Search

Clostridioides difficile infection (CDI) is the leading cause of antibiotic-associated diarrhea, and its clinical symptoms can span from asymptomatic colonization to pseudomembranous colitis and even death. The current standard of care for CDI is antibiotic treatment to achieve bacterial clearance; however, 15 to 35% of patients experience recurrence after initial response to antibiotics. We have conducted a comprehensive, global colonic transcriptomics analysis of a 10-day study in mice to provide new insights on the local host response during CDI and identify novel host metabolic mechanisms with therapeutic potential. The analysis indicates major alterations of colonic gene expression kinetics at the acute infection stage, that are restored during the recovery phase. At the metabolic level, we observe a biphasic response pattern characterized by upregulated glycolytic metabolism during the peak of inflammation, while mitochondrial metabolism predominates during the recovery/healing stage. Inhibition of glycolysis via 2-Deoxy-d-glucose (2-DG) administration during CDI decreases disease severity, protects from mortality, and ameliorates colitis in vivo. Additionally, 2-DG also protects intestinal epithelial cells from C. difficile toxin damage, preventing loss of barrier integrity and secretion of proinflammatory mediators. These data postulate the pharmacological targeting of host immunometabolic pathways as novel treatment modalities for CDI. [ABSTRACT FROM AUTHOR]

Clostridioides difficile infection (CDI) is the leading cause of antibiotic-associated diarrhea, and its clinical symptoms can span from asymptomatic colonization to pseudomembranous colitis and even death. The current standard of care for CDI is antibiotic treatment to achieve bacterial clearance; however, 15 to 35% of patients experience recurrence after initial response to antibiotics. We have conducted a comprehensive, global colonic transcriptomics analysis of a 10-day study in mice to provide new insights on the local host response during CDI and identify novel host metabolic mechanisms with therapeutic potential. The analysis indicates major alterations of colonic gene expression kinetics at the acute infection stage, that are restored during the recovery phase. At the metabolic level, we observe a biphasic response pattern characterized by upregulated glycolytic metabolism during the peak of inflammation, while mitochondrial metabolism predominates during the recovery/healing stage. Inhibition of glycolysis via 2-Deoxy-d-glucose (2-DG) administration during CDI decreases disease severity, protects from mortality, and ameliorates colitis in vivo. Additionally, 2-DG also protects intestinal epithelial cells from C. difficile toxin damage, preventing loss of barrier integrity and secretion of proinflammatory mediators. These data postulate the pharmacological targeting of host immunometabolic pathways as novel treatment modalities for CDI. [ABSTRACT FROM AUTHOR]

Helicobacter pylori is a gram-negative bacterium that persistently colonizes the human stomach by inducing immunoregulatory responses. We have used a novel platform that integrates a bone marrow-derived macrophage and live H. pylori co-culture with global time-course transcriptomics analysis to identify new regulatory genes based on expression patterns resembling those of genes with known regulatory function. We have used filtering criteria based on cellular location and novelty parameters to select 5 top lead candidate targets. Of these, Plexin domain containing 2 (Plxdc2) was selected as the top lead immunoregulatory target. Loss of function studies with in vivo models of H. pylori infection as well as a chemically-induced model of colitis, confirmed its predicted regulatory function and significant impact on modulation of the host immune response. Our integrated bioinformatics analyses and experimental validation platform has enabled the discovery of new immunoregulatory genes. This pipeline can be used for the identification of genes with therapeutic applications for treating infectious, inflammatory, and autoimmune diseases. [ABSTRACT FROM AUTHOR]

Background Helicobacter pylori causes gastric cancer in 1–2% of cases but is also beneficial for protection against allergies and gastroesophageal diseases. An estimated 85% of H. pylori –colonized individuals experience no detrimental effects. To study the mechanisms promoting host tolerance to the bacterium in the gastrointestinal mucosa and systemic regulatory effects, we investigated the dynamics of immunoregulatory mechanisms triggered by H. pylori using a high-performance computing–driven ENteric Immunity SImulator multiscale model. Immune responses were simulated by integrating an agent-based model, ordinary, and partial differential equations. Results The outputs were analyzed using 2 sequential stages: the first used a partial rank correlation coefficient regression–based and the second a metamodel-based global sensitivity analysis. The influential parameters screened from the first stage were selected to be varied for the second stage. The outputs from both stages were combined as a training dataset to build a spatiotemporal metamodel. The Sobol indices measured time-varying impact of input parameters during initiation, peak, and chronic phases of infection. The study identified epithelial cell proliferation and epithelial cell death as key parameters that control infection outcomes. In silico validation showed that colonization with H. pylori decreased with a decrease in epithelial cell proliferation, which was linked to regulatory macrophages and tolerogenic dendritic cells. Conclusions The hybrid model of H. pylori infection identified epithelial cell proliferation as a key factor for successful colonization of the gastric niche and highlighted the role of tolerogenic dendritic cells and regulatory macrophages in modulating the host responses and shaping infection outcomes. [ABSTRACT FROM AUTHOR]

Helicobacter pylori, the dominant member of the human gastric microbiota, elicits immunoregulatory responses implicated in protective versus pathological outcomes. To evaluate the role of macrophages during infection, we employed a system with a shifted proinflammatory macrophage phenotype by deleting PPARγ in myeloid cells and found a 5- to 10-fold decrease in gastric bacterial loads. Higher levels of colonization in wild-type mice were associated with increased presence of mononuclear phagocytes and in particular with the accumulation of CD11b+F4/80hiCD64+CX3CR1+ macrophages in the gastric lamina propria. Depletion of phagocytic cells by clodronate liposomes in wild-type mice resulted in a reduction of gastric H. pylori colonization compared with nontreated mice. PPARγ-deficient and macrophage-depleted mice presented decreased IL-10-mediated myeloid and T cell regulatory responses soon after infection. IL-10 neutralization during H. pylori infection led to increased IL-17-mediated responses and increased neutrophil accumulation at the gastric mucosa. In conclusion, we report the induction of IL-10-driven regulatory responses mediated by CD11b+F4/80hiCD64+CX3CR1+ mononuclear phagocytes that contribute to maintaining high levels of H. pylori loads in the stomach by modulating effector T cell responses at the gastric mucosa. [ABSTRACT FROM AUTHOR]

Immune responses to Helicobacter pylori are orchestrated through complex balances of host-bacterial interactions, including inflammatory and regulatory immune responses across scales that can lead to the development of the gastric disease or the promotion of beneficial systemic effects. While inflammation in response to the bacterium has been reasonably characterized, the regulatory pathways that contribute to preventing inflammatory events during H. pylori infection are incompletely understood. To aid in this effort, we have generated a computational model incorporating recent developments in the understanding of H. pylori-host interactions. Sensitivity analysis of this model reveals that a regulatory macrophage population is critical in maintaining high H. pylori colonization without the generation of an inflammatory response. To address how this myeloid cell subset arises, we developed a second model describing an intracellular signaling network for the differentiation of macrophages. Modeling studies predicted that LANCL2 is a central regulator of inflammatory and effector pathways and its activation promotes regulatory responses characterized by IL-10 production while suppressing effector responses. The predicted impairment of regulatory macrophage differentiation by the loss of LANCL2 was simulated based on multiscale linkages between the tissue-level gastric mucosa and the intracellular models. The simulated deletion of LANCL2 resulted in a greater clearance of H. pylori, but also greater IFNγ responses and damage to the epithelium. The model predictions were validated within a mouse model of H. pylori colonization in wild-type (WT), LANCL2 whole body KO and myeloid-specific LANCL2-/- (LANCL2Myeloid) mice, which displayed similar decreases in H. pylori burden, CX3CR1+ IL-10-producing macrophages, and type 1 regulatory (Tr1) T cells. This study shows the importance of LANCL2 in the induction of regulatory responses in macrophages and T cells during H. pylori infection. [ABSTRACT FROM AUTHOR]

Helicobacter pylori colonizes half of the world’s population as the dominant member of the gastric microbiota resulting in a lifelong chronic infection. Host responses toward the bacterium can result in asymptomatic, pathogenic or even favorable health outcomes; however, mechanisms underlying the dual role of H. pylori as a commensal versus pathogenic organism are not well characterized. Recent evidence suggests mononuclear phagocytes are largely involved in shaping dominant immunity during infection mediating the balance between host tolerance and succumbing to overt disease. We combined computational modeling, bioinformatics and experimental validation in order to investigate interactions between macrophages and intracellular H. pylori. Global transcriptomic analysis on bone marrow-derived macrophages (BMDM) in a gentamycin protection assay at six time points unveiled the presence of three sequential host response waves: an early transient regulatory gene module followed by sustained and late effector responses. Kinetic behaviors of pattern recognition receptors (PRRs) are linked to differential expression of spatiotemporal response waves and function to induce effector immunity through extracellular and intracellular detection of H. pylori. We report that bacterial interaction with the host intracellular environment caused significant suppression of regulatory NLRC3 and NLRX1 in a pattern inverse to early regulatory responses. To further delineate complex immune responses and pathway crosstalk between effector and regulatory PRRs, we built a computational model calibrated using time-series RNAseq data. Our validated computational hypotheses are that: 1) NLRX1 expression regulates bacterial burden in macrophages; and 2) early host response cytokines down-regulate NLRX1 expression through a negative feedback circuit. This paper applies modeling approaches to characterize the regulatory role of NLRX1 in mechanisms of host tolerance employed by macrophages to respond to and/or to co-exist with intracellular H. pylori. [ABSTRACT FROM AUTHOR]

T helper (Th) cells play a major role in the immune response and pathology at the gastric mucosa during Helicobacter pylori infection. There is a limited mechanistic understanding regarding the contributions of CD4+ T cell subsets to gastritis development during H. pylori colonization. We used two computational approaches: ordinary differential equation (ODE)-based and agent-based modeling (ABM) to study the mechanisms underlying cellular immune responses to H. pylori and how CD4+ T cell subsets influenced initiation, progression and outcome of disease. To calibrate the model, in vivo experimentation was performed by infecting C57BL/6 mice intragastrically with H. pylori and assaying immune cell subsets in the stomach and gastric lymph nodes (GLN) on days 0, 7, 14, 30 and 60 post-infection. Our computational model reproduced the dynamics of effector and regulatory pathways in the gastric lamina propria (LP) in silico. Simulation results show the induction of a Th17 response and a dominant Th1 response, together with a regulatory response characterized by high levels of mucosal Treg) cells. We also investigated the potential role of peroxisome proliferator-activated receptor γ (PPARγ) activation on the modulation of host responses to H. pylori by using loss-of-function approaches. Specifically, in silico results showed a predominance of Th1 and Th17 cells in the stomach of the cell-specific PPARγ knockout system when compared to the wild-type simulation. Spatio-temporal, object-oriented ABM approaches suggested similar dynamics in induction of host responses showing analogous T cell distributions to ODE modeling and facilitated tracking lesion formation. In addition, sensitivity analysis predicted a crucial contribution of Th1 and Th17 effector responses as mediators of histopathological changes in the gastric mucosa during chronic stages of infection, which were experimentally validated in mice. These integrated immunoinformatics approaches characterized the induction of mucosal effector and regulatory pathways controlled by PPARγ during H. pylori infection affecting disease outcomes. [ABSTRACT FROM AUTHOR]

Background: Enteroaggregative Escherichia coli (EAEC) is recognized as an emerging cause of persistent diarrhea and enteric disease worldwide. Mucosal immunity towards EAEC infections is incompletely understood due in part to the lack of appropriate animal models. This study presents a new mouse model and investigates the role of peroxisome proliferator-activated receptor gamma (PPARγ) in the modulation of host responses to EAEC in nourished and malnourished mice. Methods/Principal Findings: Wild-type and T cell-specific PPARγ null C57BL/6 mice were fed protein-deficient diets at weaning and challenged with 5×109cfu EAEC strain JM221 to measure colonic gene expression and immune responses to EAEC. Antigen-specific responses to E. coli antigens were measured in nourished and malnourished mice following infection and demonstrated the immunosuppressive effects of malnutrition at the cellular level. At the molecular level, both pharmacological blockade and deletion of PPARγ in T cells resulted in upregulation of TGF-β, IL-6, IL-17 and anti-microbial peptides, enhanced Th17 responses, fewer colonic lesions, faster clearance of EAEC, and improved recovery. The beneficial effects of PPARγ blockade on weight loss and EAEC clearance were abrogated by neutralizing IL-17 in vivo. Conclusions: Our studies provide in vivo evidence supporting the beneficial role of mucosal innate and effector T cell responses on EAEC burden and suggest pharmacological blockade of PPARγ as a novel therapeutic intervention for EAEC infection. [ABSTRACT FROM AUTHOR]

Background: There is an inverse secular trend between the incidence of obesity and gastric colonization with Helicobacter pylori, a bacterium that can affect the secretion of gastric hormones that relate to energy homeostasis. H. pylori strains that carry the cag pathogenicity island (PAI) interact more intimately with gastric epithelial cells and trigger more extensive host responses than cag- strains. We hypothesized that gastric colonization with H. pylori strains differing in cag PAI status exert distinct effects on metabolic and inflammatory phenotypes. Methodology/Principal Findings: To test this hypothesis, we examined metabolic and inflammatory markers in db/db mice and mice with diet-induced obesity experimentally infected with isogenic forms of H. pylori strain 26695: the cag PAI wild-type and its cag PAI mutant strain 99-305. H. pylori colonization decreased fasting blood glucose levels, increased levels of leptin, improved glucose tolerance, and suppressed weight gain. A response found in both wild-type and mutant H. pylori strain-infected mice included decreased white adipose tissue macrophages (ATM) and increased adipose tissue regulatory T cells (Treg) cells. Gene expression analyses demonstrated upregulation of gastric PPAR γ-responsive genes (i.e., CD36 and FABP4) in H. pylori-infected mice. The loss of PPAR γ in immune and epithelial cells in mice impaired the ability of H. pylori to favorably modulate glucose homeostasis and ATM infiltration during high fat feeding. Conclusions/Significance: Gastric infection with some commensal strains of H. pylori ameliorates glucose homeostasis in mice through a PPAR γ-dependent mechanism and modulates macrophage and Treg cell infiltration into the abdominal white adipose tissue. [ABSTRACT FROM AUTHOR]

Background: Inflammatory bowel disease (IBD) increases the risk of colorectal cancer. Probiotic bacteria produce immunoregulatory metabolites in vitro such as conjugated linoleic acid (CLA), a polyunsaturated fatty acid with potent anticarcinogenic effects. This study aimed to investigate the cellular and molecular mechanisms underlying the efficacy of probiotic bacteria in mouse models of cancer. Methodology/Principal Findings: The immune modulatory mechanisms of VSL#3 probiotic bacteria and CLA were investigated in mouse models of inflammation-driven colorectal cancer. Colonic specimens were collected for histopathology, gene expression and flow cytometry analyses. Immune cell subsets in the mesenteric lymph nodes (MLN), spleen and colonic lamina propria lymphocytes (LPL) were phenotypically and functionally characterized. Mice treated with CLA or VSL#3 recovered faster from the acute inflammatory phase of disease and had lower disease severity in the chronic, tumor-bearing phase of disease. Adenoma and adenocarcinoma formation was also diminished by both treatments. VSL#3 increased the mRNA expression of TNF-a, angiostatin and PPAR c whereas CLA decreased COX-2 levels. Moreover, VSL#3-treated mice had increased IL-17 expression in MLN CD4+ T cells and accumulation of Treg LPL and memory CD4+ T cells. Conclusions/Significance: Both CLA and VSL#3 suppressed colon carcinogenesis, although VSL#3 showed greater anticarcinogenic and anti-inflammatory activities than CLA. Mechanistically, CLA modulated expression of COX-2 levels in the colonic mucosa, whereas VSL#3 targeted regulatory mucosal CD4+ T cell responses. [ABSTRACT FROM AUTHOR]

Background: Inflammatory bowel disease (IBD) therapies are modestly successful and associated with significant side effects. Thus, the investigation of novel approaches to prevent colitis is important. Probiotic bacteria can produce immunoregulatory metabolites in vitro such as conjugated linoleic acid (CLA), a polyunsaturated fatty acid with potent antiinflammatory effects. This study aimed to investigate the cellular and molecular mechanisms underlying the antiinflammatory efficacy of probiotic bacteria using a mouse model of colitis. Methodology/Principal Findings: The immune modulatory mechanisms of VSL#3 probiotic bacteria and CLA were investigated in a mouse model of DSS colitis. Colonic specimens were collected for histopathology, gene expression and flow cytometry analyses. Immune cell subsets in the mesenteric lymph nodes (MLN), spleen, blood and colonic lamina propria cells were phenotypically and functionally characterized. Fecal samples and colonic contents were collected to determine the effect of VSL#3 and CLA on gut microbial diversity and CLA production. CLA and VSL#3 treatment ameliorated colitis and decreased colonic bacterial diversity, a finding that correlated with decreased gut pathology. Colonic CLA concentrations were increased in response to probiotic bacterial treatment, but without systemic distribution in blood. VSL#3 and CLA decreased macrophage accumulation in the MLN of mice with DSS colitis. The loss of PPAR γ in myeloid cells abrogated the protective effect of probiotic bacteria and CLA in mice with DSS colitis. Conclusions/Significance: Probiotic bacteria modulate gut microbial diversity and favor local production of CLA in the colon that targets myeloid cell PPAR γ to suppress colitis. [ABSTRACT FROM AUTHOR]

Our goal in this study was to determine the potential for dietary fibers to prevent gut inflammation in IL-10 -deficient (IL-10-/-) mice. C57BL/6J wild-type (WT) mice (n = 90) and IL-10-/- mice (n = 185) were assigned to a control diet or diets supplemented with PROMITOR soluble corn fiber (SCFI, STA-LITE III polydextrose (PDX), Biogum (BG), PulIulan (Pl-20), PROMITOR resistant starch-75 (RS-75), SCF&BG, RS-75&BG, and inulin (4 g fiber/100 g diet). On d 47, spleen, mesenteric lymph nodes (MLN), duodenum, jejunum, ileum, and colon were macroscopically and histologically evaluated. The spleen and Peyer's patches IPP) were collected for isolating mononuclear cells and measuring the percentages of regulatory T cells (Treg) and cytokines produced by CD4+ T cells (i.e. IFNγ and IL-10). Dietary supplementation with RS-75, SCE, RS-75&BG, and inulin ameliorated disease activity on d 47. Dietary RS-75 and inulin supplementation decreased ileal and colonic inflammatory lesions. RS-75, SCF, and inulin decreased IFNγ production by effector CD4+ T cells from PP and RS-75 increased the IL-10-expressing cells in spleen of WT mice. Dietary SCF, PDX, BG, P1-20, and RS-75 upregulated colonic PPARγ expression in WT mice and SCF upregulated Supressor of cytokine signaling 3 in IL-10-/- mice. These data suggest that soluble fibers and resistant starch influence Treg cells, IFNγ, and colonic PPARy expression to suppress gut inflammation. [ABSTRACT FROM AUTHOR]

Abscisic acid (ABA) has shown efficacy in the treatment of diabetes and inflammation; however, its molecular targets and the mechanisms of action underlying its immunomodulatory effects remain unclear. This study investigates the role of peroxisome proliferator-activated receptor γ (PPAR γ) and lanthionine synthetase C-like 2 (LANCL2) as molecular targets for ABA. We demonstrate that ABA increases PPAR γ reporter activity in RAW 264.7 macrophages and increases ppar γ expression in vivo, although it does not bind to the ligand-binding domain of PPAR γ. LANCL2 knockdown studies provide evidence that ABA-mediated activation of macrophage PPAR γ is dependent on lancl2 expression. Consistent with the association of LANCL2 with G proteins, we provide evidence that ABA increases cAMP accumulation in immune cells. ABA suppresses LPS-induced prostaglandin E2 and MCP-1 production via a PPAR γ-dependent mechanism possibly involving activation of PPAR γ and suppression of NF-?B and nuclear factor of activated T cells. LPS challenge studies in PPAR γ-expressing and immune cell-specific PPAR γ null mice demonstrate that ABA down-regulates toll-like receptor 4 expression in macrophages and T cells in vivo through a PPAR γ-dependent mechanism. Global transcriptomic profiling and confirmatory quantitative RT-PCR suggest novel candidate targets and demonstrate that ABA treatment mitigates the effect of LPS on the expression of genes involved in inflammation, metabolism, and cell signaling, in part, through PPAR γ. In conclusion, ABA decreases LPS-mediated inflammation and regulates innate immune responses through a bifurcating pathway involving LANCL2 and an alternative, ligand-binding domain-independent mechanism of PPAR γ activation. [ABSTRACT FROM AUTHOR]

The prevalence of obesity and its associated comorbidities has grown to epidemic proportions in the US and worldwide. Thus, developing safe and effective therapeutic approaches against these widespread and debilitating diseases is important and timely. Activation of peroxisome proliferator-activated receptors (PPARs) α, γ, and δ through several classes of pharmaceuticals can prevent or treat a variety of metabolic and inflammatory diseases, including type II diabetes (T2D). Thus, PPARs represent important molecular targets for developing novel and better treatments for a wide range of debilitating and widespread obesity-related diseases and disorders. However, available PPAR γ agonistic drugs such as Avandia have significant adverse side effects, including weight gain, fluid retention, hepatotoxicity, and congestive heart failure. An alternative to synthetic agonists of PPAR y is the discovery and development of naturally occurring and safer nutraceuticals that may be dual or pan PPAR agonists. The purpose of this paper is to summarize the health effects of three plant-derived PPAR agonists: abscisic acid (ABA), punicic acid (PUA), and catalpic acid (CAA) in the prevention and treatment of chronic inflammatory and metabolic diseases and disorders. [ABSTRACT FROM AUTHOR]

Abstract: Inflammatory bowel disease (IBD) is an immunoinflammatory illness of the gut initiated by an immune response to bacteria in the microflora. The resulting immunopathogenesis leads to lesions in epithelial lining of the colon through which bacteria may infiltrate the tissue causing recurring bouts of diarrhea, rectal bleeding, and malnutrition. In healthy individuals such immunopathogenesis is avoided by the presence of regulatory cells that inhibit the inflammatory pathway. Highly relevant to the search for treatment strategies is the identification of components of the inflammatory pathway that allow regulatory mechanisms to be overridden and immunopathogenesis to proceed. In vitro techniques have identified cellular interactions involved in inflammation-regulation crosstalk. However, tracing immunological mechanisms discovered at the cellular level confidently back to an in vivo context of multiple, simultaneous interactions has met limited success. To explore the impact of specific interactions, we have constructed a system of 29 ordinary differential equations representing different phenotypes of T-cells, macrophages, dendritic cells, and epithelial cells as they move and interact with bacteria in the lumen, lamina propria, and lymphoid tissue of the colon. Simulations revealed the positive inflammatory feedback loop formed by inflammatory M1 macrophage activation of T-cells as a driving force underlying the immunopathology of IBD. Furthermore, strategies that remove M1 from the site of infection, by either (i) increasing its potential to switch to a regulatory M2 phenotype or (ii) increasing the rate of reversion (for M1 and M2 alike) to a resting state, cease immunopathogenesis even as bacteria are eliminated by other inflammatory cells. Based on these results, we identify macrophages and their mechanisms of plasticity as key targets for mucosal inflammation intervention strategies. In addition, we propose that the primary mechanism behind the association of mutation with IBD is its ability to mediate the M1 to M2 switch. [Copyright &y& Elsevier]

Virtual screening (VS) is a discovery technique to identify novel compounds with therapeutic and preventive efficacy against disease. Our current focus is on the in silico screening and discovery of novel peroxisome proliferator-activated receptor- gamma (PPARγ) agonists. It is well recognized that PPARγ agonists have therapeutic applications as insulin sensitizers in type 2 diabetes or as anti-inflammatories. VS is a cost- and time-effective means for identifying small molecules that have therapeutic potential. Our long-term goal is to devise computational approaches for testing the PPARγ-binding activity of extensive naturally occurring compound libraries prior to testing agonist activity using ligand-binding and reporter assays. This review summarizes the high potential for obtaining further fundamental understanding of PPARγ biology and development of novel therapies for treating chronic inflammatory diseases through evolution and implementation of computational screening processes for immunotherapeutics in conjunction with experimental methods for calibration and validation of results. [ABSTRACT FROM AUTHOR]

Abstract: Macrophage infiltration into adipose tissue is a hallmark of obesity. We recently reported two phenotypically distinct subsets of adipose tissue macrophages (ATM) based on the surface expression of the glycoprotein F4/80 and responsiveness to treatment with a peroxisome proliferator-activated receptor (PPAR) γ agonist. Hence, we hypothesized that F4/80hi and F4/80lo ATM differentially express PPAR γ. This study phenotypically and functionally characterizes F4/80hi and F4/80lo ATM subsets during obesity. Changes in gene expression were also examined on sorted F4/80lo and F4/80hi ATM by quantitative real-time RT-PCR. We show that while F4/80lo macrophages predominate in adipose tissue of lean mice, obesity causes accumulation of both F4/80lo and F4/80hi ATM. Moreover, accumulation of F4/80hi ATM in adipose tissue is associated with impaired glucose tolerance. Phenotypically, F4/80hi ATM express greater amounts of CD11c, MHC II, CD49b, and CX3CR1 and produce more TNF-α, MCP-1, and IL-10 than F4/80lo ATM. Gene expression analyses of the sorted populations revealed that only the F4/80lo population produced IL-4, whereas the F4/80hi ATM expressed greater amounts of PPAR γ, δ, CD36 and toll-like receptor-4. In addition, the deficiency of PPAR γ in immune cells favors expression of M1 and impairs M2 macrophage marker expression in adipose tissue. Thus, PPAR γ is differentially expressed in F4/80hi versus F4/80low ATM subsets and its deficiency favors a predominance of M1 markers in WAT. [Copyright &y& Elsevier]

The spirochaeteBrachyspira hyodysenteriaecauses swine dysentery, a severe colitis characterized by mucosal enlargement as a result of crypt elongation and epithelial necrosis. Most efforts to understand the pathogenesis of this disease have focused on the aetiological agent and its virulence factors. However, the host immune response has been considered an important factor in disease development. Previous research has shown thatB. hyodysenteriaeinduces systemic CD4+ andγδ T-cell responses after intramuscular immunization. Here, we have evaluated changes in the CD4+ andγδ T-cell composition and distribution the different compartments of the colonic mucosa of pigs challenged withB. hyodysenteriae. We report that, in infected pigs,γδ T cells were significantly depleted from the epithelial layer, although their numbers were maintained in the lamina propria. In addition, CD4+ T cells aggregated in clusters located in the lamina propria and submucosa.Ex vivoanalyses of CD4+ T-cell responses toB. hyodysenteriaeantigens correlated with the changes in the mucosal CD4+ T-cell distribution observed in infected pigs; CD4+ T cells recovered from peripheral blood and colonic lymph nodes of infected pigs proliferated toB. hyodysenteriaeantigens, whereas no differences were found in theγδ T-cell responses between challenged and control groups. In addition, colonic lymph node CD4+ T cells had a predominant memory/activated phenotype. These results indicate that infection withB. hyodysenteriaeinduces a mucosal CD4+ T-cell response and points to CD4+ T cells being important contributors to the immunopathogenesis of swine dysentery. [ABSTRACT FROM AUTHOR]

We investigated the cellular and molecular immunoregulatory actions of conjugated linoleic acid (CLA) of relevance to viral disease pathogenesis and antiviral responses. To test the hypothesis that CLA ameliorates viral disease, we developed a viral challenge model by infecting pigs with type-2 porcine circovirus (PCV2). After 42 d of dietary supplementation with either soybean oil (n = 16) or CLA (n = 16), half of the pigs in each group were challenged with PCV2. We examined the effect of CLA on the development of lesions (i.e., lymphoid depletion and pneumonia) and observed the kinetics of the immune responses against PCV2. The viral infection depleted immature B cells (IgM+SWC3+) and favored proapoptotic mRNA expression profiles [i.e., suppressed B-cell leukemia/lymphoma-xl (Bcl-xl) and stimulated Bcl-2 homologous antagonist/killer (Bak)] in the external inguinal lymph nodes. B-cell depletion was more accentuated in pigs fed the control diet, whereas interleukin (IL)-2 mRNA expression was downregulated. Histopathological examination of the lungs revealed that the interstitial pneumonia tended to be more severe in infected pigs fed the control diet, which were also affected by growth retardation. CD8+ T cells were the primary cellular targets of CLA action in peripheral blood (CD8+CD29low and CD8+CD45RC+) and thymus (CD8+ and CD4+CD8+). CLA interacted with PCV2 to increase the proliferation of CD8+ T cells and to suppress PCV2-specific interferon (IFN)-gamma production in CD4+ T cells. At the molecular level, these cellular immunoregulatory properties were associated with differential patterns of peroxisome proliferator-activated receptor (alpha and gamma) mRNA expression between diets in virally infected pigs. [ABSTRACT FROM AUTHOR]

αβ+ and γδ+ T cells have different mechanisms of epitope recognition and are stimulated by antigens of different chemical nature. An immunization model with antigens from the spirochete Brachyspira hyodysenteriae was used to examine the requirements for proliferation of circulating porcine CD4+ and γδ+ T cells in mixed lymphocyte cultures. CD4+ T cells only responded to stimulation with B. hyodysenteriae antigens, whereas γδ+ T cells proliferated when cultures were stimulated with either spirochetal antigens or interleukin-2 (IL-2). T cells that had proliferated expressed high levels of IL-2-receptor-α (IL-2Rα). Furthermore, neutralization of IL-2 at the beginning of the culture period was more efficient in blocking γδ+ than CD4+ T cell proliferation. Immunization induced interferon-γ (IFN-γ) production by CD4+ T cells, whereas only a small fraction of the antigen-stimulated γδ+ T cells produced this cytokine. Our results indicate that, under the same environmental conditions, CD4+ T cell functions are more tightly regulated when compared to γδ+ T cells. We conclude that these differences are due, in part, to the enhanced γδ+ T cell responsiveness to IL-2. [Copyright &y& Elsevier]

Focuses on a study which demonstrated that porcine CD8... lymphocytes are essential for the development of cell-mediated protection against intracellular pathogens and neoplasic cells. Materials and methods; Effects of dietary conjugated linoleic acid on CD8... cell subsets; Comparison of the influence of lipid nutrition on porcine cellular immunity with the effects induced by vaccination.

While the immunomodulatory effects of feeding either arachidonic acid (AA) or docosahexaenoic acid (DHA) separately have been previously investigated, little is known about their immunomodulatory efficacy when fed in combination as infant formula ingredients. This study investigates the ability of AA/DHA-enriched infant formula to modulate immune responses in the neonate in response to an inactivated influenza virus vaccine. Neonatal piglets (n=48) were weaned on day 2 of age and distributed into 16 blocks of 3 littermate piglets each. Within each block, piglets were randomly assigned to a control formula, AA/DHA-enriched formula (0.63% AA and 0.34% DHA) or sow's milk for 30 days. On d 9, 8 blocks of piglets were immunized with an inactivated influenza virus vaccine. On days 0, 9, 16, 23 and 30 post-weaning we measured influenza virus-specific proliferation. Dietary AA/DHA-supplementation and sow's milk modulated influenza virus-specific CD4+ and CD8+ T cell ex vivo lymphoproliferative responses on d 23 post-immunization in comparison to the , unsupplemented control formula; The immunomodulatory effects of AA/DHA-enriched formulas were consistent with upregulation of interleukin-10 (IL-10) in PBMC. Overall, the AA/DHA-enriched formula modulated antigen-specific T cell responses through an IL-10-dependent mechanism. [ABSTRACT FROM AUTHOR]

Conjugated linoleic acid (CLA) is a mixture of positional and geometric isomers of linoleic acid. This family of polyunsaturated fatty acids has drawn significant attention in the last three decades for its variety of biologically beneficial properties and health effects. CLA has been shown to exert various potent protective functions such as anti-inflammatory, anticarcinogenic, antiadipogenic, antidiabetic and antihypertensive properties in animal models of disease. Therefore, CLA represents a nutritional avenue to prevent lifestyle diseases or metabolic syndrome. Initially, the overall effects of CLA were thought to be the result of interactions between its two major isomers: cis-9, trans-11 and trans-10, cis-12. However, later evidence suggests that such physiological effects of CLA might be different between the isomers: t-10, c-12-CLA is thought to be anticarcinogenic, antiobesity and antidiabetic, whereas c-9, t-11-CLA is mainly anti-inflammatory. Although preclinical data support a benefit of CLA supplementation, human clinical findings have yet to show definitive evidence of a positive effect. The purpose of this review is to comprehensively summarize the mechanisms of action and anti-inflammatory properties of dietary CLA supplementation and evaluate the potential uses of CLA in human health and disease. [ABSTRACT FROM AUTHOR]

We built a computational model of complex mechanisms at the intersection of immunity and metabolism that regulate CD4+ T cell effector and regulatory functions by using coupled ordinary differential equations. The model provides an improved understanding of how CD4+ T cells are shaping the immune response during Clostridioides difficile infection (CDI), and how they may be targeted pharmacologically to produce a more robust regulatory (Treg) response, which is associated with improved disease outcomes during CDI and other diseases. LANCL2 activation during CDI decreased the effector response, increased regulatory response, and elicited metabolic changes that favored Treg. Interestingly, LANCL2 activation provided greater immune and metabolic modulation compared to the addition of exogenous IL-2. Additionally, we identified gluconeogenesis via PEPCK-M as potentially responsible for increased immunosuppressive behavior in Treg cells. The model can perturb immune signaling and metabolism within a CD4+ T cell and obtain clinically relevant outcomes that help identify novel drug targets for infectious, autoimmune, metabolic, and neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

Peroxisome proliferator-activated receptor-gamma (PPARγ) is a nuclear hormone receptor involved in regulating various metabolic and immune processes. The PPAR family of receptors possesses a large binding cavity that imparts promiscuity of ligand binding not common to other nuclear receptors. This feature increases the challenge of using computational methods to identify PPAR ligands that will dock favorably into a structural model. Utilizing both ligand- and structure-based pharmacophore methods, we sought to improve agonist prediction by grouping ligands according to pharmacophore features, and pairing models derived from these features with receptor structures for docking. For 22 of the 33 receptor structures evaluated we observed an increase in true positive rate (TPR) when screening was restricted to compounds sharing molecular features found in rosiglitazone. A combination of structure models used for docking resulted in a higher TPR (40 %) when compared to docking with a single structure model (<20 %). Prediction was also improved when specific protein-ligand interactions between the docked ligands and structure models were given greater weight than the calculated free energy of binding. A large-scale screen of compounds using a marketed drug database verified the predictive ability of the selected structure models. This study highlights the steps necessary to improve screening for PPARγ ligands using multiple structure models, ligand-based pharmacophore data, evaluation of protein-ligand interactions, and comparison of docking datasets. The unique combination of methods presented here holds potential for more efficient screening of compounds with unknown affinity for PPARγ that could serve as candidates for therapeutic development. [ABSTRACT FROM AUTHOR]

Peroxisome-proliferator activated receptor-γ (PPARγ) is a nuclear hormone receptor that forms a heterodimeric complex with retinoid X receptor-α (RXRα) to regulate transcription of genes involved in fatty acid storage and glucose metabolism. PPARγ is a target for pharmaceutical intervention in type 2 diabetes, and insight into interactions between PPARγ, RXRα, and DNA is of interest in understanding the function and regulation of this complex. Phosphorylation of PPARγ by cyclin-dependent kinase 5 (Cdk5) has been shown to dysregulate the expression of metabolic regulation genes, an effect that is counteracted by PPARγ ligands. We applied molecular dynamics (MD) simulations to study the relationship between the ligand-binding domains of PPARγ and RXRα with their respective DNA-binding domains. Our results reveal that phosphorylation alters collective motions within the PPARγ-RXRα complex that affect the LBD-LBD dimerization interface and the AF-2 coactivator binding region of PPARγ. [ABSTRACT FROM AUTHOR]

Nucleotide-binding oligomerization domain, leucine rich repeat containing X1 (NLRX1) is an emerging therapeutic target for a spectrum of human diseases. NX-13 is a small molecule therapeutic designed to target and activate NLRX1 to induce immunometabolic changes resulting in lower inflammation and therapeutic responses in inflammatory bowel disease (IBD). This study investigates the safety of NX-13 in a seven-day, repeat-dose general toxicity study in male and female Sprague Dawley rats at oral doses of 500 and 1000 mg/kg. Weights, clinical signs, functional observational battery, clinical pathology and histopathology were used for evaluation. Daily oral dosing of NX-13 up to 1000 mg/kg did not result in any changes in weight, abnormal clinical signs or behavior. No significant differences were observed between treated and control rats in hematology or blood biochemistry. Histopathological evaluation of 12 tissues demonstrated no differences between controls and treated rats. There were no changes in weights of brain, heart, kidney, liver or spleen. Pharmacokinetic analysis of a single oral dose of NX-13 at 10 mg/kg in Sprague Dawley rats provided a maximum plasma concentration of 57 ng/mL at 0.5 h post-dose. Analysis of colon tissue after oral dosing with 1 and 10 mg/kg indicated high peak concentrations (10 and 100 µg/g, respectively) that scale in a dose-proportional manner. These experiments suggest that NX-13 is safe and well-tolerated in rats given oral doses as high as 1000 mg/kg with a favorable gastrointestinal localized pharmacokinetic profile, confirming NX-13 as a promising therapeutic for Crohn's disease and ulcerative colitis. [ABSTRACT FROM AUTHOR]

Psoriasis (PsO) is a complex immune-mediated disease that afflicts 100 million people. Omilancor is a locally-acting, small molecule that selectively activates the Lanthionine Synthetase C-like 2 (LANCL2) pathway, resulting in immunoregulatory effects at the intersection of immunity and metabolism. Topical omilancor treatment in an imiquimod-induced mouse model of PsO ameliorates disease severity, epidermal hyperplasia and acanthosis. Further, pharmacological activation of LANCL2 results in significant downregulation of proinflammatory markers including local reduction of IL17, and infiltration of proinflammatory cell subsets. These therapeutic effects were further validated in an IL-23 PsO model. This model reported increased preservation of homeostatic skin structure, accompanied by a decreased infiltration of proinflammatory T cell subsets. In CD4+ T cells and Th17 cells, the LANCL2 pathway regulates proinflammatory cytokine production, proliferation and glucose metabolism. Metabolically, the loss of Lancl2 resulted in increased glycolytic rates, lactate production and upregulated enzymatic activity of hexokinase and lactate dehydrogenase (LDH). Inhibition of LDH activity abrogated the increased proliferation rate in Lancl2−/− CD4+ T cells. Additionally, topical omilancor treatment decreased the metabolic upregulation in keratinocytes, keratinocyte hyperproliferation and expression of inflammatory markers. Omilancor is a promising topical, LANCL2-targeting therapeutic candidate for the treatment of PsO and other dermatology indications. [ABSTRACT FROM AUTHOR]

Aspergillus fumigatus is an opportunistic fungal pathogen of immunocompromised patient populations. Mortality is thought to be context-specific and occurs via both enhanced fungal growth and immuno-pathogenesis. NLRX1 is a negative regulator of immune signaling and metabolic pathways implicated in host responses to microbes, cancers, and autoimmune diseases. Our study indicates loss of Nlrx1 results in enhanced fungal burden, pulmonary inflammation, immune cell recruitment, and mortality across immuno-suppressed and immuno-competent models of IPA using two clinically derived isolates (AF293, CEA10). We observed that the heightened mortality is due to enhanced recruitment of CD103+ dendritic cells (DCs) that produce elevated amounts of IL-4 resulting in a detrimental Th2-mediated immune response. Adoptive transfer of Nlrx1-/- CD103+ DCs in neutropenic NRG mice results in enhanced mortality that can be ablated using IL-4 neutralizing antibodies. In vitro analysis of CD103+ DCs indicates loss of Nlrx1 results in enhanced IL-4 production via elevated activation of the JNK/JunB pathways. Interestingly, loss of Nlrx1 also results in enhanced recruitment of monocytes and neutrophils. Chimeras of irradiated Nlrx1-/- mice reconstituted with wild type bone marrow have enhanced neutrophil recruitment and survival during models of IPA. This enhanced immune cell recruitment in the absence of Nlrx1 is mediated by excessive production of CXCL8/IL-8 family of chemokines and IL-6 via early and enhanced activation of P38 in response to A. fumigatus conidia as shown in BEAS-2B airway epithelial cells. In summary, our results point strongly towards the cell-specific and contextual function of Nlrx1 during invasive pulmonary aspergillosis and may lead to novel therapeutics to reduce Th2 responses by CD103+ DCs or heightened recruitment of neutrophils. Author summary: Fungal infections are mitigated and controlled in part by a robust immune response and generation of reactive oxygen species. In certain instances, the immune response may become harmful to the host. Nlrx1 is a known negative regulator of inflammatory aspects of the immune system in response to viruses, bacteria, and cancers. In this study we describe the novel importance of Nlrx1 in controlling and fighting fungal infections in two different host cell populations through two distinct mechanisms. Nlrx1 may function as a future target to mitigate inflammation and immunopathogenesis during fungal pulmonary infection as well as enhance beneficial neutrophil recruitment. [ABSTRACT FROM AUTHOR]

Abscisic acid is a phytohormone found in fruits and vegetables and is endogenously produced in mammals. In humans and mice, lanthionine synthetase C-like 2 (LANCL2) has been characterized as the natural receptor for ABA. Herein, we characterize the efficacy of a fig fruit extract of ABA in promoting glycemic control. This ABA-enriched extract, at 0.125 µg ABA/kg body weight, improves glucose tolerance, insulin sensitivity and fasting blood glucose in diet-induced obesity (DIO) and db/db mouse models. In addition to decreasing systemic inflammation and providing glycemic control without increasing insulin, ABA extract modulates the metabolic activity of muscle. ABA increases expression of important glycogen synthase, glucose, fatty acid and mitochondrial metabolism genes and increases direct measures of fatty acid oxidation, glucose oxidation and metabolic flexibility in soleus muscle cells from ABA-treated mice with DIO. Glycolytic and mitochondrial ATP production were increased in ABA-treated human myotubes. Further, ABA synergized with insulin to dramatically increase the rate of glycogen synthesis. The loss of LANCL2 in skeletal muscle abrogated the effect of ABA extract in the DIO model and increased fasting blood glucose levels. This data further supports the clinical development of ABA in the treatment of pre-diabetes, type 2 diabetes and metabolic syndrome. [ABSTRACT FROM AUTHOR]

Inflammatory bowel disease (IBD) is a complex autoimmune disease with dysfunction in pattern-recognition responses, including within the NLR family. Nucleotide-binding oligomerization domain, leucine rich repeat containing X1 (NLRX1) is a unique NLR with regulatory and anti-inflammatory functions resulting in protection from IBD in mouse models. NX-13 is an orally active, gut-restricted novel drug candidate that selectively targets and activates the NLRX1 pathway locally in the gut. In vitro and in vivo efficacy of NLRX1 activation by NX-13 was examined. Oral treatment with NX-13 alleviates disease severity, colonic leukocytic infiltration, and cytokine markers of inflammation in three mouse models of IBD (dextran sulfate sodium, Mdr1a-/-, and CD45RBhi adoptive transfer). Treatment of naive CD4+ T cells with NX-13 in vitro decreases differentiation into Th1 and Th17 subsets with increased oxidative phosphorylation and decreased NF-kB activation and reactive oxygen species. With stimulation by PMA/ionomycin, TNF-α, or H2O2, PBMCs from ulcerative colitis patients treated with NX-13 had decreased NF-kB activity, TNF-α+ and IFN-γ+ CD4+ T cells and overall production of IL-6, MCP1, and IL-8. NX-13 activates NLRX1 to mediate a resistance to both inflammatory signaling and oxidative stress in mouse models and human primary cells from ulcerative colitis patients with effects on NF-kB activity and oxidative phosphorylation. NX-13 is a promising oral, gut-restricted NLRX1 agonist for treating IBD. [ABSTRACT FROM AUTHOR]

Inflammatory bowel disease (IBD) is an expanding autoimmune disease afflicting millions that remains difficult to treat due to the accumulation of multiple immunological changes. BT-11 is an investigational new drug for IBD that is orally active, gut restricted, and targets the lanthionine synthetase C-like 2 immunometabolic pathway. CD25+ FOXP3+ CD4+ T cells are increased locally within the colon of BT-11-treated mice in Citrobacter rodentium and IL-10-/- mouse models of colitis. The maintained efficacy of BT-11 in the absence of IL-10 combined with the loss of efficacy when direct cell-cell interactions are prevented suggest that the regulatory T cell (Treg)-related elements of suppression are cell contact-mediated. When PD-1 is inhibited, both in vitro and in vivo, the efficacy of BT-11 is reduced, validating this assertion. The depletion of CD25+ cells in vivo abrogated the retention of therapeutic efficacy postdiscontinuation of treatment, indicating that Tregs are implicated in the maintenance of tolerance mediated by BT-11. Furthermore, the involvement of CD25 suggested a role of BT-11 in IL-2 signaling. Cotreatment with BT-11 and IL-2 greatly enhances the differentiation of CD25+ FOXP3+ cells from naive CD4+ T cells relative to either alone. BT-11 enhances phosphorylation of STAT5, providing a direct linkage to the regulation of FOXP3 transcription. Notably, when STAT5 is inhibited, the effects of BT-11 on the differentiation of Tregs are blocked. BT-11 effectively enhances the IL-2/STAT5 signaling axis to induce the differentiation and stability of CD25+ FOXP3+ cells in the gastrointestinal mucosa to support immunoregulation and immunological tolerance in IBD. [ABSTRACT FROM AUTHOR]

BT-11 is an orally active, gut-restricted investigational therapeutic targeting the lanthionine synthetase C-like 2 pathway with lead indications in ulcerative colitis (UC) and Crohn disease (CD), 2 manifestations of inflammatory bowel disease (IBD). In 5 mouse models of IBD, BT-11 is effective at oral doses of 8 mg/kg. BT-11 was also efficacious at nanomolar concentrations in primary human samples from patients with UC and CD. BT-11 was tested under Good Laboratory Practice conditions in 90-day repeat-dose general toxicity studies in rats and dogs, toxicokinetics, respiratory, cardiovascular and central nervous system safety pharmacology, and genotoxicity studies. Oral BT-11 did not cause any clinical signs of toxicity, biochemical or hematological changes, or macroscopic or microscopic changes to organs in 90-day repeat-dose toxicity studies in rats and dogs at doses up to 1,000 mg/kg/d. Oral BT-11 resulted in low systemic exposure in both rats (area under the curve exposure from t = 0 to t = 8 hours [AUC0-8] of 216 h × ng/mL) and dogs (650 h × ng/mL) and rapid clearance with an average half-life of 3 hours. BT-11 did not induce changes in respiratory function, electrocardiogram parameters, or behavior with single oral doses of 1,000 mg/kg/d. There was no evidence of mutagenic or genotoxic potential for BT-11 up to tested limit doses using an Ames test, chromosomal aberration assay in human peripheral blood lymphocytes, or micronucleus assay in rats. Therefore, nonclinical studies show BT-11 to be a safe and well-tolerated oral therapeutic with potential as a potent immunometabolic therapy for UC and CD with no-observed adverse effect level >1,000 mg/kg in in vivo studies. [ABSTRACT FROM AUTHOR]

The current treatment paradigm in Clostridium difficile infection is the administration of antibiotics contributing to the high rates of recurrent infections. Recent alternative strategies, such as fecal microbiome transplantation and anti-toxin antibodies, have shown similar efficacy in the treatment of C. difficile associated disease (CDAD). However, barriers exist for either treatment or other novel treatments to displace antibiotics as the standard of care. To aid in the comparison of these and future treatments in CDAD, we developed an in silico pipeline to predict clinical efficacy with nonclinical results. The pipeline combines an ordinary differential equation (ODE)-based model, describing the immunological and microbial interactions in the gastrointestinal (GI) mucosa, with machine learning algorithms to translate simulated output quantities (i.e. time of clearance, quantity of commensal bacteria, T cell ratios) into clinical predictions based on prior preclinical, translational and clinical trial data. As a use case, we compare the efficacy of lanthionine synthetase C-like 2 (LANCL2), a novel immunoregulatory target with promising efficacy in inflammatory bowel disease (IBD), activation with antibiotics, fecal microbiome transplantation and anti-toxin antibodies in the treatment of CDAD. We further validate the potential of LANCL2 pathway activation, in a mouse model of C. difficile infection in which it displays an ability to decrease weight loss and inflammatory cell types while protecting against mortality. The computational pipeline can serve as an important resource in the development of new treatment modalities. [ABSTRACT FROM AUTHOR]

Nucleotide oligomerization domain-like receptor X1 (NLRX1) has been implicated in viral response, cancer progression, and inflammatory disorders; however, its role as a dual modulator of CD4+ T cell function and metabolism has not been defined. The loss of NLRX1 results in increased disease severity, populations of Th1 and Th17 cells, and inflammatory markers (IFN-γ, TNF-α, and IL-17) in mice with dextran sodium sulfate-induced colitis. To further characterize this phenotype, we used in vitro CD4+ T cell-differentiation assays and show that NLRX1-deficient T cells have a greater ability to differentiate into an inflammatory phenotype and possess greater proliferation rates. Further, NLRX1-/- cells have a decreased responsiveness to immune checkpoint pathways and greater rates of lactate dehydrogenase activity. When metabolic effects of the knockout are impaired, NLRX1-deficient cells do not display significant differences in differentiation or proliferation. To confirm the role of NLRX1 specifically in T cells, we used an adoptive-transfer model of colitis. Rag2-/- mice receiving NLRX1-/- naive or effector T cells experienced increased disease activity and effector T cell populations, whereas no differences were observed between groups receiving wildtype or NLRX1-/- regulatory T cells. Metabolic effects of NLRX1 deficiency are observed in a CD4-specific knockout of NLRX1 within a Citrobacter rodentium model of colitis. The aerobic glycolytic preference in NLRX1-/- effector T cells is combined with a decreased sensitivity to immunosuppressive checkpoint pathways to provide greater proliferative capabilities and an inflammatory phenotype bias leading to increased disease severity. [ABSTRACT FROM AUTHOR]

Emergent responses of the immune system result from the integration of molecular and cellular networks over time and across multiple organs. High-content and high-throughput analysis technologies, concomitantly with data-driven and mechanistic modeling, hold promise for the systematic interrogation of these complex pathways. However, connecting genetic variation and molecular mechanisms to individual phenotypes and health outcomes has proven elusive. Gaps remain in data, and disagreements persist about the value of mechanistic modeling for immunology. Here, we present the perspectives that emerged from the National Institute of Allergy and Infectious Disease (NIAID) workshop ‘Complex Systems Science, Modeling and Immunity’ and subsequent discussions regarding the potential synergy of high-throughput data acquisition, data-driven modeling, and mechanistic modeling to define new mechanisms of immunological disease and to accelerate the translation of these insights into therapies. [ABSTRACT FROM AUTHOR]

Human immunodeficiency virus (HIV)-infected patients are at an increased risk of co-infection with human papilloma virus (HPV), and subsequent malignancies such as oral cancer. To determine the role of HIV-associated immune suppression on HPV persistence and pathogenesis, and to investigate the mechanisms underlying the modulation of HPV infection and oral cancer by HIV, we developed a mathematical model of HIV/HPV co-infection. Our model captures known immunological and molecular features such as impaired HPV-specific effector T helper 1 (Th1) cell responses, and enhanced HPV infection due to HIV. We used the model to determine HPV prognosis in the presence of HIV infection, and identified conditions under which HIV infection alters HPV persistence in the oral mucosa system. The model predicts that conditions leading to HPV persistence during HIV/HPV co-infection are the permissive immune environment created by HIV and molecular interactions between the two viruses. The model also determines when HPV infection continues to persist in the short run in a co-infected patient undergoing antiretroviral therapy. Lastly, the model predicts that, under efficacious antiretroviral treatment, HPV infections will decrease in the long run due to the restoration of CD4+ T cell numbers and protective immune responses. [ABSTRACT FROM AUTHOR]

Lanthionine synthetase C-like 2 (LANCL2), a novel therapeutic target for inflammatory and autoimmune diseases and diabetes, exerts anti-inflammatory and insulin-sensitizing effects. This study reports the first LANCL2-based therapeutics for inflammatory bowel disease (IBD). Analogues of 1 (ABA) and 2 (NSC61610) were screened by molecular docking, then synthesized and analyzed for binding to LANCL2 by surface plasmon resonance. Piperazine-1,4-diylbis(6-benzo[d]imidazole-2-yl)pyridine-2-yl)methanone, 7, was identified as the lead LANCL2-binding compound for treating IBD. The oral treatment with 7 (8 mg/kg/d) in a mouse model of IBD resulted in lowering the disease activity index, decreasing colonic inflammatory lesions by 4-fold, and suppressing inflammatory markers (e.g., TNF-α, and interferon-γ) in the gut. Furthermore, studies in LANCL2-/- mice demonstrated that loss of LANCL2 abrogated beneficial actions of 7, suggesting high selectivity for the target. In conclusion, 7 merits continued development as a LANCL2-based, first-in-class orally active therapeutic for IBD. [ABSTRACT FROM AUTHOR]

Lanthionine synthetase cyclase-like receptor 2 (LANCL2) is a novel therapeutic target for Crohn’s disease (CD). BT-11 is a small molecule that binds LANCL2, is orally active, and has demonstrated therapeutic efficacy in 3 validated mouse models of colitis at doses as low as 8 mg/kg/d. Exploratory experiments evaluated BT-11 in male Harlan Sprague Dawley rats with a single oral dose of 500 mg/kg and 80 mg/kg/d for 14 days (n = 10 rats dosed/group). Treated and control rats were observed for behavioral detriments, and blood and tissues were collected for clinical pathology and histopathological examination. A functional observational battery demonstrated no differences between treated and control groups over multiple times of observation for quantal, categorical, and continuous end points, including posture, in cage activity, approach, response to touch, weight, grip strength, body temperature, and time on a rotarod. Histopathological examination of the brain, kidney, liver, adrenal gland, testes, stomach, small and large intestines, duodenum, pancreas, heart, lungs, spleen, thymus, and rib found no significant differences between the groups. Plasma enzymes associated with liver function were transiently elevated 2 to 4 days after the 500 mg/kg single dose but returned to normal values by 8 days and were not observed at any time in rats given 80 mg/kg/d for 14 days. One hour after oral administration of a single dose of 80 mg/kg, BT-11 had a maximal concentration of 21 ng/mL; the half-life was 3 hours. These experimental results demonstrated that BT-11 is well tolerated in rats, and, with further testing, may hold promise as an orally active therapeutic for CD. [ABSTRACT FROM AUTHOR]

T follicular helper (Tfh) cells are a highly plastic subset of CD4+ T cells specialized in providing B cell help and promoting inflammatory and effector responses during infectious and immune-mediate diseases. Helicobacter pylori is the dominant member of the gastric microbiota and exerts both beneficial and harmful effects on the host. Chronic inflammation in the context of H. pylori has been linked to an upregulation in T helper (Th)1 and Th17 CD4+ T cell phenotypes, controlled in part by the cytokine, interleukin-21. This study investigates the differentiation and regulation of Tfh cells, major producers of IL-21, in the immune response to H. pylori challenge. To better understand the conditions influencing the promotion and inhibition of a chronically elevated Tfh population, we used top-down and bottom-up approaches to develop computational models of Tfh and T follicular regulatory (Tfr) cell differentiation. Stability analysis was used to characterize the presence of two bi-stable steady states in the calibrated Tfh/Tfr models. Stochastic simulation was used to illustrate the ability of the parameter set to dictate two distinct behavioral patterns. Furthermore, sensitivity analysis helped identify the importance of various parameters on the establishment of Tfh and Tfr cell populations. The core network model was expanded into a more comprehensive and predictive model by including cytokine production and signaling pathways. From the expanded network, the interaction between TGFB-Induced Factor Homeobox 1 (Tgif1) and the retinoid X receptor (RXR) was displayed to exert control over the determination of the Tfh response. Model simulations predict that Tgif1 and RXR respectively induce and curtail Tfh responses. This computational hypothesis was validated experimentally by assaying Tgif1, RXR and Tfh in stomachs of mice infected with H. pylori . [ABSTRACT FROM AUTHOR]

Nucleotide-binding domain and leucine-rich repeat containing (NLR) family are intracellular sentinels of cytosolic homeostasis that orchestrate immune and inflammatory responses in infectious and immune-mediated diseases. NLRX1 is a mitochondrial-associated NOD-like receptor involved in the modulation of immune and metabolic responses. This study utilizes molecular docking approaches to investigate the structure of NLRX1 and experimentally assesses binding to naturally occurring compounds from several natural product and lipid databases. Screening of compound libraries predicts targeting of NLRX1 by conjugated trienes, polyketides, prenol lipids, sterol lipids, and coenzyme A-containing fatty acids for activating the NLRX1 pathway. The ligands of NLRX1 were identified by docking punicic acid (PUA), eleostearic acid (ESA), and docosahexaenoic acid (DHA) to the C-terminal fragment of the human NLRX1 (cNLRX1). Their binding and that of positive control RNA to cNLRX1 were experimentally determined by surface plasmon resonance (SPR) spectroscopy. In addition, the ligand binding sites of cNLRX1 were predicted in silico and validated experimentally. Target mutagenesis studies demonstrate that mutation of 4 critical residues ASP677, PHE680, PHE681, and GLU684 to alanine resulted in diminished affinity of PUA, ESA, and DHA to NLRX1. Consistent with the regulatory actions of NLRX1 on the NF-κB pathway, treatment of bone marrow derived macrophages (BMDM)s with PUA and DHA suppressed NF-κB activity in a NLRX1 dependent mechanism. In addition, a series of pre-clinical efficacy studies were performed using a mouse model of dextran sodium sulfate (DSS)-induced colitis. Our findings showed that the regulatory function of PUA on colitis is NLRX1 dependent. Thus, we identified novel small molecules that bind to NLRX1 and exert anti-inflammatory actions. [ABSTRACT FROM AUTHOR]

Background: Modeling of the immune system - a highly non-linear and complex system - requires practical and efficient data analytic approaches. The immune system is composed of heterogeneous cell populations and hundreds of cell types, such as neutrophils, eosinophils, macrophages, dendritic cells, T cells, and B cells. Each cell type is highly diverse and can be further differentiated into subsets with unique and overlapping functions. For example, CD4+ T cells can be differentiated into Th1, Th2, Th17, Th9, Th22, Treg, Tfh, as well as Tr1. Each subset plays different roles in the immune system. To study molecular mechanisms of cell differentiation, computational systems biology approaches can be used to represent these processes; however, the latter often requires building complex intracellular signaling models with a large number of equations to accurately represent intracellular pathways and biochemical reactions. Furthermore, studying the immune system entails integration of complex processes which occur at different time and space scales. Methods: This study presents and compares four supervised learning methods for modeling CD4+ T cell differentiation: Artificial Neural Networks (ANN), Random Forest (RF), Support Vector Machines (SVM), and Linear Regression (LR). Application of supervised learning methods could reduce the complexity of Ordinary Differential Equations (ODEs)-based intracellular models by only focusing on the input and output cytokine concentrations. In addition, this modeling framework can be efficiently integrated into multiscale models. Results: Our results demonstrate that ANN and RF outperform the other two methods. Furthermore, ANN and RF have comparable performance when applied to in silico data with and without added noise. The trained models were also able to reproduce dynamic behavior when applied to experimental data; in four out of five cases, model predictions based on ANN and RF correctly predicted the outcome of the system. Finally, the running time of different methods was compared, which confirms that ANN is considerably faster than RF. Conclusions: Using machine learning as opposed to ODE-based method reduces the computational complexity of the system and allows one to gain a deeper understanding of the complex interplay between the different related entities. [ABSTRACT FROM AUTHOR]

Agent-based models (ABM) are widely used to study immune systems, providing a procedural and interactive view of the underlying system. The interaction of components and the behavior of individual objects is described procedurally as a function of the internal states and the local interactions, which are often stochastic in nature. Such models typically have complex structures and consist of a large number of modeling parameters. Determining the key modeling parameters which govern the outcomes of the system is very challenging. Sensitivity analysis plays a vital role in quantifying the impact of modeling parameters in massively interacting systems, including large complex ABM. The high computational cost of executing simulations impedes running experiments with exhaustive parameter settings. Existing techniques of analyzing such a complex system typically focus on local sensitivity analysis, i.e. one parameter at a time, or a close “neighborhood” of particular parameter settings. However, such methods are not adequate to measure the uncertainty and sensitivity of parameters accurately because they overlook the global impacts of parameters on the system. In this article, we develop novel experimental design and analysis techniques to perform both global and local sensitivity analysis of large-scale ABMs. The proposed method can efficiently identify the most significant parameters and quantify their contributions to outcomes of the system. We demonstrate the proposed methodology for ENteric Immune SImulator (ENISI), a large-scale ABM environment, using a computational model of immune responses to Helicobacter pylori colonization of the gastric mucosa. [ABSTRACT FROM AUTHOR]

Computational modeling techniques are playing increasingly important roles in advancing a systems-level mechanistic understanding of biological processes. Computer simulations guide and underpin experimental and clinical efforts. This study presents ENteric Immune Simulator (ENISI), a multiscale modeling tool for modeling the mucosal immune responses. ENISI's modeling environment can simulate in silico experiments from molecular signaling pathways to tissue level events such as tissue lesion formation. ENISI's architecture integrates multiple modeling technologies including ABM (agent-based modeling), ODE (ordinary differential equations), SDE (stochastic modeling equations), and PDE (partial differential equations). This paper focuses on the implementation and developmental challenges of ENISI. A multiscale model of mucosal immune responses during colonic inflammation, including CD4+ T cell differentiation and tissue level cell-cell interactions was developed to illustrate the capabilities, power and scope of ENISI MSM. Background: Computational techniques are becoming increasingly powerful and modeling tools for biological systems are of greater needs. Biological systems are inherently multiscale, from molecules to tissues and from nano-seconds to a lifespan of several years or decades. ENISI MSM integrates multiple modeling technologies to understand immunological processes from signaling pathways within cells to lesion formation at the tissue level. This paper examines and summarizes the technical details of ENISI, from its initial version to its latest cutting-edge implementation. Implementation: Object-oriented programming approach is adopted to develop a suite of tools based on ENISI. Multiple modeling technologies are integrated to visualize tissues, cells as well as proteins; furthermore, performance matching between the scales is addressed. Conclusion: We used ENISI MSM for developing predictive multiscale models of the mucosal immune system during gut inflammation. Our modeling predictions dissect the mechanisms by which effector CD4+ T cell responses contribute to tissue damage in the gut mucosa following immune dysregulation. [ABSTRACT FROM AUTHOR]

Clostridium difficile infections are associated with the use of broad-spectrum antibiotics and result in an exuberant inflammatory response, leading to nosocomial diarrhea, colitis and even death. To better understand the dynamics of mucosal immunity during C. difficile infection from initiation through expansion to resolution, we built a computational model of the mucosal immune response to the bacterium. The model was calibrated using data from a mouse model of C. difficile infection. The model demonstrates a crucial role of T helper 17 (Th17) effector responses in the colonic lamina propria and luminal commensal bacteria populations in the clearance of C. difficile and colonic pathology, whereas regulatory T (Treg) cells responses are associated with the recovery phase. In addition, the production of anti-microbial peptides by inflamed epithelial cells and activated neutrophils in response to C. difficile infection inhibit the re-growth of beneficial commensal bacterial species. Computational simulations suggest that the removal of neutrophil and epithelial cell derived anti-microbial inhibitions, separately and together, on commensal bacterial regrowth promote recovery and minimize colonic inflammatory pathology. Simulation results predict a decrease in colonic inflammatory markers, such as neutrophilic influx and Th17 cells in the colonic lamina propria, and length of infection with accelerated commensal bacteria re-growth through altered anti-microbial inhibition. Computational modeling provides novel insights on the therapeutic value of repopulating the colonic microbiome and inducing regulatory mucosal immune responses during C. difficile infection. Thus, modeling mucosal immunity-gut microbiota interactions has the potential to guide the development of targeted fecal transplantation therapies in the context of precision medicine interventions. [ABSTRACT FROM AUTHOR]

The anti-inflammatory phytohormone abscisic acid (ABA) modulates immune and inflammatory responses in mouse models of colitis and obesity. ABA has been identified as a ligand of lanthionine synthetase C-like 2, a novel therapeutic target upstream of the peroxisome proliferator-activated receptor γ (PPARγ) pathway. The goal of this study was to investigate the immune modulatory mechanisms underlying the anti-inflammatory efficacy of ABA against influenza-associated pulmonary inflammation. Wild-type (WT) and conditional knockout mice with defective PPARγ expression in lung epithelial and hematopoietic cells (cKO) treated orally with or without ABA (100 mg/kg diet) were challenged with influenza A/Udorn (H3N2) to assess ABA's impact in disease, lung lesions and gene expression. Dietary ABA ameliorated disease activity and lung inflammatory pathology, accelerated recovery and increased survival in WT mice. ABA suppressed leukocyte infiltration and monocyte chemotactic protein 1 mRNA expression in WT mice through PPARγ since this effect was abrogated in cKO mice. ABA ameliorated disease when administered therapeutically on the same day of the infection to WT but not mice lacking PPARγ in myeloid cells. We also show that ABA's greater impact is between days 7 and 10 postchallenge when it regulates the expression of genes involved in resolution, like 5-lipoxygenase and other members of the 5-lipoxygenase pathway. Furthermore, ABA significantly increased the expression of the immunoregulatory cytokine interleukin-10 in WT mice. Our results show that ABA, given preventively or therapeutically, ameliorates influenza-virus-induced pathology by activating PPARγ in pulmonary immune cells, suppressing initial proinflammatory responses and promoting resolution. [ABSTRACT FROM AUTHOR]

Abstract: The anti-inflammatory phytohormone abscisic acid (ABA) modulates immune and inflammatory responses in mouse models of colitis and obesity. ABA has been identified as a ligand of lanthionine synthetase C-like 2, a novel therapeutic target upstream of the peroxisome proliferator-activated receptor γ (PPARγ) pathway. The goal of this study was to investigate the immune modulatory mechanisms underlying the anti-inflammatory efficacy of ABA against influenza-associated pulmonary inflammation. Wild-type (WT) and conditional knockout mice with defective PPARγ expression in lung epithelial and hematopoietic cells (cKO) treated orally with or without ABA (100 mg/kg diet) were challenged with influenza A/Udorn (H3N2) to assess ABA''s impact in disease, lung lesions and gene expression. Dietary ABA ameliorated disease activity and lung inflammatory pathology, accelerated recovery and increased survival in WT mice. ABA suppressed leukocyte infiltration and monocyte chemotactic protein 1 mRNA expression in WT mice through PPARγ since this effect was abrogated in cKO mice. ABA ameliorated disease when administered therapeutically on the same day of the infection to WT but not mice lacking PPARγ in myeloid cells. We also show that ABA''s greater impact is between days 7 and 10 postchallenge when it regulates the expression of genes involved in resolution, like 5-lipoxygenase and other members of the 5-lipoxygenase pathway. Furthermore, ABA significantly increased the expression of the immunoregulatory cytokine interleukin-10 in WT mice. Our results show that ABA, given preventively or therapeutically, ameliorates influenza-virus-induced pathology by activating PPARγ in pulmonary immune cells, suppressing initial proinflammatory responses and promoting resolution. [Copyright &y& Elsevier]

Abstract: Inflammatory bowel disease (IBD) is a debilitating and widespread immune-mediated illness characterized by excessive inflammatory and effector mucosal responses leading to tissue destruction at the gastrointestinal tract. Interactions among the immune system, the commensal microbiota and the host genotype are thought to underlie the pathogenesis of IBD. However, the precise etiology of IBD remains unknown. Diet-induced changes in the composition of the gut microbiome can modulate the induction of regulatory versus effector immune responses at the gut mucosa and improve health outcomes. Therefore, manipulation of gut microbiota composition and the local production of microbial-derived metabolites by using prebiotics, probiotics and dietary fibers is being explored as a promising avenue of prophylactic and therapeutic intervention against gut inflammation. Prebiotics and fiber carbohydrates are fermented by resident microflora into short chain fatty acids (SCFAs) in the colon. SCFAs then activate peroxisome proliferator-activated receptor (PPAR)γ, a nuclear transcription factor with widely demonstrated anti-inflammatory efficacy in experimental IBD. The activation of PPARγ by naturally ocurring compounds such as conjugated linoleic acid, pomegranate seed oil-derived punicic acid, eleostearic acid and abscisic acid has been explored as nutritional interventions that suppress colitis by directly modulating the host immune response. The aim of this review is to summarize the status of innovative nutritional interventions against gastrointestinal inflammation, their proposed mechanisms of action, preclinical and clinical efficacy as well as bioinformatics and computational modeling approaches that accelerate discovery in nutritional and mucosal immunology research. [Copyright &y& Elsevier]