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Suppression of CD8~+T Cell Proliferation and Activation by Regulatory Dendritic Cells and the Underlying Mechanisms

Author: DingYuanYuan
Tutor: CaoXueTao
School: Second Military Medical University
Course: Immunology
Keywords: Regulatory dendritic cells CD8 ~ T cells Nitric oxide Immunomodulatory
CLC: R392
Type: Master's thesis
Year: 2010
Downloads: 200
Quote: 1
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Abstract

The body functions as far as we know, the strongest professional antigen-presenting cells (Antigen-presenting cell, APC), dendritic cells (Dendritic cell, DC) in antigen recognition, start the immune response or induce immune tolerance plays an important role. DC as a heterogeneous population of cells, due to the presence of anatomical sites and functional status, divided into many subsets. These subgroups at different stages of maturity can be rendered a different phenotype, expression of multiple cytokines, and its function diversity. The previous DC awareness is limited to activate the function of the immune aspects, but in recent years a large number of studies have shown that DC also has negative regulation of immune function and the type of DC called regulatory DC (Regulatory DC) of the experimental chamber found in previous studies, was considered to be terminally differentiated cells mature DC can further proliferation, and differentiation as a new class of regulatory dendritic cells in co-culture with the stromal cells of the spleen, the name for the differentiation of dendritic cells (Differentiated the dendritic cells diffDC). DC The group can inhibit the proliferation of CD4 T cells through the secretion of NO, but not significant induction of regulatory T cells generated. diffDC discovery, breaking the previous standard definition of terminally differentiated cells: mature DC after the completion of the antigen-presenting tasks in secondary lymphoid organ microenvironment can further proliferation and differentiation was given new life and function. T cells are important effector cells in the immune system. Identification associated antigen naive T cells in dendritic cells and other antigen presenting cells, occurred cascade amplified, to stimulate the antigen-specific cellular immunity and humoral immunity, and thus play an important role in the resilience pathogens . , CD8 T cells are the main cellular immune effector cells. CD8 cytotoxic T cells (cytotoxic T lymphocyte, CTL) cell subsets, immune killing effect can specifically recognize the antigen, under limited conditions in MHCI molecules direct, continuous and specifically kill target cells. But in the immune response, effector CD8 T cells unlimited amplification will lead to adverse consequences such as autoimmune diseases, the study of the regulation of CD8 T cell immune negative is particularly important. DC in the proliferation and activation of CD8 T cells play a regulatory role in how and what the specific mechanism is not yet fully understood. Around the laboratory in the past found regulatory of DC (diffDC) on the effects and mechanism of regulation of CD8 T cells. First, we studied the diffDC proliferation of OVA-specific CD8 T cells, the activation of the results found diffDC able to inhibit the proliferation and activation of CD8 T cells. After that, we further inhibition mechanism of CD8 T cells of diffDC preliminary discussion. A regulatory dendritic cells inhibit the proliferation and activation of CD8 T cells in this part of the experiment, we studied the regulatory role of dendritic cells in the regulation of CD8 T cell function. OT-I transgenic mice source of CD8 T cells specifically recognize OVA257-264 peptide, a good reaction cells in vitro validation the single antigen peptide-specific antigen-presenting reaction. The we adopt OT-I mice Source of CD8 T cells as responder cells, C57BL / 6 (H-2Kb) mice Source load OVA257-264 polypeptide DC as stimulator cells. For to detect diffDC function in the system, we will different proportions diffDC Join in the co-culture system (maDC/CD8), diffDC significant inhibition of CD8 T cells induced by mature DC (mature DC, maDC) proliferation, and such inhibition was diffDC cell number-dependent. CFSE-labeled CD8 T cells have also been the same experimental results. Analysis of cell cycle status also found similar results: Join diffDC after co-culture system in reducing the proportion of the active phase of the cell cycle cell, CD8 T cells. These results suggest that diffDC inhibit the proliferation of CD8 T cells. Activated phenotype CD8 T cells in the co-culture system and culture supernatant cytokine analysis, we found that the suppression of CD8 T cell activation phenotype CD25 upregulation diffDC and reduced CD8 T cell CD62L expression levels. CD8 T cell secretion of cytokines IL-2 is also decreased. We also make use of real-time quantitative PCR detected the after co-cultured CD8 T cells granzyme B and perforation prime the expression level, found that total training system added diffDC, CD8 T cells expressed granzyme B and perforation prime significantly with reducing. These results indicate that diffDC inhibit the activation of CD8 T cells. The most significant features of the regulatory T cells that inhibit the proliferation of other T cells receive antigen stimulation. Then, diffDC through the induction of regulatory T cells generated thereby inhibiting the proliferation and activation of CD8 T? Our detection ability of CD122 CD8 regulatory T cells generated diffDC induced results found, diffDC did not induce regulatory T cells in CD122 CD8 . The above results suggest that the proliferation and activation of diffDC specific CD8 T cells may play an inhibitory effect. This inhibition is likely to maintain a relatively tolerant immune status of CD8 T cells, in order to maintain the balance and stability of the immune system. Regulatory dendritic cells suppress CD8 T cell proliferation and activation mechanism study in this section, we diffDC how to suppress the CD8 T cell proliferation and activation mechanism. The most significant features of the regulatory T cells that inhibit the proliferation of other T cells receive antigen stimulation. Then, diffDC can induce regulatory T cells generated thereby indirectly inhibit the proliferation and activation of CD8 T? We have detected diffDC induced regulatory T cells generated in CD122 CD8 ability, found diffDC did not induce CD122 CD8 adjustment T cells produce. CD8 T cell proliferation was inhibited, it may be because diffDC promote T cell apoptosis. We detected. The results showed that the case with diffDC presence, and no significant increase in the proportion of CD8 T-7-AAD, apoptotic cells. T cells after deactivation, will lose the reactivity to the antigen. Play the role of one of the mechanisms of regulatory DC induced T cells disability or low reactivity. Therefore, we have such a role diffDC whether studied. We first specific CD8 T and load the OVA257-264 maDC or a diffDC of culture, 2 days later with beads male selected CD8 T cells, in the presence of IL-2 under the conditions of the resting 2 days after restimulation with DC as a result, not affected by diffDC after induction of CD8 T again stimulated proliferation ability, did not significantly reduce the RNA levels of the intracellular granzyme B. The description diffDC can not induce CD8 T cells disability. To determine whether diffDC produce soluble molecules or membrane molecules, or membrane molecules with maDC or activation of CD8 T cells on the role of membrane molecules diffDC the soluble factors produced thereby inhibiting its role in CD8 T cell-specific proliferation and activation, we use the Transwell system will diffDC maDC/CD8T spaced then incubated and observed results found diffDC with maDC/CD8T cells spaced, which inhibit the ability of the OVA-specific CD8 T cell activation reduces, but inhibit CD8 T proliferation ability is not completely lost, to show that diffDC for it is required to inhibit the activation of CD8 T cells role in cell-cell contact mediated, namely: directly by membrane molecules diffDC, or indirectly, by the film on a molecule in the diffDC generated after maDC or activated CD8 T cells, the role of the soluble factor mediated; inhibit the proliferation of CD8 T cells is required to double the role of the soluble molecules and membrane molecules. diffDC an important surface markers is high expression of CD11b. CD11b blocking antibody added to the proliferation of CD8 T costimulatory system in diffDC suppression, no effective to reverse the diffDC proliferation of antigen-specific CD8 T CD11b may not be directly involved in diffDC of antigen-specific The inhibitory effect of CD8 T. We found in the experiment, co-cultured with diffDC CD8 T cell surface expression of FasL and there is no difference between the control group, but increased the expression of Fas think diffDC constitutively high expression of FasL characteristics guess it may be caused diffDC of antigen reasons for the suppression of proliferation of specific CD8 T cells. However, in the experiment, we found that FasL blocking antibody was added in the reaction system, can not to reversal diffDC inhibited the proliferation of antigen-specific CD8 T. diffDC high expression of FasL perhaps more significance to play an important role in other aspects. Studies have reported that NO can block IL-2 downstream signaling pathways, thereby inhibiting the proliferation of T cells, and thereby be inspired to detect each of the co-culture system of NO content was found, diffDC/maDC/CD8T co-culture system The high amount of NO secretion, while the NO in the control group is much lower than the group, which suggests that NO may be inhibition of T-cell proliferation effector molecule. We use the L-arginine analogue L-NAME blocking the synthesis of NO, the results show that the proliferation of CD8 T cell inhibition is reversed. Transwell chamber diffDC and maDC/CD8T spaced, the NO concentration in the system is greatly reduced. These results suggest that NO produced by the cell membrane diffDC and maDC/CD8T contact mediated by CD8 T cells specific of diffDC of maDC induced proliferation inhibition. These results suggest that inhibition diffDC CD8 T cell activation depends on the direct contact between diffDC mature DC and CD8 T CD8 T cell proliferation negative regulation of the dual role of the cells in direct contact and soluble molecules. The diffDC can be generated through the induction of NO to the proliferation of CD8 T cell response plays a negative regulatory role summary, diffDC NO produced by cell contact to suppress the proliferation and activation of specific CD8 T cells play a negative immune regulatory functions. The design and application of the results of this study will contribute to a better understanding of the DC regulatory mechanisms involved in immune regulation, for the study of the body's self-tolerance, explore oncology and autoimmune diseases, such as clinical disease pathogenesis and immunotherapy provides a new research approaches and theoretical basis.

Suppression of CD8~+T Cell Proliferation and Activation by Regulatory Dendritic Cells and the Underlying Mechanisms

Abstract

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