Duodenal mucosa of untreated celiac disease patients has altered expression of the GAS6 and PROS1 and the negative regulator tyrosine kinase TAM receptors subfamily.

Celiac disease (CD) is an immune-driven disease characterized by tissue damage in the small intestine of genetically-susceptible individuals. We evaluated here a crucial immune regulatory pathway involving TYRO3, AXL, and MERTK (TAM) receptors and their ligands PROS1 and GAS6 in duodenal biopsies of controls and CD patients. We found increased GAS6 expression associated with downregulation of PROS1 and variable TAM receptors levels in duodenum tissue of CD patients. Interestingly, CD3+ lymphocytes, CD68+, CD11c+ myeloid and epithelial cells, showed differential expressions of TAM components comparing CD vs controls. Principal component analysis revealed a clear segregation of two groups of CD patients based on TAM components and IFN signaling. In vitro validation demonstrated that monocytes, T lymphocytes and epithelial cells upregulated TAM components in response to IFN stimulation. Our findings highlight a dysregulated TAM axis in CD related to IFN signaling and contribute to a deeper understanding of the pathophysiology of CD.

Coexistence of apoptosis, pyroptosis, and necroptosis pathways in celiac disease.

Usually, the massive elimination of cells under steady-state conditions occurs by apoptosis, which is also acknowledged to explain the loss of enterocytes in the small intestine of celiac disease (CD) patients. However, little is known about the role of proinflammatory cell death pathways in CD. Here, we have used confocal microscopy, western blot, and RT-qPCR analysis to assess the presence of regulated cell death pathways in the duodenum of CD patients. We found an increased number of dead (TUNEL+) cells in the lamina propria of small intestine of CD patients, most of them are plasma cells (CD138+). Many dying cells expressed FAS and were in close contact with CD3+ T cells. Caspase-8 and caspase-3 expression was increased in CD, confirming the activation of apoptosis. In parallel, caspase-1, IL-1ß, and GSDMD were increased in CD samples indicating the presence of inflammasome-dependent pyroptosis. Necroptosis was also present, as shown by the increase of RIPK3 and phosphorylate MLKL. Analysis of published databases confirmed that CD has an increased expression of regulated cell death -related genes. Together, these results reveal that CD is characterized by cell death of different kinds. In particular, the presence of proinflammatory cell death pathways may contribute to mucosal damage.

Probiotic Lactobacilli Isolated from Kefir Promote Down-Regulation of Inflammatory Lamina Propria T Cells from Patients with Active IBD.

Ulcerative colitis and Crohn's disease, the two main forms of inflammatory bowel disease (IBD), are immunologically mediated disorders. Several therapies are focused on activated T cells as key targets. Although Lactobacillus kefiri has shown anti-inflammatory effects in animal models, few studies were done using human mucosal T cells. The aim of this work was to investigate the immunomodulatory effects of this bacterium on intestinal T cells from patients with active IBD. Mucosal biopsies and surgical samples from IBD adult patients (n = 19) or healthy donors (HC; n = 5) were used. Lamina propria mononuclear cells were isolated by enzymatic tissue digestion, and entero-adhesive Escherichia coli-specific lamina propria T cells (LPTC) were expanded. The immunomodulatory properties of L. kefiri CIDCA 8348 strain were evaluated on biopsies and on anti-CD3/CD28-activated LPTC. Secreted cytokines were quantified by ELISA, and cell proliferation and viability were assessed by flow cytometry. We found that L. kefiri reduced spontaneous release of IL-6 and IL-8 from inflamed biopsies ex vivo. Activated LPTC from IBD patients showed low proliferative rates and reduced secretion of TNF-α, IL-6, IFN-γ and IL-13 in the presence of L. kefiri. In addition, L. kefiri induced an increased frequency of CD4+FOXP3+ LPTC along with high levels of IL-10. This is the first report showing an immunomodulatory effect of L. kefiri CIDCA 8348 on human intestinal cells from IBD patients. Understanding the mechanisms of interaction between probiotics and immune mucosal cells may open new avenues for treatment and prevention of IBD.

IL-33 Alarmin and Its Active Proinflammatory Fragments Are Released in Small Intestine in Celiac Disease.

Initially described as Th2 promoter cytokine, more recently, IL-33 has been recognized as an alarmin, mainly in epithelial and endothelial cells. While localized in the nucleus acting as a gene regulator, it can be also released after injury, stress or inflammatory cell death. As proinflammatory signal, IL-33 binds to the surface receptor ST2, which enhances mast cell, Th2, regulatory T cell, and innate lymphoid cell type 2 functions. Besides these Th2 roles, free IL-33 can activate CD8+ T cells during ongoing Th1 immune responses to potentiate its cytotoxic function. Celiac Disease (CD) is a chronic inflammatory disorder characterized by a predominant Th1 response leading to multiple pathways of mucosal damage in the proximal small intestine. By immunofluorescence and western blot analysis of duodenal tissues, we found an increased expression of IL-33 in duodenal mucosa of active CD (ACD) patients. Particularly, locally digested IL-33 releases active 18/21kDa fragments which can contribute to expand the proinflammatory signal. Endothelial (CD31+) and mesenchymal, myofibroblast and pericyte cells from microvascular structures in villi and crypts, showed IL-33 nuclear location; while B cells (CD20+) showed a strong cytoplasmic staining. Both ST2 forms, ST2L and sST2, were also upregulated in duodenal mucosa of CD patients. This was accompanied by increased number of CD8+ST2+ T cells and the expression of T-bet in some ST2+ intraepithelial lymphocytes and lamina propria cells. IL-33 and sST2 mRNA levels correlated with IRF1, an IFN induced factor relevant in responses to viral infections and interferon mediated proinflammatory responses highly represented in duodenal tissues in ACD. These findings highlight the potential contribution of IL-33 and its fragments to exacerbate the proinflammatory circuit and potentiate the cytotoxic activity of CD8+ T cells in CD pathology.

Treatment Adherence in Inflammatory Bowel Disease Patients from Argentina: A Multicenter Study.

METHODS: A multicenter cross-sectional study involving seven referral centers from three cities of Argentina was undertaken. Patients with a diagnosis of ulcerative colitis (UC), Crohn's disease (CD), or indeterminate colitis (IBDU/IC) were invited to answer an anonymous survey, which included a 5-point Likert scale to evaluate adherence to therapies. Independent variables associated with inadequate adherence were evaluated. RESULTS: Overall, 447 UC/IBDU and 135 CD patients were enrolled. Median age was 37 years (range 21-72); 39.8% were male; median time from diagnosis was 6 years (0.5-35). 91.4% were under treatment with at least one oral medication; 50.3% of patients reported inadequate adherence to oral medications. Patients with UC/IBDU had a lower risk of inadequate adherence when compared to patients with CD (OR 0.57 (0.37-0.87)). 21.8% reported inadequate adherence to biologics; subcutaneous administration was significantly associated with inadequate adherence to biologics (OR 4.8 (1.57-14.66)). CONCLUSION: Inadequate treatment adherence is common among patients with IBD, and potentially modifiable factors were identified.