Eric Bernasconi

Interleukin-17 Is a Critical Mediator of Vaccine-Induced Reduction of Helicobacter Infection in the Mouse Model

BACKGROUND & AIMS Despite the proven ability of immunization to reduce Helicobacter infection in mouse models, the precise mechanism of protection has remained elusive. This study explores the possibility that interleukin (IL)-17 plays a role in the reduction of Helicobacter infection following vaccination of wild-type animals or in spontaneous reduction of bacterial infection in IL-10-deficient mice. METHODS In mice, reducing Helicobacter infection, the levels and source of IL-17 were determined and the role of IL-17 in reduction of Helicobacter infection was probed by neutralizing antibodies. RESULTS Gastric IL-17 levels were strongly increased in mice mucosally immunized with urease plus cholera toxin and challenged with Helicobacter felis as compared with controls (654 +/- 455 and 34 +/- 84 relative units for IL-17 messenger RNA expression [P < .01] and 6.9 +/- 8.4 and 0.02 +/- 0.04 pg for IL-17 protein concentration [P < .01], respectively). Flow cytometry analysis showed that a peak of CD4(+)IL-17(+) T cells infiltrating the gastric mucosa occurred in immunized mice in contrast to control mice (4.7% +/- 0.3% and 1.4% +/- 0.3% [P < .01], respectively). Gastric mucosa-infiltrating CD4(+)IL-17(+) T cells were also observed in IL-10-deficient mice that spontaneously reduced H felis infection (4.3% +/- 2.3% and 2% +/- 0.6% [P < .01], for infected and noninfected IL-10-deficient mice, respectively). In wild-type immunized mice, intraperitoneal injection of anti-IL-17 antibodies significantly inhibited inflammation and the reduction of Helicobacter infection in comparison with control antibodies (1 of 12 mice vs 9 of 12 mice reduced Helicobacter infection [P < .01], respectively). CONCLUSIONS IL-17 plays a critical role in the immunization-induced reduction of Helicobacter infection from the gastric mucosa.

Granulocyte‐macrophage colony‐stimulating factor elicits bone marrow‐derived cells that promote efficient colonic mucosal healing

Background: Granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) therapy is effective in treating some Crohns disease (CD) patients and protects mice from colitis induced by dextran sulfate sodium (DSS) administration. However, its mechanisms of action remain elusive. We hypothesized that GM‐CSF affects intestinal mucosal repair. Methods: DSS colitic mice were treated with daily pegylated GM‐CSF or saline and clinical, histological, and inflammatory parameters were kinetically evaluated. Further, the role of bone marrow‐derived cells in the impact of GM‐CSF therapy on DSS colitis was addressed using cell transfers. Results: GM‐CSF therapy reduced clinical signs of colitis and the release of inflammatory mediators. GM‐CSF therapy improved mucosal repair, with faster ulcer reepithelialization, accelerated hyperproliferative response of epithelial cells in ulcer‐adjacent crypts, and lower colonoscopic ulceration scores in GM‐CSF‐administered mice relative to untreated mice. We observed that GM‐CSF‐induced promotion of mucosal repair is timely associated with a reduction in neutrophil numbers and increased accumulation of CD11b+ monocytic cells in colon tissues. Importantly, transfer of splenic GM‐CSF‐induced CD11b+ myeloid cells into DSS‐exposed mice improved colitis, and lethally irradiated GM‐CSF receptor‐deficient mice reconstituted with wildtype bone marrow cells were protected from DSS‐induced colitis upon GM‐CSF therapy. Lastly, GM‐CSF‐induced CD11b+ myeloid cells were shown to promote in vitro wound repair. Conclusions: Our study shows that GM‐CSF‐dependent stimulation of bone marrow‐derived cells during DSS‐induced colitis accelerates colonic tissue repair. These data provide a putative mechanism for the observed beneficial effects of GM‐CSF therapy in Crohns disease. (Inflamm Bowel Dis 2010;)

Macrophages promote epithelial repair through hepatocyte growth factor secretion

Macrophages play a critical role in intestinal wound repair. However, the mechanisms of macrophage‐assisted wound repair remain poorly understood. We aimed to characterize more clearly the repair activities of murine and human macrophages. Murine macrophages were differentiated from bone marrow cells and human macrophages from monocytes isolated from peripheral blood mononuclear cells of healthy donors (HD) or Crohns disease (CD) patients or isolated from the intestinal mucosa of HD. In‐vitro models were used to study the repair activities of macrophages. We found that murine and human macrophages were both able to promote epithelial repair in vitro. This function was mainly cell contact‐independent and relied upon the production of soluble factors such as the hepatocyte growth factor (HGF). Indeed, HGF‐silenced macrophages were less capable of promoting epithelial repair than control macrophages. Remarkably, macrophages from CD patients produced less HGF than their HD counterparts (HGF level: 84 ± 27 pg/mg of protein and 45 ± 34 pg/mg of protein, respectively, for HD and CD macrophages, P < 0·009) and were deficient in promoting epithelial repair (repairing activity: 90·1 ± 4·6 and 75·8 ± 8·3, respectively, for HD and CD macrophages, P < 0·0005). In conclusion, we provide evidence that macrophages act on wounded epithelial cells to promote epithelial repair through the secretion of HGF. The deficiency of CD macrophages to secrete HGF and to promote epithelial repair might contribute to the impaired intestinal mucosal healing in CD patients.

Chronic lung allograft dysfunction: a systematic review of mechanisms

Abstract Chronic lung allograft dysfunction (CLAD) is the major limitation of long-term survival after lung transplantation. Chronic lung allograft dysfunction manifests as bronchiolitis obliterans syndrome or the recently described restrictive allograft syndrome. Although numerous risk factors have been identified so far, the physiopathological mechanisms of CLAD remain poorly understood. We investigate here the immune mechanisms involved in the development of CLAD after lung transplantation. We explore the innate or adaptive immune reactions induced by the allograft itself or by the environment and how they lead to allograft dysfunction. Because current literature suggests bronchiolitis obliterans syndrome and restrictive allograft syndrome as 2 distinct entities, we focus on the specific factors behind one or the other syndromes. Chronic lung allograft dysfunction is a multifactorial disease that remains irreversible and unpredictable so far. We thus finally discuss the potential of systems-biology approach to predict its occurrence and to better understand its underlying mechanisms.

PAR2 Promotes Vaccine-Induced Protection Against Helicobacter Infection in Mice

BACKGROUND & AIMS Protective immunization limits Helicobacter infection of mice by undetermined mechanisms. Protease-activated receptor 2 (PAR2) signaling is believed to regulate immune and inflammatory responses. We investigated the role of PAR2 in vaccine-induced immunity against Helicobacter infection. METHODS Immune responses against Helicobacter infection were compared between vaccinated PAR2-/- and wild-type (WT) mice. Bacterial persistence, gastric pathology, and inflammatory and cellular responses were assessed using the rapid urease test (RUT), histologic analyses, quantitative polymerase chain reaction, and flow cytometry, respectively. RESULTS Following vaccination, PAR2-/- mice did not have reductions in Helicobacter felis infection (RUT values were 0.01±0.01 for WT mice and 0.11±0.13 for PAR2-/- mice; P<.05). The vaccinated PAR2-/- mice had reduced inflammation-induced stomach tissue damage (tissue damage scores were 8.83±1.47 for WT mice and 4.86±1.35 for PAR2-/- mice; P<.002) and reduced T-helper (Th)17 responses, based on reduced urease-induced interleukin (IL)-17 secretion by stomach mononuclear cells (5182 ± 1265 pg/mL for WT mice and 350±436 pg/mL for PAR2-/- mice; P<.03) and reduced recruitment of CD4+ IL-17+ T cells into the gastric mucosa of PAR2-/- mice following bacterial challenge (3.7%±1.5% for WT mice and 2.6%±1.1% for PAR2-/- mice; P<.05). In vitro, H felis-stimulated dendritic cells (DCs) from WT mice induced greater secretion of IL-17 by ovalbumin-stimulated OT-II transgenic CD4+ T cells compared with DCs from PAR2-/- mice (4298±347 and 3230±779; P<.04), indicating that PAR2-/- DCs are impaired in priming of Th17 cells. Adoptive transfer of PAR2+/+ DCs into vaccinated PAR2-/- mice increased vaccine-induced protection (RUT values were 0.11±0.10 and 0.26±0.15 for injected and noninjected mice, respectively; P<.03). CONCLUSIONS PAR2 activates DCs to mediate vaccine-induced protection against Helicobacter infection in mice.

Airway microbiota signals anabolic and catabolic remodeling in the transplanted lung

Background: Homeostatic turnover of the extracellular matrix conditions the structure and function of the healthy lung. In lung transplantation, long‐term management remains limited by chronic lung allograft dysfunction, an umbrella term used for a heterogeneous entity ultimately associated with pathological airway and/or parenchyma remodeling. Objective: This study assessed whether the local cross‐talk between the pulmonary microbiota and host cells is a key determinant in the control of lower airway remodeling posttransplantation. Methods: Microbiota DNA and host total RNA were isolated from 189 bronchoalveolar lavages obtained from 116 patients post lung transplantation. Expression of a set of 11 genes encoding either matrix components or factors involved in matrix synthesis or degradation (anabolic and catabolic remodeling, respectively) was quantified by real‐time quantitative PCR. Microbiota composition was characterized using 16S ribosomal RNA gene sequencing and culture. Results: We identified 4 host gene expression profiles, among which catabolic remodeling, associated with high expression of metallopeptidase‐7, ‐9, and ‐12, diverged from anabolic remodeling linked to maximal thrombospondin and platelet‐derived growth factor D expression. While catabolic remodeling aligned with a microbiota dominated by proinflammatory bacteria (eg, Staphylococcus, Pseudomonas, and Corynebacterium), anabolic remodeling was linked to typical members of the healthy steady state (eg, Prevotella, Streptococcus, and Veillonella). Mechanistic assays provided direct evidence that these bacteria can impact host macrophage‐fibroblast activation and matrix deposition. Conclusions: Host‐microbes interplay potentially determines remodeling activities in the transplanted lung, highlighting new therapeutic opportunities to ultimately improve long‐term lung transplant outcome. Graphical abstract Figure. No caption available.

Critical role of the GM-CSF signaling pathway in macrophage pro-repair activities.

Objective: Macrophages play a critical role in intestinal wound repair. However, the molecular pathways that regulate macrophage wound repair activities remain poorly understood. The aim of this study was to evaluate the role of GM-CSF receptor signaling in the wound repair activities of macrophages. Methods: Murine macrophages were differentiated from bone marrow cells and human macrophages from monocytes isolated from peripheral blood mononuclear cells of Crohns disease (CD) patients. In vitro models were used to study the repair activities of macrophages. Results: We provide evidence that GM-CSF receptor signaling is required for murine macrophages to promote epithelial repair. In addition, we demonstrate that the deficient repair properties of macrophages from CD patients with active disease can be recovered via GM-CSF therapy. Conclusion: Our data support a critical role of the GM-CSF signaling pathway in the pro-repair activities of mouse and human macrophages.

Gingival Tissue Inflammation Promotes Increased Matrix Metalloproteinase-12 Production by CD200Rlow Monocyte-Derived Cells in Periodontitis

Irreversible tissue recession in chronic inflammatory diseases is associated with dysregulated immune activation and production of tissue degradative enzymes. In this study, we identified elevated levels of matrix metalloproteinase (MMP)-12 in gingival tissue of patients with the chronic inflammatory disease periodontitis (PD). The source of MMP12 was cells of monocyte origin as determined by the expression of CD14, CD68, and CD64. These MMP12-producing cells showed reduced surface levels of the coinhibitory molecule CD200R. Similarly, establishing a multicellular three-dimensional model of human oral mucosa with induced inflammation promoted MMP12 production and reduced CD200R surface expression by monocyte-derived cells. MMP12 production by monocyte-derived cells was induced by CSF2 rather than the cyclooxygenase-2 pathway, and treatment of monocyte-derived cells with a CD200R ligand reduced CSF2-induced MMP12 production. Further, MMP12-mediated degradation of the extracellular matrix proteins tropoelastin and fibronectin in the tissue model coincided with a loss of Ki-67, a protein strictly associated with cell proliferation. Reduced amounts of tropoelastin were confirmed in gingival tissue from PD patients. Thus, this novel association of the CD200/CD200R pathway with MMP12 production by monocyte-derived cells may play a key role in PD progression and will be important to take into consideration in the development of future strategies to diagnose, treat, and prevent PD.

Microbial and host immune factors as drivers of COPD

Compartmentalisation of the respiratory tract microbiota in patients with different chronic obstructive pulmonary disease (COPD) severity degrees needs to be systematically investigated. In addition, it is unknown if the inflammatory and emphysematous milieux in patients with COPD are associated with changes in the respiratory tract microbiota and host macrophage gene expression. We performed a cross-sectional study to compare non-COPD controls (n=10) to COPD patients (n=32) with different disease severity degrees. Samples (n=187) were obtained from different sites of the upper and lower respiratory tract. Microbiota analyses were performed by 16S ribosomal RNA gene sequencing and host gene expression analyses by quantitative real-time PCR of distinct markers of bronchoalveolar lavage cells. Overall, the microbial communities of severe COPD (Global Initiative for Chronic Obstructive Lung Disease (GOLD) grade 3/4) patients clustered significantly differently to controls and less severe COPD (GOLD 1/2) patients (permutational multivariate ANOVA (MANOVA), p=0.001). However, we could not detect significant associations between the different sampling sites in the lower airways. In addition, the chosen set of host gene expression markers significantly separated COPD GOLD 3/4 patients, and we found correlations between the composition of the microbiota and the host data. In conclusion, this study demonstrates associations between host gene expression and microbiota profiles that may influence the course of COPD. Associations of the host immune response and a disordered microbiota in patients with different COPD severity degrees http://ow.ly/h2mW30k9Nua