Generation and Regulation of CD8＋ Regulatory T Cells

Research into the suppressive activity of CD4＋FoxP3＋ T regulatory cells （Treg） has defined a sublineage of CD4＋ cells that contribute to self-tolerance and resistance to autoimmune disease. Much less attention has been given to the potential contribution of regulatory sublineages of CD8＋ cells. Analysis of a small fraction of CD8＋ cells that target autoreactive CD4＋ cells through recognition of the MHC class Ib molecule Qa-1 in mouse and HLA-E in human has revitalized interest in CD8＋ Treg. Here we summarize recent progress and future directions of research into the role of this CD8＋ sublineage in resistance to autoimmune disease. Cellular ＆ Molecular Immunology. 2008; 5（6）：401-406.     

Bacillus Calmette-Guérin Suppresses Asthmatic Responses via CD4+CD25+ Regulatory T Cells and Dendritic Cells

Purpose

Bacillus Calmette-Guérin (BCG) is known to suppress the asthmatic responses in a murine model of asthma and to induce dendritic cells (DCs) maturation. Mature DCs play a crucial role in the differentiation of regulatory T cells (Tregs), which are known to regulate allergic inflammatory responses. To investigate whether BCG regulates Tregs in a DCs-mediated manner, we analyzed in a murine model of asthma.

Methods

BALB/c mice were injected intraperitoneally with BCG or intravenously with BCG-stimulated DCs and then sensitized and challenged with ovalbumin (OVA). Mice were analysed for bronchial hyperresponsiveness (BHR), the influx of inflammatory cells in the bronchoalveolar lavage (BAL) fluid, and histopathological changes in the lung. To identify the mechanisms, IgE, IgG₁ and IgG_2a in the serum were analysed and the CD25₊ Tregs in the mice were depleted with anti-CD25 monoclonal antibody (mAb).

Results

BCG and the transfer of BCG-stimulated DCs both suppressed all aspects of the asthmatic responses, namely, BHR, the production of total IgE and OVA-specific IgE and IgGs, and pulmonary eosinophilic inflammation. Anti-CD25mAb treatment reversed these effects.

Conclusions

BCG can attenuate the allergic inflammation in a mouse model of asthma by a Tregs-related mechanism that is mediated by DCs.     

Pleural Mesothelial Cells Promote Expansion of IL-17–Producing CD8+ T Cells in Tuberculous Pleural Effusion

IL-17–producing CD8⁺ T lymphocytes (Tc17 cells) have recently been detected in many cancers and autoimmune diseases. However, the possible implication of Tc17 cells in tuberculous pleural effusion remains unclarified. In this study, distribution and phenotypic features of Tc17 cells in both tuberculous pleural effusion (TPE) and peripheral blood from patients with tuberculosis were determined. The effects of proinflammatory cytokines and local accessory cells (pleural mesothelial cells) on Tc17 cell expansion were also explored. We found that TPE contained more Tc17 cells than the blood. Compared with IFN-γ–producing CD8⁺ T cells, Tc17 cells displayed higher expression of chemokine receptors (CCRs) and lower expression of cytotoxic molecules. In particularly, Tc17 cells in TPE exhibited high expression levels of CCR6, which could migrate in response to CCL20. Furthermore, IL-1β, IL-6, IL-23, or their various combinations could promote Tc17 cell expansion from CD8⁺ T cells, whereas the proliferative response of Tc17 cells to above cytokines was lower than that of Th17 cells. Pleural mesothelial cells (PMCs) were able to stimulate Tc17 cell expansion via cell contact in an IL-1β/IL-6/IL-23 independent fashion. Thus this study demonstrates that Tc17 cells marks a subset of non-cytotoxic, CCR6⁺ CD8⁺ T lymphocytes with low proliferative capacity. The overrepresentation of Tc17 cells in TPE may be due to Tc17 cell expansion stimulated by pleural proinflammatory cytokines and to recruitment of Tc17 cells from peripheral blood. Additionally, PMCs may promote the production of IL-17 by CD8⁺ T cells at sites of TPE via cell–cell interactions.     

Naturally Occurring Self-Reactive CD4^＋CD25^＋ Regulatory T Cells： Universal Immune Code

Naturally occurring thymus-arisen CD4^＋CD25^＋ regulatory T （Treg） cells are considered to play a central role in self-tolerance. Precise signals that promote the development of Treg cells remain elusive, but considerable evidence suggests that costimulatory molecules, cytokines, the nature of the TCR and the niche or the context in which the T cell encounters antigen in the thymus play important roles. Analysis of TCR from Treg cells has demonstrated that a large proportion of this population has a higher avidity to self-antigen in comparison with TCR from CD4^＋CD25^＋ cells and that peripheral antigen is required for their development, maintenance, or expansion. Treg cells have been shown to undergo expansion in the periphery, likely regulated by the presence of self-antigen. Many studies have shown that the involvement of Treg cells in the tolerance induction is antigen-specific, even with MHC-mismatched, in transplantation/graft versus host disease （GVHD）, autoimmunity, cancer, and pregnancy. Theses studies concluded a vital role for self-reactive Treg cells in maintenance of the body integrity. Based on those studies, we hypothesize that self-reactive Treg cells are shared among all healthy individuals and recognize same self-antigens and their TCR encodes for few dominant antigens of each organ which defines the healthy self. These dominant self antigens can be regarded as ＂universal immune code＂. Cellular ＆ Molecular Immunology.     

The ex vivo Microenviroments in MLTC of Poorly Immunogenic Tumor Cells Facilitate Polarization of CD4^＋ CD25^＋ Regulatory T Cells

CD4^＋CD25^＋ regulatory T （TR） cells play an important role in maintaining a balanced peripheral immune system. Recent studies have shown that TR cells may also play a key role in suppressing anti-tumor immune response. In order to investigate the tumor immune microenvironment and its influence on TR polarization, poorly immunogenic tumor cell line Ds （C57BL/6, H-2^b）, immunogenic tumor cell lines FBL3 （C57BL/6, H-2^b） and H22 BALB/c, H-2^d） were used to establish the syngeneic/allogeneic, poorly immunogenic/immunogenic mixed lymphocytes-tumor cell culture （MLTC）. Our results revealed that the proportion of CD4^＋CD25^＋ T cells in MLTC of syngeneic primed splenocytes stimulated with D5 tumor cells was higher than that with H22 cells （0.43% vs 0.044%, and the similar results appeared in allogeneic splenocytes stimulated with D5 tumor cells （0.39% vs 0.04%）. The splenocytes stimulated with supernatant from syngeneic MLTC of D5 tumor cells demonstrated higher proportion of CD4^＋CD25^＋ cells than that from allogeneic MLTC of D5 tumor cells, and the splenocytes stimulated with supernatant from syngeneic or allogeneic MLTC of H22 tumor cells generated lower proportion of CD4^＋CD25^＋ T cells than that of D5 tumor cells. The TGF-β1 and Th2-oriented cytokines （IL-4 and IL-10） were dominated in supernatants of syngeneic MLTC of poorly immunogenic tumor cells. Our results provided useful information for studying the mechanisms underlying tumor immune surveillance as well as for the tumor immunotherapy.     

Epigenomic-Guided Mass Cytometry Profiling Reveals Disease-Specific Features of Exhausted CD8 T Cells

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CD4+LAP+ and CD4+CD25+Foxp3+ Regulatory T Cells Induced by Nasal Oxidized Low-Density Lipoprotein Suppress Effector T Cells Response and Attenuate Atherosclerosis in ApoE−/− Mice

Increasing studies have demonstrated that atherosclerosis is a chronic immunoinflammatory disease, and that oxidized low-density lipoprotein (oxLDL)-specific T cells contribute to the autoimmune process in atherosclerosis. Oral administration of oxLDL, which was identified as a candidate autoantigen in atherosclerosis, was shown to induce tolerance and suppress atherogenesis. However, the precise mechanisms of mucosal tolerance induction, in particular nasal tolerance, remain unknown. In this study, we explored the effect of nasal oxLDL on atherosclerosis as well as the cellular and molecular mechanisms leading to atheroprotective responses, and then found that nasal oxLDL drastically ameliorate the initiation (47.6 %, p?p?=?0.001) of atherosclerosis. Most importantly, a significant 35.8 % reduction of the progression of atherosclerosis was observed in the enhanced immunization group (p?+ latency-associated peptide (LAP)⁺ regulatory T cells (Tregs) and CD4⁺CD25⁺Foxp3⁺ Tregs in spleens and cervical lymph nodes, together with increased transforming growth factor (TGF)-β production and suppressed T-helper cells type 1, 2, and 17 immune responses. Surprisingly, neutralization of TGF-β in vivo partially counteracted the protective effect of nasal oxLDL treatment, indicating that the presence of TGF-β was indispensable to CD4⁺LAP⁺ Tregs and CD4⁺CD25⁺Foxp3⁺ Tregs to acquire regulatory properties. Our studies suggest that CD4⁺LAP⁺ Tregs and CD4⁺CD25⁺Foxp3⁺ Tregs induced by nasal delivery of oxLDL can inhibit oxLDL-specific T cells response and ameliorate atherosclerosis process.     

CD4+CD25+CD127low/? T Cells: A More Specific Treg Population in Human Peripheral Blood

The quantitative identification and enrichment of viable regulatory T cells (Treg) requires reliable surface markers that are selectively expressed on Treg. Foxp3 is the accepted marker of nTreg, but it cannot be used to isolate cells for functional studies. In this study, we compared four staining profiles of Treg, including CD4⁺CD25^high T cells, CD4⁺CD39⁺ T cells, CD4⁺CD73⁺ T cells, and CD4⁺CD25⁺CD127^low/? T cells. We found that CD4⁺CD25⁺CD127^low/? T cells expressed the highest level of Foxp3 and had the strongest correlation with CD4⁺CD25⁺Foxp3⁺ T cells, the accepted identifying characteristics for ??real?? nTreg cells. Moreover, functional data showed that CD4⁺CD25⁺CD127^low/? T cells could effectively suppress the proliferation of CD4⁺CD25^? T cells, suggesting that compared with the other three populations, CD4⁺CD25⁺CD127^low/? T cells best fit the definition of naturally occurring regulatory T cells in human peripheral blood. Finally, we showed that CD4⁺CD25⁺CD127^low/? can be used to quantitate Treg cells in individuals with systemic lupus erythematosus supporting the use of CD4⁺CD25⁺CD127^low/? to identify human Treg cells.     

Self-Reactive CD4+ T Cells and B Cells in the Blood in Health and Autoimmune Disease: Increased Frequency of Thyroglobulin-Reactive Cells in Graves’ Disease

The mechanisms underlying activation of potentially self-reactive circulating B cells and T cells remain unclear. We measured the uptake of a self-antigen, thyroglobulin, by antigen presenting cells, and the subsequent proliferation of CD4⁺ T cells and B cells from healthy controls and patients with autoimmune thyroiditis. In Hashimoto's thyroiditis, B cells bound increased amounts of thyroglobulin in a complement- and autoantibody-dependent manner, and the thyroglobulin-elicited proliferation of CD4⁺ T cells and B cells was complement dependent. Increased proportions of Tg-responsive CD4⁺ T cells and B cells were found in patients with Graves’ disease. Notably, both patient groups and healthy controls exhibited higher proliferative responses to thyroglobulin than to a foreign recall antigen, tetanus toxoid. Our results suggest that self-tolerance can be broken by exposure of circulating lymphocytes to high local concentrations of self-antigen, and that complement plays a role in the maintenance of autoimmune processes, at least in Hashimoto's thyroiditis.     

Thymic Nurse Cells Support CD4^-CD8^＋ Thymocytes to Differentiate into CD4^＋CD8^＋ Cells

Thymic nurse cells （TNCs） represent a unique microenvironment in the thymus for T cell maturation. In order to investigate the role of thymic nurse cells during T cell differentiation, a TNC clone, RWTE-1, which formed a typical complex with fetal thymocytes in vitro was established from normal Wistar rat. Hanging drop culture method was applied to reveal the interaction between TNCs and thymocytes. Our result revealed that eighty percent of immature CD4^-CD8^＋ cells differentiated into CD4^＋CD8^＋ cells after a 12-hour hanging drop culture with RWTE-1. However, in a 12-hour culture of immature CD4^-CD8^＋ cells with or without RWTE-1 supernatant, only 30% of the cells differentiated into CD4^＋CD8^＋ cells spontaneously. This observation led to the conclusion that RWTE-1 cell has the capacity to facilitate immature CD4^-CD8^＋ thymocytes to differentiate into CD4^＋CD8^＋ T cells by direct interaction.     

Type 1 CD8^＋ T Cells are Superior to Type 2 CD8^＋ T Cells in Tumor Immunotherapy due to Their Efficient Cytotoxicity, Prolonged Survival and Type 1 Immune Modulation

CD8^＋ cytotoxic T （Tc） cells play a crucial role in host immune responses to cancer, and in this context, adoptive CD8^＋ Tc cell therapy has been studied in numerous animal tumor models. Its antitumor efficacy is, to a large extent, determined by the ability of Tc cells to survive and infiltrate tumors. In clinical trials, such in vitro-activated T cells often die within hours to days, and this greatly limits their therapeutic efficacy. CD8^＋ Tc cells fall into two subpopulations based upon their differential cytokine secretion. In this study, we in vitro generated that ovalbumin （OVA）-pulsed dendritic cell （DCovA）-activated CD8^＋ type 1 Tc （Tcl） cells secreting IFN-T, and CD8^＋ type 2 Tc （Tc2） cells secreting IL-4, IL-5 and IL-10, which were derived from OVA-specific T cell receptor （TCR） transgenic OT I mice. We then systemically investigated the in vitro and in vivo effector function and survival of Tcl and Tc2 cells, and then assessed their survival kinetics after adoptively transferred into C57BL/6 mice, respectively. We demonstrated that, when compared to CD8^＋ Tc2, Tcl cells were significantly more effective in perforin-mediated cytotoxicity to tumor cells, had a significantly higher capacity for in vivo survival after the adoptive T cell transfer, and had a significantly stronger therapeutic effect on eradication of well-established tumors expressing OVA in animal models. In addition, CD8^＋ Tcl and Tc2 cells skewed the phenotype of CD4^＋ T cells toward Thl and Th2 type, respectively. Therefore, the information regarding the differential effector function, survival and immune modulation of CD8^＋ Tcl and Tc2 cells may provide useful information when preparing in vitro DC-activated CD8^＋ T cells for adoptive T cell therapy of cancer.     

How do Regulatory T Cells Work?

CD4⁺ T cells are commonly divided into regulatory T (Treg) cells and conventional T helper (Th) cells. Th cells control adaptive immunity against pathogens and cancer by activating other effector immune cells. Treg cells are defined as CD4⁺ T cells in charge of suppressing potentially deleterious activities of Th cells. This review briefly summarizes the current knowledge in the Treg field and defines some key questions that remain to be answered. Suggested functions for Treg cells include: prevention of autoimmune diseases by maintaining self-tolerance; suppression of allergy, asthma and pathogen-induced immunopathology; feto-maternal tolerance; and oral tolerance. Identification of Treg cells remains problematic, because accumulating evidence suggests that all the presently-used Treg markers (CD25, CTLA-4, GITR, LAG-3, CD127 and Foxp3) represent general T-cell activation markers, rather than being truly Treg-specific. Treg-cell activation is antigen-specific, which implies that suppressive activities of Treg cells are antigen-dependent. It has been proposed that Treg cells would be self-reactive, but extensive TCR repertoire analysis suggests that self-reactivity may be the exception rather than the rule. The classification of Treg cells as a separate lineage remains controversial because the ability to suppress is not an exclusive Treg property. Suppressive activities attributed to Treg cells may in reality, at least in some experimental settings, be exerted by conventional Th cell subsets, such as Th1, Th2, Th17 and T follicular (Tfh) cells. Recent reports have also demonstrated that Foxp3⁺ Treg cells may differentiate in vivo into conventional effector Th cells, with or without concomitant downregulation of Foxp3.     

CD4+ T Cells and IFN-γ Are Required for the Development of Pneumocystis-Associated Pulmonary Hypertension

Pulmonary hypertension (PH) is a disease of diverse etiology. Although primary PH can develop in the absence of prior disease, PH more commonly develops in conjunction with other pulmonary pathologies. We previously reported a mouse model in which PH occurs as a sequela of Pneumocystis infection in the context of transient CD4 depletion. Here, we report that instead of the expected Th2 pathways, the Th1 cytokine IFN-γ is essential for the development of PH, as wild-type mice developed PH but IFN-γ knockout mice did not. Because gene expression analysis showed few strain differences that were not immune-function related, we focused on those responses as potential pathologic mechanisms. In addition to dependence on IFN-γ, we found that when CD4 cells were continuously depleted, but infection was limited by antibiotic treatment, PH did not occur, confirming that CD4 T cells are required for PH development. Also, although CD8 T-cells are implicated in the pathology of Pneumocystis pneumonia, they did not have a role in the onset of PH. Finally, we found differences in immune cell phenotypes that correlated with PH, including elevated CD204 expression in lung CD11c⁺ cells, but their role remains unclear. Overall, we demonstrate that a transient, localized, immune response requiring IFN-γ and CD4-T cells can disrupt pulmonary vascular function and promote lingering PH.Pulmonary hypertension (PH) is a devastating disease with complex etiology and, in all likelihood, diverse mechanisms of pathology. A recent reclassification of the types of PH involves five major divisions, including forms associated with specific causative agents (such as drugs), hypoxia, and infectious agents (such as schistosomes).¹ A common feature of many of these agents is that they initiate local inflammation, which may act as a trigger for the development of PH,^2–4 even if the inflammation does not persist after the manifestation of PH. However, there does not seem to be any single inflammatory mediator responsible for the inflammatory initiation of PH. For example, several immune cell types (T cells, B cells, and macrophages) and inflammatory cytokines (TGF-β, IL-1β, IL-6, RANTES, and IL-13) have been implicated in various forms of PH.^5–9In the T helper 1 and T helper 2 (Th1 and Th2) paradigm, CD4⁺ Th2 cells drive an immune response characterized by the production of cytokines such as IL-4, IL-5, and IL-13, as well as by secreted antibody (in particular, IgE).¹⁰ In several studies using animal models, a strong case has been made for a role of Th2 immune responses as instigators of PH. For example, a Th2 response associated with sensitization to an antigen and subsequent challenge with that antigen can result in muscularization of smaller pulmonary arteries, and this response is associated with CD4⁺ cells and IL-13.¹¹ The protein resistin-like alpha (Retnla; alias cysteine-rich secreted protein FIZZ1) can be induced by hypoxia (which is associated with vascular remodeling¹²), but it is also induced in Th2 immune responses; in some cases, the Th2-associated molecule Retn1a appears to have a strong association with vascular remodeling and resultant PH,^13,14 which may be related to its induction by hypoxia, itself a potent stimulator of vascular remodeling.¹⁴ An interesting mouse model of PH associated with repeated inhalation of spores of the fungus Stachybotrys chartarum also is associated with the Th2 cytokines IL-4 and IL-5, but not the Th1 cytokine IFN-γ.¹⁵ Finally, in what is probably one of the best-known examples of PH in conjunction with an infectious agent, schistosomiasis-induced PH appears to be associated with the Th2 cytokine IL-13.¹⁶ Furthermore, IL-13 is implicated in several other forms of PH.¹⁷In contrast, Th1 immune responses, which are characterized by the secretion of cytokines such as IFN-γ and TNF-α and activation of phagocytic macrophages, appear to have little connection with the development of vascular remodeling and PH. Despite a few reports of elevated TNF-α in conjunction with clinical syndromes that include PH,^7,18 there is very little association of PH with the canonical Th1 cytokine, IFN-γ, although there is one report of IFN-γ having a synergistic effect with other cytokines on in vitro pulmonary vascular cell remodeling.¹⁹ Indeed, this lack of effect is illustrated by the fact that IFN-γ has been used in clinical treatment of idiopathic pulmonary fibrosis,²⁰ although with little effectiveness, even though ≤40% of patients with idiopathic pulmonary fibrosis also exhibit PH.²¹Recently, we reported that PH developed in the aftermath of a resolved Pneumocystis pneumonia in mice, in the context of a transient depletion of CD4⁺ cells.²² At the time, it was unclear which immune responses are involved in the development of PH in that mouse model; although there were elevated levels of Retnla (FIZZ1) in the bronchoalveolar lavage fluid (BALF) of mice that developed PH, as well as some perivascular fibrosis, IL-4 signaling was not required for these developments.²² In the present study, surprisingly, we found that the Th1 cytokine IFN-γ is absolutely required for the development of PH in this mouse model, and that perivascular fibrosis does not appear to be a cause of PH. Furthermore, having previously established that onset of PH in these mice is correlated with the resurgence of CD4⁺ T cells after depletion,²² with the present study we have demonstrated that CD4⁺ cells are also absolutely required for the development of PH.     

Neuropilin-1: The Glue between Regulatory T Cells and Dendritic Cells?

The interaction between dendritic cells and regulatory T cells is critical for the maintenance of self-tolerance. In this issue of Immunity, Sarris et al. (2008) find that Neuropilin-1 contributes to the prolonged interaction of regulatory T cells with dendritic cells.     

CD1限制性T细胞

在检测感染递呈抗原的细胞浆质时，CD1分子能与外来的脂类抗原结合。与T细胞识别CD1限制性外来脂质不同，CD1限制性T细胞具有自身抗原反应功能，作为自身效应器它可快速被激活，在CD1表达的递呈抗原细胞之间起到辅助和效应器功能。CD1l限制性T细胞亚群及其通路之间功能上的差异表现在这些细胞既能影响树突状细胞(DC)的功能和耐受作用，同时还能激活自然杀伤(NK)细胞和其它淋巴细胞，从而揭示了CD1限制性T细胞如何调节抗微生物应答、抗肿瘤免疫和平衡免疫耐受、自身免疫。     

CD1限制性T细胞

在检测感染递呈抗原的细胞浆质时,CD1分子能与外来的脂类抗原结合。与T细胞识别CD1限制性外来脂质不同,CD1限制性T细胞具有自身抗原反应功能,作为自身效应器它可快速被激活,在CD1表达的递呈抗原细胞之间起到辅助和效应器功能。CD1限制性T细胞亚群及其通路之间功能上的差异表现在这些细胞既能影响树突状细胞(DC)的功能和耐受作用,同时还能激活自然杀伤(NK)细胞和其它淋巴细胞,从而揭示了CD1限制性T细胞如何调节抗微生物应答、抗肿瘤免疫和平衡免疫耐受、自身免疫。