Microbiota-Induced TNF-like Ligand 1A Drives Group 3 Innate Lymphoid Cell-Mediated Barrier Protection and Intestinal T Cell Activation during Colitis.

Inflammatory bowel disease (IBD) results from a dysregulated interaction between the microbiota and a genetically susceptible host. Genetic studies have linked TNFSF15 polymorphisms and its protein TNF-like ligand 1A (TL1A) with IBD, but the functional role of TL1A is not known. Here, we found that adherent IBD-associated microbiota induced TL1A release from CX3CR1+ mononuclear phagocytes (MNPs). Using cell-specific genetic deletion models, we identified an essential role for CX3CR1+MNP-derived TL1A in driving group 3 innate lymphoid cell (ILC3) production of interleukin-22 and mucosal healing during acute colitis. In contrast to this protective role in acute colitis, TL1A-dependent expression of co-stimulatory molecule OX40L in MHCII+ ILC3s during colitis led to co-stimulation of antigen-specific T cells that was required for chronic T cell colitis. These results identify a role for ILC3s in activating intestinal T cells and reveal a central role for TL1A in promoting ILC3 barrier immunity during colitis.

Transferable Immunoglobulin A-Coated Odoribacter splanchnicus in Responders to Fecal Microbiota Transplantation for Ulcerative Colitis Limits Colonic Inflammation.

BACKGROUND & AIMS: Fecal microbiota transplantation (FMT) is an emerging treatment modality for ulcerative colitis (UC). Several randomized controlled trials have shown efficacy for FMT in the treatment of UC, but a better understanding of the transferable microbiota and their immune impact is needed to develop more efficient microbiome-based therapies for UC. METHODS: Metagenomic analysis and strain tracking was performed on 60 donor and recipient samples receiving FMT for active UC. Sorting and sequencing of immunoglobulin (Ig) A-coated microbiota (called IgA-seq) was used to define immune-reactive microbiota. Colonization of germ-free or genetically engineered mice with patient-derived strains was performed to determine the mechanism of microbial impact on intestinal immunity. RESULTS: Metagenomic analysis defined a core set of donor-derived transferable bacterial strains in UC subjects achieving clinical response, which predicted response in an independent trial of FMT for UC. IgA-seq of FMT recipient samples and gnotobiotic mice colonized with donor microbiota identified Odoribacter splanchnicus as a transferable strain shaping mucosal immunity, which correlated with clinical response and the induction of mucosal regulatory T cells. Colonization of mice with O splanchnicus led to an increase in Foxp3+/RORγt+ regulatory T cells, induction of interleukin (IL) 10, and production of short chain fatty acids, all of which were required for O splanchnicus to limit colitis in mouse models. CONCLUSIONS: This work provides the first evidence of transferable, donor-derived strains that correlate with clinical response to FMT in UC and reveals O splanchnicus as a key component promoting both metabolic and immune cell protection from colitis. These mechanistic features will help enable strategies to enhance the efficacy of microbial therapy for UC. Clinicaltrials.gov ID NCT02516384.

Cooperation of Gastric Mononuclear Phagocytes with Helicobacter pylori during Colonization.

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.

Bistability analyses of CD4+ T follicular helper and regulatory cells during Helicobacter pylori infection.

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.

Preclinical studies of amixicile, a systemic therapeutic developed for treatment of Clostridium difficile infections that also shows efficacy against Helicobacter pylori.

Amixicile shows efficacy in the treatment of Clostridium difficile infections (CDI) in a mouse model, with no recurrence of CDI. Since amixicile selectively inhibits the action of a B vitamin (thiamine pyrophosphate) cofactor of pyruvate:ferredoxin oxidoreductase (PFOR), it may both escape mutation-based drug resistance and spare beneficial probiotic gut bacteria that do not express this enzyme. Amixicile is a water-soluble derivative of nitazoxanide (NTZ), an antiparasitic therapeutic that also shows efficacy against CDI in humans. In comparative studies, amixicile showed no toxicity to hepatocytes at 200 µM (NTZ was toxic above 10 µM); was not metabolized by human, dog, or rat liver microsomes; showed equivalence or superiority to NTZ in cytochrome P450 assays; and did not activate efflux pumps (breast cancer resistance protein, P glycoprotein). A maximum dose (300 mg/kg) of amixicile given by the oral or intraperitoneal route was well tolerated by mice and rats. Plasma exposure (rats) based on the area under the plasma concentration-time curve was 79.3 h · µg/ml (30 mg/kg dose) to 328 h · µg/ml (100 mg/kg dose), the maximum concentration of the drug in serum was 20 µg/ml, the time to the maximum concentration of the drug in serum was 0.5 to 1 h, and the half-life was 5.6 h. Amixicile did not concentrate in mouse feces or adversely affect gut populations of Bacteroides species, Firmicutes, segmented filamentous bacteria, or Lactobacillus species. Systemic bioavailability was demonstrated through eradication of Helicobacter pylori in a mouse infection model. In summary, the efficacy of amixicile in treating CDI and other infections, together with low toxicity, an absence of mutation-based drug resistance, and excellent drug metabolism and pharmacokinetic metrics, suggests a potential for broad application in the treatment of infections caused by PFOR-expressing microbial pathogens in addition to CDI.

Systems modeling of molecular mechanisms controlling cytokine-driven CD4+ T cell differentiation and phenotype plasticity.

Differentiation of CD4+ T cells into effector or regulatory phenotypes is tightly controlled by the cytokine milieu, complex intracellular signaling networks and numerous transcriptional regulators. We combined experimental approaches and computational modeling to investigate the mechanisms controlling differentiation and plasticity of CD4+ T cells in the gut of mice. Our computational model encompasses the major intracellular pathways involved in CD4+ T cell differentiation into T helper 1 (Th1), Th2, Th17 and induced regulatory T cells (iTreg). Our modeling efforts predicted a critical role for peroxisome proliferator-activated receptor gamma (PPARγ) in modulating plasticity between Th17 and iTreg cells. PPARγ regulates differentiation, activation and cytokine production, thereby controlling the induction of effector and regulatory responses, and is a promising therapeutic target for dysregulated immune responses and inflammation. Our modeling efforts predict that following PPARγ activation, Th17 cells undergo phenotype switch and become iTreg cells. This prediction was validated by results of adoptive transfer studies showing an increase of colonic iTreg and a decrease of Th17 cells in the gut mucosa of mice with colitis following pharmacological activation of PPARγ. Deletion of PPARγ in CD4+ T cells impaired mucosal iTreg and enhanced colitogenic Th17 responses in mice with CD4+ T cell-induced colitis. Thus, for the first time we provide novel molecular evidence in vivo demonstrating that PPARγ in addition to regulating CD4+ T cell differentiation also plays a major role controlling Th17 and iTreg plasticity in the gut mucosa.

The gut microbiome regulates the clinical efficacy of sulfasalazine therapy for IBD-associated spondyloarthritis.

Sulfasalazine is a prodrug known to be effective for the treatment of inflammatory bowel disease (IBD)-associated peripheral spondyloarthritis (pSpA), but the mechanistic role for the gut microbiome in regulating its clinical efficacy is not well understood. Here, treatment of 22 IBD-pSpA subjects with sulfasalazine identifies clinical responders with a gut microbiome enriched in Faecalibacterium prausnitzii and the capacity for butyrate production. Sulfapyridine promotes butyrate production and transcription of the butyrate synthesis gene but in F. prausnitzii in vitro, which is suppressed by excess folate. Sulfasalazine therapy enhances fecal butyrate production and limits colitis in wild-type and gnotobiotic mice colonized with responder, but not non-responder, microbiomes. F. prausnitzii is sufficient to restore sulfasalazine protection from colitis in gnotobiotic mice colonized with non-responder microbiomes. These findings reveal a mechanistic link between the efficacy of sulfasalazine therapy and the gut microbiome with the potential to guide diagnostic and therapeutic approaches for IBD-pSpA.

Helicobacter pylori infection in a pig model is dominated by Th1 and cytotoxic CD8+ T cell responses.

Helicobacter pylori infection is the leading cause for peptic ulcer disease and gastric adenocarcinoma. Mucosal T cell responses play an important role in mediating H. pylori-related gastric immunopathology. While induced regulatory T (iTreg) cells are required for chronic colonization without disease, T helper 1 (Th1) effector responses are associated with lower bacterial loads at the expense of gastric pathology. Pigs were inoculated with either H. pylori strain SS1 or J99. Phenotypic and functional changes in peripheral blood mononuclear cell (PBMC) populations were monitored weekly, and mucosal immune responses and bacterial loads were assessed up to 2 months postinfection. Both H. pylori strains elicited a Th1 response characterized by increased percentages of CD4(+)Tbet(+) cells and elevated gamma interferon (IFN-γ) mRNA in PBMCs. A subset of CD8(+) T cells expressing Tbet and CD16 increased following infection. Moreover, a significant increase in perforin and granzyme mRNA expression was observed in PBMCs of infected pigs, indicating a predominant cytotoxic immune response. Infiltration of B cells, myeloid cells, T cells expressing Treg- and Th17-associated transcription factors, and cytotoxic T cells was found in the gastric lamina propria of both infected groups. Interestingly, based on bacterial reisolation data, strain SS1 showed greater capacity to colonize and/or persist in the gastric mucosa than did strain J99. This novel pig model of infection closely mimics human gastric pathology and presents a suitable avenue for studying effector and regulatory responses toward H. pylori described in humans.

Mutations to essential orphan response regulator HP1043 of Helicobacter pylori result in growth-stage regulatory defects.

Helicobacter pylori establishes lifelong infections of the gastric mucosa, a niche considered hostile to most microbes. While responses to gastric acidity and local inflammation are understood, little is known as to how they are integrated into homeostatic control of cell division and growth-stage gene expression. Here we investigate the essential orphan response regulator HP1043, a member of the OmpR/PhoB subfamily of transcriptional regulators that is unique to the Epsilonproteobacteria and that lacks phosphorylation domains. To test the hypothesis that conformational changes in the homodimer might lead to defects in gene expression, we sought mutations that might alter DNA-binding efficiency. Two introduced mutations (C215S, C221S) C terminal to the DNA-binding domain of HP1043 (HP1043CC11) resulted in a 2-fold higher affinity for its own promoter by footprinting. Modeling studies with the crystal structure of HP1043 suggested that C215S might affect the helix-turn-helix domain. Genomic replacement of the hp1043 allele with the hp1043CC11 mutant allele resulted in a 2-fold decrease in protein levels, despite a dramatic increase in mRNA. The mutations did not affect in vitro growth rates or colonization efficiency in a mouse model. Proteomic profiling (CC11 mutant strain versus wild type) identified many expression differences, and quantitative PCR further revealed that 11 out of 12 examined genes had lost growth-stage regulation and that 6 of the genes contained HP1043 binding consensus sequences within the promoter regions (fur, cagA, cag23, flhA, flip, and napA). Our studies show that mutations that affect DNA-binding affinity can be used to identify new members of the HP1043 regulon.

Agr2-associated ER stress promotes adherent-invasive E. coli dysbiosis and triggers CD103+ dendritic cell IL-23-dependent ileocolitis.

Endoplasmic reticulum (ER) stress is associated with Crohn's disease (CD), but its impact on host-microbe interaction in disease pathogenesis is not well defined. Functional deficiency in the protein disulfide isomerase anterior gradient 2 (AGR2) has been linked with CD and leads to epithelial cell ER stress and ileocolitis in mice and humans. Here, we show that ileal expression of AGR2 correlates with mucosal Enterobactericeae abundance in human inflammatory bowel disease (IBD) and that Agr2 deletion leads to ER-stress-dependent expansion of mucosal-associated adherent-invasive Escherichia coli (AIEC), which drives Th17 cell ileocolitis in mice. Mechanistically, our data reveal that AIEC-induced epithelial cell ER stress triggers CD103+ dendritic cell production of interleukin-23 (IL-23) and that IL-23R is required for ileocolitis in Agr2-/- mice. Overall, these data reveal a specific and reciprocal interaction of the expansion of the CD pathobiont AIEC with ER-stress-associated ileocolitis and highlight a distinct cellular mechanism for IL-23-dependent ileocolitis.

Soluble fibers and resistant starch ameliorate disease activity in interleukin-10-deficient mice with inflammatory bowel disease.

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 (SCF), STA-LITE III polydextrose (PDX), Biogum (BG), Pullulan (PI-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 (PP) 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, SCF, 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, PI-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 PPARγ expression to suppress gut inflammation.

Decoding the matrix: multiomics reveals host-microbe biomarker for inflammatory bowel disease.

Inflammatory bowel disease (IBD) is a chronic inflammatory disease of the intestine associated with genetic susceptibility and alterations in the intestinal microbiome. Multiomics data developed and analyzed over the last several decades have yielded an unprecedented amount of genetic and microbial data. But how do we pinpoint mechanistic insight into the host-microbe relationship that will ultimately enable better care for patients with IBD? In this issue of the JCI, Grasberger et al. undertook a major decoding effort to decipher this multiomic data matrix. The authors analyzed anonymized data from more than 2800 individuals to discover a link between heterozygous carriers of deleterious DUOX2 variants and high levels of plasma IL-17C. These findings provide an example of how harnessing big data can drive mechanistic discovery to define disease biomarkers that have the potential to improve clinical care in IBD.

Adherent-invasive E. coli metabolism of propanediol in Crohn's disease regulates phagocytes to drive intestinal inflammation.

Adherent-invasive E. coli (AIEC) are enriched in the intestinal microbiota of patients with Crohn's disease (CD) and promote intestinal inflammation. Yet, how AIEC metabolism of nutrients impacts intestinal homeostasis is poorly defined. Here, we show that AIEC encoding the large subunit of propanediol dehydratase (PduC), which facilitates the utilization of fucose fermentation product 1,2-propanediol, are increased in the microbiome of CD patients and drive AIEC-induced intestinal T cell inflammation. In murine models, CX3CR1+ mononuclear phagocytes (MNP) are required for PduC-dependent induction of T helper 17 (Th17) cells and interleukin-1ß (IL-1ß) production that leads to AIEC-induced inflammatory colitis. Activation of this inflammatory cascade requires the catalytic activity of PduC to generate propionate, which synergizes with lipopolysaccharide (LPS) to induce IL-1ß by MNPs. Disrupting fucose availability limits AIEC-induced propionate production and intestinal inflammation. These findings identify MNPs as metabolic sensors linking AIEC metabolism with intestinal inflammation and identify microbial metabolism as a potential therapeutic target in Crohn's disease treatment.

Identification of new regulatory genes through expression pattern analysis of a global RNA-seq dataset from a Helicobacter pylori co-culture system.

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.

Single Delivery of High-Diversity Fecal Microbiota Preparation by Colonoscopy Is Safe and Effective in Increasing Microbial Diversity in Active Ulcerative Colitis.

BACKGROUND: Recent trials suggest fecal microbiota transplantation (FMT) with repeated enemas and high-diversity FMT donors is a promising treatment to induce remission in ulcerative colitis. METHODS: We designed a prospective, open-label pilot study to assess the safety, clinical efficacy, and microbial engraftment of single FMT delivery by colonoscopy for active ulcerative colitis using a 2-donor fecal microbiota preparation (FMP). Safety and clinical endpoints of response, remission, and mucosal healing at week 4 were assessed. Fecal DNA and rectal biopsies were used to characterize the microbiome and mucosal CD4 T cells, respectively, before and after FMT. RESULTS: Of the 20 patients enrolled in this study, 7 patients (35%) achieved a clinical response by week 4. Three patients (15%) were in remission at week 4 and 2 of these patients (10%) achieved mucosal healing. Three patients (15%) required escalation of care. No serious adverse events were observed. Microbiome analysis revealed that restricted diversity of recipients pre-FMT was significantly increased by high-diversity 2-donor FMP. The microbiome of recipients post-transplant was more similar to the donor FMP than the pretransplant recipient sample in both responders and nonresponders. Notably, donor composition correlated with clinical response. Mucosal CD4 T-cell analysis revealed a reduction in both Th1 and regulatory T-cells post-FMT. CONCLUSIONS: High-diversity, 2-donor FMP delivery by colonoscopy seems safe and effective in increasing fecal microbial diversity in patients with active ulcerative colitis. Donor composition correlated with clinical response and further characterization of immunological parameters may provide insight into factors influencing clinical outcome.

IgA-coated E. coli enriched in Crohn's disease spondyloarthritis promote TH17-dependent inflammation.

Peripheral spondyloarthritis (SpA) is a common extraintestinal manifestation in patients with active inflammatory bowel disease (IBD) characterized by inflammatory enthesitis, dactylitis, or synovitis of nonaxial joints. However, a mechanistic understanding of the link between intestinal inflammation and SpA has yet to emerge. We evaluated and functionally characterized the fecal microbiome of IBD patients with or without peripheral SpA. Coupling the sorting of immunoglobulin A (IgA)-coated microbiota with 16S ribosomal RNA-based analysis (IgA-seq) revealed a selective enrichment in IgA-coated Escherichia coli in patients with Crohn's disease-associated SpA (CD-SpA) compared to CD alone. E. coli isolates from CD-SpA-derived IgA-coated bacteria were similar in genotype and phenotype to an adherent-invasive E. coli (AIEC) pathotype. In comparison to non-AIEC E. coli, colonization of germ-free mice with CD-SpA E. coli isolates induced T helper 17 cell (TH17) mucosal immunity, which required the virulence-associated metabolic enzyme propanediol dehydratase (pduC). Modeling the increase in mucosal and systemic TH17 immunity we observed in CD-SpA patients, colonization of interleukin-10-deficient or K/BxN mice with CD-SpA-derived E. coli lead to more severe colitis or inflammatory arthritis, respectively. Collectively, these data reveal the power of IgA-seq to identify immunoreactive resident pathosymbionts that link mucosal and systemic TH17-dependent inflammation and offer microbial and immunophenotype stratification of CD-SpA that may guide medical and biologic therapy.

Modulation of inflammation and immunity by dietary conjugated linoleic acid.

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.

Systems-wide analyses of mucosal immune responses to Helicobacter pylori at the interface between pathogenicity and symbiosis.

Helicobacter pylori is the dominant member of the gastric microbiota in over half of the human population of which 5-15% develop gastritis or gastric malignancies. Immune responses to H. pylori are characterized by mixed T helper cell, cytotoxic T cell and NK cell responses. The presence of Tregs is essential for the control of gastritis and together with regulatory CX3CR1+ mononuclear phagocytes and immune-evasion strategies they enable life-long persistence of H. pylori. This H. pylori-induced regulatory environment might contribute to its cross-protective effect in inflammatory bowel disease and obesity. Here we review host-microbe interactions, the development of pro- and anti-inflammatory immune responses and how the latter contribute to H. pylori's role as beneficial member of the gut microbiota. Furthermore, we present the integration of existing and new data into a computational/mathematical model and its use for the investigation of immunological mechanisms underlying initiation, progression and outcomes of H. pylori infection.

Modeling the Role of Lanthionine Synthetase C-Like 2 (LANCL2) in the Modulation of Immune Responses to Helicobacter pylori Infection.

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.

Modeling the Regulatory Mechanisms by Which NLRX1 Modulates Innate Immune Responses to Helicobacter pylori Infection.

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.