Hepatic AAV Gene Transfer of Cytoplasmic Transgene Induces Transgene Product-Specific T Cell Activation Initially in the Liver and Celiac Lymph Node Preceding Treg Induction

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3241-3241
Author(s):  
Roland W. Herzog ◽  
George Q. Perrin
Keyword(s):  
T Cells ◽  
Lymph Node ◽  
T Cell ◽  
Gene Transfer ◽  
Antigen Presentation ◽  
T Cell Activation ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Lymphoid Organs ◽  
Activated T Cells

Abstract In several published studies, we have shown induction of immune tolerance to coagulations factors by hepatic gene transfer to animals with hemophilia. Tolerance induction is influenced by a number of complex factors, most notably T cell activation and induction of antigen-specific CD4+CD25+FoxP3+ regulatory T cells (Treg). We sought to better understand antigen presentation to CD4+ T cells and the dynamics of the resulting T cell response. To characterize the interaction of adeno-associated virus (AAV) antigen expression in the liver with immune cells, we used an AAV8 vector, which have a high tropism for murine liver, expressing cytoplasmic ovalbumin (AAV8-Cyto-Ova) from the EF1α promoter. Use of AAV8-Cyto-Ova allowed us to eliminate effects from systemic antigen delivery. Vector was injected into the tail vein of DO11.10-transgenic RAG-/- mice, which contain exclusively Ova-specific CD4+ T cells and lack Treg. AAV8-Cyto-Ova caused upregulation of the very early activation marker CD69 on the CD4+ T cells as early as 2 weeks after gene transfer, with induced Treg emerging at about 3 weeks. CD69+CD4+ T cells were first observed in greatest numbers in the liver and celiac lymph node (LN), one of the liver-draining LN. This T cell activation persisted for several weeks. To better define the sites of T cell activation, we used the compound FTY720, which is an agonist of sphingosine-1-phosphate receptors and prevents migration of lymphocytes but does not alter T cell function. Two weeks after AAV8-Cyto-Ova, FTY720 sequestered activated T cells mostly in the liver and celiac LN, when compared to other lymphoid organs, indicating that these are the initial sites of T cell activation. At the 3-week time point, there were fewer activated T cells in the liver and celiac LN in mice that received FTY720, while instead accumulating in the blood. Most likely, activated T cells were prevented from reentering the lymphoid organs from the circulation, where they were sequestered. We conclude that T cells are first activated by AAV8-Cyto-Ova in the liver and celiac LN after two weeks, where they subsequently egress into the circulation and re-enter lymphoid tissues, with many returning to the liver and celiac LN. FTY720 given at 2 weeks prevented the newly activated T cells from leaving the liver and celiac LN. These results strongly suggest that antigen presentation and CD4+ T cell activation occur first in the liver and celiac LN, beginning about 2 weeks after vector administration. Consistent with this conclusion, adoptively transferred Ova-specific CD4+ T cells proliferated first and to a much greater degree in the celiac LN of AAV8-Cyto-Ova transduced mice. Inactiviating Kupffer cells with gadolinium chloride significantly reduced antigen-specific proliferation, illustrating the requirement for professional resident liver antigen-presenting cells. Furthermore, we show that - in contrast to the AAV expression of secreted Ova - Treg are exclusively extrathymically induced after AAV8-Cyto-Ova vector administration. These Treg are found in high numbers in the blood after 2 weeks in mice given the FTY720 compound, suggesting that these peripherally induced Treg quickly enter the circulation. In conclusion, the liver and its draining celiac LN are key sites for antigen presentation and T cell activation in response to transgene expression directed by hepatic gene transfer. Presentation of antigen derived from a non-secreted transgene product induces FoxP3+ Treg that rapidly distribute through the circulation. Disclosures Herzog: Novo Nordisk: Research Funding; Spark Therapeutics: Patents & Royalties: Patent licenses.

scholarly journals Isolevuglandin-Modified Cardiac Proteins Drive CD4+ T Cell Activation in the Heart and Promote Cardiac Dysfunction

Circulation ◽  
2021 ◽  
Author(s):  
Njabulo Ngwenyama ◽  
Annet Kirabo ◽  
Mark Aronovitz ◽  
Francisco Velázquez ◽  
Francisco Carrillo-Salinas ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Antigen Presentation ◽  
T Cell Activation ◽  
Cardiac Dysfunction ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Cd4 T Cell ◽  
Myocardial Oxidative Stress

Background: Despite the well-established association between T cell-mediated inflammation and non-ischemic heart failure (HF), the specific mechanisms triggering T cell activation during the progression of HF and the antigens involved are poorly understood. We hypothesized that myocardial oxidative stress induces the formation of isolevuglandin (IsoLG)-modified proteins that function as cardiac neoantigens to elicit CD4+ T cell receptor (TCR) activation and promote HF. Methods: We used transverse aortic constriction (TAC) in mice to trigger myocardial oxidative stress and T cell infiltration. We profiled the TCR repertoire by mRNA sequencing of intramyocardial activated CD4+ T cells in Nur77 GFP reporter mice, which transiently express GFP upon TCR engagement. We assessed the role of antigen presentation and TCR specificity in the development of cardiac dysfunction using antigen presentation-deficient MhcII -/- mice, and TCR transgenic OTII mice that lack specificity for endogenous antigens. We detected IsoLG-protein adducts in failing human hearts. We also evaluated the role of reactive oxygen species (ROS) and IsoLGs in eliciting T cell immune responses in vivo by treating mice with the antioxidant TEMPOL, and the IsoLG scavenger 2-hydroxybenzylamine (2-HOBA) during TAC, and ex-vivo in mechanistic studies of CD4+ T cell proliferation in response to IsoLG-modified cardiac proteins. Results: We discovered that TCR antigen recognition increases in the left ventricle (LV) as cardiac dysfunction progresses, and identified a limited repertoire of activated CD4+ T cell clonotypes in the LV. Antigen presentation of endogenous antigens was required to develop cardiac dysfunction since MhcII -/- mice reconstituted with CD4+ T cells, and OTII mice immunized with their cognate antigen were protected from TAC-induced cardiac dysfunction despite the presence of LV-infiltrated CD4+ T cells. Scavenging IsoLGs with 2-HOBA reduced TCR activation and prevented cardiac dysfunction. Mechanistically, cardiac pressure overload resulted in ROS dependent dendritic cell accumulation of IsoLG-protein adducts which induced robust CD4+ T cell proliferation. Conclusions: Collectively, our study demonstrates an important role of ROS-induced formation of IsoLG-modified cardiac neoantigens that lead to TCR-dependent CD4+ T cell activation within the heart.

scholarly journals MHCII glycosylation modulates Bacteroides fragilis carbohydrate antigen presentation

2011 ◽  
Vol 208 (5) ◽  
pp. 1041-1053 ◽  
Author(s):  
Sean O. Ryan ◽  
Jason A. Bonomo ◽  
Fan Zhao ◽  
Brian A. Cobb
Keyword(s):  
T Cells ◽  
T Cell ◽  
Antigen Presentation ◽  
T Cell Activation ◽  
Cell Activation ◽  
Bacteroides Fragilis ◽  
Carbohydrate Antigen ◽  
Activated T Cells ◽  
Mhc Molecules

N-linked glycans are thought to protect class II major histocompatibility complex (MHC) molecules (MHCII) from proteolytic cleavage and assist in arranging proteins within the immune synapse, but were not thought to directly participate in antigen presentation. Here, we report that antigen-presenting cells (APCs) lacking native complex N-glycans showed reduced MHCII binding and presentation of the T cell activating glycoantigen (GlyAg) polysaccharide A from Bacteroides fragilis but not conventional peptides. APCs lacking native N-glycans also failed to mediate GlyAg-driven T cell activation but activated T cells normally with protein antigen. Mice treated with the mannosidase inhibitor kifunensine to prevent the formation of complex N-glycans were unable to expand GlyAg-specific T cells in vivo upon immunization, yet adoptive transfer of normally glycosylated APCs into these animals overcame this defect. Our findings reveal that MHCII N-glycosylation directly impacts binding and presentation of at least one class of T cell–dependent antigen.

scholarly journals Quantitative Proteomics Reveals Protein Dysregulation During T Cell Activation in Multiple Sclerosis Patients Compared to Healthy Controls

2021 ◽  
Author(s):  
Chiara Cappelletti ◽  
Anna Maria Eriksson ◽  
Ina Skaara Brorson ◽  
Ingvild S. Leikfoss ◽  
Oda Glomstad Kråbøl ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Susceptibility Genes ◽  
Healthy Controls ◽  
Activated T Cells ◽  
Multiple Sclerosis Patients

Abstract Background: Multiple sclerosis (MS) is an autoimmune, neurodegenerative disorder with a strong genetic component that acts in a complex interaction with environmental factors for disease development. CD4 + T cells are pivotal players in MS pathogenesis, where peripherally activated T cells migrate to the central nervous system leading to demyelination and axonal degeneration. Through a proteomic approach, we aim at identifying dysregulated pathways in activated T cells from MS patients as compared to healthy controls. Methods: CD4 + T cells were purified from peripheral blood from MS patients and healthy controls by magnetic separation. Cells were left unstimulated or stimulated in vitro through the TCR and costimulatory CD28 receptor for 24 hours prior to sampling. Electrospray liquid chromatographytandem mass spectrometry was used to measure protein abundances. Results: Upon T cell activation the abundance of 1,801 proteins was changed. Among these proteins, we observed an enrichment of proteins expressed by MS-susceptibility genes. When comparing protein abundances in T cell samples from healthy controls and MS patients, 18 and 33 proteins were differentially expressed in unstimulated and stimulated CD4 + T cells, respectively. Moreover, 353 and 304 proteins were identified as proteins exclusively induced upon T cell activation in healthy controls and MS patients, respectively and dysregulation of the Nur77 pathway was observed only in samples from MS patients. Conclusions: Our study highlights the importance of CD4 + T cell activation for MS, as proteins that change in abundance upon T cell activation are enriched for proteins encoded by MS susceptibility genes. The results provide evidence for proteomic disturbances in T cell activation in MS, and pinpoint to dysregulation of the Nur77 pathway, a biological pathway known to limit aberrant effector T cell responses.

scholarly journals The Ca2+-activated K+ channel KCa3.1 compartmentalizes in the immunological synapse of human T lymphocytes

2007 ◽  
Vol 292 (4) ◽  
pp. C1431-C1439 ◽  
Author(s):  
Stella A. Nicolaou ◽  
Lisa Neumeier ◽  
YouQing Peng ◽  
Daniel C. Devor ◽  
Laura Conforti
Keyword(s):  
T Cells ◽  
T Cell ◽  
Antigen Presentation ◽  
T Cell Activation ◽  
Cell Activation ◽  
Immunological Synapse ◽  
Cell Receptor ◽  
K Channel ◽  
Activated T Cells ◽  
Barr Virus

T cell receptor engagement results in the reorganization of intracellular and membrane proteins at the T cell-antigen presenting cell interface forming the immunological synapse (IS), an event required for Ca2+ influx. KCa3.1 channels modulate Ca2+ signaling in activated T cells by regulating the membrane potential. Nothing is known regarding KCa3.1 membrane distribution during T cell activation. Herein, we determined whether KCa3.1 translocates to the IS in human T cells using YFP-tagged KCa3.1 channels. These channels showed electrophysiological and pharmacological properties identical to wild-type channels. IS formation was induced by either anti-CD3/CD28 antibody-coated beads for fixed microscopy experiments or Epstein-Barr virus-infected B cells for fixed and live cell microscopy. In fixed microscopy experiments, T cells were also immunolabeled for F-actin or CD3ε, which served as IS formation markers. The distribution of KCa3.1 was determined with confocal and fluorescence microscopy. We found that, upon T cell activation, KCa3.1 channels localize with F-actin and CD3ε to the IS but remain evenly distributed on the cell membrane when no stimulus is provided. Detailed imaging experiments indicated that KCa3.1 channels are recruited in the IS shortly after antigen presentation and are maintained there for at least 15–30 min. Interestingly, pretreatment of activated T cells with the specific KCa3.1 blocker TRAM-34 blocked Ca2+ influx, but channel redistribution to the IS was not prevented. These results indicate that KCa3.1 channels are a part of the signaling complex that forms at the IS upon antigen presentation.

Azacitidine Induces A Shift In Th1/Th17 Ratio and FoxP3 Expression In Anti-CD3 Stimulated CD4+ T-Cells; Implications for the Treatment of Myelodysplastic Syndromes

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4026-4026
Author(s):  
Hetty J Bontkes ◽  
Jurjen M Ruben ◽  
Theresia M. Westers ◽  
Gert J. Ossenkoppele ◽  
Arjan A. Van de Loosdrecht
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cd4 T Cells ◽  
Th17 Cells ◽  
Dna Methyltransferase ◽  
Cell Activation ◽  
Foxp3 Expression ◽  
Cd4 T Cell ◽  
Activated T Cells

Abstract Abstract 4026 Aberrant DNA methylation and other epigenetic changes play a role in the development of myelodysplastic syndromes (MDS). Epigenetic drugs such as DNA methyltransferase inhibitors are therefore increasingly employed in MDS treatment regimens. Recent studies show that gene methylation processes also regulate T-cell function. Here we analyzed the in vitro effects of the DNA methyltransferase inhibitor ‘5-azacitidine (Aza) on CD4+ T-cell activation. We confirmed the previously described inhibition of proliferation and increased expression of FoxP3, the regulatory T-cell (Treg) marker, by anti-CD3 stimulated T-cells in the presence of 1mM Aza. Here we have sorted CD4+ T-cells isolated form healthy donor peripheral blood into CD25neg resting, CD25dim recently activated and CD25hi Treg cells. Aza facilitated the induction of CD25hiFoxP3+ T-cells from CD25neg (4.7% of vehicle treated cells versus 17.3% of Aza treated cells p=0.0007, n=9) and to a lesser extend from CD25dim (1.2% versus 8.6%, p=0.0015, n=7) CD4+ T-cells, while Aza had no effect on FoxP3 expression in CD25hi sorted cells, FoxP3 expression remained high. In addition, cytokine producing T-cells were enumerated after stimulation with phorbol-12-myristate-13-acetate (PMA) and ionomycin in the presence of Brefeldin A. Aza treatment increased the number of IFNγ producing cells in the total CD4+ population (19.1% versus 40.8%; p<0.0001, n=10) as well as among the CD25neg (5.7% vs 41.2%; p=0.001, n=8) and CD25dim CD4+ T-cell populations (28.4% versus 46.6%; p=0.06, n=7). TNFα producing cells were increased in the total CD4+ (36.4% versus 51.1%; p=0.011, n=9) and CD4posCD25dim (36.7% versus 52.3%; p=0.033, n=6) populations but not in the CD4posCD25neg cells (50.9% versus 51.1%; p=0.9, n=7). This increase in pro-inflammatory cytokine production indicates that Aza induces T-cell activation and that the increase in FoxP3 expression may reflect T-cell activation rather than an increase in bona fide Treg by Aza treatment. Indeed a proportion of the FoxP3+ cells was positive for TNFa or IFNg, suggesting that these are activated T-cells rather than Treg. However, the proportion of FoxP3+IFNγ- and FoxP3+TNFα- cells was significantly higher among Aza treated CD4+ cells (p=0.0037 and 0.0018 respectively, n=5), suggesting an increase in Treg as well. Functional assays to demonstrate that these FoxP3+ cells are indeed regulatory T-cells are currently being set up. Next to IFNγ and TNFα producing Th1 cells, the more recently described IL-17 committed Th17 cells have been described to play a role in low risk MDS. Furthermore, it has been shown that Treg can differentiate into IL-17 producing cells. We, therefore evaluated the effect of Aza on Th17 cells. Interestingly, in contrast to IFNγ and TNFα producing cells, the proportion of IL-17+IFNγ- Th17 cells among the total CD4+ population was reduced by Aza treatment (1.8% versus 1.1% p=0.035, n=10), leading to a significant increase in the Th1/Th17 ratio (14.0 versus 47.9, p=0.0005, n=10). In contrast, although the numbers were small, the proportion of Th17 cells was increased by Aza in the CD25neg population (0.08% versus 0.20%, p=0.028, n=8), suggesting that Aza may have differential effects on resting and recently activated T-cells. In conclusion, our data show that Aza increases the induction of FoxP3+ Treg and Th1 cells but inhibits IL-17 production, particularly by previously activated T-cells. Aza may therefore particularly be beneficial in pathogenic immune disorders characterized by increased Th17 numbers accompanied by reduced Treg frequencies, such as low-risk MDS. Disclosures: No relevant conflicts of interest to declare.

Induction of Mouse Naive T Cells Transition into CD4+ CD2+ FoxP3+ Regulatory T Cells in a Modified Culture Medium As a Potential Armament in Graft-Versus-Host Disease

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 5426-5426
Author(s):  
Tzeon-Jye Chiou ◽  
Tan-Hwa Chu ◽  
Sin-Tak Chu ◽  
Woan-Fang Tzeng
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
T Cell Activation ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Foxp3 Expression ◽  
Treg Cells ◽  
Activated T Cells ◽  
Allogeneic Hsct

Abstract Allogeneic hematopoietic stem cell transplantation (HSCT) has been used to treat some of hematological malignancies and inherited or acquired non-malignant disorders. Unfortunately, graft-versus-host disease (GVHD) occurred approximately 15% in transplant recipients and impacts on the outcome of allogeneic HSCT. At present, no effective modality could completely prevent the GVHD from allogeneic HSCT patients. CD4+ CD25+ FoxP3+ regulatory T cells (Tregs) have been shown to be important in maintaining immune homeostasis and preventing autoimmunity. However, 5% to 10% Tregs could be measured in human CD4+ T cells and few Tregs would convert to conventional activated T cells because of losing FoxP3 expression orn Tregs in suppression of T cell activation. It had been reported to correlate with the occurrence and severity of GVHD in some study. In order to study the potential use of CD4+ CD25+ FoxP3+ Tregs for the prevention of GVHD, we attempt to evaluate the better efficient method to increase the number of induced Treg cells (i Tregs) in donor and stabilize the FoxP3 ini Treg cells. Using mouse as a model, the splenocytes were prepared from mouse spleen. Before having biological function,i Treg cells need to stabilize the FoxP3 protein expression. Using retinoic acid (RA, 0.1-5ng/ml) as a stabilizer of the FoxP3 protein expression can keep thei Treg cells in stable. The endogenous regulatory T cells (n Treg) can inhibit T cell activation, thereby affecting T cells intoi Treg efficiency. We should remove the n Treg cells from the CD4+ T cells. Therefore, CD4+ T cells were isolated by negative selection, and then using the n Treg removing kit, we harvested the CD4+ CD62L+ naïve T cells fori Treg cell induction. For this purpose, naïve CD4+ cells were harvested, and then activated with anti-CD3/CD28 Dynabeads in the presence of IL-2, TGF-β1 and retinoic acid (RA) containing RPMI1640 medium. During the Tregs induction, the activated T cells were performed under low nutrient supplement (5% FBS) for three days then refreshed the cells into the full nutrient supplement (10% FBS) for another four days. The harvested cells were analyzed by flow cytometry method with fluorescence-conjugated CD-antibodies, including CD4, CD25, CD127, CD62L and FoxP3. Currently, the removal of n Treg cells could improve the efficiency of i Treg cell formation from 15% to 70-80% under this modified culture method (Fig.1). Further improvement of human peripheral blood regulatory T cell generation efficiency is our ongoing target. Our study showed that the combination of IL-2, TGF-β1 and RA in 3-day-nutrient-deprived medium could convert naïve CD4+ CD62L+ T cells to CD4+ CD25+ FoxP3+i Treg cells and stabilize FoxP3 expression in thei Treg cells efficiently. Further, we will develop thei Treg suppression assay to clarify the biological function ofi Tregs in vitro. GVHD mouse model will be established by using allogeneic HSCT to verify the function of i Tregs in vivo, too. Disclosures No relevant conflicts of interest to declare.

scholarly journals Antigen Display, T-Cell Activation, and Immune Evasion during Acute and Chronic Ehrlichiosis

2009 ◽  
Vol 77 (10) ◽  
pp. 4643-4653 ◽  
Author(s):  
Bisweswar Nandi ◽  
Madhumouli Chatterjee ◽  
Kathryn Hogle ◽  
Maura McLaughlin ◽  
Katherine MacNamara ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Antigen Presentation ◽  
Cell Line ◽  
T Cell Activation ◽  
Cd4 T Cells ◽  
Chronic Infection ◽  
Cell Activation ◽  
Cd4 T Cell ◽  
Anatomical Location

ABSTRACT How spatial and temporal changes in major histocompatibility complex/peptide antigen presentation to CD4 T cells regulate CD4 T-cell responses during intracellular bacterial infections is relatively unexplored. We have shown that immunization with an ehrlichial outer membrane protein, OMP-19, protects mice against fatal ehrlichial challenge infection, and we identified a CD4 T-cell epitope (IAb/OMP-19107-122) that elicited CD4 T cells following either immunization or infection. Here, we have used an IAb/OMP-19107-122-specific T-cell line to monitor antigen display ex vivo during acute and chronic infection with Ehrlichia muris, a bacterium that establishes persistent infection in C57BL/6 mice. The display of IAb/OMP-19107-122 by host antigen-presenting cells was detected by measuring intracellular gamma interferon (IFN-γ) production by the T-cell line. After intravenous infection, antigen presentation was detected in the spleen, peritoneal exudate cells, and lymph nodes, although the kinetics of antigen display differed among the tissues. Antigen presentation and bacterial colonization were closely linked in each anatomical location, and there was a direct relationship between antigen display and CD4 T-cell effector function. Spleen and lymph node dendritic cells (DCs) were efficient presenters of IAb/OMP-19107-122, demonstrating that DCs play an important role in ehrlichial infection and immunity. Chronic infection and antigen presentation occurred within the peritoneal cavity, even in the presence of highly activated CD4 T cells. These data indicated that the ehrlichiae maintain chronic infection not by inhibiting antigen presentation or T-cell activation but, in part, by avoiding signals mediated by activated T cells.

scholarly journals 643 Vasoactive intestinal peptide as a novel immune checkpoint molecule in activated T cells

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A672-A672
Author(s):  
Sruthi Ravindranathan ◽  
Tenzin Passang Fnu ◽  
Edmund Waller
Keyword(s):  
T Cells ◽  
T Cell ◽  
Vasoactive Intestinal Peptide ◽  
T Cell Activation ◽  
Cd8 T Cells ◽  
Immune Checkpoint ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Checkpoint Inhibitors ◽  
Activated T Cells

BackgroundOnly a fraction of cancer patients responds to current antibody-based immune checkpoint inhibitors.1 Our lab has identified vasoactive intestinal peptide-receptor (VIP-R) signaling as a targetable immune checkpoint pathway in cancer. VIP is a small neuropeptide with known immunosuppressive effects on T cells, in particular, CD4+ T cells.2–5 However, little is known about VIP-R signaling in CD8+ T cells. To define mechanisms by which VIP limits T cell activation and function, we studied the regulation of VIP and VIP receptors (VIP-R) in T cells following their activation in vitro and in mouse models of cancer.MethodsT cells from healthy human donors and murine splenocytes were activated using anti-CD3 coated plates. Western blots measured intracellular pre-pro-VIP, along with its cognate receptors; VPAC1 and VPAC2. Purified cultures of CD4+ and CD8+ T cells were used to interrogate the protein expression on specific T cell subsets. Activation and chemokine receptor expression was assessed by flow cytometry to evaluate T cell response to VIP-R antagonists in vitro and in tumor-bearing mice engrafted with pancreatic cancer cell lines.ResultsBoth murine and human T cells upregulate pre-pro-VIP following TCR stimulation with similar kinetics of VIP receptors between species. VIP expression is upregulated in vivo following treatment of tumor-bearing mice with anti-PD1 MoAb. VIP expression is temporally correlated with the upregulation of other co-inhibitory molecules. VPAC1 expression modestly increased in activated T cells while VPAC2 expression decreased. A non-canonical high molecular weight (HMW) form of VPAC2-related protein robustly and transiently increase in activated T cells. Expression of HMW form of VPAC2 is only detected in activated CD4+ T cells. Of note, activated CD4+ but not CD8+ T cells upregulate pre-pro-VIP. Pharmacological inhibition of VIP-R signaling significantly increased CD69+, OX40+, Lag3+, and PD1+ expression in CD4+ subsets compared to activated T cells without VIP-R antagonists (p < 0.05). In contrast, CD8+ T cells upregulate VPAC1 but not VPAC2 receptor following activation. VIP-R antagonist treatment of activated CD8+ T cells significantly decreased CXCR4+ expression (p < 0.05). CXCR3 and CXCR5 expression were not affected by VIP-R antagonist treatment.ConclusionsVIP-R signaling is a novel immune autocrine and paracrine checkpoint pathway in activated CD4+ T cells. Activated CD4+ and CD8+ T cells demonstrate different kinetics of VPAC1 and VPAC2 expression, suggesting different immune-regulatory responses to VIP-R antagonists. Understanding VIP-R signaling induced during T cell activation can lead to specific drugs that target VIP-R pathways to enhance cancer immunotherapy.AcknowledgementsWe thank healthy volunteers for blood samples. The authors also thank the shared resources at Emory University, namely, Emory Flow Cytometry Core (EFCC) and Integrated Cellular Imaging Core (ICI) and Yerkes Nonhuman Primate Genomics Core that provided services or instruments at subsidized cost to conduct some of the reported experiments. This work was supported in part by Katz Foundation funding, Georgia Research Alliance, and Emory School of Medicine Dean's Imagine, Innovate and Impact (I3) venture award to Edmund K. Waller.ReferencesDarvin P, Toor SM, Sasidharan Nair V, Elkord E. Immune checkpoint inhibitors: recent progress and potential biomarkers. Experimental and Molecular Medicine 2018.Wang HY, Jiang XM, Ganea D. The Neuropeptides VIP and PACAP Inhibit IL-2 Transcription by Decreasing c-Jun and Increasing JunB Expression in T Cells. J Neuroimmunol 2000;104(1):68–78.Delgado M. Vasoactive intestinal peptide generates CD4+CD25+ regulatory T Cells in Vivo. J Leukoc Biol 2005.Anderson P, Gonzalez-Rey E. Vasoactive intestinal peptide induces cell cycle arrest and regulatory functions in human T cells at multiple levels. Mol Cell Biol 2010.Delgado M, Ganea D. Vasoactive intestinal peptide: a neuropeptide with pleiotropic immune functions. Amino Acids. NIH Public Access July 2013, 25–39.Ethics ApprovalDe-identified blood samples from consented healthy volunteers (IRB 00046063) were obtained with approval from Institutional Review Boards.

scholarly journals Probiotic supplementation reduces inflammatory profiles but does not prevent oral immune perturbations during SIV infection

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Rhianna Jones ◽  
Kyle Kroll ◽  
Courtney Broedlow ◽  
Luca Schifanella ◽  
Scott Smith ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Rhesus Macaques ◽  
Oral Microbiome ◽  
Probiotic Supplementation ◽  
Siv Infection ◽  
Anti Inflammatory

AbstractHIV/SIV infections lead to massive loss of mucosal CD4 + T cells and breakdown of the epithelial mucosa resulting in severe microbial dysbiosis and chronic immune activation that ultimately drive disease progression. Moreover, disruption of one of the most understudied mucosal environments, the oral cavity, during HIV-induced immunosuppression results in significant microbial and neoplastic co-morbidities and contributes to and predicts distal disease complications. In this study we evaluated the effects of oral probiotic supplementation (PBX), which can stimulate and augment inflammatory or anti-inflammatory pathways, on early SIV infection of rhesus macaques. Our study revealed that similar to the GI mucosae, oral CD4 + T cells were rapidly depleted, and as one of the first comprehensive analyses of the oral microflora in SIV infection, we also observed significant modulation among two genera, Porphyromonas and Actinobacillus, early after infection. Interestingly, although PBX therapy did not substantially protect against oral dysbiosis or ameliorate cell loss, it did somewhat dampen inflammation and T cell activation. Collectively, these data provide one of the most comprehensive evaluations of SIV-induced changes in oral microbiome and CD4 + T cell populations, and also suggest that oral PBX may have some anti-inflammatory properties in lentivirus infections.

scholarly journals Novel self-amplificatory loop between T cells and tenocytes as a driver of chronicity in tendon disease

2021 ◽  
pp. annrheumdis-2020-219335
Author(s):  
Emma Garcia-Melchor ◽  
Giacomo Cafaro ◽  
Lucy MacDonald ◽  
Lindsay A N Crowe ◽  
Shatakshi Sood ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Migration ◽  
T Cell ◽  
Inflammatory Cytokines ◽  
T Cell Activation ◽  
Cell Activation ◽  
Conditioned Media ◽  
Tissue Remodelling ◽  
Activated T Cells ◽  
T Cell Migration

ObjectivesIncreasing evidence suggests that inflammatory mechanisms play a key role in chronic tendon disease. After observing T cell signatures in human tendinopathy, we explored the interaction between T cells and tendon stromal cells or tenocytes to define their functional contribution to tissue remodelling and inflammation amplification and hence disease perpetuation.MethodsT cells were quantified and characterised in healthy and tendinopathic tissues by flow cytometry (FACS), imaging mass cytometry (IMC) and single cell RNA-seq. Tenocyte activation induced by conditioned media from primary damaged tendon or interleukin-1β was evaluated by qPCR. The role of tenocytes in regulating T cell migration was interrogated in a standard transwell membrane system. T cell activation (cell surface markers by FACS and cytokine release by ELISA) and changes in gene expression in tenocytes (qPCR) were assessed in cocultures of T cells and explanted tenocytes.ResultsSignificant quantitative differences were observed in healthy compared with tendinopathic tissues. IMC showed T cells in close proximity to tenocytes, suggesting tenocyte–T cell interactions. On activation, tenocytes upregulated inflammatory cytokines, chemokines and adhesion molecules implicated in T cell recruitment and activation. Conditioned media from activated tenocytes induced T cell migration and coculture of tenocytes with T cells resulted in reciprocal activation of T cells. In turn, these activated T cells upregulated production of inflammatory mediators in tenocytes, while increasing the pathogenic collagen 3/collagen 1 ratio.ConclusionsInteraction between T cells and tenocytes induces the expression of inflammatory cytokines/chemokines in tenocytes, alters collagen composition favouring collagen 3 and self-amplifies T cell activation via an auto-regulatory feedback loop. Selectively targeting this adaptive/stromal interface may provide novel translational strategies in the management of human tendon disorders.

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