Instructive Cues of Thymic T Cell Selection.

A high diversity of αß T cell receptors (TCRs), capable of recognizing virtually any pathogen but also self-antigens, is generated during T cell development in the thymus. Nevertheless, a strict developmental program supports the selection of a self-tolerant T cell repertoire capable of responding to foreign antigens. The steps of T cell selection are controlled by cortical and medullary stromal niches, mainly composed of thymic epithelial cells and dendritic cells. The integration of important cues provided by these specialized niches, including (a) the TCR signal strength induced by the recognition of self-peptide-MHC complexes, (b) costimulatory signals, and (c) cytokine signals, critically controls T cell repertoire selection. This review discusses our current understanding of the signals that coordinate positive selection, negative selection, and agonist selection of Foxp3+ regulatory T cells. It also highlights recent advances that have unraveled the functional diversity of thymic antigen-presenting cell subsets implicated in T cell selection.

A regulatory circuit controlled by extranuclear and nuclear retinoic acid receptor α determines T cell activation and function.

Ligation of retinoic acid receptor alpha (RARα) by RA promotes varied transcriptional programs associated with immune activation and tolerance, but genetic deletion approaches suggest the impact of RARα on TCR signaling. Here, we examined whether RARα would exert roles beyond transcriptional regulation. Specific deletion of the nuclear isoform of RARα revealed an RARα isoform in the cytoplasm of T cells. Extranuclear RARα was rapidly phosphorylated upon TCR stimulation and recruited to the TCR signalosome. RA interfered with extranuclear RARα signaling, causing suboptimal TCR activation while enhancing FOXP3+ regulatory T cell conversion. TCR activation induced the expression of CRABP2, which translocates RA to the nucleus. Deletion of Crabp2 led to increased RA in the cytoplasm and interfered with signalosome-RARα, resulting in impaired anti-pathogen immunity and suppressed autoimmune disease. Our findings underscore the significance of subcellular RA/RARα signaling in T cells and identify extranuclear RARα as a component of the TCR signalosome and a determinant of immune responses.

Highlight of 2023: Tregs in the lungs.

In this article for the Highlights of the 2023 Series, we discuss recent discoveries on regulatory T cells in the lungs and their multifaceted roles in various contexts. Key advancements in Treg immunology have broadened our understanding of lung tissue homeostasis and the potential role of Tregs in pathological processes.

Stage-dependent immunity orchestrates AQP4 antibody-guided NMOSD pathology: a role for netting neutrophils with resident memory T cells in situ.

Neuromyelitis optica spectrum disorder (NMOSD) is an autoimmune disease of the CNS characterized by the production of disease-specific autoantibodies against aquaporin-4 (AQP4) water channels. Animal model studies suggest that anti-AQP4 antibodies cause a loss of AQP4-expressing astrocytes, primarily via complement-dependent cytotoxicity. Nonetheless, several aspects of the disease remain unclear, including: how anti-AQP4 antibodies cross the blood-brain barrier from the periphery to the CNS; how NMOSD expands into longitudinally extensive transverse myelitis or optic neuritis; how multiphasic courses occur; and how to prevent attacks without depleting circulating anti-AQP4 antibodies, especially when employing B-cell-depleting therapies. To address these knowledge gaps, we conducted a comprehensive 'stage-dependent' investigation of immune cell elements in situ in human NMOSD lesions, based on neuropathological techniques for autopsied/biopsied CNS materials. The present study provided three major findings. First, activated or netting neutrophils and melanoma cell adhesion molecule-positive (MCAM+) helper T (TH) 17/cytotoxic T (TC) 17 cells are prominent, and the numbers of these correlate with the size of NMOSD lesions in the initial or early-active stages. Second, forkhead box P3-positive (FOXP3+) regulatory T (Treg) cells are recruited to NMOSD lesions during the initial, early-active or late-active stages, suggesting rapid suppression of proinflammatory autoimmune events in the active stages of NMOSD. Third, compartmentalized resident memory immune cells, including CD103+ tissue-resident memory T (TRM) cells with long-lasting inflammatory potential, are detected under "standby" conditions in all stages. Furthermore, CD103+ TRM cells express high levels of granzyme B/perforin-1 in the initial or early-active stages of NMOSD in situ. We infer that stage-dependent compartmentalized immune traits orchestrate the pathology of anti-AQP4 antibody-guided NMOSD in situ. Our work further suggests that targeting activated/netting neutrophils, MCAM+ TH17/TC17 cells, and CD103+ TRM cells, as well as promoting the expansion of FOXP3+ Treg cells, may be effective in treating and preventing relapses of NMOSD.

Foxp3+ regulatory T cells reside within the corneal epithelium and co-localize with limbal stem cells.

In this study we investigated the presence of resident Foxp3+ regulatory T cells (Tregs) within the cornea and assessed the role of resident Tregs in corneal epithelial wound healing. Using a mouse model, we showed that in the steady state Foxp3+Tregs are either in close proximity or co-localize with ABCG2+ limbal stem cells. We also showed that these Tregs reside within the epithelial layer and not the corneal stroma. In addition, using a mouse model of mechanical injury, we demonstrated that depletion of Tregs from the cornea prior to corneal mechanical injury, using subconjunctival injection of anti-CD25, was associated with delayed epithelial healing. These results suggest a role for cornea resident Tregs in corneal epithelial cell function and wound healing and opens doors for further exploration of the role of Tregs in limbal stem cell function and survival.

Investigating the T regulatory cells and Sirtuin-I serum level in immunotherapy treated house dust mite allergic asthma patients.

OBJECTIVES: House dust mite aeroallergens are predominant triggers of frequent asthma attacks among adults and children. The intensity of asthma and immune reaction necessitates treatment alternatives based on adjusting chosen immunity biomarkers to control the exacerbation of symptoms and establish long-term immune tolerance. In this study, we selected CD4+CD25+Foxp3+ regulatory T cells (Tregs), FOXP3, and Sirtuin-1 as they are known to have a potential role in the immune reaction in different allergic diseases. We investigated their interplay during HDM allergic asthma and its respective immunotherapy. METHODS: Eighty-four subjects were divided into 3 groups; healthy controls (CT), HDM asthma patients without immunotherapy (WOIT), and HDM asthma patients treated with subcutaneous immunotherapy for 6 months before recruitment (WIT). They were enrolled according to the pulmonary function, skin prick tests, and HDM-specific IgE. CD4+ CD25+ and CD4+ CD25+ FOXP3+hi T cells Cell percentages, FOXP3 gene expression, and Sirtuin-1 (Sirt1) serum level were analyzed. RESULTS: We found that there is a significant difference between WOIT and WIT groups in the CD4+ CD25+ and CD4+ CD25+ FOXP3+hi T cell percentages. While there is no statistically significant difference between WOIT and WIT groups in FOXP3 level. On the controversy, the SIRT1 level in the CT group (4.53 ± 3.880) significantly decreased in the WOIT and WIT groups. CONCLUSION: This study revealed that both CD4 CD25 and CD4 CD25 high FOXP3 cell percentages increased in the WIT group and declined in the WOIT group. While, FOXP3 gene expression increased in both groups. In addition, the Sirt1 serum level showed some improvement in WIT group after a serious drop in the WOIT group comparing with the CT group. The modulation of these biomarkers for the remission and control of allergic asthma can be a prognostic outcome of immunotherapy which needs to be confirmed by larger scale studies.

Anti-TNFR2 enhanced the antitumor activity of a new HMGN1/3M-052 stimulated dendritic cell vaccine in a mouse model of colon cancer.

Immunotherapy is the new approach for cancer treatment that can be achieved through several strategies, one of which is dendritic cells (DCs) vaccine therapy. However, traditional DC vaccination lacks accurate targeting, so DC vaccine preparation needs to be optimized. Immunosuppressive CD4+Foxp3+ regulatory T cells (Tregs) in the tumor microenvironment can promote tumor immune escape. Therefore, targeting Tregs has become a strategy for tumor immunotherapy. In this study, we found that HMGN1 (N1, a dendritic cell-activating TLR4 agonist) and 3M-052 (a newly synthesized TLR7/8 agonist) synergistically stimulate DCs maturation and increase the production of proinflammatory cytokines TNFα and IL-12. In a colon cancer mice model, vaccination with N1 and 3M-052 stimulated and tumor antigen-loaded DCs combined with anti-TNFR2 inhibited tumor growth in mice, and the antitumor effect was mainly achieved through stimulation of cytotoxic CD8 T cell activation and depletion of Tregs. Overall, the combinating of DC activation by N1 and 3M-052 with inhibition of Tregs by antagonizing TNFR2 as a therapeutic strategy may represent a more effective strategy for cancer treatment.

Soluble CD25 imposes a low-zone IL-2 signaling environment that favors competitive outgrowth of antigen-experienced CD25high regulatory and memory T cells.

This study focused on soluble (s)CD25-mediated regulation of IL-2 signaling in murine and human CD4+ T cells. Recombinant sCD25 reversibly sequestered IL-2 to limit acute maximal proliferative responses while preserving IL-2 bioavailability to subsequently maintain low-zone IL-2 signaling during prolonged culture. By inhibiting IL-2 signaling during acute activation, sCD25 suppressed T-cell growth and inhibited IL-2-evoked transmembrane CD25 expression, thereby resulting in lower prevalence of CD25high T cells. By inhibiting IL-2 signaling during quiescent IL-2-mediated growth, sCD25 competed with transmembrane CD25, IL2Rßγ, and IL2Rαßγ receptors for limited pools of IL-2 such that sCD25 exhibited strong or weak inhibitory efficacy in IL-2-stimulated cultures of CD25low or CD25high T cells, respectively. Preferential blocking of IL-2 signaling in CD25low but not CD25high T cells caused competitive enrichment of CD25high memory/effector and regulatory FOXP3+ subsets. In conclusion, sCD25 modulates IL-2 bioavailability to limit CD25 expression during acute activation while enhancing CD25highT-cell dominance during low-zone homeostatic IL-2-mediated expansion, thereby 'flattening' the inflammatory curve over time.

Recirculating Foxp3+ regulatory T cells are restimulated in the thymus under Aire control.

Thymically-derived Foxp3+ regulatory T cells (Treg) critically control immunological tolerance. These cells are generated in the medulla through high affinity interactions with medullary thymic epithelial cells (mTEC) expressing the Autoimmune regulator (Aire). Recent advances have revealed that thymic Treg contain not only developing but also recirculating cells from the periphery. Although Aire is implicated in the generation of Foxp3+ Treg, its role in the biology of recirculating Treg remains elusive. Here, we show that Aire regulates the suppressive signature of recirculating Treg independently of the remodeling of the medullary 3D organization throughout life where Treg reside. Accordingly, the adoptive transfer of peripheral Foxp3+ Treg in AireKO recipients led to an impaired suppressive signature upon their entry into the thymus. Furthermore, recirculating Treg from AireKO mice failed to attenuate the severity of multiorgan autoimmunity, demonstrating that their suppressive function is altered. Using bone marrow chimeras, we reveal that mTEC-specific expression of Aire controls the suppressive signature of recirculating Treg. Finally, mature mTEC lacking Aire were inefficient in stimulating peripheral Treg both in polyclonal and antigen-specific co-culture assays. Overall, this study demonstrates that Aire confers to mTEC the ability to restimulate recirculating Treg, unravelling a novel function for this master regulator in Treg biology.

Elevated apoptosis and abnormal apoptosis signaling of regulatory T cells in patients with systemic lupus erythematosus.

In systemic lupus erythematosus (SLE), immune tolerance is influenced by defects in naturally occurring T cells (Tregs). To investigate the apoptosis rate of Tregs and their suppressive activity in patients with SLE and then to recognize the genes and signaling pathways that cause Treg apoptosis. FACS was used to assess the frequency and apoptosis rates of Tregs in 48 SLE patients and 28 normal controls (NCs). Coculture of Tregs with CD4+CD25-CD127dim/- T cells was used to assess the suppressive activity of Tregs. Microarray analysis was used to generate unstimulated Tregs gene expression profiles from very high activity patients with SLE and NCs. Real-time PCR was used to confirm differential gene expression. In patients with SLE, the frequency of Tregs was substantially reduced compared to Tregs from NCs. Furthermore, Tregs from SLE patients had an elevated rate of apoptosis and a lower suppressing ability than Tregs from NCs. Tregs apoptosis was negatively associated with the total count of Tregs and positively related to disease activity. Unstimulated Tregs gene expression profiles from patients with recent-onset SLE revealed a biological response that can cause apoptosis, partially triggered by stress, DNA damage, and cytokine stimulation. The discovery of pathway-specific expression signatures is a significant step forward in understanding how Tregs defects contribute to the pathogenesis of SLE. Our findings may contribute to the development of new strategies for treating SLE based on abnormal Tregs apoptosis and restoring immune homeostasis in patients with SLE.

TNF-TNFR2 Signal Plays a Decisive Role in the Activation of CD4+Foxp3+ Regulatory T Cells: Implications in the Treatment of Autoimmune Diseases and Cancer.

The puzzling biphasic or dual roles of tumor necrosis factor α (TNF) in the inflammatory and immune responses are likely to be mediated by distinct signaling pathways transduced by one of its two receptors, e.g., TNF receptor type I (TNFR1) and TNF receptor type II (TNFR2). Unlike TNFR1 that is ubiquitously expressed on almost all types of cells, the expression of TNFR2 is rather restricted to certain types of cells, such as T lymphocytes. There is now compelling evidence that TNFR2 is preferentially expressed by CD4+Foxp3+ regulatory T cells (Tregs), and TNFR2 plays a decisive role in the activation, expansion, in vivo function, and phenotypical stability of Tregs. In this chapter, the current understanding of the molecular basis and signaling pathway of TNF-TNFRs signal is introduced. Latest studies that have further supported and substantiated the pivotal role of TNF-TNFR2 interaction in Tregs biology and its molecular basis are discussed. The research progress regarding TNFR2-targeting treatment for autoimmune diseases and cancer is analyzed. Future study should focus on the further understanding of molecular mechanism underlying Treg-stimulatory effect of TNFR2 signal, as well as on the translation of research findings into therapeutic benefits of human patients with autoimmune diseases, allergy, allograft rejection, and cancer.

Heavy Metal to Rock the Immune Infiltrate.

Two resource articles recently published in Cell demonstrate that the elevated phenotypic complexity of the immune infiltrate in human lung adenocarcinomas and renal cell carcinomas can be reliably dissected with mass cytometry. These findings may pave the way to a new era of precision cancer immunotherapy.

Treg-specific IL-27Rα deletion uncovers a key role for IL-27 in Treg function to control autoimmunity.

Dysregulated Foxp3+ Treg functions result in uncontrolled immune activation and autoimmunity. Therefore, identifying cellular factors modulating Treg functions is an area of great importance. Here, using Treg-specific Il27ra-/- mice, we report that IL-27 signaling in Foxp3+ Tregs is essential for Tregs to control autoimmune inflammation in the central nervous system (CNS). Following experimental autoimmune encephalomyelitis (EAE) induction, Treg-specific Il27ra-/- mice develop more severe EAE. Consistent with the severe disease, the numbers of IFNγ- and IL-17-producing CD4 T cells infiltrating the CNS tissues are greater in these mice. Treg accumulation in the inflamed CNS tissues is not affected by the lack of IL-27 signaling in Tregs, suggesting a functional defect of Il27ra-/- Tregs. IL-10 production by conventional CD4 T cells and their CNS accumulation are rather elevated in Treg-specific Il27ra-/- mice. Analysis with Treg fate-mapping reporter mice further demonstrates that IL-27 signaling in Tregs may control stability of Foxp3 expression. Finally, systemic administration of recombinant IL-27 in Treg-specific Il27ra-/- mice fails to ameliorate the disease even in the presence of IL-27-responsive conventional CD4 T cells. These findings uncover a previously unknown role of IL-27 in regulating Treg function to control autoimmune inflammation.

Synergistic tumoricidal effect of combined hPD-L1 vaccine and HER2 gene vaccine.

Immunotherapy is gathering momentum as a kind of important therapy for cancer patients. However, monotherapies have limited efficacy in improving outcomes and benefit only in a small subset of patients. Combination therapies targeting multiple pathways often can augment an immune response to improve survival further. Here, the tumoricidal effects of the dual hPD-L1(human programmed cell death ligand 1) vaccination/HER2(human epidermal growth factor receptor 2) gene vaccination immunotherapy against the established HER2-expressed cancers were observed. Animals treated with combination therapy using hPD-L1 vaccine and HER2 gene vaccine had significantly improved survival in a mammary carcinoma model. We observed an increase in tumor growth inhibition following treatment. The percentage of the tumor-free mice (%) was much higher in the combined PD-L1/HER2 group. Furthermore, under the tumor-burden condition, hPD-L1 vaccine enhanced humoral immunity of HER2 gene vaccine. And the combination treatment increased the IFN-γ-producing effector T cells. Additionally, splenocytes from the combined PD-L1/HER2 group immunized mice possessed higher CTL activity. Notably, vaccination with combination therapy induced a significant decrease in the percentage of CD4+CD25+ Treg cells. Collectively, these data demonstrate that PD-L1/HER2 gene vaccine combination therapy synergistically generates marked tumoricidal effects against established HER2-expressing cancers.

CD4+ Foxp3+ regulatory T cell-mediated immunomodulation by anti-depressants inhibiting acid sphingomyelinase.

Acid sphingomyelinase (ASM) is the rate-limiting enzyme cleaving sphingomyelin into ceramide and phosphorylcholin. CD4+ Foxp3+ regulatory T (Treg) cells depend on CD28 signaling for their survival and function, a receptor that activates the ASM. Both, basal and CD28-induced ASM activities are higher in Treg cells than in conventional CD4+ T (Tconv) cells. In ASM-deficient (Smpd1-/-) as compared to wt mice, membranes of T cells contain 7-10-fold more sphingomyelin and two- to three-fold more ceramide, and are in a state of higher order than membranes of T cells from wt mice, which may facilitate their activation. Indeed, the frequency of Treg cells among CD4+ T cells in ASM-deficient mice and their suppressive activity in vitro are increased. Moreover, in vitro stimulation of ASM-deficient T cells in the presence of TGF-ß and IL-2 leads to higher numbers of induced Treg cells. Pharmacological inhibition of the ASM with a clinically used tricyclic antidepressant such as amitriptyline in mice or in tissue culture of murine or human T cells induces higher frequencies of Treg cells among CD4+ T cells within a few days. This fast alteration of the balance between T cell populations in vitro is due to the elevated cell death of Tconv cells and protection of the CD25high Treg cells by IL-2. Together, these findings suggest that ASM-inhibiting antidepressants, including a fraction of the serotonin re-uptake inhibitors (SSRIs), are moderately immunosuppressive and should be considered for the therapy of inflammatory and autoimmune disorders.

MHC-mismatched mixed chimerism augments thymic regulatory T-cell production and prevents relapse of EAE in mice.

Multiple sclerosis (MS) is an autoimmune inflammatory disease of the central nervous system with demyelination, axon damage, and paralysis. Induction of mixed chimerism with allogeneic donors has been shown to not cause graft-versus-host disease (GVHD) in animal models and humans. We have reported that induction of MHC-mismatched mixed chimerism can cure autoimmunity in autoimmune NOD mice, but this approach has not yet been tested in animal models of MS, such as experimental autoimmune encephalomyelitis (EAE). Here, we report that MHC-mismatched mixed chimerism with C57BL/6 (H-2(b)) donor in SJL/J (H-2(s)) EAE recipients eliminates clinical symptoms and prevents relapse. This cure is demonstrated by not only disappearance of clinical signs but also reversal of autoimmunity; elimination of infiltrating T, B, and macrophage cells in the spinal cord; and regeneration of myelin sheath. The reversal of autoimmunity is associated with a marked reduction of autoreactivity of CD4(+) T cells and significant increase in the percentage of Foxp3(+) Treg among host-type CD4(+) T cells in the spleen and lymph nodes. The latter is associated with a marked reduction of the percentage of host-type CD4(+)CD8(+) thymocytes and an increase of Treg percentage among the CD4(+)CD8(+) and CD4(+)CD8(-) thymocytes. Thymectomy leads to loss of prevention of EAE relapse by induction of mixed chimerism, although there is a dramatic expansion of host-type Treg cells in the lymph nodes. These results indicate that induction of MHC-mismatched mixed chimerism can restore thymic negative selection of autoreactive CD4(+) T cells, augment production of Foxp3(+) Treg, and cure EAE.

Disruption of de novo fatty acid synthesis via acetyl-CoA carboxylase 1 inhibition prevents acute graft-versus-host disease.

Upon antigen-specific or allogeneic activation, T cells sharply increase their metabolic activity to cope with augmented needs for proliferation and effector functions. Therefore, enzymes involved in energy metabolism constitute attractive targets to modulate the activity of pathogenic effector T cells in the setting of graft-versus-host-disease (GVHD). Here, we show that T cells deficient for acetyl-CoA carboxylase 1 (TACC1) are dramatically less pathogenic than wild-type (WT) T cells in a lethal C57BL/6 into BALB/c model of acute GVHD and permitted sustained survival of recipient mice. In line with this clinical observation, higher frequencies of GVHD-suppressing Foxp3(+) regulatory T (Treg) cells were detected in the colon of TACC T-cell recipients. In vitro, T-cell stimulation with allogeneic DCs induced higher proportions of Treg cells but also led to diminished proliferation of TACC1 T cells compared to WT T cells. Furthermore, TACC1 T cells activated by allogeneic DCs showed impaired glycolysis and lipid synthesis. Thus, targeting de novo fatty acid synthesis via acetyl-CoA carboxylase inhibition may be a promising new strategy to prevent GVHD.

Rapamycin results in Bim-mediated loss of thymic regulatory T cells during development in organ culture.

Thymus-derived regulatory T cells (Tregs) are essential for the maintenance of immunological tolerance. Diverse signalling pathways contribute to thymic Treg cells (tTregs) development; however, the role of mammalian target of rapamycin (mTOR) remains unclear. Rapamycin is a well-characterized inhibitor of mTOR complex 1 signalling and a potent inducer of Treg cells in the periphery. However, the effect of rapamycin on the development of tTregs is poorly defined. Here we have used thymic organ culture to investigate the effect of rapamycin on tTreg development. We show that, contrary to its effect in the periphery, rapamycin inhibits the development of tTregs in wild-type thymi. The inhibition of tTregs by rapamycin could be rescued by a deficiency of Bim. However, rapamycin did not inhibit the development of antigen-specific TCR transgenic tTregs in response to exogenous peptide antigen, indicating that the development of thymic Foxp3+CD4+ cells was not intrinsically inhibited by rapamycin. Collectively our data demonstrate that rapamycin results in a reduction of tTregs because of Bim-mediated apoptosis of immature tTregs via a cell extrinsic mechanism. These findings are important not only for understanding the mechanism of tTreg induction but also for an appreciation of the impact of the clinical application of rapamycin.

Exogenous interleukin-33 targets myeloid-derived suppressor cells and generates periphery-induced Foxp3⁺ regulatory T cells in skin-transplanted mice.

Interleukin-33 (IL-33) has been a focus of study because of its variety of functions shaping CD4(+) T-cell biology. In the present work, we evaluated the modulatory effect of IL-33 on suppressor cells in an in vivo transplantation model. C57BL/6 wild-type mice were grafted with syngeneic or allogeneic skin transplants and treated with exogenous IL-33 daily. After 10 days of treatment, we analysed draining lymph node cellularity and found in allogeneic animals an increment in myeloid-derived suppressor cells, which co-express MHC-II, and become enriched upon IL-33 treatment. In line with this observation, inducible nitric oxide synthase and arginase 1 expression were also increased in allogeneic animals upon IL-33 administration. In addition, IL-33 treatment up-regulated the number of Foxp3(+) regulatory T (Treg) cells in the allogeneic group, complementing the healthier integrity of the allografts and the increased allograft survival. Moreover, we demonstrate that IL-33 promotes CD4(+) T-cell expansion and conversion of CD4(+)  Foxp3(-) T cells into CD4(+)  Foxp3(+) Treg cells in the periphery. Lastly, the cytokine pattern of ex vivo-stimulated draining lymph nodes indicates that IL-33 dampens interferon-γ and IL-17 production, stimulating IL-10 secretion. Altogether, our work complements previous studies on the immune-modulatory activity of IL-33, showing that this cytokine affects myeloid-derived suppressor cells at the cell number and gene expression levels. More importantly, our research demonstrates for the first time that IL-33 allows for in vivo Foxp3(+) Treg cell conversion and favours an anti-inflammatory or tolerogenic state by skewing cytokine production. Therefore, our data suggest a potential use of IL-33 to prevent allograft rejection, bringing new therapeutics to the transplantation field.

Expression of TNFR2 by regulatory T cells in peripheral blood is correlated with clinical pathology of lung cancer patients.

CD4(+)FoxP3(+) regulatory T cells (Tregs) represent a major cellular mediator of cancer immune evasion. The expression of tumor necrosis factor receptor type II (TNFR2) on Tregs is reported to identify the maximally suppressive Treg population in both mice and human. We therefore investigated the phenotype and function of TNFR2(+) Tregs present in the peripheral blood (PB) of 43 lung cancer patients. Further, the association of TNFR2 expression on Tregs with clinicopathological factors was analyzed. The results showed that in the PB of lung cancer patients, Tregs expressed markedly higher levels of TNFR2 than conventional T cells (Tconvs). Expression of TNFR2 appeared to correlate better than CD25(+) and CD127(-) with FoxP3 expression. PB TNFR2(+) Tregs in lung cancer patients were more proliferative and expressed higher levels of the immunosuppressive molecule CTLA-4, and consequently more potently suppressed IFNγ production by cocultured CD8 CTLs. More importantly, higher TNFR2 expression levels on Tregs were associated with lymphatic invasion, distant metastasis and more advanced clinical stage of lung cancer patients. Therefore, our study suggests that TNFR2(+) Tregs play a role in promoting tumor progressive metastasis and expression of TNFR2 by PB Tregs may prove to be a useful prognostic marker in lung cancer patients.