BIOCHEMICAL MECHANISMS UNDERLYING THE CELIAC CELLULAR PHENOTYPE

CELIAC DISEASE (CD) IS AN ENTEROPATHY CHARACTERISED BY AN IMMUNE RESPONSE TO DIETARY GLUTEN. EMERGING EVIDENCE HIGHLIGHTS THE EXISTENCE OF A CONSTITUTIVE CELLULAR PHENOTYPE THAT PERSISTS INDEPENDENTLY OF GLUTEN EXPOSURE. THIS THESIS PROVIDES A COMPREHENSIVE BIOCHEMICAL AND MOLECULAR CHARACTERIZATION OF THIS PHENOTYPE USING, AS STUDY MODELS, PRIMARY FIBROBLASTS DERIVED FROM SKIN AND INTESTINAL BIOPSIES OF CD PATIENTS, COMPARED WITH MATCHED HEALTHY CONTROLS. SKIN FIBROBLASTS FROM CD PATIENTS DISPLAYED REDUCED SENSITIVITY TO THE ER STRESS-INDUCER THAPSIGARGIN, PROLONGED UNFOLDED PROTEIN RESPONSE, AND IMPAIRED AUTOPHAGIC FLUX, LINKED TO ALTERED CALCIUM HOMEOSTASIS AND DECREASED ACTIVATION OF TYPE 2 TRANSGLUTAMINASE 2 (TG2). SIMILAR DEFECTS WERE OBSERVED IN INTESTINAL FIBROBLASTS, INDICATING THAT THESE ALTERATIONS MAY BE SYSTEMIC AND NOT TISSUE RESTRICTED. OMICS APPROACHES FURTHER HIGHLIGHTED SHARED MOLECULAR SIGNATURES IN BOTH MODELS. PROTEOMICS IDENTIFIED AN CD CELLS CONSISTENT DYSREGULATION OF PROTEINS LINKED TO EXTRACELLULAR MATRIX ORGANIZATION, EXOSOMES, VESICULAR TRAFFICKING, CYTOSKELETAL REMODELLING, CA²⁺ HOMEOSTASIS, UPR SIGNALLING, INTERESTINGLY, AMONG PROTEINS DOWNREGULATED IN BOTH CELL SYSTEMS, WE ALSO FOUND TG2, THUS UNDERLINING THE CENTRAL ROLE OF THIS ENZYME IN THE CELIAC PHENOTYPE. IN ADDITION, SEVERAL PROTEINS APPEARED DIFFERENTIALLY PHOSPHORYLATED IN SKIN FIBROBLASTS. ON THE OTHER HAND, A METABOLOMIC ANALYSIS, CONDUCTED ON INTESTINAL FIBROBLASTS, REVEALED DISTINCT ENERGY METABOLISM DEFECTS, INCLUDING ALTERED LEVELS OF GLUCOSE, GLUTAMINE, AND DIHYDROXYACETONE, SUPPORTING IMPAIRED GLYCOLYSIS AND AMINO ACID METABOLISM. FINALLY, TRANSCRIPTOMIC PROFILING OF CD INTESTINAL FIBROBLASTS TREATED WITH THE CLINICALLY RELEVANT IRREVERSIBLE TG2 INHIBITOR ZED1227 COULD PROVIDE NEW INSIGHTS INTO TG2-DEPENDENT TRANSCRIPTIONAL NETWORKS, WITH DIFFERENTIAL EXPRESSION ANALYSIS CURRENTLY UNDERWAY. ALTOGETHER, THESE FINDINGS UNCOVER A MULTI-LAYERED CELLULAR PHENOTYPE IN CD ACROSS DISTINCT TISSUE-DERIVED FIBROBLAST POPULATIONS, COMPRISING ALTERED STRESS RESPONSES, AUTOPHAGY, CA²⁺ HOMEOSTASIS, VESICULAR TRAFFICKING, AND METABOLIC PATHWAYS. IN CONCLUSION, THIS THESIS CONTRIBUTES TO INCREASE KNOWLEDGE ABOUT THE CELIAC CELLULAR PHENOTYPE E MAY PAVE THE WAY FOR THE RESEARCH OF NEW DISEASE BIOMARKERS AND THERAPEUTIC TARGETS.