Abstract Streptococcus pneumoniae is a major cause of pneumonia and a leading cause of death world-wide. Antibody-mediated immune responses can offer protection against repeated exposure to S. pneumoniae , yet vaccines only offer partial protection. Patients with Activated PI3Kδ Syndrome (APDS) are highly susceptible to S. pneumoniae . We generated a conditional knockin mouse model of this disease and identified a CD19 + B220 − B cell subset that is induced by PI3Kδ signaling, is resident in the lungs, and which promotes increased susceptibility to S. pneumoniae during the early phase of infection via an antibody-independent mechanism. We show that an inhaled PI3Kδ inhibitor improves survival rates following S. pneumoniae infection in wild-type mice and in mice with activated PI3Kδ. These results suggest that a subset of B cells in the lung can promote the severity of S. pneumoniae infection, representing a novel therapeutic target.
We have demonstrated augmentation of the CD3−CD56+ natural killer (NK) and CD8+CD56− T-cell–mediated tumor cell cytotoxicity by neem leaf glycoprotein (NLGP). These NK and T cells were isolated from the peripheral blood of head and neck squamous cell carcinoma patients with a state of immunosuppression. NLGP induces TCRαβ-associated cytotoxic T lymphocyte (CTL) reaction to kill oral cancer (KB) cells. This CTL reaction is assisted by NLGP-mediated up-regulation of CD28 on T cells and HLA-ABC, CD80/86 on monocytes. CTL-mediated killing of KB cells and NK-cell–mediated killing of K562 (erythroleukemic) cells are associated with activation of these cells by NLGP. This activation is evidenced by increased expression of early activation marker CD69 with altered expression of CD45RO/CD45RA. NLGP is a strong inducer of IFNγ from both T and NK cells; however, IFNγ regulates the T-cell–mediated cytotoxicity only without affecting NK-cell–mediated one. Reason of this differential regulation may lie within up-regulated expression of IFNγ-receptor on T-cell surface, not on NK cells. This NLGP-induced cytotoxicity is dependent on up-regulated perforin/granzyme B expression in killer cells, which is again IFNγ dependent in T cells and independent in NK cells. Although, FasL expression is increased by NLGP, it may not be truly linked with the cytotoxic functions, as brefeldin A could not block such NLGP-mediated cytotoxicity, like, concanamycin A, a perforin inhibitor. On the basis of these results, we conclude that NLGP might be effective to recover the suppressed cytotoxic functions of NK and T cells from head and neck squamous cell carcinoma patients.
Severe asthma (SA) is a significant problem both clinically and economically, given its poor response to corticosteroids (CS). We recently reported a complex type 1-dominated (IFN-γ-dominated) immune response in more than 50% of severe asthmatics despite high-dose CS treatment. Also, IFN-γ was found to be critical for increased airway hyperreactivity (AHR) in our model of SA. The transcription factor IRF5 expressed in M1 macrophages can induce a Th1/Th17 response in cocultured human T cells. Here we show markedly higher expression of IRF5 in bronchoalveolar lavage (BAL) cells of severe asthmatics as compared with that in cells from milder asthmatics or healthy controls. Using our SA mouse model, we demonstrate that lack of IRF5 in lymph node migratory DCs severely limits their ability to stimulate the generation of IFN-γ- and IL-17-producing CD4+ T cells and IRF5-/- mice subjected to the SA model displayed significantly lower IFN-γ and IL-17 responses, albeit showing a reciprocal increase in Th2 response. However, the absence of IRF5 rendered the mice responsive to CS with suppression of the heightened Th2 response. These data support the notion that IRF5 inhibition in combination with CS may be a viable approach to manage disease in a subset of severe asthmatics.
Abstract Dendritic cells (DCs) play a central role in regulating immune tolerance and activation but the molecular axis within DCs that dictates immune outcome is unknown. We investigated the role of Peroxisome proliferator-activated receptor-gamma (PPARγ) in DCs in establishment of immune tolerance in the airways using an experimental model. Using mice with selective deletion of PPARγ in CD11c+ cells (PPARγΔ) to ablate PPARγ expression in DCs, we show that DC-specific PPARγ expression exerts a dual role whose collective goal in response to inhaled antigen is to not only promote de novo Foxp3 expression in T cells but to actively dampen T effector cell development. We demonstrate that PPARγ plays a crucial role in enhancing aldh1a2 expression in lung CD103+ DCs under tolerogenic conditions and when tolerized PPARγΔ mice were antigen-challenged, a complete loss of immune tolerance with increased neutrophil-dominated airway inflammation was observed. Absence of PPARγ promoted the expression of multiple pro-inflammatory cytokines such as IL-6 and IL-23 in DCs, limited Foxp3 induction but augmented IL-17 production in T cells with significant increase in the frequency of dual Foxp3+RORγt+ CD4+ T cells in PPARγΔ mice. In summary, our study identifies PPARγ as a central regulator in DC-mediated programming of the Foxp3/RORγt balance in CD4+ T cells dictating tolerance versus inflammation upon antigen provocation.
We have studied the immunomodulatory effects of interferon-α2b (IFN-α2b) in rectification of the dysregulated IFN-γ-dependent chemokines and their receptor CXCR3 splice variants in head and neck squamous cell carcinoma–peripheral blood mononuclear cells (HNSCC-PBMC). CXCR3 expression was upregulated in HNSCC-PBMC, with the demonstration of poor chemotactic function. CXCR3 upregulation possibly represents the increased synthesis of CXCR3B splice variant, without significant change in CXCR3A. Upregulated expression of CXCR3B was downregulated following in vitro IFN-α2b treatment of HNSCC-PBMC. Upregulation of CXCR3A+B by IFN-α2b with downregulation of the CXCR3B indirectly suggests that the upregulation of the CXCR3A splice variant induces cellular migration. On the other hand, the stimulation of PBMC with IFN-α2b maintains physiological homeostasis of CXCR3 ligands, CXCL10 and CXCL9, and increases the secretion of IFN-γ. The suppressed chemotactic ability of HNSCC-PBMC could be restored either by in vitro treatment of PBMC with IFN-α2b or during the use of IFN-α2b stimulated PBMC supernatant as a chemoattractant. Chemoattraction process is guided at the level of both receptor and its ligands, as confirmed by neutralization studies. IFN-α2b possibly controls chemotaxis by regulating the interaction between CXCL10 and CXCR3A. Neutralization of IFN-γ downregulates the IFN-α2b mediated CXCL10 release, suggesting the active role of IFN-γ in the transduction of chemotactic signal for the migration of cytotoxic T/NK cells at the tumor site.
ABSTRACT PD-1 acts as a negative regulator of T cell-mediated immune responses in the setting of persistent antigen expression, including cancer and chronic pathogen infections. Antibody-mediated blockade of the PD-1/PD-L1 axis benefits a subset of patients with highly immunogenic malignancies; however, many patients fail to respond due to a requirement for expression of the cell surface ligand PD-L1 within the tumor microenvironment. CISH is a member of a new class of intra-cellular immune checkpoint molecules that function downstream of the T cell receptor to regulate antigen-specific effector functions, including reactivity to cancer neoantigens. Herein, we employed multiplex CRISPR editing of primary human T cells to systematically compare the function of CISH deletion relative to PDCD1 (the gene encoding PD-1) and/or VSIG9 (the gene encoding TIGIT) in a model of neoantigen-mediated cancer cell cytolysis. PD-1 and TIGIT disruption enhanced cytolytic activity exclusively in the setting of high PD-L1 expression. In contrast, CISH inactivation enhanced antigen-specific cytolysis of tumor cells regardless of PD-L1 expression, including outperforming PD-1 and TIGIT disruption even in the presence of high PD-L1 tumor cells. Furthermore, we observed a synergistic increase in tumor cell killing when CISH and PD-1 or TIGIT are inactivated in combination, supporting the notion that these immune checkpoints regulate non-redundant pathways of T cell activation. Together, these data demonstrate that the intra-cellular immune checkpoint protein CISH can potentially enhance anti-tumor responses against a broad range of cancer types regardless of PD-L1 biomarker status.
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