Immune-checkpoint Protein VISTA Critically Regulates the IL-23/IL-17 Inflammatory Axis

V-domain Immunoglobulin Suppressor of T cell Activation (VISTA) is an inhibitory immune-checkpoint molecule that suppresses CD4+ and CD8+ T cell activation when expressed on antigen-presenting cells. Vsir−/− mice developed loss of peripheral tolerance and multi-organ chronic inflammatory phenotypes. Vsir−/− CD4+ and CD8+ T cells were hyper-responsive towards self- and foreign antigens. Whether or not VISTA regulates innate immunity is unknown. Using a murine model of psoriasis induced by TLR7 agonist imiquimod (IMQ), we show that VISTA deficiency exacerbated psoriasiform inflammation. Enhanced TLR7 signaling in Vsir−/− dendritic cells (DCs) led to the hyper-activation of Erk1/2 and Jnk1/2, and augmented the production of IL-23. IL-23, in turn, promoted the expression of IL-17A in both TCRγδ+ T cells and CD4+ Th17 cells. Furthermore, VISTA regulates the peripheral homeostasis of CD27− γδ T cells and their activation upon TCR-mediated or cytokine-mediated stimulation. IL-17A-producing CD27− γδ T cells were expanded in the Vsir−/− mice and amplified the inflammatory cascade. In conclusion, this study has demonstrated that VISTA critically regulates the inflammatory responses mediated by DCs and IL-17-producing TCRγδ+ and CD4+ Th17 T cells following TLR7 stimulation. Our finding provides a rationale for therapeutically enhancing VISTA-mediated pathways to benefit the treatment of autoimmune and inflammatory disorders

Oral Selective TLR8 Agonist Selgantolimod Induces Multiple Immune Cell Responses in Humans

TLR8 agonists have the potential for use as immunomodulatory components in therapeutic modalities for viral infections such as chronic HBV (CHB) and HIV. In this study, using peripheral blood samples from a phase 1a clinical trial, we examined the acute effects of a single oral administration of a selective TLR8 agonist on immune cell phenotypes. Administration of the TLR8 agonist selgantolimod (SLGN) in healthy individuals resulted in alteration in frequencies of peripheral blood monocytes, pDCs, mDCs and MAIT cells. Frequencies of mDCs and lymphoid cells significantly reduced after 8 h of SLGN administration, whereas pDC frequencies significantly increased, with changes possibly reflecting migration of different cell types between peripheral and tissue compartments in response to the agonist. Myeloid cell activation was evident by an upregulated expression of co-stimulatory molecules CD40 and CD86 accompanied by the production of IL-6 and IL-18 from these cells. Concomitantly, there was induction of the early activation marker CD69 on innate and adaptive lymphoid cells, including MAIT and NK cell subsets. Further, these activated lymphoid cells had enhanced expression of the effector molecules granzyme B and perforin. Microarray analysis of isolated lymphocytes and monocytes from baseline and post-SLGN treatment revealed changes in expression of genes involved in cellular response to cytokine stimulus, innate immune response, myeloid cell differentiation and antigen receptor-mediated signaling pathway. In a preliminary analysis of samples from CHB patients treated with selgantolimod, activation of innate and adaptive lymphocytes was evident. In conclusion, this first in-human study shows that selgantolimod administration in humans results in activation of multiple immune cell responses with antiviral potential

Virus-Like Particles Activate Type I Interferon Pathways to Facilitate Post-Exposure Protection against Ebola Virus Infection

<div><p>Ebola virus (EBOV) causes a severe hemorrhagic disease with high fatality. Virus-like particles (VLPs) are a promising vaccine candidate against EBOV. We recently showed that VLPs protect mice from lethal EBOV infection when given before or after viral infection. To elucidate pathways through which VLPs confer post-exposure protection, we investigated the role of type I interferon (IFN) signaling. We found that VLPs lead to accelerated induction of IFN stimulated genes (ISGs) in liver and spleen of wild type mice, but not in <i>Ifnar^-/-</i> mice. Accordingly, EBOV infected <i>Ifnar^-/-</i> mice, unlike wild type mice succumbed to death even after VLP treatment. The ISGs induced in wild type mice included anti-viral proteins and negative feedback factors known to restrict viral replication and excessive inflammatory responses. Importantly, proinflammatory cytokine/chemokine expression was much higher in WT mice without VLPs than mice treated with VLPs. In EBOV infected <i>Ifnar^-/-</i> mice, however, uninhibited viral replication and elevated proinflammatory factor expression ensued, irrespective of VLP treatment, supporting the view that type I IFN signaling helps to limit viral replication and attenuate inflammatory responses. Further analyses showed that VLP protection requires the transcription factor, IRF8 known to amplify type I IFN signaling in dendritic cells and macrophages, the probable sites of initial EBOV infection. Together, this study indicates that VLPs afford post-exposure protection by promoting expeditious initiation of type I IFN signaling in the host.</p></div

VLP post-exposure protection requires the transcription factor IRF8.

<p>(A) <i>Irf8</i>^<i>+/+</i> (WT) and <i>Irf8</i>^<i>-/-</i> mice (n = 5/group) were infected with EBOV followed by VLP treatment as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118345#pone.0118345.g001" target="_blank">Fig. 1A</a>. Mortality is expressed as the percentage survival. (B) EBOV GP levels in liver were tested by qRT-PCR on day2. (C) Expression of indicated ISGs in <i>Irf8</i>^<i>+/+</i> and <i>Irf8</i>^<i>-/-</i> liver was tested by qRT-PCR on day 2. Data represent the mean of duplicate samples ± SEM and a representative of three independent experiments. *** indicates <i>P</i> ≤ 0.001, ** <i>P</i> ≤ 0.01 and *<i>P</i> ≤ 0.05.</p

A model for VLP mediated post-exposure protection against EBOV infection.

<p>Top: EBOV infection in macrophages and DCs inhibits induction of type I IFNs and delays induction of ISGs via the mechanism of anti-IFN antagonism. Impaired type I IFN signaling delays induction of antiviral ISGs, and hinders maturation of macrophages and DCs and development of antigen presentation activity. It also inhibits prompt induction of negative regulatory ISGs that leads to exaggerated inflammatory responses. As a result, adaptive immunity is not adequately induced. Bottom: VLPs trigger timely induction of type I IFNs and ISGs by overcoming EBOV’s anti-IFN antagonism. This prompts rapid onset of anti-viral activity, simultaneously limiting exaggerated inflammatory responses and allowing maturation of antigen presentation function, which results in robust adaptive immune responses.</p

VLPs attenuate proinflammatory cytokine induction in EBOV infected mice.

<p>RNA from liver of WT mice (n = 5/group) infected with EBOV with or without VLP treatment was tested for expression of proinflammatory cytokines (TNFα, IL-6 and IL-1β), chemokines (MCP-1, MIP-1α, MIP-1β and KC (CXCL1)) and Inflammasome gene NLRP3 on indicated days post-infection. Values represent the average of duplicate samples from three independent experiments ± SEM. *<i>p</i> ≤ 0.05, **<i>p</i> ≤ 0.01, ***<i>p</i> ≤ 0.001 indicates statistical significance.</p

Accelerated ISG induction in VLP treated mice.

<p>WT mice were injected with VLPs 24 h post-EBOV challenge (n = 5/group) and transcripts of indicated ISGs in liver (A) and spleen (B) were measured on days 0, 1.5 and 2 post-infection. Ifit1, Mx1, Oas1a and Stat1 represent ISGs with anti-viral activity, while Irgm1, Usp18, Trim21 and Trim30 are ISGs with negative regulatory activities. Values represent the mean of duplicate samples from three to five independent experiments ± SEM. *** denotes <i>P</i> ≤ 0.001, ** <i>P</i> ≤ 0.01 and *<i>P</i> ≤ 0.05.</p

<i>Ifnar</i>^<i>-/-</i> macrophages elicit exaggerated proinflammatory cytokine responses following LPS and IFNβ stimulation.

<p>Bone marrow-derived macrophages from <i>Ifnar</i>^<i>+/+</i> and <i>Ifnar</i>^<i>-/-</i> mice (1x10⁶ cells/well) were primed with IFNγ (100U/ml) followed by co-stimulation with LPS (200ng/ml) and IFNβ (100U/ml) for 3 h. Expression of indicated ISGs, proinflammatory cytokines and chemokines were assessed by qRT-PCR. Data represent the average of duplicate samples from three individual experiments ± SEM. Asterisks denote *<i>p</i> ≤ 0.05, **<i>p</i> ≤ 0.01.</p