OBJECTIVETo investigate the enhancing effect of Chinese medicine-Xuebijing injection on lipopolysaccharide (LPS) -induced apoptosis of CD4+ CD25+ regulatory T cells (Tregs) and polarization of helper T cells (Th).

METHODSCD4+ CD25+ Tregs collected from rat spleen in vitro by immunomagnetic beads assay were divided into the control group, anti-CD3/CD28 group, anti-CD3/CD28 + LPS group, anti-CD3/CD28 + "Xuebijing injection" group and anti-CD3/CD28 + LPS + "Xuebijing injection" group. Tregs apoptosis rate and expression of winged helix transcription factor (Foxp3) in Tregs were detected by flow cytometry on 3rd post culture day. CD4+ CD25- T cells were co-cultured with CD4+ CD25- Tregs (1:1) for 68 hours with canavalin A stimulation. Interferon gamma (gamma-IFN), interleukin (IL)-4 and IL-17 in supernatants, which respectively was secreted by Th1, Th2 and Th17, were measured by ELISA.

RESULTSTregs apoptosis rate of anti-CD3/CD28 + LPS + "Xuebijing injection" group (45.1 +/- 2.7%) was significantly higher than that of anti-CD3/CD28 + LPS group (29.4 +/- 1.6%, P < 0.01). Meanwhile, Foxp3 expressions in Tregs in above 2 groups were 95 +/- 9 and 140 +/- 18 respectively, showing statistically significant difference between them (P < 0.01). Gamma-IFN levels secreted in anti-CD3/CD28 + LPS + "Xuebijing injection" group were significantly higher than those in anti-CD3/CD28 + LPS group (P < 0.01), while IL-4 levels had an opposite tendency compared with gamma-IFN (P < 0.05), resulting in a marked increase in the ra- tio of gamma-IFN/IL-4 in anti-CD3/CD28 + LPS + "Xuebijing injection" group (P < 0.01). In anti-CD3/ CD28 + "Xuebijing injection" group, IL-17 secretion levels were significantly decreased compared with anti-CD3/CD28 group (P < 0.05).

CONCLUSIONSActivation of CD4+ CD25+ Tregs induced by LPS may mediate Th1 shift to Th2 response. "Xuebijing injection" can effectively regulate immune function of T cells, increase the LPS-induced apoptosis of CD4+ CD25+ Tregs as well as enhance the polarization of Th2 to Th1, thereby abating the suppressive state of cell-mediated immunity.

Animals ; Apoptosis ; Drugs, Chinese Herbal ; pharmacology ; Endotoxins ; Lipopolysaccharides ; Male ; Rats ; Rats, Wistar ; T-Lymphocytes, Helper-Inducer ; drug effects ; immunology ; T-Lymphocytes, Regulatory ; drug effects ; immunology

OBJECTIVETo determine whether supplementation of zinc and vitamin A may improve the function of T cells in human PBMC.

METHODST cells were separated and cultured in vitro, supplemented with either Zn or vitamin A alone, or both Zn and vitamin A (10(-6) mol/L, 10(-5) mol/L, 10(-4) mol/L). After harvesting, cell proliferation, cell cycle, apoptosis, expression or function of cell-surface molecules, such as CD3+, CD4+, and CD8+ were detected.

RESULTSHigher proliferation level and lower apoptosis level were observed in cells supplemented with both Zn and vitamin A, showing the strongest effect (P < 0.05). Zn-supplement increased the CD4+/CD3+ cell percentage, and simultaneously decreased the CD8+/CD3+ cell population. VA-supplement showed the opposite effect in comparison with Zn-supplement.

CONCLUSIONT-cell function can be improved, depending on the zinc and/or vitamin A supplemented.

Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Drug Synergism ; Humans ; T-Lymphocytes ; drug effects ; Vitamin A ; pharmacology ; Zinc ; pharmacology

Adult ; Benzene ; toxicity ; Female ; Humans ; Male ; Occupational Exposure ; T-Lymphocytes, Regulatory ; drug effects ; Young Adult

OBJECTIVETo establish a method for isolating exosomes from dendritic cells (DC), and to analyse its biological characteristics and function in antitumor immunity.

METHODSImmature DCs (im-DC) from human peripheral blood mononuclear cells were loaded with the antigen of K562 tumor cells, then exosomes were secreted from imDC and lipopolysaccharide (LPS) induced mature DC (mDC). The exosomes from imDC and mDC were isolated separately by ultracentrifugation and ultrafiltration. The exosomes diameter was determined, their profile was observed by electron microscope, and the surface molecules were detected by Western blot. To analyse the effect of exosomes on antitumor immunity, the proliferation, IFN-gamma expression, CD69 up-regulation and cytotoxicity of antigen-specific T cells were measured.

RESULTSExosomes were small flattened sphere vesicles with an average diameter of 72.3 nm and expressed CD80, CD86, HLA-DR, FasL, CD54 and MFG-E8 molecules. As compared to immature exosomes, exosomes from mDC were proved to express more CD80 and less MFG-E8, to be more potent for inducing antigen-specific T cells proliferation and immunity respond in vitro: at its optimum concentration, the absorption value of T cell proliferation test was 0.50 +/- 0.01, CD69 was up-regulated and (13.4 +/- 5.8)% of T cells was in proliferating, (22.8 +/-2.4)% of T cells expressed IFN-gamma, and (21.3 +/-8.6)% of tumor cells were killed.

CONCLUSIONA simple and quick method to isolate and analyse exosomes is established. The exosomes can induce antitumor immunity respond.

Cells, Cultured ; Dendritic Cells ; immunology ; secretion ; Exosomes ; immunology ; Humans ; Lymphocyte Activation ; drug effects ; T-Lymphocytes ; drug effects ; immunology

OBJECTIVETo investigate the levels of T cell receptor rearrangement excision DNA circles (TRECs) within peripheral blood from workers exposed to lead, and thereby to evaluate the number of naive T cells and recent thymic output function.

METHODSQuantitative detection of TRECs in peripheral blood mononuclear cells (PBMNC) from 10 cases of workers exposed to lead was performed by real time PCR analysis. 11 workers without exposure to lead served as unexposed controls. In addition, the relationship between TRECs, age, length of service, blood lead, urea lead, blood ZPP and urea delta-ALA was investigated.

RESULTSThe mean value of TRECs in workers exposed to lead was (2.44 +/- 1.87)/1000 PBMC, significantly under (5.60 +/- 3.96)/1000 PBMC in unexposed controls. A significant negative correlation was found between the TRECs and urea-ALA. But there was no significant correlation between them after controlling for blood lead, urea lead.

CONCLUSIONLead exposure may damage thymic output naive T cells function. Furthermore, low-level exposure to lead may damage immune system and earlier than expected.

Adult ; DNA ; drug effects ; Gene Rearrangement, T-Lymphocyte ; drug effects ; Humans ; Lead ; toxicity ; Leukocytes, Mononuclear ; Male ; Occupational Exposure ; adverse effects ; Receptors, Antigen, T-Cell ; genetics ; immunology ; T-Lymphocytes ; drug effects ; immunology ; Thymus Gland ; immunology

OBJECTIVETo investigate the cellular immune regulation of the long-term intervention of moxa smoke.

METHODSThirty-two Wistar rats were randomly divided into a blank group, a low concentration group, a medium concentration group and a high concentration group, 8 cases in each group. In addition to the blank group, rats in the other groups were exposed to the corresponding concentration moxa smoke for 20 min every day, the T lymphocyte subsets and proportion of the CD4+ CD25+ Treg in CD4+ T cells in peripheral blood were tested by flow cytometry after 6 months.

RESULTSCompared with the blank group, the proportions of CD3+ CD4+, CD3+ CD8+ T cells and CD3+ CD4/CD3+ CD8+ in the other 3 moxa smoke groups were not significantly different (P > 0.05), while the proportions of the CD4+ CD25+ Treg in CD4+ T cells were significantly lower (P < 0.05), but no statistically significant differences among those 3 moxa smoke intervention groups (P > 0.05).

CONCLUSIONLong-term moxa smoke intervention has no significant effect on the proportions of CD3+ CD4+, CD3+ CD8+ T cells and CD3+ CD4+/CD3+ CD8+, but it can decrease the proportions of the CD4+ CD25+ Treg in CD4+ T cells in peripheral blood of rats. The way produced by pretreatment with moxa smoke may play immunomodulatory effect.

Animals ; Lymphocyte Count ; Male ; Moxibustion ; Rats ; Rats, Wistar ; Smoke ; analysis ; T-Lymphocyte Subsets ; drug effects ; immunology ; T-Lymphocytes, Regulatory ; drug effects ; immunology ; Time Factors

OBJECTIVETo explore the partial therapeutic mechanism of Ginkgo Biloba extract (GBE) in treating asthma.

METHODSFourteen SD rats were randomly divided into two groups, 7 rats were sensitized as the asthmatic model group and the others taken as the healthy control group. T lymphocytes were isolated from peripheral blood mononuclear cells (PBMCs) of the rats, and were cultured in vitro with Ginkgolide B (BN-52021 group) or Ginkgo Biloba extract 761 (EGb761 group) in different concentrations or without any of them (control group). T lymphocytes proliferation in groups were measured by using MTT assay and the effect of BN-52021 on T lymphocytes apoptosis was analyzed by flow cytometry at various times.

RESULTSCompared with the control group, BN-52021 could significantly inhibit the proliferation of T lymphocytes in both healthy and asthmatic rats in vitro (P <0. 05). The effects were enhanced as the concentration increasing and the time prolonging, the effects to the latter were higher than those to the former, showing significant difference between them ( P <0.05 ). However, the effect of EGb761 was varied with the concentrations. EGb761 could promote T lymphocytes proliferation at low concentration but inhibit it at high concentration, there was a significant difference as compared with that in the control group ( all P < 0. 05). The apoptotic rate of T lymphocytes rose as the concentration of BN-52021 increasing (P < 0. 01).

CONCLUSIONGBE has different effects on T lymphocytes proliferation since the different ingredients and the concentrations in vitro, and it also has different effects between healthy and asthmatic rats. Ginkgolide B is the main active ingredient among them, it can not only inhibit T lymphocytes proliferation but also increase the apoptotic rate.

Animals ; Apoptosis ; drug effects ; Asthma ; drug therapy ; immunology ; pathology ; Cell Proliferation ; drug effects ; Ginkgo biloba ; Plant Extracts ; pharmacology ; therapeutic use ; Rats ; Rats, Sprague-Dawley ; T-Lymphocytes ; drug effects

OBJECTIVETo explore an alternative method for easier and more effective establishment of allergen-specific T-cell clones (TCC) from peripheral blood monouclear cells (PBMCs) of allergic asthma patients with allogeneic feeding cells.

METHODSTo determine the optimal condition for T cell growth and effective dose and time of mitomycin-C (MMC) treatment of the feeding cells to prevent their proliferation, the PBMCs were treated with PHA, IL-2 or MMC at different concentrations, and the proliferation rate of the treated cells was analyzed by MTT assay. The effect of IL-4 on the growth and subset selection of TCC was also analyzed. Allergen-specific TCC was established by limiting dilution method with allogeneic PBMCs as the feeding cells, and the proliferation of the allergen-specific TCC was observed to evaluate the feasibility of the feeding cells.

RESULTSPHA at 25 microg/ml and IL-2 at 27 U/ml achieved optimal growth of the T cells, while MMC treatment at the dose of 60 microg/ml for 80 min effectively enriched the non-proliferative feeding cell from the PBMCs. IL-4 could not promote the survival of the TCC, but promoted the formation of CD(4)(+) TCC. Allergen-specific TCC obtained using allogeneic feeding cells required the presence of PHA, but the allergen reactivity of the TCC remained unpredictable.

CONCLUSIONIL-4 can promote the formation of CD(4)(+) TCC, but allogeneic feeding cells may fail to produce TCC with high allergen specificity.

Allergens ; immunology ; Asthma ; blood ; Cell Culture Techniques ; Cell Proliferation ; drug effects ; Cells, Cultured ; Clone Cells ; cytology ; drug effects ; immunology ; Humans ; Interleukin-2 ; pharmacology ; Leukocytes, Mononuclear ; cytology ; drug effects ; immunology ; Mitomycin ; pharmacology ; T-Lymphocyte Subsets ; cytology ; drug effects ; immunology ; T-Lymphocytes ; cytology ; drug effects ; immunology ; T-Lymphocytes, Regulatory ; cytology ; drug effects ; immunology

OBJECTIVETo study the effect of rapamycin in inducing naïve murine effector T cell (Teff) convert to regulatory T cell (Treg) in vitro.

METHODSThe forkhead box protein 3 (FoxP3) negative Teff were isolated and purified from the spleen and lymph node of C57 BL/6 murines aged 6-8 weeks, then Teff were cultured in three groups with mature dendritic cells (mDC), B cells, and plate coated Anti-CD3. In addition, the control wells and the test wells were prepared in each group, rapamycin were not added in the control wells but added in the test wells with concentrations of 1, 10, 50, and 100 nmol/L. Percentages of FoxP3 positive Treg were examined by flow cytometry after 4 days in Anti-CD3 group and after 6 days in the other two groups.

RESULTSAs shown by the flow cytometry, the percentages of FoxP3 positive Treg were as follows in three group: in the mDC group, it was 0.01% in the control well and 0.39%, 0.47%, 0.34%, and 0.26% in test wells; in B cell group, it was 0.01% in the control wells and 5.56%, 5.89%, 7.15%, and 4.72% in the test wells; in Anti-CD3 group, it was 0.93% in the control wells and 1.35%, 1.07%, 1.02%, and 1.19% in test wells. No significant difference was found between the test wells and control wells in the mDC group and Anti-CD3 group; however, the percentages of FoxP3 positive Treg was significantly different between the test wells and control wells in the B cell group (P < 0.01).

CONCLUSIONWhen B cell is acted as the antigen-presenting cell, rapamycin can effectively induce Teff convert to Treg in vitro.

Animals ; B-Lymphocytes ; cytology ; drug effects ; immunology ; Cell Differentiation ; drug effects ; Cells, Cultured ; Dendritic Cells ; cytology ; drug effects ; immunology ; Flow Cytometry ; Forkhead Transcription Factors ; immunology ; Mice ; Mice, Inbred C57BL ; Mice, Inbred DBA ; Precursor Cells, T-Lymphoid ; cytology ; drug effects ; immunology ; Sirolimus ; pharmacology ; T-Lymphocyte Subsets ; cytology ; drug effects ; immunology ; T-Lymphocytes, Regulatory ; cytology ; drug effects ; immunology

The essential effect of vitamin A on immune function occurs through various mechanisms including direct effect on Th1-Th2 balance modulation. However, it is unclear whether or not vitamin A can regulate Th1-Th2 balance under a strong Th1-polarizing condition. Therefore, the purpose of our study was to examine the effect of vitamin A metabolite all-trans retinoic acid (ATRA) on Th1-Th2 differentiation in CD4+ T cells under GATA-3 deficiency, which can induce Th1-polarizing condition. In the present study, GATA-3 deficiency T cells were induced by siRNA and checked by real-time quantitative PCR and western blot. GATA-3 deficiency CD4+ T cells and normal CD4+ T were treated for 48 h with or without ATRA. The expression of Th1 and Th2 cytokines were detected by qPCR and ELISA. The results would contribute to clarify the knowledge of the role of vitamin A in regulating Th1-Th2 balance under some special conditions, and help to explain the mechanism of immune regulatory function of vitamin A.
CD4-Positive T-Lymphocytes ; drug effects ; Cell Differentiation ; drug effects ; Cells, Cultured ; GATA3 Transcription Factor ; deficiency ; Humans ; Th1-Th2 Balance ; drug effects ; Tretinoin ; pharmacology