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

This study focused on soluble (s)CD25-mediated regulation of IL-2 signaling in murine and human CD4 T cells. Recombinant sCD25 reversibly sequestered IL-2 to limit acute maximal proliferative responses while preserving IL-2 bioavailability to subsequently maintain low-zone IL-2 signaling during prolonged culture. By inhibiting IL-2 signaling during acute activation, sCD25 suppressed T-cell growth and inhibited IL-2-evoked transmembrane CD25 expression, thereby resulting in lower prevalence of CD25 T cells.

Hypoxia-inducible factor-1 drives divergent immunomodulatory functions in the pathogenesis of autoimmune diseases.

Hypoxia-inducible factor-1 (HIF-1) is a heterodimeric (HIF-1α/ HIF-1β) transcription factor in which the oxygen-sensitive HIF-1α subunit regulates gene transcription to mediate adaptive tissue responses to hypoxia. HIF-1 is a key mediator in both regulatory and pathogenic immune responses, because ongoing inflammation in localized tissues causes increased oxygen consumption and consequent hypoxia within the inflammatory lesions. In autoimmune diseases, HIF-1 plays complex and divergent roles within localized inflammatory lesions by orchestrating a critical immune interplay sponsoring the pathogenesis of the disease.

Low-Zone IL-2 Signaling: Fusion Proteins Containing Linked CD25 and IL-2 Domains Sustain Tolerogenic Vaccination and Promote Dominance of FOXP3 Tregs .

Low-zone IL-2 signaling is key to understanding how CD4 CD25 FOXP3 regulatory T cells (Tregs) exhibit dominance and overgrow conventional effector T cells (Tcons) that typically express lower levels of the IL-2 receptor alpha chain (i.e., CD25).

Tolerogenic vaccines: Targeting the antigenic and cytokine niches of FOXP3 regulatory T cells.

FOXP3 regulatory T cells (Tregs) constitute a critical barrier that enforces tolerance to both the self-peptidome and the extended-self peptidome to ensure tissue-specific resistance to autoimmune, allergic, and other inflammatory disorders. Here, we review intuitive models regarding how T cell antigen receptor (TCR) specificity and antigen recognition efficiency shape the Treg and conventional T cell (Tcon) repertoires to adaptively regulate T cell maintenance, tissue-residency, phenotypic stability, and immune function in peripheral tissues. Three zones of TCR recognition efficiency are considered, including Tcon recognition of specific low-efficiency self MHC-ligands, Treg recognition of intermediate-efficiency agonistic self MHC-ligands, and Tcon recognition of cross-reactive high-efficiency agonistic foreign MHC-ligands.

A GM-CSF-neuroantigen tolerogenic vaccine elicits inefficient antigen recognition events below the CD40L triggering threshold to expand CD4 CD25 FOXP3 Tregs that inhibit experimental autoimmune encephalomyelitis (EAE).

Background: Tolerogenic vaccines represent antigen-specific interventions designed to re-establish self-tolerance and thereby alleviate autoimmune diseases, which collectively comprise over 100 chronic inflammatory diseases afflicting more than 20 million Americans. Tolerogenic vaccines comprised of single-chain GM-CSF-neuroantigen (GMCSF-NAg) fusion proteins were shown in previous studies to prevent and reverse disease in multiple rodent models of experimental autoimmune encephalomyelitis (EAE) by a mechanism contingent upon the function of CD4 CD25 FOXP3 regulatory T cells (Tregs). GMCSF-NAg vaccines inhibited EAE in both quiescent and inflammatory environments in association with low-efficiency T cell receptor (TCR) signaling events that elicited clonal expansion of immunosuppressive Tregs.

A GMCSF-Neuroantigen Tolerogenic Vaccine Elicits Systemic Lymphocytosis of CD4 CD25 FOXP3 Regulatory T Cells in Myelin-Specific TCR Transgenic Mice Contingent Upon Low-Efficiency T Cell Antigen Receptor Recognition.

Previous studies showed that single-chain fusion proteins comprised of GM-CSF and major encephalitogenic peptides of myelin, when injected subcutaneously in saline, were potent tolerogenic vaccines that suppressed experimental autoimmune encephalomyelitis (EAE) in rats and mice. These tolerogenic vaccines exhibited dominant suppressive activity in inflammatory environments even when emulsified in Complete Freund's Adjuvant (CFA). The current study provides evidence that the mechanism of tolerance was dependent upon vaccine-induced regulatory CD25 T cells (Tregs), because treatment of mice with the Treg-depleting anti-CD25 mAb PC61 reversed tolerance.

Partial CD25 Antagonism Enables Dominance of Antigen-Inducible CD25 FOXP3 Regulatory T Cells As a Basis for a Regulatory T Cell-Based Adoptive Immunotherapy.

FOXP3 regulatory T cells (Tregs) represent a promising platform for effective adoptive immunotherapy of chronic inflammatory disease, including autoimmune diseases such as multiple sclerosis. Successful Treg immunotherapy however requires new technologies to enable long-term expansion of stable, antigen-specific FOXP3 Tregs in cell culture. Antigen-specific activation of naïve T cells in the presence of TGF-β elicits the initial differentiation of the FOXP3 lineage, but these Treg lines lack phenotypic stability and rapidly transition to a conventional T cell (Tcon) phenotype during propagation.

IFN-β Facilitates Neuroantigen-Dependent Induction of CD25+ FOXP3+ Regulatory T Cells That Suppress Experimental Autoimmune Encephalomyelitis.

This study introduces a flexible format for tolerogenic vaccination that incorporates IFN-β and neuroantigen (NAg) in the Alum adjuvant. Tolerogenic vaccination required all three components, IFN-β, NAg, and Alum, for inhibition of experimental autoimmune encephalomyelitis (EAE) and induction of tolerance. Vaccination with IFN-β + NAg in Alum ameliorated NAg-specific sensitization and inhibited EAE in C57BL/6 mice in pretreatment and therapeutic regimens.

Depletion of CD4+ CD25+ regulatory T cells confers susceptibility to experimental autoimmune encephalomyelitis (EAE) in GM-CSF-deficient Csf2-/- mice.

Previous studies established that GM-CSF-deficient (Csf2-deficient) mice exhibit profound resistance to experimental autoimmune encephalomyelitis. This study addressed whether the resistance of Csf2-deficient mice was a result of a requirement for GM-CSF in controlling the functional balance between effector and regulatory T cell subsets during experimental autoimmune encephalomyelitis. The main observation was that treatment with the anti-CD25 mAb PC61 rendered Csf2-deficient mice fully susceptible to severe, chronic experimental autoimmune encephalomyelitis, with disease incidences and severities equivalent to that of C57BL/6 mice.

The extracellular domain of myelin oligodendrocyte glycoprotein elicits atypical experimental autoimmune encephalomyelitis in rat and Macaque species.

Atypical models of experimental autoimmune encephalomyelitis (EAE) are advantageous in that the heterogeneity of clinical signs appears more reflective of those in multiple sclerosis (MS). Conversely, models of classical EAE feature stereotypic progression of an ascending flaccid paralysis that is not a characteristic of MS. The study of atypical EAE however has been limited due to the relative lack of suitable models that feature reliable disease incidence and severity, excepting mice deficient in gamma-interferon signaling pathways.

GM-CSF-neuroantigen fusion proteins reverse experimental autoimmune encephalomyelitis and mediate tolerogenic activity in adjuvant-primed environments: association with inflammation-dependent, inhibitory antigen presentation.

Single-chain fusion proteins comprised of GM-CSF and neuroantigen (NAg) are potent, NAg-specific inhibitors of experimental autoimmune encephalomyelitis (EAE). An important question was whether GMCSF-NAg tolerogenic vaccines retained inhibitory activity within inflammatory environments or were contingent upon steady-state conditions. GM-CSF fused to the myelin oligodendrocyte glycoprotein MOG35-55 peptide (GMCSF-MOG) reversed established paralytic disease in both passive and active models of EAE in C57BL/6 mice.

Tolerogenic vaccines for Multiple sclerosis.

Tolerogenic vaccines represent a new class of vaccine designed to re-establish immunological tolerance, restore immune homeostasis, and thereby reverse autoimmune disease. Tolerogenic vaccines induce long-term, antigen-specific, inhibitory memory that blocks pathogenic T cell responses via loss of effector T cells and gain of regulatory T cell function. Substantial advances have been realized in the generation of tolerogenic vaccines that inhibit experimental autoimmune encephalomyelitis in a preclinical setting, and these vaccines may be a prequel of the tolerogenic vaccines that may have therapeutic benefit in Multiple Sclerosis.

Cytokine-neuroantigen fusion proteins as a new class of tolerogenic, therapeutic vaccines for treatment of inflammatory demyelinating disease in rodent models of multiple sclerosis.

Myelin-specific induction of tolerance represents a promising means to modify the course of autoimmune inflammatory demyelinating diseases such as multiple sclerosis (MS). Our laboratory has focused on a novel preclinical strategy for the induction of tolerance to the major encephalitogenic epitopes of myelin that cause experimental autoimmune encephalomyelitis (EAE) in rats and mice. This novel approach is based on the use of cytokine-NAg (neuroantigen) fusion proteins comprised of the native cytokine fused either with or without a linker to a NAg domain.

Neuroantigen-specific, tolerogenic vaccines: GM-CSF is a fusion partner that facilitates tolerance rather than immunity to dominant self-epitopes of myelin in murine models of experimental autoimmune encephalomyelitis (EAE).

Background: Vaccination strategies that elicit antigen-specific tolerance are needed as therapies for autoimmune disease. This study focused on whether cytokine-neuroantigen (NAg) fusion proteins could inhibit disease in chronic murine models of experimental autoimmune encephalomyelitis (EAE) and thus serve as potential therapeutic modalities for multiple sclerosis.

Results: A fusion protein comprised of murine GM-CSF as the N-terminal domain and the encephalitogenic MOG35-55 peptide as the C-terminal domain was tested as a tolerogenic, therapeutic vaccine (TTV) in the C57BL/6 model of EAE.

Autoimmunity and asthma: The dirt on the hygiene hypothesis.

Self peptides shape T-cell development through selectional processes in the thymus and secondary lymphoid organs to promote a diverse and balanced repertoire of conventional and regulatory T cells. Foreign proteins and their derivative peptides permeate our mucosal tissues to constitute another diverse array of peptides that may specify and diversify the mucosal T-cell repertoire. Indeed, the distinction between self peptides and environmental foreign peptides may be academic if both are present constantly within the body.

Vaccinia virus decreases major histocompatibility complex (MHC) class II antigen presentation, T-cell priming, and peptide association with MHC class II.

Vaccinia virus (VACV) is the current live virus vaccine used to protect humans against smallpox and monkeypox, but its use is contraindicated in several populations because of its virulence. It is therefore important to elucidate the immune evasion mechanisms of VACV. We found that VACV infection of antigen-presenting cells (APCs) significantly decreased major histocompatibility complex (MHC) II antigen presentation and decreased synthesis of 13 chemokines and cytokines, suggesting a potent viral mechanism for immune evasion.

A GMCSF-neuroantigen fusion protein is a potent tolerogen in experimental autoimmune encephalomyelitis (EAE) that is associated with efficient targeting of neuroantigen to APC.

Cytokine-NAg fusion proteins represent an emerging platform for specific targeting of self-antigen to particular APC subsets as a means to achieve antigen-specific immunological tolerance. This study focused on cytokine-NAg fusion proteins that targeted NAg to myeloid APC. Fusion proteins contained GM-CSF or the soluble extracellular domain of M-CSF as the N-terminal domain and the encephalitogenic 69-87 peptide of MBP as the C-terminal domain.

Experimental autoimmune encephalomyelitis in Lewis rats: IFN-beta acts as a tolerogenic adjuvant for induction of neuroantigen-dependent tolerance.

Cytokine-Ag fusion proteins represent a novel approach for induction of Ag-specific tolerance and may constitute an efficient therapy for autoimmune disease. This study addressed whether a fusion protein containing rat IFN-beta and the encephalitogenic 73-87 determinant of myelin basic protein (i.e.

Experimental autoimmune encephalomyelitis in the rat.

There are several diverse rat models of experimental autoimmune encephalomyelitis (EAE) that can be used to investigate the pathogenesis and regulation of autoimmunity against CNS myelin. The disease course of these models ranges from an acute monophasic disease with limited demyelination to a chronic relapsing or chronic progressive course marked by severe demyelination. These models enable the study of encephalitogenic T cells and demyelinating antibody specific for major neuroantigens such as myelin basic protein (MBP), myelin oligodendrocyte glycoprotein (MOG), or proteolipid protein (PLP), among other important CNS autoantigens.

A fusion protein consisting of IL-16 and the encephalitogenic peptide of myelin basic protein constitutes an antigen-specific tolerogenic vaccine that inhibits experimental autoimmune encephalomyelitis.

To test a novel concept for the generation of tolerogenic vaccines, fusion proteins were constructed encompassing a tolerogenic or biasing cytokine and the major encephalitogenic peptide of guinea pig myelin basic protein (GPMBP; i.e., neuroantigen or NAg).

IL-2/neuroantigen fusion proteins as antigen-specific tolerogens in experimental autoimmune encephalomyelitis (EAE): correlation of T cell-mediated antigen presentation and tolerance induction.

The purpose of this study was to assess whether the Ag-targeting activity of cytokine/neuroantigen (NAg) fusion proteins may be associated with mechanisms of tolerance induction. To assess this question, we expressed fusion proteins comprised of a N-terminal cytokine domain and a C-terminal NAg domain. The cytokine domain comprised either rat IL-2 or IL-4, and the NAg domain comprised the dominant encephalitogenic determinant of the guinea pig myelin basic protein.

Cytokine-neuroantigen fusion proteins: new tools for modulation of myelin basic protein (MBP)-specific T cell responses in experimental autoimmune encephalomyelitis.

Fusion proteins incorporating anti-inflammatory cytokines and immunodominant self antigen as separate domains of a single protein may hold promise for development of antigen-specific tolerogenic vaccines. Proteins incorporating rat sequences of IL-1RA, IL-2, IL-4, IL-10, or IL-13 were expressed as fusion proteins containing the major encephalitogenic region of myelin basic protein (MBP). These fusion proteins were expressed via baculovirus (bv) expression systems and were shown to have cytokine-dependent and antigen-specific biological activity.

Activation-dependent phases of T cells distinguished by use of optical tweezers and near infrared Raman spectroscopy.

Near-infrared Raman spectroscopy may provide a highly sensitive, noninvasive means to identify activation status of leukocytes. The purpose of the current study was to establish Raman spectroscopic characteristics of T cell activation. Activation of the RsL.

MHC class II biosynthesis by activated rat CD4+ T cells: development of repression in vitro and modulation by APC-derived signals.

This study focused on synthesis of MHC class II glycoproteins (MHCII) by rat CD4(+) T-helper cells. During activation in Con A and IL-2, purified rat splenic CD4(+) T cells expressed abundant surface MHCII together with transcripts for I-A alpha/beta, invariant chain, and the type III and type IV MHC class II transactivator (CIITA). Activated thymic CD8(+)CD4(-) and CD8(+)CD4(+) T cells exhibited essentially the same phenotype.