Differentiation, phenotype, and function of interleukin-17–producing human Vγ9Vδ2 T cells

Abstract In healthy adults, the major peripheral blood γδ T-cell subset expresses the Vγ9Vδ2 TCR and displays pleiotropic features. Here we report that coculture of naive Vγ9Vδ2 T cells with phosphoantigens and a cocktail of cytokines (IL-1-β, TGF-β, IL-6, and IL-23), leads to selective expression of the transcription factor RORγt and polarization toward IL-17 production. IL-17+ Vγ9Vδ2 T cells express the chemokine receptor CCR6 and produce IL-17 but neither IL-22 nor IFN-γ; they have a predominant terminally differentiated (CD27−CD45RA+) phenotype and express granzyme B, TRAIL, FasL, and CD161. On antigen activation, IL-17+ Vγ9Vδ2 T cells rapidly induce CXCL8-mediated migration and phagocytosis of neutrophils and IL-17–dependent production of β-defensin by epithelial cells, indicating that they may be involved in host immune responses against infectious microorganisms. Accordingly, an increased percentage of IL-17+ Vγ9Vδ2 lymphocytes is detected in the peripheral blood and at the site of disease in children with bacterial meningitis, and this pattern was reversed after successful antibacterial therapy. Most notably, the phenotype of IL-17+ Vγ9Vδ2 T cells in children with meningitis matches that of in vitro differentiated IL-17+ Vγ9Vδ2 T cells. Our findings delineate a previously unknown subset of human IL-17+ Vγ9Vδ2 T lymphocytes implicated in the pathophysiology of inflammatory responses during bacterial infections.

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Interleukin-12 Is the Optimum Cytokine To Expand Human Th17 Cells In Vitro

Clinical and Vaccine Immunology ◽

10.1128/cvi.00022-09 ◽

2009 ◽

Vol 16 (6) ◽

pp. 798-805 ◽

Cited By ~ 6

Author(s):

Soad Nady ◽

James Ignatz-Hoover ◽

Mohamed T. Shata

Keyword(s):

T Cells ◽

Peripheral Blood ◽

Th17 Cells ◽

Interleukin 12 ◽

Interleukin 17 ◽

Functional Evaluation ◽

Optimum Conditions ◽

T Helper ◽

Ifn Γ

ABSTRACT Recently, a new lineage of CD4+ T cells in humans and in mice has been reported. This T helper cell secretes interleukin-17 (IL-17) and has been defined as T helper 17 (Th17). Th17 cells express the IL-23 receptor (IL-23R) and play an important pathogenic role in different inflammatory conditions. In this study, our aim was to characterize the optimum conditions for isolation and propagation of human peripheral blood Th17 cells in vitro and the optimum conditions for isolation of Th17 clones. To isolate Th17 cells, two steps were taken. Initially, we negatively isolated CD4+ T cells from peripheral blood mononuclear cells of a normal human blood donor. Then, we isolated the IL-23R+ cells from the CD4+ T cells. Functional studies revealed that CD4+ IL-23R+ cells could be stimulated ex vivo with anti-CD3/CD28 to secrete both IL-17 and gamma interferon (IFN-γ). Furthermore, we expanded the CD4+ IL-23R+ cells for 1 week in the presence of anti-CD3/CD28, irradiated autologous feeder cells, and different cytokines. Our data indicate that cytokine treatment increased the number of propagated cells 14- to 99-fold. Functional evaluation of the expanded number of CD4+ IL-23R+ cells in the presence of different cytokines with anti-CD3/CD28 revealed that all cytokines used (IL-2, IL-7, IL-12, IL-15, and IL-23) increased the amount of IFN-γ secreted by IL-23R+ CD4+ cells at different levels. Our results indicate that IL-7 plus IL-12 was the optimum combination of cytokines for the expansion of IL-23R+ CD4+ cells and the secretion of IFN-γ, while IL-12 preferentially stimulated these cells to secrete predominately IL-17.

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Monomethyl fumarate treatment impairs maturation of human myeloid dendritic cells and their ability to activate T cells

Multiple Sclerosis Journal ◽

10.1177/1352458517740213 ◽

2017 ◽

Vol 25 (1) ◽

pp. 63-71 ◽

Cited By ~ 9

Author(s):

Maria Antonietta Mazzola ◽

Radhika Raheja ◽

Keren Regev ◽

Vanessa Beynon ◽

Felipe von Glehn ◽

...

Keyword(s):

T Cells ◽

Dendritic Cells ◽

Dimethyl Fumarate ◽

Nuclear Factor Κb ◽

Interleukin 17 ◽

Myeloid Dendritic Cells ◽

Gm Csf ◽

Monomethyl Fumarate ◽

And Function

Background: Dimethyl fumarate (DMF) and its active metabolite monomethyl fumarate (MMF) effectively lead to reduction in disease relapses and active magnetic resonance imaging (MRI) lesions. DMF and MMF are known to be effective in modulating T- and B-cell responses; however, their effect on the phenotype and function of human myeloid dendritic cells (mDCs) is not fully understood. Objective: To investigate the role of MMF on human mDCs maturation and function. Methods: mDCs from healthy controls were isolated and cultured in vitro with MMF. The effect of MMF on mDC gene expression was determined by polymerase chain reaction (PCR) array after in vitro MMF treatment. The ability of mDCs to activate T cells was assessed by in vitro co-culture system. mDCs from DMF-treated multiple sclerosis (MS) patients were analyzed by flow cytometry and PCR. Results: MMF treatment induced a less mature phenotype of mDCs with reduced expression of major histocompatibility complex class II (MHC-II), co-stimulatory molecules CD86, CD40, CD83, and expression of nuclear factor κB (NF-κB) subunits RELA and RELB. mDCs from DMF-treated MS patients also showed the same immature phenotype. T cells co-cultured with MMF-treated mDCs showed reduced proliferation with decreased production of interferon gamma (IFN-γ), interleukin-17 (IL-17), and granulocyte-macrophage colony-stimulating factor (GM-CSF) compared to untreated cells. Conclusion: We report that MMF can modulate immune response by affecting human mDC function.

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HLA-G expression defines a novel regulatory T-cell subset present in human peripheral blood and sites of inflammation

Blood ◽

10.1182/blood-2006-11-057125 ◽

2007 ◽

Vol 110 (2) ◽

pp. 568-577 ◽

Cited By ~ 122

Author(s):

Ute Feger ◽

Eva Tolosa ◽

Yu-Hwa Huang ◽

Anne Waschbisch ◽

Tilo Biedermann ◽

...

Keyword(s):

Skeletal Muscle ◽

T Cells ◽

Cerebrospinal Fluid ◽

Cd8 T Cells ◽

Peripheral Blood ◽

Inflammatory Responses ◽

Human Peripheral Blood ◽

Cell Subset ◽

Single Positive

Abstract Regulatory T cells can inhibit harmful immunopathologic responses directed against self and foreign antigens and play a major role in controlling autoimmunity. Here we have identified and characterized a subpopulation of CD4 and CD8 T cells in human peripheral blood expressing the immune tolerizing molecule HLA-G. HLA-G–expressing T cells are hypoproliferative, are CD25- and FOXP3-negative, and exhibit potent suppressive properties that are partially mediated by HLA-G. HLA-G–positive (HLA-Gpos) T cells are found at low percentages among CD4 and CD8 single-positive thymocytes, suggesting a thymic origin. The presence of HLA-Gpos T cells at sites of inflammation such as inflamed skeletal muscle in myositis or the cerebrospinal fluid of patients with acute neuroinflammatory disorders suggests an important function in modulating parenchymal inflammatory responses in vivo.

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High-Fat Diet Alters Immunogenic Properties of Circulating and Adipose Tissue-Associated Myeloid-Derived CD45+DDR2+ Cells

Mediators of Inflammation ◽

10.1155/2019/1648614 ◽

2019 ◽

Vol 2019 ◽

pp. 1-15

Author(s):

Sara J. Sidles ◽

Ying Xiong ◽

M. Rita I. Young ◽

Amanda C. LaRue

Keyword(s):

T Cells ◽

Adipose Tissue ◽

T Cell ◽

Peripheral Blood ◽

Cell Activity ◽

Cell Subset ◽

Total Body Weight ◽

T Cell Subsets ◽

Activation Markers

Chronic inflammation is evident in the adipose tissue and periphery of patients with obesity, as well as mouse models of obesity. T cell subsets in obese adipose tissue are skewed towards Th1- and Th17-associated phenotypes and their secreted cytokines contribute to obesity-associated inflammation. Our lab recently identified a novel, myeloid-derived CD45+DDR2+ cell subset that modulates T cell activity. The current study sought to determine how these myeloid-derived CD45+DDR2+ cells are altered in the adipose tissue and peripheral blood of preobese mice and how this population modulates T cell activity. C57BL/6 mice were fed with a diet high in milkfat (60%·kcal, HFD) ad libitum until a 20% increase in total body weight was reached, and myeloid-derived CD45+DDR2+ cells and CD4+ T cells in visceral adipose tissue (VAT), mammary gland-associated adipose tissue (MGAT), and peripheral blood (PB) were phenotypically analyzed. Also analyzed was whether mediators from MGAT-primed myeloid-derived CD45+DDR2+ cells stimulate normal CD4+ T cell cytokine production. A higher percentage of myeloid-derived CD45+DDR2+ cells expressed the activation markers MHC II and CD80 in both VAT and MGAT of preobese mice. CD4+ T cells were preferentially skewed towards Th1- and Th17-associated phenotypes in the adipose tissue and periphery of preobese mice. In vitro, MGAT from HFD-fed mice triggered myeloid-derived CD45+DDR2+ cells to induce CD4+ T cell IFN-γ and TNF-α production. Taken together, this study shows that myeloid-derived CD45+DDR2+ cells express markers of immune activation and suggests that they play an immune modulatory role in the adipose tissue of preobese mice.

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TNF-Alpha Priming Enhances CD4+ FoxP3+ Regulatory T Cell Suppressive Function in GvHD Prevention and Treatment

Blood ◽

10.1182/blood.v126.23.1885.1885 ◽

2015 ◽

Vol 126 (23) ◽

pp. 1885-1885

Author(s):

Antonio Pierini ◽

Caitlin Moffett ◽

Dominik Schneidawind ◽

Jeanette Baker ◽

Hidekazu Nishikii ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Mouse Model ◽

Peripheral Blood ◽

Activation Markers ◽

Suppressive Function ◽

Prevention And Treatment ◽

And Function

Abstract CD4+ CD25+ FoxP3+ regulatory T cells (Treg) have been shown to effectively prevent graft versus host disease (GvHD) when adoptively transferred in murine models of hematopoietic cell transplantation (HCT) and phase I/II clinical trials. Critical limitations to the clinical application of Treg are the paucity of cells and limited knowledge of the mechanism(s) of in vivo function. In physiologic conditions Treg regulate immune responses during inflammation. We hypothesized that inflammatory conditions in GvHD modify Treg characteristics and function. To test this hypothesis, we primed Treg with irradiated (3000 cGy) peripheral blood from acute GvHD (aGvHD) affected mice for 20-24 hours and then transferred these cells in a mouse model of GvHD where allogeneic T cell depleted bone marrow (TCD BM) from C57BL/6 mice was transplanted into lethally irradiated (8 Gy) BALB/c recipients together with 7.5x105 to 1x106 /animal donor derived conventional CD4+ and CD8+ T cells (Tcon). C57BL/6 Treg primed with irradiated aGvHD peripheral blood were injected at day 0 after HCT for preventing GvHD or at day +7 or +8 as GvHD treatment. Their adoptive transfer resulted in improved survival in comparison to unprimed natural occurring Treg when used for both GvHD prevention (p=0.01) and treatment (p=0.04). Moreover treatment with irradiated aGvHD peripheral blood-primed Treg did not impact graft versus tumor effects in a mouse model of T cell mediated tumor killing. BLI demonstrated that injected allogeneic Tcon completely cleared previously infused luc+ A20 tumor cells even in the presence of primed Treg (primed Treg + Tcon + A20 vs A20 alone p<0.001). Irradiated aGvHD peripheral blood-primed Treg express increased levels of activation markers with suppressive function such as CTLA4 (p<0.001) and LAG3 (p<0.05) in comparison to unprimed Treg in vitro. We also found that Treg primed with irradiated cells of aGvHD affected animals after removing the serum did not enhance the expression of the same markers (p>0.05) demonstrating that serum from aGvHD animals is required for Treg priming and function. We further tested the ability of several inflammatory cytokines that are normally secreted during GvHD such as IFN-γ, IL-6, IL-12 and TNFα to induce similar in vitro Treg activation. We found that TNFɑ selectively activated Treg without impacting CD4+ FoxP3- T cells. TNFɑ-primed Treg have increased expression of activation markers such as CD69 (p<0.0001), CD25 (p<0.0001), and LAG3 (p=0.0002), produce a greater amount of suppressive cytokines such as IL-10 (p=0.03) and TGF-β (p=0.02), and enhance the expression of homing markers such as CD62L (p=0.005) that are required for in vivo function. TNFɑ-primed Treg had increased ability to proliferate (p=0.02) and, at the same time, to suppress Tcon proliferation (p=0.04) in a mixed lymphocyte reaction against irradiated allogeneic splenocytes, while, on the contrary, TNFɑ-primed Tcon had reduced ability to proliferate in similar conditions in comparison to unprimed Tcon (p=0.0004). To test the effect of TNFɑ priming on in vivo Tcon proliferation we injected TNFɑ-primed and unprimed luc+ Tcon in allogeneic BALB/c Rag2-/- γ-chain-/- immune deficient animals that were sublethally irradiated (400 cGy). BLI at day +7 after Tcon injection revealed reduced TNFɑ-primed Tcon in vivo proliferation (p=0.01) that resulted in milder GvHD symptoms (p=0.02). Finally, in a GvHD prevention mouse model TNFɑ-primed Treg infused at 1:10 Treg/Tcon ratio resulted in improved animal survival as compared to unprimed Treg (p=0.02), demonstrating enhanced efficacy of TNFɑ priming in the in vivo function of Treg. In summary, our study demonstrates that Treg respond to TNFɑ acquiring an activated status resulting in increased function. As TNFɑ is produced by several immune cells during inflammation, our work elucidates aspects of the physiologic mechanisms of Treg function. Furthermore TNFɑ priming of Treg in vitro provides a new tool to optimize Treg cellular therapies also allowing for the use of a reduced cell number for GvHD prevention and treatment. Disclosures No relevant conflicts of interest to declare.

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Interleukin-17 activates human renal epithelial cells in vitro and is expressed during renal allograft rejection.

Journal of the American Society of Nephrology ◽

10.1681/asn.v981526 ◽

1998 ◽

Vol 9 (8) ◽

pp. 1526-1534

Author(s):

C Van Kooten ◽

J G Boonstra ◽

M E Paape ◽

F Fossiez ◽

J Banchereau ◽

...

Keyword(s):

T Cells ◽

Epithelial Cells ◽

Inflammatory Responses ◽

Biological Activities ◽

Critical Role ◽

Interleukin 17 ◽

Tubular Epithelial Cells ◽

Proximal Tubular Epithelial Cells ◽

Proximal Tubular

Local production of cytokines plays a critical role in the regulation of pathophysiologic processes leading to rejection of transplanted organs. In the present study, the possible role of interleukin-17 (IL-17), a recently identified cytokine with unique properties, was investigated. IL-17 is specifically produced by activated T cells, whereas biological activities are restricted to the activation of nonhematopoietic cells. In vitro, IL-17 induced primary human proximal tubular epithelial cells, a type of cell regulating local interstitial inflammatory responses, to secrete higher levels of IL-6, IL-8, and monocyte chemoattractant protein-1, but not of the chemokine RANTES. The effect was specific for IL-17, because it was completely abrogated by a neutralizing anti-IL-17 antibody and was demonstrated to be dose- and time-dependent. In addition, IL-17 increased the production of complement component C3 by human proximal tubular epithelial cells, but not of other complement components. Immunofluorescence showed expression of IL-17 in kidney biopsies from patients suffering from graft rejection (8 of 8 positive), whereas pretransplant biopsies and normal kidneys were negative (0 of 6). Analysis of whole kidney isolates confirmed the presence of IL-17 mRNA by reverse transcription-PCR. IL-17 expression could also be found in in vitro cultured and activated graft-infiltrating T cells. These results represent the first demonstration of IL-17 protein expression in pathologic conditions and suggest that IL-17 might be important in the regulation of local inflammatory responses.

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SLAM Negatively Regulates IL-21 Production in Tfh-Like Cells from Allergic Rhinitis Patients

10.21203/rs.3.rs-93330/v1 ◽

2020 ◽

Author(s):

Jun Yang ◽

Lina Geng ◽

Yongming Ma ◽

Xinyi Tang ◽

Huiyong Peng ◽

...

Keyword(s):

T Cells ◽

Allergic Rhinitis ◽

B Cells ◽

Cell Subset ◽

Follicular Helper ◽

Th Cell ◽

Signaling Lymphocytic Activation Molecule ◽

And Function ◽

Humoral Responses

Abstract Background: Allergic rhinitis (AR) is characterized by type Ⅰ hypersensitivity that is mediated by IgE-induced humoral responses. It is known that follicular helper T cells (Tfh) comprise the key Th cell subset that promotes antibody production. Signaling lymphocytic activation molecule (SLAM) participates in the regulation of the differentiation and function of Tfh cells, but whether this regulation is involved in the pathogenesis of AR is unknown.Methods: CD4+CXCR5+ Tfh-like cells from peripheral blood were detected by flow cytometry. The levels of IL-21 and IgE in serum were tested by ELISA. Blood CD4+CXCR5+ Tfh-like cells were sorted and cultured with anti-SLAM mAb in vitro.Results: The frequencies of circulating CD4+CXCR5+ Tfh-like cells appeared virtually unchanged in AR patients, but the expression of SLAM and SLAM-associated protein (SAP) on circulating Tfh-like cells was significantly decreased. Meanwhile, the level of serum IL-21 was increased in AR patients, and there was a negative correlation between the levels of IL-21 and the expression of SLAM or SAP in CD4+CXCR5+ T cells. Treatment with anti-SLAM mAb resulted in the reduced production of IL-21 by Tfh-like cells in vitro. Additionally, the expression of SLAM on B cells significantly decreased, although the percentages of B cells were increased in AR patients.Conclusions: SLAM negatively regulates the production of IL-21 in CD4+CXCR5+ Tfh-like cells, which contributes to the pathogenesis of AR.

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Impact of Superantigen-Producing Bacteria on T Cells from Tonsillar Hyperplasia

Pathogens ◽

10.3390/pathogens8030090 ◽

2019 ◽

Vol 8 (3) ◽

pp. 90 ◽

Cited By ~ 1

Author(s):

Fiona J Radcliff ◽

Sharon Waldvogel-Thurlow ◽

Fiona Clow ◽

Murali Mahadevan ◽

James Johnston ◽

...

Keyword(s):

T Cells ◽

Peripheral Blood ◽

Group A Streptococcus ◽

Mait Cells ◽

Tonsillar Hyperplasia ◽

Close Proximity ◽

Group A ◽

Tonsil Tissue ◽

And Function

Staphylococcus aureus and Group A Streptococcus (GAS) are common occupants of the tonsils and many strains produce potent exotoxins (mitogens) that directly target T cells, which could be a driver for tonsillar hyperplasia. Tonsil tissues from 41 patients were tested for these bacteria in conjunction with profiling of B and T cells by flow cytometry. S. aureus and GAS were detected in tonsil tissue from 44% and 7%, respectively, of patients by bacteriological culture; immuno-histology showed bacteria in close proximity to both B and T lymphocytes. The presence of tonsillar S. aureus did not alter B or T cell populations, whereas peripheral blood mucosal-associated invariant T (MAIT) cells were significantly increased in S. aureus culture positive individuals (p < 0.006). Alterations of tonsil CD4+ TCR Vβ family members relative to peripheral blood were evident in 29 patients. Three patients had strong TCR Vβ skewing indicative of recent exposure to superantigens, their tonsils contained mitogenic bacteria, and supernatants from these bacteria were used to partially recapitulate the skewing profile in vitro, supporting the notion that superantigens can target tonsillar T cells in situ. Tonsils are a reservoir for superantigen-producing bacteria with the capacity to alter the composition and function of key immune cells.

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CD8 Treg-Mediated Suppression of Naive CD4+ T Cell Differentiation into Follicular Helper T Cells

International Archives of Allergy and Immunology ◽

10.1159/000521427 ◽

2021 ◽

pp. 1-11

Author(s):

Taissa M. Kasahara ◽

Sudhir Gupta

Keyword(s):

T Cells ◽

Cell Differentiation ◽

T Cell ◽

Inflammatory Responses ◽

Treg Cells ◽

Follicular Helper ◽

And Function ◽

Proliferation In Vitro

Background: The regulatory CD8+ T (CD8+ Treg) cells play an important role in immune tolerance and have been implicated in several human autoimmune diseases. In this context, follicular helper T (TFH) cells contribute by controlling the antibody production. In mice, CD8+ Treg cells control the number and function of TFH cells however the role of human CD8+ Treg cells on the differentiation of naive CD4+ T cells into TFH cells has not been studied. Objectives: Here, we evaluated the ability of human CD183+ CD8+ Treg cells to suppress TFH cell differentiation in vitro. Methods: Activated CD183+CCR7+CD45RA−CD8+ Treg and CD183+CD25highICOS+CD8+ Treg cells were sorted and cocultured with naïve CD4+ T cells under TFH differentiation condition. The differentiation of TFH cells was evaluated by flow cytometry. Results: Our results showed that activated CD183+CD8+ Treg cells upregulated the expression of Forkhead box P3 transcription factor, inducible T-cell co-stimulator (ICOS), and CD25 compared to CD183−CD8+ T cells. The CD183+CD25highICOS+CD8+ Treg cells suppressed TFH cell differentiation and CD4+ T cell proliferation in vitro which was not observed when CD183+CCR7+CD45RA−CD8+ Treg were cocultured with naïve CD4+ T cells under TFH cell differentiation condition. Conclusion: These results suggest that CD25highICOS+CD183+CD8+ Treg cells may regulate autoimmune and inflammatory responses mediated by TFH cells.

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Suppressor CD4+ T cells expressing HLA-G are expanded in the peripheral blood from patients with acute decompensation of cirrhosis

Gut ◽

10.1136/gutjnl-2021-324071 ◽

2021 ◽

pp. gutjnl-2021-324071

Author(s):

Wafa Khamri ◽

Cathrin Gudd ◽

Tong Liu ◽

Rooshi Nathwani ◽

Marigona Krasniqi ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Peripheral Blood ◽

Cytotoxic T Lymphocyte ◽

Cell Subset ◽

Infectious Complications ◽

Decompensated Cirrhosis ◽

T Helper 17 ◽

Acute Decompensation

ObjectiveIdentifying components of immuneparesis, a hallmark of chronic liver failure, is crucial for our understanding of complications in cirrhosis. Various suppressor CD4+ T cells have been established as potent inhibitors of systemic immune activation. Here, we establish the presence, regulation and mechanism of action of a suppressive CD4+ T cell subset expressing human leucocyte antigen G (HLA-G) in patients with acute decompensation of cirrhosis (AD).DesignFlow cytometry was used to determine the proportion and immunophenotype of CD4+HLA-G+ T cells from peripheral blood of 20 healthy controls (HCs) and 98 patients with cirrhosis (28 with stable cirrhosis (SC), 20 with chronic decompensated cirrhosis (CD) and 50 with AD). Transcriptional and functional signatures of cell-sorted CD4+HLA-G+ cells were delineated by NanoString technology and suppression assays, respectively. The role of immunosuppressive cytokine interleukin (IL)-35 in inducing this population was investigated through in vitro blockade experiments. Immunohistochemistry (IHC) and cultures of primary human Kupffer cells (KCs) were performed to assess cellular sources of IL-35. HLA-G-mediated T cell suppression was explored using neutralising antibodies targeting co-inhibitory pathways.ResultsPatients with AD were distinguished by an expansion of a CD4+HLA-G+CTLA-4+IL-35+ immunosuppressive population associated with disease severity, clinical course of AD, infectious complications and poor outcome. Transcriptomic analyses excluded the possibility that these were thymic-derived regulatory T cells. IHC analyses and in vitro cultures demonstrate that KCs represent a potent source of IL-35 which can induce the observed HLA-G+ phenotype. These exert cytotoxic T lymphocyte antigen-4-mediated impaired responses in T cells paralleled by an HLA-G-driven downregulation of T helper 17-related cytokines.ConclusionWe have identified a cytokine-driven peripherally derived suppressive population that may contribute to immuneparesis in AD.

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