ELISA.

RESULTSThe CD(69) expression rates of CD(4)(+) and of CD(8)(+) cells in SAA patients were (8.96 +/- 7.23)% and (10.67 +/- 7.58)%, respectively, and that of CD(8)(+) cells in CAA patients was (7.36 +/- 5.49)% before PHA stimulation. The CD(69) expression rates of CD(4)(+) and of CD(8)(+) cells in SAA patients were (71.73 +/- 11.91)% and (61.74 +/- 13.44)% and in CAA (59.35 +/- 10.15)% and (48.78 +/- 8.25)% respectively, and were significantly elevated after PHA stimulation. CD(69) expression on CD(4)(+) cells was much higher than that on CD(8)(+) cells after stimulation. The levels of the two sTNF-R (sTNF-R1 and sTNF-R2) in peripheral blood and bone marrow of SAA patients were elevated and in the bone marrow of CAA patients were also increased. The serum levels of sTNF-R2 were positively related to the CD(69) expression rates of CD(8)(+) cells before PHA stimulation.

CONCLUSIONIncreased early activation and activated potentials of T lymphocytes, along with abnormally elevated immunologically active molecules might play a major role in the pathogenesis of AA.

Adolescent ; Adult ; Aged ; Anemia, Aplastic ; immunology ; Antigens, CD ; blood ; Antigens, Differentiation, T-Lymphocyte ; blood ; CD4-Positive T-Lymphocytes ; chemistry ; CD8-Positive T-Lymphocytes ; chemistry ; Child ; Female ; Humans ; Lectins, C-Type ; Male ; Middle Aged ; Receptors, Tumor Necrosis Factor ; blood ; Receptors, Tumor Necrosis Factor, Type I ; Receptors, Tumor Necrosis Factor, Type II

OBJECTIVETo investigate the significance of detecting TCR gene clonal rearrangement in the diagnosis of mycosis fungoides (MF) and to optimize the primers used for detecting the TCR gene clonal rearrangement with PCR in paraffin embedded tissues of MF.

METHODSNineteen cases of MF were enrolled into the study. A panel of 10 antibodies were used for immunophenotypic analysis and polymerase chain reaction for TCR-γ and TCR-β gene rearrangement detection in this study.

RESULTSTCR gene clonal rearrangements were detected in all 19 cases, in which 84.2% cases (16/19) had TCR-γ gene clonal rearrangements. The positive rates of the primers T(VG)/T(JX), V(2-5)/V(8-12)/JGT(1) and BIOMED-2-TCR-γ were 47.4%, 78.9% and 31.6%, respectively. The positive rate of V(2-5)/V(8-12)/JGT(1) was statistically significantly higher than that of T(VG)/T(JX) and BIOMED-2-TCR-γ (P < 0.05). No TCR gene clonal rearrangement was detected using the primers V(γ11)/V(γ101)/Jγ12 and V(γ11)/V(γ101)/J(p12). TCR-β gene clonal rearrangement was detected in 31.6% (6/19) cases.

CONCLUSIONSTCR gene clonal rearrangement analysis is a useful tool in the diagnosis of MF and TCR-γ gene is a good target gene for the detection. The primers T(VG)/T(JX), V(2-5)/V(8-12)/JGT(1) and BIOMED-2-TCR-γ can be used in clinicopathologic detection for TCR gene clonal rearrangement and V(2-5)/V(8-12)/JGT(1) may be the first choice.

Adolescent ; Adult ; Aged ; Antigens, CD7 ; metabolism ; Base Sequence ; CD2 Antigens ; metabolism ; CD3 Complex ; metabolism ; CD4 Antigens ; metabolism ; Child ; Child, Preschool ; Female ; Gene Rearrangement, beta-Chain T-Cell Antigen Receptor ; Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor ; Humans ; Leukocyte Common Antigens ; metabolism ; Male ; Middle Aged ; Molecular Sequence Data ; Mycosis Fungoides ; diagnosis ; genetics ; metabolism ; pathology ; Paraffin Embedding ; Receptors, Antigen, T-Cell, alpha-beta ; genetics ; Receptors, Antigen, T-Cell, gamma-delta ; genetics ; Skin Neoplasms ; diagnosis ; genetics ; metabolism ; pathology ; Young Adult

OBJECTIVETo explore the mechanisms of Chinese herbal medicine Sanqi Oral Liquid, composed of Astragalus membranaceus and Panpax notoginseng, in alleviating renal injury by observing its effect on the expressions of CD4(+), CD8(+) and CD68(+) cells in 5/6 nephrectomized rats with chronic renal failure.

METHODSA total of 102 SD rats were randomly divided into six groups: three treatment groups were administrated with high, medium and low dosage of Sanqi Oral Liquid respectively by gavage; a normal group, a 5/6 nephrectomized model group, and a group treated with coated aldehyde oxygenstarch were used as controls. Following oral administration of Sanqi Oral Liquid for 12 weeks, the general condition and renal pathological changes were observed, and the renal function, platelet count (PLT) and the expressions of CD4(+), CD8(+) and CD68(+) cells were determined for each group.

RESULTSThere were proliferation of mesangial matrix, renaltubularnecrosis and obvious tubulointerstitial fibrosis in the model group, and they were much milder in the treatment groups. Compared with the model group, the amounts of blood urea nitrogen (BUN), serum creatinine (Scr) and PLT in the treatment groups decreased (P<0.05 for all); and in the group administrated of medium dosage of Sanqi Oral Liquid, the expression of CD4(+) cells was up-regulated and those of CD8(+) and CD68(+) cells were down-regulated (P<0.05 for all), leading to an increased ratio of CD4(+)/CD8(+)(P<0.01).

CONCLUSIONSanqi Oral Liquid has a significant effect on regulating lymphocyte subsets, reducing the infiltration of macrophages in renal tissues and alleviating tubulointerstitial fibrosis, and this may be one of mechanisms of Sanqi Oral Liquid in delaying the progression of chronic kidney diseases.

Administration, Oral ; Animals ; Antigens, CD ; metabolism ; Antigens, Differentiation, Myelomonocytic ; metabolism ; Astragalus membranaceus ; chemistry ; CD4-Positive T-Lymphocytes ; drug effects ; pathology ; physiology ; CD8-Positive T-Lymphocytes ; drug effects ; pathology ; physiology ; Drug Evaluation, Preclinical ; Drugs, Chinese Herbal ; administration & dosage ; pharmacology ; Kidney Failure, Chronic ; drug therapy ; immunology ; pathology ; surgery ; Lymphocyte Count ; Male ; Nephrectomy ; Panax notoginseng ; chemistry ; Rats ; Rats, Sprague-Dawley ; Solutions