OBJECTIVETo investigate the influence of interferon-alpha on the variation of cytotoxic T lymphocytes (Tc) and suppressor T lymphocytes (Ts) in the peripheral blood of 32 patients with chronic hepatitis B, and to analyse the relationship between the efficacy of interferon-alpha and the variation of Tc and Ts cells.

METHODSThe peripheral blood Tc and Ts cells were detected by the double-staining immunocytochemistry technique.

RESULTSAt the end of the treatment with interferon-alpha, there were 9 complete responders (CR), 12 partial responders (PR) and 11 non-responders (NR). Tc cells significantly increased and Ts cells markedly decreased in CR or PR group compared with the healthy control group. There was no significant difference in the level of Tc and Ts cells between CR and PR groups, and no significant difference in the level of Tc cells in NR, CR and PR groups, The Ts cells was significantly higher in NR group than in CR or PR group.

CONCLUSIONSThe treatment of interferon-alpha can result in the change of Tc and Ts cells in patients with chronic hepatitis B. The variation of Ts cells may play an important role in the efficacy of interferon-alpha against hepatitis B virus.

Adult ; Antiviral Agents ; therapeutic use ; Female ; Hepatitis B, Chronic ; blood ; drug therapy ; Humans ; Interferon-alpha ; therapeutic use ; Lymphocyte Count ; Male ; Middle Aged ; T-Lymphocytes, Cytotoxic ; cytology ; drug effects ; T-Lymphocytes, Regulatory ; cytology ; drug effects ; Time Factors ; Treatment Outcome

This study was purposed to investigate the effects of thalidomide on proliferation of peripheral blood mononuclear cells (PBMNCs), levels of lymphocyte subsets, secretion of cytokines and its killing activity, and to elucidate the immunoregulation mechanisms in treatment of multiple myeloma with thalidomide. The method of MTT was used to detect the effects of thalidomide on the proliferations and the cytotoxic activity of PBMNC; the flow cytometer was used to analyze the lymphocyte subsets; the ELISA was used to measure the concentrations of cytokines in culture supernatants. The results showed that thalidomide enhanced the proliferations of the CD8+ T, NK cells in PHA-stimulated PBMNC from healthy volunteers, increased the secretion of IL-6 significantly, and decreased the secretion of IFN-gamma, and the secretions of IL-2 and IL-10 were not affected. Compared with control group, at the same ratio of effectors to targets the thalidomide (5 microg/ml) could enhance the cytotoxic activity of PBMNC (P < 0.01), the cytotoxic activity was maximal when the ratio of effectors to targets was 40:1. It is concluded that thalidomide preferentially enhances the proliferations of CD8+ T, NK cells in PHA-stimulated PBMNC from healthy volunteers, and enhances the cytotoxic activity of PBMNC by increasing the secretion of IL-6 significantly, in short, thalidomide can exert anti-myeloma effects by increasing cellular immune function.
Adjuvants, Immunologic ; pharmacology ; Adult ; CD8-Positive T-Lymphocytes ; cytology ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Cytokines ; biosynthesis ; Female ; Humans ; Immunity, Cellular ; drug effects ; Killer Cells, Natural ; cytology ; immunology ; Leukocytes, Mononuclear ; cytology ; drug effects ; immunology ; Lymphocyte Subsets ; drug effects ; immunology ; Male ; Middle Aged ; Multiple Myeloma ; pathology ; T-Lymphocytes, Cytotoxic ; drug effects ; Thalidomide ; pharmacology ; Tumor Cells, Cultured

OBJECTIVETo investigate the specific antitumor immunity induced by antigen peptide mixture prepared from different T lymphocytic leukaemia cells and the cross-reaction among the mixtures of different cell lines.

METHODSAntigen peptide mixtures were prepared from different leukaemia cell lines and then bound with Hsp70 in vitro. The activation and proliferation of peripheral blood mononuclear cell (PBMC) were observed after the stimulation by different Hsp70-peptide complexes. The cytotoxicity of such activated PBMCs to different target cells was assayed.

RESULTSThe antigen peptides from different leukaemia cell lines were mixed ones, which could activate PBMC effectively with Hsp70 and stimulate the activated PBMC to proliferate. The proliferative PBMC had specific cytotoxicity to the corresponding leukaemia cells. To Hut-78 cell, Molt-4 cell and Jurkat cell, the cytotoxicity of PBMC activated by either Hut78-peptides or Molt-4-peptides was significantly stronger than that of PBMC activated by HL-60-peptides (P < 0.05). The cytotoxicity to Jurkat cell of PBMC activated by Hut78/Molt-4-peptides was significantly stronger than that of PBMC activated by Hut78-peptides or Molt-4-peptides alone (P < 0.05).

CONCLUSIONAntigen peptide mixture from T lymphocytic leukaemia cells is able to induce specific antitumor immunity. There is a cross-reactivity among antigen peptide mixtures from different T lymphocytic leukaemia cell lines, with the more crossed antigen peptides obtained from the mixtures of different antigen peptides from different T lymphocytic leukaemia cell lines, which suggests that the antigen peptide mixture with broad antigenic spectrum could possibly be prepared by using multiple leukaemia cell lines.

Antigens, Neoplasm ; metabolism ; pharmacology ; Cross Reactions ; HL-60 Cells ; HSP70 Heat-Shock Proteins ; metabolism ; Humans ; Leukemia, T-Cell ; immunology ; Leukocytes, Mononuclear ; cytology ; drug effects ; Peptides ; pharmacology ; T-Lymphocytes, Cytotoxic ; drug effects ; immunology ; Tumor Cells, Cultured ; chemistry ; immunology

OBJECTIVETo investigate the pathogenesis of cytotoxic T cell (CTL) dysfunction in patients with HCV infection.

METHODSBALB/c mice were immunized by subcutaneous injection of polypeptides from HCV core region, and the CTL activity of mouse spleen cells was detected by the LDH release test. Two polypeptides which can enhance CTL function and two polypeptides which can inhibit CTL function were selected and cross-combined. BALB/c mice were immunized using the combined polypeptides and the CTL activities were detected afterwards.

RESULTSCTL activity was inhibited by CPA9 (39-74 amino acids), CPB7 (67-76 amino acids) and CPB8 (71-80 amino acids), and promoted by CPA10 (5-23 amino acids), CPB6 (63-72 amino acids) and CPB2 (131-140 amino acids). Using single factor analysis of variance, the CTL activity in the mice could be enhanced by polypeptides from the HCV core region, CPB2+CPB8, CPB6+CPB8, respectively. There was no obvious difference between CPB2+CPB7, CPB6+CPB7 and negative control.

CONCLUSIONSCPA9, CPB7, and CPB8, the 3 polypeptides from HCV core region play an inhibition role and CPA10, CPB6, and CPB2 play an enhancement role in CTL activity in mice. The inhibition and enhancement functions of the polypeptides from HCV core region interact each other.

Animals ; Cytotoxicity Tests, Immunologic ; Cytotoxicity, Immunologic ; drug effects ; Mice ; Mice, Inbred BALB C ; Peptide Fragments ; administration & dosage ; immunology ; Spleen ; cytology ; drug effects ; immunology ; T-Lymphocytes, Cytotoxic ; cytology ; immunology ; Viral Core Proteins ; administration & dosage ; chemistry ; immunology

OBJECTIVETo investigate the cytotoxic effect of γδ T cells from osteosarcoma patients against interferon-γ (IFN-γ)-treated osteosarcoma cells in vitro.

METHODSHuman γδ T cells were amplified by zoledronate from peripheral blood cells of osteosarcoma patients. The expression of Fas on the osteosarcoma cells were measured by flow cytometry and quantitative real-time PCR analysis before and after IFN-γ treatment. The cytotoxicity of γδ T cells against osteosarcoma cells was evaluated with LDH assay.

RESULTSIFN-γ significantly enhanced the susceptibility of the osteosarcoma cell lines HOS and U2OS to the cytotoxicity of γDelta; T cells from osteosarcoma patients (P<0.01). IFN-γ obviously up-regulated the expression of Fas in HOS and U2OS cells (P<0.01). Anti-FasL mAb failed to inhibit the cytotoxicity of γδ T cells in untreated osteosarcoma targets (P>0.05), but significantly impaired γδ T cell cytotoxicity in IFN-γ pre-treated osteosarcoma targets (P<0.01).

CONCLUSIONIFN-γ can enhance the cytotoxic effect of human γδ T cells from osteosarcoma patients against osteosarcoma cells in vitro.

Bone Neoplasms ; metabolism ; Cell Line, Tumor ; Cytotoxicity, Immunologic ; Humans ; Interferon-gamma ; pharmacology ; Osteosarcoma ; immunology ; metabolism ; Receptors, Antigen, T-Cell, gamma-delta ; immunology ; T-Lymphocytes, Cytotoxic ; cytology ; drug effects ; immunology ; fas Receptor ; metabolism

This study was aimed to explore the effects of interleukin 21 (IL-21) on the anti-leukemia activity of cytotoxic T lymphocytes (CTL) induced by dendritic cells (DCs) in vitro. The peripheral mononuclear cells from leukemia patients in complete remission were cultured with the specific cytokines to induce the production of DCs. The DCs loaded with RNA from autologous leukemic cells as antigen, and co-cultured with autologous T lymphocytes to get leukemia specific CTL. The cytotoxic activity of CTL against autologous leukemic cells was measured by LDH release method. The concentration of IFN-gamma and TNF-alpha in the culture supernatant was measured by enzyme immunoassay. The effects of IL-21 on the mature DCs were also studied by the measurement of the phenotype of DC and the allogenic mixed lymphocytic reactions induced by DCs. Experiments were divided into 2 groups: test group in which IL-21 (200 ng/ml) was added in coculture of DC/CTL and control group in which no IL-21 (200 ng/ml) was added. The results showed that when cultured with IL-21, the quantity of CTL increased from (56.73 +/- 10.21)% (control group) to (73.43 +/- 18.01)% (p < 0.01); The concentration of IFN-gamma and TNF-alpha in the culture supernatant increased from (154.91 +/- 67.20) ng/L (control group) to (310.62 +/- 141.15) ng/L (p < 0.01) and from (8.77 +/- 5.09) microg/L (control group) to (15.25 +/- 6.56) microg/L (p < 0.01) respectively. At the effector: target ratio of 20:1, the cytotoxic activity against autologous leukemic cells by CTL increased from (50.22 +/- 5.07)% (control group) to (75.38 +/- 9.47)% (p < 0.01). IL-21 had neither effect on the phenotype (CD1a, CD83, CD86, CD80 and HLA-DR) of mature DCs nor the allogeneic mixed lymphocytic reactions induced by DCs. It is concluded that IL-21 can strengthen the proliferation of CTL, and improve the production of IFN-gamma and TNF-alpha, thus enhance the anti-leukemia activity of CTL. Nevertheless, there is no effect of IL-21 on the function of mature DCs. These data indicate that IL-21 has a potential clinical value in the enhancement of anti-leukemia immunotherapy.
Adult ; CD8-Positive T-Lymphocytes ; drug effects ; Cell Proliferation ; Cytotoxicity, Immunologic ; drug effects ; Dendritic Cells ; cytology ; drug effects ; Female ; Humans ; Interferon-gamma ; immunology ; Interleukins ; pharmacology ; K562 Cells ; Leukemia ; drug therapy ; immunology ; Male ; Middle Aged ; T-Lymphocytes, Cytotoxic ; drug effects ; Tumor Necrosis Factor-alpha ; immunology ; Young Adult

OBJECTIVETo explore the in vitro anti-tumor effect and mechanism of dendritic cell (DC) tumor vaccine induced by astragalus polysacharin (APS).

METHODSPeripheral blood mononuclear cells (PBMCs) isolated from human peripheral blood. DCs obtained from human peripheral blood were cultivated and added with culture solution for in vitro inducing them to immature DCs. On the 5th day of culture, 100 microg/mL (as the final concentration) APS was added to cells in the APS group. DCs were induced to mature in the cytokine groups by adding 20 ng/mL rhTNF-alpha (as the final concentration). Changes of morphology and phenotype of DCs were observed. Mature DCs were sensitized with tumor antigen SGC-7901 and co-cultured with allogeneic T cells. The proliferative function of T lymphocytes was detected by MTT assay. Levels of IL-12 and IFN-gamma in co-cultured supernatant were detected by ELISA. Cytotoxic lymphocytes (CTL) activated by DC were co-cultured with tumor cell SGC-7901. The specific killing capacity of CTL to target cells was detected by LDH release assay.

RESULTSThe morphological observation and phenotypic identification of APS induced DCs were in accordance with the characteristics of mature DCs. APS induced mature DCs could stimulate the proliferation of allogeneic T lymphocytes. The proliferation index of T cells increased with increased ratio of stimulator cells to effector cells (P < 0.05). Levels of IL-12 and IFN-gamma in co-culture supernatant significantly increased in a time-dependent manner (P < 0.05). CTL cells activated by sensitization of DCs could significantly kill tumor cells, and the killing effect increased along with increased effector-to-target ratio.

CONCLUSIONAPS could in vitro induce DCs to mature, promote its antigen-presenting capacity, effectively activate CTLs, and enhance anti-tumor function of the organism.

Antigen-Presenting Cells ; cytology ; drug effects ; immunology ; Cancer Vaccines ; immunology ; Cell Line ; Cell Proliferation ; drug effects ; Coculture Techniques ; Dendritic Cells ; cytology ; drug effects ; immunology ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Interferon-gamma ; immunology ; Interleukin-12 ; immunology ; Leukocytes, Mononuclear ; cytology ; immunology ; Lymphocyte Activation ; T-Lymphocytes, Cytotoxic ; cytology ; drug effects

The study was to understand the impact on the proliferation and cytotoxicity of donor's T cells during mobilization with rhG-CSF. The peripheral blood mononuclear cells (PBMNC) were collected from 15 donors before mobilization and on fifth day of mobilization with rhG-CSF. After the PBMNC were activated with 500 ng/ml of CD3 monoclonal antibody and 500 microg/ml of rhIL-2 for 96 hours, the activated T cells were collected for testing proliferation, cytotoxicity, Fas expression, perforin and Fas ligand (FasL) mRNA expression, the IFN-gamma concentration in the culture medium of the activated T cells was determined by radioimmunoassay. The results showed that the proliferation activity of T lymphocytes and the cytotoxicity of T cells activated with CD3 monoclonal antibody and rhIL-2 were reduced markedly after mobilization with rhG-CSF (P < 0.05). The Fas molecule expression in the activated T cells was very high both before and after mobilization with rhG-CSF (P > 0.10). The activated T cells expressed perforin mRNA and didn't express FasL mRNA both before and after mobilization with rhG-CSF. The concentration of IFN-gamma in the culture medium of the activated T cells decreased significantly after mobilization with rhG-CSF (P < 0.01). It is concluded that activity of proliferation and cytotoxicity of donor's T cells is impaired after mobilization with rhG-CSF.
Adolescent ; Adult ; Cell Proliferation ; drug effects ; Cells, Cultured ; Fas Ligand Protein ; biosynthesis ; genetics ; Female ; Granulocyte-Macrophage Colony-Stimulating Factor ; administration & dosage ; pharmacology ; Hematopoietic Stem Cell Mobilization ; methods ; Humans ; Male ; Middle Aged ; RNA, Messenger ; biosynthesis ; genetics ; Recombinant Proteins ; T-Lymphocytes ; cytology ; drug effects ; T-Lymphocytes, Cytotoxic ; drug effects ; immunology ; fas Receptor ; biosynthesis ; genetics

Dendritic cells (DCs) are potent antigen-presenting cells. OK432 (Picibanil(R)) was introduced as a potent stimulator of DC maturation in combination with prostaglandin-E2 and interferon-alpha. We compared the efficacy of a DC-prostate cancer vaccine using early-mature DCs stimulated with OK432, PGE2 and INF-alpha (OPA) with that of vaccines using other methods. On days 3 or 7 of DC culture, TNF-alpha (T), TNF-alpha and LPS (TL) or OPA were employed as maturation stimulators. DU145 cells subjected to heat stress were hybridized with mature DCs using polyethyleneglycol. T cells were sensitized by the hybrids, and their proliferative and cytokine secretion activities and cytotoxicity were measured. The yields of early-mature DCs were higher, compared to yields at the conventional maturation time (P<0.05). In the early maturation setting, the mean fusion ratios, calculated from the fraction of dual-positive cells, were 13.3%, 18.6%, and 39.9%, respectively (P=0.051) in the T only, TL, and OPA-treated groups. The function of cytotoxic T cells, which were sensitized with the hybrids containing DCs matured early with OPA, was superior to that using other methods. The antitumor effects of DC-DU145 hybrids generated with DCs subjected to early maturation with the OPA may be superior to that of the hybrids using conventional maturation methods.
Cancer Vaccines/*immunology ; Cell Line, Tumor ; Dendritic Cells/cytology/drug effects/*immunology ; Dinoprostone/*pharmacology ; Humans ; Immunologic Factors/*pharmacology ; Interferon-alpha/*pharmacology ; Lipopolysaccharides/toxicity ; Male ; Neoplasms, Hormone-Dependent/*immunology ; Phenotype ; Picibanil/*pharmacology ; Prostatic Neoplasms/*immunology ; T-Lymphocytes, Cytotoxic/immunology

We evaluated the effectiveness of rhamnogalacturonan II (RG-II)-stimulated bone marrow-derived dendritic cells (BMDCs) vaccination on the induction of antitumor immunity in a mouse lymphoma model using EG7-lymphoma cells expressing ovalbumin (OVA). BMDCs treated with RG-II had an activated phenotype. RG-II induced interleukin (IL)-12, IL-1beta, tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) production during dendritic cell (DC) maturation. BMDCs stimulated with RG-II facilitate the proliferation of CD8+ T cells. Using BMDCs from the mice deficient in Toll-like receptors (TLRs), we revealed that RG-II activity is dependent on TLR4. RG-II showed a preventive effect of immunization with OVA-pulsed BMDCs against EG7 lymphoma. These results suggested that RG-II expedites the DC-based immune response through the TLR4 signaling pathway.
Acute-Phase Proteins/metabolism ; Adaptor Proteins, Vesicular Transport/metabolism ; Animals ; Antigens, CD14/metabolism ; Bone Marrow Cells/cytology/drug effects ; CD8-Positive T-Lymphocytes/*immunology ; Carrier Proteins/metabolism ; Cell Differentiation/drug effects ; Cell Nucleus/drug effects/metabolism ; Cell Proliferation/drug effects ; Cytokines/biosynthesis ; Dendritic Cells/cytology/drug effects/enzymology/*immunology ; Enzyme Activation/drug effects ; Lymphocyte Activation/*drug effects ; Membrane Glycoproteins/metabolism ; Mice ; Mice, Inbred C57BL ; Mice, Knockout ; Mitogen-Activated Protein Kinases/metabolism ; Myeloid Differentiation Factor 88/metabolism ; NF-kappa B/metabolism ; Neoplasms/immunology/*pathology ; Pectins/*pharmacology ; Phenotype ; Protein Transport/drug effects ; Receptors, Chemokine/metabolism ; Signal Transduction/drug effects ; T-Lymphocytes, Cytotoxic/cytology/drug effects ; Toll-Like Receptor 4/*agonists/metabolism