scholarly journals In vivo delivery of a microRNA-regulated transgene induces antigen-specific regulatory T cells and promotes immunologic tolerance

Blood ◽  
2009 ◽  
Vol 114 (25) ◽  
pp. 5152-5161 ◽  
Author(s):  
Andrea Annoni ◽  
Brian D. Brown ◽  
Alessio Cantore ◽  
Lucia Sergi Sergi ◽  
Luigi Naldini ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Fluorescent Protein ◽  
Lentiviral Vector ◽  
Antigen Expression ◽  
Immunologic Tolerance ◽  
Green Fluorescent ◽  
Antigen Presenting ◽  
In Vivo Delivery

Abstract We previously showed that incorporating target sequences for the hematopoietic-specific microRNA miR-142 into an antigen-encoding transgene prevents antigen expression in antigen-presenting cells (APCs). To determine whether this approach induces immunologic tolerance, we treated mice with a miR-142–regulated lentiviral vector encoding green fluorescent protein (GFP), and subsequently vaccinated the mice against GFP. In contrast to control mice, no anti-GFP response was observed, indicating that robust tolerance to the transgene-encoded antigen was achieved. Furthermore, injection of the miR-142–regulated vector induced a population of GFP-specific regulatory T cells. Interestingly, an anti-GFP response was observed when microRNA miR-122a was inserted into the vector and antigen expression was detargeted from hepatocytes as well as APCs. This demonstrates that, in the context of lentiviral vector-mediated gene transfer, detargeting antigen expression from professional APCs, coupled with expression in hepatocytes, can induce antigen-specific immunologic tolerance.

scholarly journals Targeting Antigen Presenting   Cells to Treat Autoimmune  Inflammation

2021 ◽  
Author(s):  
◽  
Aras Toker
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Mhc Class Ii ◽  
T Cell Proliferation ◽  
Target Cells ◽  
Antigen Presenting

<p>Glatiramer acetate (GA) is approved for the treatment of relapsing-remitting multiple sclerosis (MS), and can suppress experimental autoimmune encephalomyelitis (EAE), a murine model of human MS. GA treatment is associated with the induction of anti-inflammatory TH2 responses and with the antigen specific expansion of regulatory T cells that counteract or inhibit pathogenic events in MS and EAE. These T cell mediated mechanisms of protection are considered to be a result of modulation of antigen presenting cells (APCs) by GA, rather than direct effects on T cells. However, it is unknown if GA preferentially targets a specific APC subset or can act through multiple APCs in vivo. In addition, GA-modulated innate cells may also exhibit direct antigen non-specific suppression of autoreactive cells. One objective of this study was to identify the in vivo target cell population of GA and to assess the potential of the target cells to antigen non-specifically suppress immune responses. Fluorophor-labelled GA bound to monocytes after intravenous injections, suggesting that monocytes may be the primary target of GA in vivo. In addition, intravenous GA treatment enhanced the intrinsic ability of monocytes to suppress T cell proliferation, both in vitro and in vivo. The findings of this study therefore suggest that GA-induced monocytes may contribute to GA therapy through direct mechanisms of antigen non-specific T cell immunosuppression. A further objective of this work was to investigate the potential of an in vivo drug targeting approach. This approach was hypothesised to increase the uptake of GA by the target cells and substantially improve GA treatment through antigen specific mechanisms such as induction of TH2 or regulatory T cells. Targeting antigens to professional APCs with an anti-MHC class II antibody resulted in significantly enhanced T cell proliferation in vitro. However, no EAE suppression occurred when GA was targeted to MHC class II in vivo. In addition, targeting GA specifically to monocytes also failed to suppress EAE. These findings suggest that GA treatment may selectively modulate monocytes to enhance their ability to inhibit autoreactive T cells, which could be part of the mechanism by which GA ameliorates MS. Targeting GA to a specific cell type may not be a powerful approach to improve treatment, because increased proliferation of GA specific T cells is not sufficient for disease suppression, and conjugation to antibodies may functionally reduce GA to a mere antigen devoid of immunomodulatory capacity.</p>

The immune response to lentiviral-delivered transgene is modulated in vivo by transgene-expressing antigen-presenting cells but not by CD4+CD25+ regulatory T cells

Blood ◽  
2007 ◽  
Vol 110 (6) ◽  
pp. 1788-1796 ◽  
Author(s):  
Andrea Annoni ◽  
Manuela Battaglia ◽  
Antonia Follenzi ◽  
Angelo Lombardo ◽  
Lucia Sergi-Sergi ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
Adoptive Transfer ◽  
T Regulatory Cells ◽  
Lentiviral Vector ◽  
Antigen Presenting Cells ◽  
Effector T Cells ◽  
Systemic Delivery ◽  
Antigen Presenting

Abstract Systemic delivery of lentiviral vector (LV) in immunocompetent mice leads to efficient in vivo cell transduction and expression of the encoded protein under the control of the ubiquitous promoter of human cytomegalovirus (CMV). However, antitransgene immune response results in clearance of transduced cells 4 weeks after injection. T regulatory cells (Tregs), which have been demonstrated to control immune responses in vivo, were tested for their ability to suppress antitransgene response leading to stable long-term expression. Adoptive transfer of natural CD4+CD25+ Tregs (nTregs) isolated from wild type (wt) mice or from transgene tolerant transgenic (tg) mice did not suppress the antitransgene immune response after LV delivery. These data demonstrate that neither increasing the endogenous pool of natural Tregs nor transferring nTregs selected in a transgene-expressing thymus can modulate the immune response and mediate sustained transgene expression. Conversely, adoptive transfer of antigen-presenting cells (APCs) isolated from transgene-tolerant tg mice efficiently reduced the immune response leading to stable LV-encoded protein expression in vivo. Reduction of CD8+ effector T cells was observed in LV-treated mice coinjected with transgene-expressing APCs compared with control mice. These data indicate that antitransgene immune response can be modulated by transgene-expressing APCs possibly through deletion of effector T cells.

scholarly journals Selective depletion of Foxp3+ regulatory T cells induces a scurfy-like disease

2007 ◽  
Vol 204 (1) ◽  
pp. 57-63 ◽  
Author(s):  
Katharina Lahl ◽  
Christoph Loddenkemper ◽  
Cathy Drouin ◽  
Jennifer Freyer ◽  
Jon Arnason ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Fluorescent Protein ◽  
Early Death ◽  
Artificial Chromosome ◽  
Skin Inflammation ◽  
Lymphoproliferative Disease ◽  
Control Element ◽  
Protein Fusion

The scurfy mutant mouse strain suffers from a fatal lymphoproliferative disease leading to early death within 3–4 wk of age. A frame-shift mutation of the forkhead box transcription factor Foxp3 has been identified as the molecular cause of this multiorgan autoimmune disease. Foxp3 is a central control element in the development and function of regulatory T cells (T reg cells), which are necessary for the maintenance of self-tolerance. However, it is unclear whether dysfunction or a lack of T reg cells is etiologically involved in scurfy pathogenesis and its human correlate, the IPEX syndrome. We describe the generation of bacterial artificial chromosome–transgenic mice termed “depletion of regulatory T cell” (DEREG) mice expressing a diphtheria toxin (DT) receptor–enhanced green fluorescent protein fusion protein under the control of the foxp3 gene locus, allowing selective and efficient depletion of Foxp3+ T reg cells by DT injection. Ablation of Foxp3+ T reg cells in newborn DEREG mice led to the development of scurfy-like symptoms with splenomegaly, lymphadenopathy, insulitis, and severe skin inflammation. Thus, these data provide experimental evidence that the absence of Foxp3+ T reg cells is indeed sufficient to induce a scurfy-like phenotype. Furthermore, DEREG mice will allow a more precise definition of the function of Foxp3+ T reg cells in immune reactions in vivo.

scholarly journals Three-Dimensional in Vivo Imaging of Green Fluorescent Protein-Expressing T Cells in Mice with Noncontact Fluorescence Molecular Tomography

2007 ◽  
Vol 6 (2) ◽  
pp. 7290.2007.00007 ◽  
Author(s):  
Anikitos Garofalakis ◽  
Giannis Zacharakis ◽  
Heiko Meyer ◽  
Eleftherios N. Economou ◽  
Clio Mamalaki ◽  
...  
Keyword(s):  
T Cells ◽  
Green Fluorescent Protein ◽  
In Vivo Imaging ◽  
Fluorescent Protein ◽  
Three Dimensional ◽  
Fluorescence Molecular Tomography ◽  
Green Fluorescent

scholarly journals The antigen-presenting activities of Ia+ dendritic cells shift dynamically from lung to lymph node after an airway challenge with soluble antigen.

1995 ◽  
Vol 181 (4) ◽  
pp. 1275-1283 ◽  
Author(s):  
W Xia ◽  
C E Pinto ◽  
R L Kradin
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
Lymph Node ◽  
Mononuclear Cells ◽  
Antigen Expression ◽  
Soluble Antigen ◽  
Regional Lymph Nodes ◽  
Antigen Presenting

Dendritic cells (DC) are widely distributed in the lung where they are distinguished by their morphology and class II major histocompatibility complex (Ia) antigen expression. Although a role for DC as pulmonary antigen-presenting cell (APC) has been suggested, little is currently known concerning how these cells respond to inhaled antigens in vivo. Hen-egg lysozyme (HEL) was injected intratracheally into Lewis rats; DC were subsequently purified from the lung and regional lymph nodes (LN) at intervals of up to 14 d and examined for their ability to stimulate the proliferation of HEL-immune T cells in vitro in the absence of added HEL. Pulmonary DC displayed APC activities at 3 h and for up to 7 d after the injection of antigen. Dendritic cells in the draining hilar LN showed APC activities that appeared at 24 h, peaked at day 3, and then diminished progressively. After the primary sensitization, HEL-immune T cells were detected in hilar LN but not in the lung. A second airway challenge with HEL at day 14 yielded an antigen-specific pulmonary immune response, characterized histologically by the accumulation of mononuclear cells around lung venules. We conclude that APC activities shift from lung to lymph node during the response to inhaled antigen.

scholarly journals Dynamic Interactions of Macrophages with T Cells during Antigen Presentation

1999 ◽  
Vol 190 (12) ◽  
pp. 1909-1914 ◽  
Author(s):  
David M. Underhill ◽  
Michael Bassetti ◽  
Alexander Rudensky ◽  
Alan Aderem
Keyword(s):  
T Cells ◽  
Antigen Presentation ◽  
Green Fluorescent Protein Expression ◽  
Fluorescent Protein ◽  
Nuclear Factor ◽  
Dynamic Interactions ◽  
Activated T Cells ◽  
Green Fluorescent ◽  
Antigen Presenting ◽  
Multiple Interactions

We have established a method for real-time video analysis of the interaction of antigen-presenting cells (APCs) with T cells. Green fluorescent protein expression controlled by a nuclear factor of activated T cells (NFAT)-responsive promoter permits the visualization of productive antigen presentation in single T cells. The readout is rapid (within 2 h) and semiquantitative and allows analysis by video microscopy and flow cytometry. Using this approach, we demonstrate that macrophages have the capacity to simultaneously activate multiple T cells. In addition, the interaction of T cells with macrophages is extraordinarily dynamic: after initial stable contact, the T cells migrate continuously on the surface of the macrophage and from APC to APC during productive antigen presentation. Thus, T cells sum up signals from multiple interactions with macrophages during stimulation.

scholarly journals Impairment of antigen-presenting function of peripheral γδ T cells in patients with sepsis

2021 ◽  
Author(s):  
Xue-Wei Yang ◽  
Hong Li ◽  
Ting Feng ◽  
Wei Zhang ◽  
Xiang-Rong Song ◽  
...  
Keyword(s):  
T Cells ◽  
Fluorescent Protein ◽  
Γδ T Cells ◽  
In Vitro Study ◽  
Healthy Individuals ◽  
Γδt Cells ◽  
Green Fluorescent ◽  
Antigen Presenting ◽  
Αβ T Cells

Abstract Impairment of antigen-presenting functions is a key mechanism contributing to sepsis-induced immunosuppression. Recently, γδ T cells have been demonstrated as professional antigen-presenting cells (APCs); however, their role in sepsis remains unknown. In this in vitro study, the APC function of human peripheral γδ T cells was assessed using samples collected from 42 patients with sepsis and 27 age-matched healthy controls. The APC-related markers HLA-DR, CD27, CD80, and CCR7 on fresh γδ T cells were significantly higher in patients with sepsis compared with matched controls; however, they responded poorly to 4-hydroxy-3-methyl-2-butenyl pyrophosphate (HMBPP) stimulation, characterised by the deactivation of these APC markers and impaired proliferation. Furthermore, the adhesion function of γδ T cells, essential for antigen presentation, was greatly reduced in patients with sepsis; for instance, in co-cultures with green fluorescent protein-expressing Escherichia coli, HMBPP-activated γδT cells from healthy individuals adhered to E. coli efficiently, whereas no such phenomenon was observed with respect to γδT cells from patients with sepsis. In line with these results, in co-cultures with isolated CD4 + αβ T cells, HMBPP-activated γδT cells of healthy individuals promoted the efficient proliferation of CD4 + αβ T cells, whereas γδT cells from patients with sepsis did not do so. In conclusion, our findings show that the antigen-presenting function of γδ T cells is severely impaired in patients with sepsis and the mechanisms behind need further study.

scholarly journals Targeting Antigen Presenting   Cells to Treat Autoimmune  Inflammation

2021 ◽  
Author(s):  
◽  
Aras Toker
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Mhc Class Ii ◽  
T Cell Proliferation ◽  
Target Cells ◽  
Antigen Presenting

<p>Glatiramer acetate (GA) is approved for the treatment of relapsing-remitting multiple sclerosis (MS), and can suppress experimental autoimmune encephalomyelitis (EAE), a murine model of human MS. GA treatment is associated with the induction of anti-inflammatory TH2 responses and with the antigen specific expansion of regulatory T cells that counteract or inhibit pathogenic events in MS and EAE. These T cell mediated mechanisms of protection are considered to be a result of modulation of antigen presenting cells (APCs) by GA, rather than direct effects on T cells. However, it is unknown if GA preferentially targets a specific APC subset or can act through multiple APCs in vivo. In addition, GA-modulated innate cells may also exhibit direct antigen non-specific suppression of autoreactive cells. One objective of this study was to identify the in vivo target cell population of GA and to assess the potential of the target cells to antigen non-specifically suppress immune responses. Fluorophor-labelled GA bound to monocytes after intravenous injections, suggesting that monocytes may be the primary target of GA in vivo. In addition, intravenous GA treatment enhanced the intrinsic ability of monocytes to suppress T cell proliferation, both in vitro and in vivo. The findings of this study therefore suggest that GA-induced monocytes may contribute to GA therapy through direct mechanisms of antigen non-specific T cell immunosuppression. A further objective of this work was to investigate the potential of an in vivo drug targeting approach. This approach was hypothesised to increase the uptake of GA by the target cells and substantially improve GA treatment through antigen specific mechanisms such as induction of TH2 or regulatory T cells. Targeting antigens to professional APCs with an anti-MHC class II antibody resulted in significantly enhanced T cell proliferation in vitro. However, no EAE suppression occurred when GA was targeted to MHC class II in vivo. In addition, targeting GA specifically to monocytes also failed to suppress EAE. These findings suggest that GA treatment may selectively modulate monocytes to enhance their ability to inhibit autoreactive T cells, which could be part of the mechanism by which GA ameliorates MS. Targeting GA to a specific cell type may not be a powerful approach to improve treatment, because increased proliferation of GA specific T cells is not sufficient for disease suppression, and conjugation to antibodies may functionally reduce GA to a mere antigen devoid of immunomodulatory capacity.</p>

scholarly journals Basophils Initiate IL-4 Production during a Memory T-dependent Response

2004 ◽  
Vol 200 (7) ◽  
pp. 857-870 ◽  
Author(s):  
Marat V. Khodoun ◽  
Tatyana Orekhova ◽  
Crystal Potter ◽  
Suzanne Morris ◽  
Fred D. Finkelman
Keyword(s):  
T Cells ◽  
Cd4 T Cells ◽  
Fluorescent Protein ◽  
Memory T Cells ◽  
Cell Types ◽  
Primary Response ◽  
Capture Assay ◽  
Green Fluorescent ◽  
Memory Cd4 T Cells

Experiments were performed to characterize and identify the cellular sources of the secondary interleukin (IL)-4 response to a T cell–dependent antigen. Mice were primed by immunization with goat anti–mouse immunoglobulin (Ig)D antibody (GaMD), which stimulates naive CD4+ T cells to secrete IL-4 in 3–4 d. When challenged with goat serum 14 d after immunization, GaMD-primed mice generated an IL-4 response that exceeded the primary response by ∼100-fold, started in &lt;2 h, and lasted for 4 d. Studies with 4get mice, in which cells with an accessible Il4 gene express a green fluorescent protein (GFP), revealed CD4+ memory T cells, natural killer T cells, basophils, mast cells, and eosinophils as possible rapid producers of IL-4. GFP+CD4+ T cells and basophils expanded more in the spleen than the other cell types during the primary response to GaMD. Quantitation of in vivo IL-4 production by the in vivo cytokine capture assay after individual cell types were selectively stimulated or deleted demonstrated that basophils and memory CD4+ T cells account for most of the secondary IL-4 response, with basophils initiating that response through IgE/FcεRI-mediated signaling but secreting IL-4 for &lt;4 h and memory T cells secreting IL-4 within 4 h and continuing to secrete this cytokine for 4 d.

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