Extracellular vesicles derived from the intestinal microbiota of patients with CGD-associated inflammatory bowel disease have pro-inflammatory effects

Chronic granulomatous disease (CGD) is an inborn error of immunity characterized by mutations in any one of the 6 subunits forming the nicotinamide-adenine dinucleotide phosphate oxidase complex 2 leading to defective phagocyte-derived ROS production. As a result, patients with CGD have recurrent infections and increased inflammation and 50% of patients with CGD have inflammatory bowel disease (CGD-IBD). Although we have previously demonstrated that patients with CGD-IBD have intestinal dysbiosis, its functional impact is poorly understood. Interestingly, intestinal microbes can produce gut microbiota-derived extracellular vesicles (GMEVs) which can cross the intestinal barrier and have systemic and/or end-organ immunomodulatory effects. Yet, the role of GMEVs in patients with CGD has never been evaluated. To investigate the impact of GMEVs isolated from patients with CGD, with or without CGD-IBD, on intestinal permeability and inflammatory responses using wild type and NOX2-deficient (CYBB-/-) models. GMEVs were isolated and characterized from fecal samples obtained from 11 patients without CGD-IBD and 11 patients with active CGD-IBD recruited at the National Institutes of Health. Stimulation with GMEVs was performed on HT29, THP1 and CYBB-/- THP1 cell lines, HT29/THP1 co-culture models, and induced pluripotent stem cell-derived human intestinal organoids. TNF-α, IL-6, IL-8, IL-1β, IP-10, GM-CSF and MCP1 gene expression and production were assessed by RT-qPCR and ELISA, respectively. Our data showed that stimulation with GMEVs from patients with active CGD-IBD increased intestinal permeability compared to cells stimulated with GMEVs from patients without CGD-IBD. In HT29 cells, increased intestinal permeability was associated with increased gene expression and production of IL-8 and IL-1β. IL-8 was similarly increased in intestinal organoids. In THP1 cells, stimulation led to increased gene expression and production of TNF-α, IL-6 and IP-10, while stimulation in CYBB-/- THP1 cells increased TNF-α and MCP1 gene expression. Finally, stimulation of HT29/THP1 cell co-cultures resulted in increased gene expression of TNF-α, IL-6 and GM-CSF. Our results suggest that GMEVs isolated from patients with CGD-IBD may induce pro-inflammatory immune responses in patients with CGD, thereby contributing to their underlying inflammatory pathology. These findings bring forward GMEVs as a possible new therapeutic target for CGD-IBD. Download : Download high-res image (111KB)Download : Download full-size imageFig. 1. Experimental model used in this work. Fig. 1. Experimental model used in this work.