TRACKING A PRIMARY BARRIER DYSFUNCTION IN CELIAC DISEASE Celiac Disease (CeD) is an autoimmune disease that develops in genetically predisposed individuals after the ingestion of gluten. It induces a malabsorption syndrome, commonly provoking diarrhea, weight loss and vitamin deficiency and the only standard treatment so far is a gluten-free diet. Celiac patients present impaired epithelial barrier function with lower TEER and increased permeability to disaccharides. In addition, tight junction strands are discontinuous and decreased in celiac patients. Changes in barrier function are mostly attributed to the immune process, however, it was shown that treated patients may present impaired barrier function, despite the lack of symptoms. Moreover, risk loci for CeD were found in genes related to cell-cell adhesion, including LPP and C1orf106. LPP is involved in focal adhesions formation and E-cadherin cell-cell adhesion. C1orf106 inhibits the degradation of E-cadherin indirectly and its depletion causes reduction in TEER. In this context, we sought out to study the effect of LPP and C1orf106 in barrier function in intestinal cell lines and in patients with CeD. Our results show that cells depleted of LPP and C1orf106 present changes in tight junction protein content and present a reduced ability to re-assemble the tight junctions after a calcium switch assay. In patients, we did not see significant changes regarding LPP or C1orf106 protein content, but further analysis of electrical resistance and RNA may provide further insights into the importance of both proteins in the barrier impairment in CeD.
THE ROLE OF OSTEOPONTIN IN THE PATHOGENESIS OF CAC IBD patients present an increased risk of developing colorectal cancer, namely colitis-associated cancer (CAC). In the case of CAC, the immune response in the IBD plays an important role in tumorigenesis and results in a different progression process than sporadic colorectal carcinoma (CRC). For example, the early mutation of APC seen in CRC does not occur as frequently in CAC and when it does, it is only at the final stages of progressions. On the other hand, p53 mutations occur very early in CAC progression, whereas in CRC it is a late finding. CAC pathogenesis is not as well understood as CRC and there is still much to clarify regarding CAC progression. Then, we decided to study CAC progression in samples from patients who underwent colectomy and performed an RNA analysis for immune-related genes. The results of this experiment showed osteopontin (OPN) as the most upregulated gene in both CAC coming from ulcerative colitis and Crohn’s disease patients, impelling us to investigate it further. OPN was found in both epithelial cells and stromal cells and one of its receptors, CD44, was also identified in both cell compartments, with a tendency for being increased in CAC epithelial cells. OPN is known to promote tumorigenesis in several cancer types, especially solid tumors, and one of its key functions is the induction of epithelial to mesenchymal transition (EMT). Indeed, findings from the RNA analysis and immunohistochemical analysis of the patients’ samples point out to the presence of the EMT process in CAC. When we sought out to study OPN effects in cell lines, OPN activated ERK1/2, but not STAT3, AKT or P-65/NFκB. However, we failed to reproduce EMT by exposing the cells to OPN. Finally, we decided to perform an RNA-Seq analysis of the cells treated with OPN and found changes in mitochondrial respiratory chain, especially downregulation of complexes III and IV. These results suggest a new role for OPN in the tumorigenesis of CAC.