ABSTRACT
Central to inflammatory bowel disease (IBD) pathogenesis is loss of mucosal barrier function. Emerging evidence implicates extracellular adenosine signaling in attenuating mucosal inflammation. We hypothesized that adenosine-mediated protection from intestinal barrier dysfunction involves tissue-specific signaling through the A2B adenosine receptor (Adora2b) at the intestinal mucosal surface. To address this hypothesis, we combined pharmacologic studies and studies in mice with global or tissue-specific deletion of the Adora2b receptor. Adora2b(-/-) mice experienced a significantly heightened severity of colitis, associated with a more acute onset of disease and loss of intestinal epithelial barrier function. Comparison of mice with Adora2b deletion on vascular endothelial cells (Adora2b(fl/fl)VeCadCre(+)) or intestinal epithelia (Adora2b(fl/fl)VillinCre(+)) revealed a selective role for epithelial Adora2b signaling in attenuating colonic inflammation. In vitro studies with Adora2b knockdown in intestinal epithelial cultures or pharmacologic studies highlighted Adora2b-driven phosphorylation of vasodilator-stimulated phosphoprotein (VASP) as a specific barrier repair response. Similarly, in vivo studies in genetic mouse models or treatment studies with an Adora2b agonist (BAY 60-6583) recapitulate these findings. Taken together, our results suggest that intestinal epithelial Adora2b signaling provides protection during intestinal inflammation via enhancing mucosal barrier responses.
Subject(s)
Colitis/pathology , Epithelial Cells/metabolism , Intestinal Mucosa/pathology , Receptor, Adenosine A2B/metabolism , Signal Transduction , Acute Disease , Animals , Blotting, Western , Colitis/metabolism , Disease Models, Animal , Epithelial Cells/pathology , Flow Cytometry , Fluorescent Antibody Technique , In Situ Nick-End Labeling , Intestinal Mucosa/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , Signal Transduction/physiologyABSTRACT
Inflammatory bowel disease (IBD) is a chronic inflammatory condition thought to reflect a failure of the enteral immune system to adequately regulate itself. Inflammatory stress drives upregulation of heat-shock proteins (HSPs), including the pro-inflammatory chaperone, HSP90. This protein sequesters the transcription factor, heat-shock factor 1 (HSF1) in the cytoplasm preventing transcription of a number of anti-inflammatory proteins. We hypothesized that inhibition of HSP90 would exert an anti-inflammatory effect and thereby attenuate intestinal inflammation in murine models of IBD. Inhibition of HSP90 with 17-allylaminogeldanamycin (17-AAG) reduced inflammation in acute dextran sodium sulfate and chronic CD45RB(High) colitis models coinciding with increased interleukin (IL)-10 production in the colon. Regulatory T cells (Tregs) from mice treated with 17-AAG demonstrated significantly greater suppressive capacity in vitro abolished in HSF1-/- or IL-10-/- cells. Finally, Tregs treated with 17-AAG exhibited increased nuclear localization of HSF1 with resultant upregulation of HSF1 response genes, including HSP70, HSP90 and IL-10.
Subject(s)
Adenosine Triphosphatases/metabolism , Cell Nucleus/metabolism , Colitis/immunology , Cytoplasm/metabolism , DNA-Binding Proteins/metabolism , HSP90 Heat-Shock Proteins/metabolism , T-Lymphocytes, Regulatory/immunology , Transcription Factors/metabolism , Animals , Benzoquinones/pharmacology , Cells, Cultured , Colitis/chemically induced , Colitis/drug therapy , DNA-Binding Proteins/genetics , Dextran Sulfate/administration & dosage , Heat Shock Transcription Factors , Humans , Interleukin-10/genetics , Interleukin-10/metabolism , Lactams, Macrocyclic/pharmacology , Leukocyte Common Antigens/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , Protein Transport/drug effects , Protein Transport/genetics , T-Lymphocytes, Regulatory/drug effects , Transcription Factors/geneticsABSTRACT
Acute lung injury (ALI) is associated with high morbidity and mortality in critically ill patients. At present, the functional contribution of airway mucins to ALI is unknown. We hypothesized that excessive mucus production could be detrimental during lung injury. Initial transcriptional profiling of airway mucins revealed a selective and robust induction of MUC5AC upon cyclic mechanical stretch exposure of pulmonary epithelia (Calu-3). Additional studies confirmed time- and stretch-dose-dependent induction of MUC5AC transcript or protein during cyclic mechanical stretch exposure in vitro or during ventilator-induced lung injury in vivo. Patients suffering from ALI showed a 58-fold increase in MUC5AC protein in their bronchoalveolar lavage. Studies of the MUC5AC promoter implicated nuclear factor κB in Muc5ac induction during ALI. Moreover, mice with gene-targeted deletion of Muc5acâ»/â» experience attenuated lung inflammation and pulmonary edema during injurious ventilation. We observed that neutrophil trafficking into the lungs of Muc5acâ»/â» mice was selectively attenuated. This implicates that endogenous Muc5ac production enhances pulmonary neutrophil trafficking during lung injury. Together, these studies reveal a detrimental role for endogenous Muc5ac production during ALI and suggest pharmacological strategies to dampen mucin production in the treatment of lung injury.
