Cells Suppress T-cell Proliferation Research Articles

Abstract PD9-01: Stromal mediators of lymphocyte exclusion and dysfunction in triple negative breast cancer

Abstract Triple negative breast cancer (TNBC) is the most aggressive breast cancer subtype with poor prognosis and limited treatment options. One of the mechanisms contributing to this is the ability of TNBC tumors to evade anti-tumour response, limiting the success of immune-checkpoint therapy. In recent studies of the tumor microenvironment, stromal cells have emerged as potential important mediators of lymphocyte function in TNBC. Multi-omics studies from our group and others have elucidated the heterogeneity of stromal cells and their interactions with immune cells in TNBC. However, the clinical relevance and functional characteristics of these relationships remain poorly explored. Our previous work revealed functionally distinct subpopulations of stromal cells in breast cancer: endothelial cells, myofibroblast-like cancer-associated fibroblasts (myCAFs), inflammatory-CAFs (iCAFs), and perivascular-like cells which resemble pericyte and smooth muscle cells in immature and differentiated states (imPVLs; dPVLs), respectively. Here we directly explored the clinical relevance of these stromal subpopulations and their association with immune evasion in a large independent TNBC cohort with long-term survival data. Using markers derived from our single-cell multi-omics studies, we performed multiplex immunofluorescence using the OPAL9 platform on tumor microarrays from 222 TNBC patients to mark myCAFs, iCAFs, dPVLs, endothelial cells, CD8 and/or PD1 positive T-cells. Digital imaging analysis (QuPath) revealed a significant negative correlation between the abundance of stromal cells and CD8 T cells. This cytotoxic T-cell exclusion is primarily driven by smooth muscle dPVLs. Parallel to this T-cell exclusion, our functional studies demonstrate that stromal cells suppress T-cell proliferation. Using multiple ex vivo co-culture models of primary TNBC CAFs and peripheral blood mononuclear cells (PBMCs) from healthy and matched donors, we show that CAFs suppress the proliferative capability of CD4 and CD8 T-cells. ScRNA-seq of these co-culture models demonstrate that CAF educated T cells are driven into a LAG3+ exhausted state enriched for canonical pathways of immunosuppressive cytokine signaling. Our findings suggest that manipulation of these stromal subpopulations could elicit a more effective immune response in a subset of patients through inhibiting T-cell dysfunction and exclusion. Citation Format: Julia Chen, Sunny Wu, Travis Ruan, Iveta Slapetova, Ewan Millar, Elgene Lim, Alex Swarbrick. Stromal mediators of lymphocyte exclusion and dysfunction in triple negative breast cancer [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr PD9-01.

Impact of p53 Knock-Down on T-Cell Proliferation and T-Cell Mediated Cytotoxicity Against AML Cell Lines Mediated By a CD33 Specific BiTE® Antibody Construct

Impact of p53 Knock-Down on T-Cell Proliferation and T-Cell Mediated Cytotoxicity Against AML Cell Lines Mediated By a CD33 Specific BiTE® Antibody Construct

Taurodeoxycholate Increases the Number of Myeloid-Derived Suppressor Cells That Ameliorate Sepsis in Mice.

Bile acids (BAs) control metabolism and inflammation by interacting with several receptors. Here, we report that intravenous infusion of taurodeoxycholate (TDCA) decreases serum pro-inflammatory cytokines, normalizes hypotension, protects against renal injury, and prolongs mouse survival during sepsis. TDCA increases the number of granulocytic myeloid-derived suppressor cells (MDSCLT) distinctive from MDSCs obtained without TDCA treatment (MDSCL) in the spleen of septic mice. FACS-sorted MDSCLT cells suppress T-cell proliferation and confer protection against sepsis when adoptively transferred better than MDSCL. Proteogenomic analysis indicated that TDCA controls chromatin silencing, alternative splicing, and translation of the immune proteome of MDSCLT, which increases the expression of anti-inflammatory molecules such as oncostatin, lactoferrin and CD244. TDCA also decreases the expression of pro-inflammatory molecules such as neutrophil elastase. These findings suggest that TDCA globally edits the proteome to increase the number of MDSCLT cells and affect their immune-regulatory functions to resolve systemic inflammation during sepsis.

Pak2 regulates myeloid-derived suppressor cell development in mice

AbstractMyeloid-derived suppressor cells (MDSCs) are CD11b+Gr1+ cells that induce T-cell hyporesponsiveness, thus impairing antitumor immunity. We have previously reported that disruption of Pak2, a member of the p21-activated kinases (Paks), in hematopoietic stem/progenitor cells (HSPCs) induces myeloid lineage skewing and expansion of CD11bhighGr1high cells in mice. In this study, we confirmed that Pak2-KO CD11bhighGr1high cells suppressed T-cell proliferation, consistent with an MDSC phenotype. Loss of Pak2 function in HSPCs led to (1) increased hematopoietic progenitor cell sensitivity to granulocyte-macrophage colony-stimulating factor (GM-CSF) signaling, (2) increased MDSC proliferation, (3) decreased MDSC sensitivity to both intrinsic and Fas-Fas ligand–mediated apoptosis, and (4) promotion of MDSCs by Pak2-deficient CD4+ T cells that produced more interferon γ, tumor necrosis factor α, and GM-CSF. Pak2 disruption activated STAT5 while downregulating the expression of IRF8, a well-described myeloid transcription factor. Together, our data reveal a previously unrecognized role of Pak2 in regulating MDSC development via both cell-intrinsic and extrinsic mechanisms. Our findings have potential translational implications, as the efficacy of targeting Paks in cancer therapeutics may be undermined by tumor escape from immune control and/or acceleration of tumorigenesis through MDSC expansion.

Induced regulatory T cells are phenotypically unstable and do not protect mice from rapidly progressive glomerulonephritis.

Regulatory T (Treg) cells are a suppressive CD4+ T-cell subset. We generated induced Treg (iTreg) cells and explored their therapeutic potential in a murine model of rapidly progressive glomerulonephritis. Polyclonal naive CD4+ T cells were cultured in vitro with interleukin-2 (IL-2), transforming growth factor-β1, all-trans-retinoic acid and monoclonal antibodies against interferon-γ and IL-4, generating Foxp3+ iTreg cells. To enhance their suppressive phenotype, iTreg cultures were modified with the addition of a monoclonal antibody against IL-12p40 or by using RORγt-/- CD4+ T cells. Induced Treg cells were transferred into models of delayed-type hypersensitivity and experimental glomerulonephritis. The iTreg cells exhibited comparable surface receptor expression and in vitro suppressive ability to natural Treg cells, but did not regulate antigen-specific delayed-type hypersensitivity or systemic inflammatory immune responses, losing Foxp3 expression in vivo. In glomerulonephritis, transferred iTreg cells did not prevent renal injury or modulate systemic T helper type 1 immune responses. Induced Treg cells cultured with anti-IL-12p40 had an enhanced suppressive phenotype in vitro and regulated dermal delayed-type hypersensitivity in vivo, but were not protective against renal injury, losing Foxp3 expression, especially in the transferred cells recruited to the kidney. Use of RORγt-/- CD4+ T cells or iTreg cells generated from sensitized CD4+ Foxp3- cells did not regulate renal or systemic inflammatory responses in vivo. In conclusion, iTreg cells suppress T-cell proliferation in vitro, but do not regulate experimental glomerulonephritis, being unstable in this inflammatory milieu in vivo.

Abstract 3239: Cancer-associated fibroblasts induce immunosuppressive macrophages in head and neck squamous cell carcinoma

Abstract Background Cancer-associated fibroblasts (CAFs) are one of main elements in the tumor microenvironment (TME). Among the innate and adaptive immune cells in the TME, tumor-associated macrophages (TAMs) are particularly abundant and play a protumoral role. However, an interaction between CAFs and TAMs in head and neck squamous cell carcinoma (HNSCC) still remains unclear. In the present study, we investigated whether CAFs skewed macrophage function toward the immune suppressive phenotype. Materials and Methods In vitro assay: CAFs were prepared from surgical tissue of HNSCC patients, and culture supernatants of CAFs were collected. Using this supernatants and peripheral blood mononuclear cells collected from healthy donors, immunological interactions between CAFs and TAMs was investigated. In vivo assay: The immunohistochemistry (IHC) of 73 patients with HNSCC was performed. Anti-α-smooth muscle actin, CD68, and CD163 antibodies were used to identify CAFs and TAMs, respectively. The relationship between immunohistochemical parameters and clinical parameters was evaluated. Results The expression levels of CD68, CD163, CD14, CD80, CD86, and HLA-G were up-regulated in CD14+ cells co-cultured with the supernatants of CAFs (CAF-educated cells) compared with control cells. Moreover, the gene expressions of ARG1, IL6, IL10 and TGFB1 were also increased in CAF-educated cells. The CAF-educated cells suppressed T-cell proliferation more intensively than control cells, and neutralization of TGF-β and IL-10 led to significant restoration of T-cell proliferation. In IHC, the intensity of CAFs correlated with the number of CD68-positive macrophages, especially with CD163-positive macrophages. It also correlated with several clinical parameters such as lymphatic invasion, vascular invasion, nodal status, tumor stages and recurrence rate. Moreover, the high intensity of CAFs correlated with poor PFS and OS. The high intensity of CAFs and M2 macrophage infiltration were independent prognostic factors of PFS and OS in HNSCC. Conclusion The present study suggests that CAFs can induce an immunosuppressive phenotype of macrophages and contribute to establishing the immunosuppressive networks in TME, resulting poor prognosis in HNSCC patients. Citation Format: Hideyuki Takahashi, Koichi Sakakura, Kazuaki Chikamatsu. Cancer-associated fibroblasts induce immunosuppressive macrophages in head and neck squamous cell carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3239.

Human periodontal ligament stem cells suppress T-cell proliferation via down-regulation of non-classical major histocompatibility complex-like glycoprotein CD1b on dendritic cells.

Periodontal ligament stem cells (PDLSCs) from the periodontal ligament tissue were recently identified as mesenchymal stem cells (MSCs). The capabilities of PDLSCs in periodontal tissue or bone regeneration have been reported, but their immunomodulatory role in T-cell immune responses via dendritic cells (DCs), known as the most potent antigen-presenting cell, has not been studied. The aim of this study is to understand the immunological function of homogeneous human STRO-1+ CD146+ PDLSCs in DC-mediated T-cell immune responses to modulate the periodontal disease process. We utilized highly purified (>95%) human STRO-1+ CD146+ PDLSCs and human bone marrow mesenchymal stem cells (BMSCs). Each stem cell was co-cultured with human monocyte-derived DCs in the presence of lipopolysaccharide isolated from Porphyromonas gingivalis, a major pathogenic bacterium responsible for periodontal disease, invitro to examine the immunological effect of each stem cell on DCs and DC-mediated T-cell proliferation. We discovered that STRO-1+ CD146+ PDLSCs, as well as BMSCs, significantly decreased the level of non-classical major histocompatibility complex glycoprotein CD1b on DCs, resulting in defective T-cell proliferation, whereas most human leukocyte antigens and the co-stimulatory molecules CD80 and CD86 in/on DCs were not significantly affected by the presence of BMSCs or STRO-1+ CD146+ PDLSCs. This study unveiled an immunomodulatory role of STRO-1+ CD146+ PDLSCs in negatively regulating DC-mediated T-cell immune responses, demonstrating their potential to be utilized in promising new stem cell therapies.

Neuroblastoma Arginase Activity Creates an Immunosuppressive Microenvironment That Impairs Autologous and Engineered Immunity.

Neuroblastoma is the most common extracranial solid tumor of childhood, and survival remains poor for patients with advanced disease. Novel immune therapies are currently in development, but clinical outcomes have not matched preclinical results. Here, we describe key mechanisms in which neuroblastoma inhibits the immune response. We show that murine and human neuroblastoma tumor cells suppress T-cell proliferation through increased arginase activity. Arginase II is the predominant isoform expressed and creates an arginine-deplete local and systemic microenvironment. Neuroblastoma arginase activity results in inhibition of myeloid cell activation and suppression of bone marrow CD34(+) progenitor proliferation. Finally, we demonstrate that the arginase activity of neuroblastoma impairs NY-ESO-1-specific T-cell receptor and GD2-specific chimeric antigen receptor-engineered T-cell proliferation and cytotoxicity. High arginase II expression correlates with poor survival for patients with neuroblastoma. The results support the hypothesis that neuroblastoma creates an arginase-dependent immunosuppressive microenvironment in both the tumor and blood that leads to impaired immunosurveillance and suboptimal efficacy of immunotherapeutic approaches.

A robust and reproducible animal serum-free culture method for clinical-grade bone marrow-derived mesenchymal stromal cells.

Efficient xenofree expansion methods to replace fetal bovine serum (FBS)-based culture methods are strongly encouraged by the regulators and are needed to facilitate the adoption of mesenchymal stromal cell (MSC)-based therapies. In the current study we established a clinically-compliant and reproducible animal serum-free culture protocol for bone marrow-(BM-) MSCs based on an optimized platelet-derived supplement. Our study compared two different platelet-derived supplements, platelet lysate PL1 versus PL2, produced by two different methods and lysed with different amounts of freeze–thaw cycles. Our study also explored the effect of a low oxygen concentration on BM-MSCs. FBS-supplemented BM-MSC culture served as control. Growth kinetics, differentiation and immunomodulatory potential, morphology, karyotype and immunophenotype was analysed. Growth kinetics in long-term culture was also studied. Based on the initial results, we chose to further process develop the PL1-supplemented culture protocol at 20 % oxygen. The results from 11 individual BM-MSC batches expanded in the chosen condition were consistent, yielding 6.60 × 109 ± 4.74 × 109 cells from only 20 ml of bone marrow. The cells suppressed T-cell proliferation, displayed normal karyotype and typical MSC differentiation potential and phenotype. The BM-MSCs were, however, consistently HLA-DR positive when cultured in platelet lysate (7.5–66.1 %). We additionally show that culture media antibiotics and sterile filtration of the platelet lysate can be successfully omitted. We present a robust and reproducible clinically-compliant culture method for BM-MSCs based on platelet lysate, which enables high quantities of HLA-DR positive MSCs at a low passage number (p2) and suitable for clinical use.

Alloreactive regulatory Tcells generated with retinoic acid prevent skin allograft rejection.

CD4(+) CD25(+) Foxp3(+) regulatory T (Treg) cells mediate immunological self-tolerance and suppress immune responses. Retinoic acid (RA), a natural metabolite of vitamin A, has been reported to enhance the differentiation of Treg cells in the presence of TGF-β. In this study, we show that the co-culture of naive Tcells from C57BL/6 mice with allogeneic antigen-presenting cells (APCs) from BALB/c mice in the presence of TGF-β, RA, and IL-2 resulted in a striking enrichment of Foxp3(+) T cells. These RA in vitro-induced regulatory T (RA-iTreg) cells did not secrete Th1-, Th2-, or Th17-related cytokines, showed a nonbiased homing potential, and expressed several cell surface molecules related to Treg-cell suppressive potential. Accordingly, these RA-iTreg cells suppressed T-cell proliferation and inhibited cytokine production by Tcells in in vitro assays. Moreover, following adoptive transfer, RA-iTreg cells maintained Foxp3 expression and their suppressive capacity. Finally, RA-iTreg cells showed alloantigen-specific immunosuppressive capacity in a skin allograft model in immunodeficient mice. Altogether, these data indicate that functional and stable allogeneic-specific Treg cells may be generated using TGF-β, RA, and IL-2. Thus, RA-iTreg cells may have a potential use in the development of more effective cellular therapies in clinical transplantation.

Both retention and recirculation contribute to long-lived regulatory T-cell accumulation in the thymus.

Natural Treg cells acquire their lineage-determining transcription factor Foxp3 during development in the thymus and are important in maintaining immunologic tolerance. Here, we analyzed the composition of the thymic Treg-cell pool using RAG2-GFP/FoxP3-RFP dual reporter mice and found that a population of long-lived GFP(-) Treg cells exists in the thymus. These long-lived Treg cells substantially increased with age, to a point where they represent >90% of the total thymic Treg-cell pool at 6 months of age. In contrast, long-lived conventional T cells remained at ∼ 15% of the total thymic pool at 6 months of age. Consistent with these studies, we noticed that host-derived Treg cells represented a large fraction (∼ 10%) of the total thymic Treg-cell pool in bone marrow chimeras, suggesting that long-lived Treg cells also reside in the thymus of these mice. The pool of long-lived Treg cells in the thymus was sustained by retention of Treg cells in the thymus and by recirculation of peripheral Treg cells back into the thymus. These long-lived thymic Treg cells suppressed T-cell proliferation to an equivalent extent to splenic Treg cells. Together, these data demonstrate that long-lived Treg cells accumulate in the thymus by both retention and recirculation.

Human placenta-derived adherent cells induce tolerogenic immune responses.

Human placenta-derived adherent cells (PDAC cells) are a culture expanded, undifferentiated mesenchymal-like population derived from full-term placental tissue, with immunomodulatory and anti-inflammatory properties. PDA-001 (cenplacel-L), an intravenous formulation of PDAC cells, is in clinical development for the treatment of autoimmune and inflammatory diseases. To elucidate the mechanisms underlying the immunoregulatory properties of PDAC cells, we investigated their effects on immune cell populations, including T cells and dendritic cells (DC) in vitro and in vivo. PDAC cells suppressed T-cell proliferation in an OT-II T-cell adoptive transfer model, reduced the severity of myelin oligodendrocyte glycoprotein peptide-induced experimental autoimmune encephalomyelitis and ameliorated inflammation in a delayed type hypersensitivity response model. In vitro, PDAC cells suppressed T-cell proliferation and inhibited Th1 and Th17 differentiation. Analysis of tissues derived from PDAC cell-treated animals revealed diminished CD86 expression on splenic DC, suggesting that they can also modulate DC populations. Furthermore, PDAC cells modulate the differentiation and maturation of mouse bone marrow-derived DC. Similarly, human DC differentiated from CD14+ monocytes in the presence of PDAC cells acquired a tolerogenic phenotype. These tolerogenic DC failed to induce allogeneic T-cell proliferation and differentiation toward Th1, but skewed T-cell differentiation toward Th2. Inhibition of cyclo-oxygenase-2 activity resulted in a significant, but not complete, abrogation of PDAC cells' effects on DC phenotype and function, implying a role for prostaglandin E2 in PDAC-mediated immunomodulation. This study identifies modulation of DC differentiation toward immune tolerance as a key mechanism underlying the immunomodulatory activities of PDAC cells.

GM-CSF Promotes the Immunosuppressive Activity of Glioma-Infiltrating Myeloid Cells through Interleukin-4 Receptor-α

Malignant gliomas are lethal cancers in the brain and heavily infiltrated by myeloid cells. Interleukin-4 receptor-α (IL-4Rα) mediates the immunosuppressive functions of myeloid cells, and polymorphisms in the IL-4Rα gene are associated with altered glioma risk and prognosis. In this study, we sought to evaluate a hypothesized causal role for IL-4Rα and myeloid suppressor cells in glioma development. In both mouse de novo gliomas and human glioblastoma cases, IL-4Rα was upregulated on glioma-infiltrating myeloid cells but not in the periphery or in normal brain. Mice genetically deficient for IL-4Rα exhibited a slower growth of glioma associated with reduced production in the glioma microenvironment of arginase, a marker of myeloid suppressor cells, which is critical for their T-cell inhibitory function. Supporting this result, investigations using bone marrow-derived myeloid cells showed that IL-4Rα mediates IL-13-induced production of arginase. Furthermore, glioma-derived myeloid cells suppressed T-cell proliferation in an IL-4Rα-dependent manner, consistent with their identification as myeloid-derived suppressor cells (MDSC). Granulocyte macrophage colony-stimulating factor (GM-CSF) plays a central role for the induction of IL-4Rα expression on myeloid cells, and we found that GM-CSF is upregulated in both human and mouse glioma microenvironments compared with normal brain or peripheral blood samples. Together, our findings establish a GM-CSF-induced mechanism of immunosuppression in the glioma microenvironment via upregulation of IL-4Rα on MDSCs.

CD4⁺CD73⁺ T cells are associated with lower T-cell activation and C reactive protein levels and are depleted in HIV-1 infection regardless of viral suppression.

The role of the adenosine (ADO) suppression pathway, specifically CD39-expressing and CD73-expressing CD4⁺ T cells in HIV-1 infection is unclear. We evaluated the frequency and numbers of CD4⁺CD39⁺ and CD4⁺CD73⁺ T cells, activated T cells, and plasma C reactive protein (CRP) levels in 36 HIV-1-positive individuals and 10 normal controls (NC). Low-level plasma viremia was evaluated using single copy assay. Mass spectrometry was used to measure hydrolysis of ATP by ectoenzyme-expressing CD4⁺ T cells, whereas cyclic adenosine monophosphate (cAMP) levels were measured using enzyme immunoassay. Suppression of T-cell function by exogenous ADO and CD4⁺CD73⁺ T cells was tested by flow cytometry. CD39 and CD73 are expressed in different CD4⁺ T-cell subsets. CD4⁺CD73⁺ T cells do not express CD25 and FOXP3, and their frequency and numbers were lower in HIV-1-positive individuals regardless of virologic suppression (P=0.005 and P<0.001, respectively). CD4⁺CD73⁺ numbers inversely correlated with CD4⁺CD38⁺DR⁺ (P=0.002), CD8⁺CD38⁺DR⁺ T-cell frequency (P=0.05), and plasma CRP levels (P=0.01). Both subsets are required for hydrolysis of exogenous ATP to ADO and can increase CD4⁺ T-cell cAMP levels when incubated with exogenous ATP. Low-level viremia did not correlate with activated T-cell frequency. In vitro, ADO suppressed T-cell activation and cytokine expression. CD4⁺CD7⁺ T cells suppressed T-cell proliferation only in the presence of exogenous 5'-AMP. The ADO-producing CD4⁺CD73⁺ subset of T cells is depleted in HIV-1-positive individuals regardless of viral suppression and may play a key role in controlling HIV-1-associated immune activation.

Abstract B6: Myeloid-derived suppressor cells from melanoma patients suppress T cell proliferation and are associated with a shorter disease-free interval.

Abstract Immune therapies such as vaccines, immune stimulators (IL-2), and immune checkpoint regulators (anti-CTLA-4) for melanoma elicit an initial robust immunologic response to the tumor in some patients, however, the majority ultimately go on to die of their disease. This suggests that tumor eventually develop escape mechanisms meant to evade or defeat the intrinsic immune response. Myeloid-derived suppressor cells (MDSCs) have been reported in melanoma and other cancer patients and may contribute to immune suppression or counter-regulatory responses. The aim of this project was to assess the potential role of circulating MDSCs in mediating immune suppression in advanced stage melanoma patients. Methods: MDSC populations (DC14+ Lin-HLA-DR- , Lin-HLA-DR- CD11b+ CD33+) were isolated from patients with (stage I n=20, stage IV n=35)or without (n=15) melanoma. Isolated MDSC's were mixed with allogeneic CFSE-labeled T cells for a 4 day mixed lymphocyte reaction (MLR). T cell activation and proliferation were quantified. Results: Stage IV melanoma patients have increased expression (2-3 times) of some MDSC populations (Lin-HLA-DR- CD11b+ CD33+) in the circulation compared to normal and stage I melanoma patients. By MLR, these MDSC's significantly suppressed T cell proliferation (40-50% reduction) and markers of activation (20%) to a greater extent than the same population of MDSC's for normal controls. Inhibiting nitric oxide failed to restore T-cell proliferation. A shorter disease free interval is associated with increased expression of CD33+ MDSC in the circulation. Conclusion: MDSC populations are increased in the peripheral circulation of stage IV melanoma patients and these cells actively suppress T cell proliferation. Efforts are underway to better understand the melanoma related factors responsible for the induction of MDSC's and the mechanism by which these cells suppress T-cell proliferation. Citation Format: Martin D. McCarter, Kimberly Jordan. Myeloid-derived suppressor cells from melanoma patients suppress T cell proliferation and are associated with a shorter disease-free interval. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B6.

What are regulatory B cells?

B cells are now acknowledged to play multiple roles in the immune response, in addition to making antibodies. Their role in regulating T-cell responses during inflammation has come into focus recently. Thus, IL-10 production by B cells has been shown to be important in limiting auto-reactive and pathogen-driven immune pathology; however, the exact identity of the regulatory B cells remains elusive; do they belong to a committed subset or can all B cells regulate given the appropriate inducing stimuli? A study in this issue of the European Journal of Immunology provides insight into the IL-10-producing B cells in humans, suggesting that many B cells have the capacity to make IL-10 when optimally stimulated via the BCR and TLR9. Despite producing significant amounts of inflammatory cytokines as well, these B cells suppress T-cell proliferation. This Commentary places this study in the context of what we think we know about regulatory B cells and more importantly highlights the questions we still need to answer.

Galectin‐9 expands immunosuppressive macrophages to ameliorate T‐cell‐mediated lung inflammation

Galectin-9 (Gal-9) plays pivotal roles in the modulation of innate and adaptive immunity to suppress T-cell-mediated autoimmune models. However, it remains unclear if Gal-9 plays a suppressive role for T-cell function in non-autoimmune disease models. We assessed the effects of Gal-9 on experimental hypersensitivity pneumonitis induced by Trichosporon asahii. When Gal-9 was given subcutaneously to C57BL/6 mice at the time of challenge with T. asahii, it significantly suppressed T. asahii-induced lung inflammation, as the levels of IL-1, IL-6, IFN-gamma, and IL-17 were significantly reduced in the BALF of Gal-9-treated mice. Moreover, co-culture of anti-CD3-stimulated CD4 T cells with BALF cells harvested from Gal-9-treated mice on day 1 resulted in diminished CD4 T-cell proliferation and decreased levels of IFN-gamma and IL-17. CD11b(+)Ly-6C(high)F4/80(+) BALF Mphi expanded by Gal-9 were responsible for the suppression. We further found in vitro that Gal-9, only in the presence of T. asahii, expands CD11b(+)Ly-6C(high)F4/80(+) cells from BM cells, and the cells suppress T-cell proliferation and IFN-gamma and IL-17 production. The present results indicate that Gal-9 expands immunosuppressive CD11b(+)Ly-6C(high) Mphi to ameliorate Th1/Th17 cell-mediated hypersensitivity pneumonitis.

Endotoxin-Induced Myeloid-Derived Suppressor Cells Inhibit Alloimmune Responses via Heme Oxygenase-1

Inflammation and cancer are associated with impairment of T-cell responses by a heterogeneous population of myeloid-derived suppressor cells (MDSCs) coexpressing CD11b and GR-1 antigens. MDSCs have been recently implicated in costimulation blockade-induced transplantation tolerance in rats, which was under the control of inducible NO synthase (iNOS). Herein, we describe CD11b+GR-1+MDSC-compatible cells appearing after repetitive injections of lipopolysaccharide (LPS) using a unique mechanism of suppression. These cells suppressed T-cell proliferation and Th1 and Th2 cytokine production in both mixed lymphocyte reaction and polyclonal stimulation assays. Transfer of CD11b+ cells from LPS-treated mice in untreated recipients significantly prolonged skin allograft survival. They produced large amounts of IL-10 and expressed heme oxygenase-1 (HO-1), a stress-responsive enzyme endowed with immunoregulatory and cytoprotective properties not previously associated with MDSC activity. HO-1 inhibition by the specific inhibitor, SnPP, completely abolished T-cell suppression and IL-10 production. In contrast, neither iNOS nor arginase 1 inhibition did affect suppression. Importantly, HO-1 inhibition before CD11b+ cell transfer prevented the delay of allograft rejection revealing a new MDSC-associated suppressor mechanism relevant for transplantation.