OBJECTIVETo investigate the telomerase activity and the telomere length in CD4(+), CD8(+) and CD19(+) lymphocytes from patients with systemic lupus erythematosus (SLE).

METHODSTelomerase activity of CD4(+), CD8(+) and CD19(+) cells from patients with SLE and normal controls was examined by telomeric repeats amplification protocol. Telomere length in those cells was measured by Southern blot method.

RESULTCD4(+), CD8(+) and CD19(+) cells in patients with SLE showed high telomerase activity, but only telomerase activity of CD19(+) cells was significantly higher than that in controls. The length of telomere in CD4(+) and CD8(+) cells was significantly shorter than that in controls, but not in CD19(+) cells.

CONCLUSIONHigher telomerase activity in CD19(+) cells and shortened telomere length in CD4(+) and CD8(+) cells of patients with SLE may be associated with pathogenesis of SLE.

Adolescent ; Adult ; Antigens, CD19 ; immunology ; CD4-Positive T-Lymphocytes ; enzymology ; CD8-Positive T-Lymphocytes ; enzymology ; Female ; Humans ; Lupus Erythematosus, Systemic ; enzymology ; immunology ; Lymphocytes ; enzymology ; Male ; Middle Aged ; Telomerase ; metabolism ; Telomere ; genetics

This study was aimed to investigate the mechanism of indoleamine 2, 3-dioxygenase (IDO) activity in acute myeloid leukemia cells contributing to tumor immune escape. Myeloid leukemia cells were isolated from bone marrow of 23 patients with acute myeloid leukemia (AML) and IDO expression was detected by immunochemistry and RT-PCR methods. Then mixed lymphocyte reaction (MLR) of one way was carried out, leukemia cells were used as stimulating cells and T-lymphocytes were used as reactive cells in culture with or without 1-MT. T-lymphocyte proliferation rate was determined by MTT assay and IDO activity in supernatant of MLR was detected by high-performance liquid chromatography (HPLC). The results showed that IDO expression was found in 17 out of 23 cases of acute myeloid leukemia cells; IDO enzyme activity in leukemia cells inhibited T-lymphocyte proliferation in MLR cultures. It is concluded that IDO activity expressing in leukemia cells can suppress T-lymphocyte proliferation responses, which may be contributing to tumor immune escape.
Cell Proliferation ; Humans ; Immune Tolerance ; Indoleamine-Pyrrole 2,3,-Dioxygenase ; metabolism ; Leukemia, Myeloid, Acute ; enzymology ; immunology ; pathology ; T-Lymphocytes ; cytology ; immunology ; Tumor Cells, Cultured ; Tumor Escape ; immunology

OBJECTIVEAcute idiopathic thrombocytopenic purpura (AITP) is a common autoimmune disease in children. Thrombocytes decrease extremely in serious patients, its pathogenesis involves abnormal activation of T lymphocytes and T cell-dependent production of autoantibody. The aim of the present study was to investigate changes of protein kinase C (PKC) activity in peripheral blood T lymphocytes in children with AITP and the relationships between PKC activity and T lymphocytes activation and thrombocytopenia.

METHODSPeripheral blood specimens were collected from children with acute ITP (n = 35) and healthy children (n = 30), and T lymphocytes were isolated and purified by using T cells Segregation Enrichment Column. PKC activity was detected by using PepTag Assay, a non-radioactive detection method. The reaction mixture, in a final volume of 25 microl, consisted of 5 microl reaction buffer, PepTag C1 5 microl (0.4 microg/microl), PKC activator solution (DG) 5 microl, peptide protection solution 1 microl and sample 9 microl. Phosphorylation reaction was allowed to continue for 30 minutes, then 25 microl reaction mixture was subjected to electrophoresis on a 0.8% agarose gel at 100 V for about 20 minutes. After electrophoresis, the PepTag C1 peptides which were phosphorylated and non-phosphorylated were separated, phosphorylated PepTag C1 peptide with negative electricity migrated toward the anode (+), but nonphosphorylated PepTag C1 peptide with positive electricity migrated toward cathode (-), the gel was photographed. Electrophoresis bands on anode represented PKC activity and were analyzed quantitatively. FasL, which is T cell activation marker, was determined by flow cytometer and platelet was counted by cell counting meter.

RESULTSCompared with healthy children, children with AITP had significantly higher PKC total activity [(0.97 +/- 0.21) nmol/(min.ml) vs. (0.55 +/- 0.13) nmol/(min.ml), (P < 0.05)]. Expression of FasL on T cell subpopulation in children with AITP was significantly higher [Th FasL: (32.7 +/- 3.4) vs. (14.7 +/- 4.2); Tc FasL: (17.3 +/- 9.7) vs. (11.6 +/- 8.5)%, (P < 0.05)]. Besides, relationships between the changes of PKC activity, Th FasL and Tc FasL had positive correlation (r(1) = 0.68, r(2) = 0.53, P < 0.05). However, PKC activity and platelet count had a significantly negative correlation (r = -0.75, P < 0.05).

CONCLUSIONIncreased PKC activity was seen in children with AITP, which can cause damage to thrombocytes and reduction of thrombocytes. PKC signal transduction pathway might play an important role in the immunopathogenesis of AITP.

Acute Disease ; Case-Control Studies ; Cell Separation ; Child ; Electrophoresis ; Flow Cytometry ; Humans ; Lymphocyte Activation ; Platelet Count ; Protein Kinase C ; metabolism ; Purpura, Thrombocytopenic, Idiopathic ; enzymology ; immunology ; T-Lymphocytes ; enzymology

OBJECTIVETo identify autoepitopes of E2 subunit of pyruvate dehydrogenase complex (PDC-E2) specific CD8+ CTL in primary biliary cirrhosis (PBC) patients.

METHODSAn online database SYFPEITHI was applied to predict HLA-A*0201 restricted epitopes which located in PDC-E2 30-50 aa and 150-190 aa where B-cell epitopes clustered with CD4+ T-cell epitopes. T2 cell line reconstitution and stabilization assay, induction of specific CTL lines from peripheral blood mononuclear cells (PBMCs) of patients with PBC and cytotoxicity of peptides-induced CTL were performed to screen the epitopes from those candidates.

RESULTSFive potential epitopes were predicted by database. Of the 5 candidates, two peptides 159-167 aa and 165-174 aa, with highly binding activity to HLA-A*0201 molecules, could stimulate PBMCs from most HLA-A*0201 positive PBC patients to proliferate and peptide-induced CTL lines showed specific cytotoxicity.

CONCLUSIONPeptides of KLSEGDLLA (159-167 aa) and LLAEIETDKA (165-174 aa) in the inner lipoyl domain of PDC-E2 are HLA-A*0201 restricted CD8+ CTL immunodominant epitopes in PBC.

Antibody-Producing Cells ; cytology ; Autoantigens ; immunology ; Autoimmunity ; CD8-Positive T-Lymphocytes ; cytology ; immunology ; metabolism ; Cell Line ; Dihydrolipoyllysine-Residue Acetyltransferase ; Epitope Mapping ; Epitopes, T-Lymphocyte ; immunology ; HLA-A Antigens ; immunology ; HLA-A2 Antigen ; Humans ; Liver Cirrhosis, Biliary ; enzymology ; genetics ; immunology ; Phenotype ; Protein Binding ; Pyruvate Dehydrogenase Complex ; genetics ; immunology ; metabolism ; T-Lymphocytes, Cytotoxic ; immunology

OBJECTIVEWe used an animal model of collagen-induced arthritis (CIA) to study changes and roles of tyrosine hydroxylase (TH) expressed by CD4+ T cell subsets, and then explore the relationship between CD4+ T cell subset-derived catecholamines and inflammatory responses in CIA.

METHODSThirty-six male DBA/1 mice were randomly divided into control group, CIA model group (day 35) and CIA model group (day 55) (n = 12). CIA model was induced by type II collagen (CII) in DBA/1 mice. On the 35th and 55th day following primary immunization, the joints of the mice were observed for clinical score of swelling and the level of anti-CII IgG antibody in serum was examined. Expression of specific transcription factors and cytokines of Th1, Th17, Th2 and Treg cells and TH in mesenteric lymph nodes was measured by means of Western blot. The changes of TH expressed by CD4+ T cell subsets in mesenteric lymph nodes were determined by flow cytometry.

RESULTSClinical score and anti-CII antibody level increased in CIA compared with that in intact mice. Specific transcription factors and cytokines expressed by Th1 and Th17 cells were upregulated and cytokines expressed by Th2 and Treg cells were downregulated in mesenteric lymph nodes in CIA mice. Expression of TH was upregulated and the increased expression of TH in CD4+ T cells was attributed to Th1 and Th17 cells in mesenteric lymph nodes of CIA.

CONCLUSIONThe increase in catecholamines from CD4+ T cell subsets in mesenteric lymph nodes of CIA may be related to inflammatory alleviation in CIA progression.

Animals ; Arthritis, Experimental ; immunology ; CD4-Positive T-Lymphocytes ; enzymology ; Collagen Type II ; adverse effects ; Cytokines ; metabolism ; Disease Models, Animal ; Lymph Nodes ; immunology ; Male ; Mice ; Mice, Inbred DBA ; T-Lymphocyte Subsets ; immunology ; Transcription Factors ; metabolism ; Tyrosine 3-Monooxygenase ; metabolism

Cytokines ; secretion ; Cytotoxicity, Immunologic ; Graft vs Leukemia Effect ; Humans ; Immunophenotyping ; Lymphocyte Activation ; Retroviridae ; genetics ; Simplexvirus ; enzymology ; T-Lymphocytes ; immunology ; Thymidine Kinase ; genetics ; Transfection

M1-type macrophages are capable of inducing lysis in various types of cancer cells, but the mechanism of action is unclear. It has been noted that an "unknown protein" produced together with protease by activated macrophages is responsible for this action. Activated M1 macrophages have been recently reported to produce family 18 chitinases, all of which have been named chitotriosidase. Our experiments have demonstrated that family 18 chitinases work together with proteases and can damage various cancer cells both in vitro and in vivo. Thus, in this article, we suggest that the 50-kDa chitotriosidase is the reported "unknown protein". In addition, we discuss how to properly stimulate activated M1 macrophages to produce 50-kDa chitotriosidases and proteases for destroying cancer cells. Because family 19 chitinase has recently been reported to kill cancer cells, we also discuss the possibility of directly using human family 18 chitotriosidase and the humanized plant family 19 chitinase for cancer treatment.
Animals ; Antineoplastic Agents, Alkylating ; pharmacology ; Chitinases ; metabolism ; Cyclophosphamide ; pharmacology ; Hexosaminidases ; metabolism ; Humans ; Immunosuppressive Agents ; pharmacology ; Macrophage Activation ; immunology ; Macrophages ; classification ; enzymology ; immunology ; pathology ; Neoplasms ; immunology ; pathology ; Peptide Hydrolases ; metabolism ; T-Lymphocytes, Regulatory ; metabolism ; Th1 Cells ; metabolism ; Th2 Cells ; metabolism

OBJECTIVETo study the effect of Extractum trametes robiniophila murr on cardiac allograft rejection in mice.

METHODSAll abdominal heterotopic heart transplantation models were divided into three groups as follows: (A) Extractum trametes robiniophila murr group. (B) Rejection group. (C) Isograft group. In each group, mean survival times (MST) of transplanted hearts and their pathologic histological changes at postoperative fifth day were observed. With fluoroimmunoassay, granzyme B and CD8(+) expressions were examined.

RESULTSThe MST of heart allografts in group A were (6.38 +/- 0.69) d, significantly shorter than that of group B [(8.31 +/- 0.59) d] (P < 0.01). In group A, acute rejection was present in advance; transplanted hearts were seriously damaged into acute rejection pathological grade 3, and CD8(+) T lymphocytes infiltrated diffusely and the expression of granzyme B increased significantly as compared with other groups.

CONCLUSIONSExclusive application of Extractum trametes robiniophila murr can promote the acute rejection of graft in early phase of postoperation, and the mechanism may be the promoted proliferation and infiltration of CD8(+) T lymphocytes and the increased expression of granzyme B.

Animals ; CD8-Positive T-Lymphocytes ; immunology ; Drugs, Chinese Herbal ; adverse effects ; Female ; Graft Rejection ; chemically induced ; Granzymes ; metabolism ; Heart Transplantation ; Male ; Mice ; Mice, Inbred C3H ; Mice, Inbred C57BL ; Myocardium ; enzymology ; immunology ; Postoperative Care

The major house-dust mite allergen, Der f 2, stimulates the phospholipase D (PLD) in T lymphocytes from Dermatophagoides farinae specific allergic individuals. PLD activity increased more than two-fold in T cells from allergic patients compared with those cells from normal controls with maximal responses within 30 min after exposure of Der f 2. A well-known PLD activator PKC-alpha was found to be translocated to membrane from cytosol in Der f 2-treated T cells from Dermatophagoides farinae specific allergic individuals. Down-regulation of PKC-alpha with phorbol myristate acetate pretreatment for 24 h abolished Der f 2-induced PLD activation. Ro 320432, PKC inhibitor also reduced the effects of Der f 2-induced PLD activation suggesting that PKC-alpha acts as upstream activator of PLD in Der f 2-treated T cells. Taken together, the present data suggest that Der f 2 can stimulate PLD activity through the PKC-alpha activation in T cells from Dermatophagoides farinae allergic individuals
Adolescent ; Adult ; Animals ; Antigens, Dermatophagoides/*immunology ; Dermatophagoides farinae/*immunology ; Female ; Humans ; Hypersensitivity, Immediate/*enzymology/*immunology ; Male ; Phospholipase D/*metabolism ; Protein Kinase C/antagonists & inhibitors/*physiology ; Research Support, Non-U.S. Gov't ; Skin Tests ; T-Lymphocytes/*enzymology/immunology ; Tetradecanoylphorbol Acetate/*analogs & derivatives/pharmacology ; Up-Regulation

OBJECTIVEEscape from the body's immune response is a basic characteristic of lung cancer, and indoleamine-2,3-dioxygenase (IDO) plays a key role in mediating immune escape of non-small-cell lung cancer, which leads to recurrence and metastasis. Feiji Recipe, a compound Chinese herbal medicine, has the effect of stabilizing lesions and prolonging survival in patients with lung cancer. The purpose of this study was to investigate the mechanisms underlying the anticancer properties of Feiji Recipe.

METHODSAn orthotopic transplant model of mouse Lewis lung cancer, with stable expression of IDO gene, was established in C57BL/6 mice. Optical imaging was used to observe the effects of Feiji Recipe in the treatment of lung cancer in vivo. The effects of Feiji Recipe on the proliferation of mouse Lewis lung cancer cell line 2LL, 2LL-enhanced green fluorescent protein (2LL-EGFP) and 2LL-EGFP-IDO were investigated, and the apoptosis of T-cells was examined by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide using flow cytometry. Chemical composition of Feiji Recipe was validated by high-performance liquid chromatography.

RESULTSCompared to the control group, the survival of animals treated with Feiji Recipe was significantly prolonged (P = 0.0074), and the IDO protein level decreased (P = 0.0072); moreover, the percentages of CD4CD25 T-cells and Foxp3 T-cells were significantly decreased (P < 0.05). The molecular mechanism of Feiji Recipe against lung cancer may relate to the regulation of immune cells, such as T-cells and regulatory T-cells.

CONCLUSIONThe molecular mechanism of Feiji Recipe in treatment of lung cancer is to restore the function of T-cells in the cancer microenvironment through interfering with the IDO pathway.

Animals ; Apoptosis ; drug effects ; Carcinoma, Lewis Lung ; drug therapy ; enzymology ; immunology ; physiopathology ; Cell Proliferation ; drug effects ; Disease Models, Animal ; Drugs, Chinese Herbal ; administration & dosage ; Growth Inhibitors ; administration & dosage ; Humans ; Indoleamine-Pyrrole 2,3,-Dioxygenase ; genetics ; immunology ; Lung Neoplasms ; drug therapy ; enzymology ; immunology ; physiopathology ; Male ; Mice ; Mice, Inbred C57BL ; T-Lymphocytes, Regulatory ; drug effects ; immunology