engleski

Modulation of TFF genes and its genetic response in animal models and humans

Members of the trefoil factor (TFF) family are small mucin-associated peptides that are predominantly expressed along the entire length of gastrointestinal (GI) tract. Trefoils play important role in mucosal defense through both protective and reparative mechanisms. The presented thesis consists of two parts. First part deals with the role of trefoils in Inflammatory Bowel Disease (IBD), a specific GI disorder coming in two forms as ulcerative colitis or as a Crohn’ s disease. Since TNF- and NF B deregulation is hallmark of IBD, we monitored their effect on trefoils expression. Our data indicated that in the HT-29 colon cell line, TNF- , through activation of NF B transcription factor, down-regulated the TFF3 gene by transcriptional repression. In the rat model of colitis, inflammation and NF B activation were associated with strong reduction of TFF3 expression. Our in vitro and in vivo data imply that TNF- causes NF B mediated transcriptional repression of the TFF3 gene. Reduction of intestinal TFF3 level due to NF B activation partially explains the impairment of wound healing in IBD. These data are presented in the manuscript “ TNF- and NF B inhibit transcription of human TFF3 encoding a gastrointestinal healing peptide” . Since several NF B consensus binding sites are present in promoter of TFF3 gene, by mutating them we wanted to elucidate which one is responsible for NF B binding and thus regulation of TFF3 expression. We found that mutation of -97 site display an effect by recovering the TFF3 expression by twofold. Additionally this site was identified as site of NF- B binding by electrophoretic mobility shift assay (EMSA). This data are subject of enclosed manuscript “ Transcriptional control of the intestinal trefoil factor (TFF3) via promoter binding sites for the nuclear factor  B and C/EBP ” . The second part of this thesis investigates the role of TFF2. For that purpose Tff2 deficient mice, generated by a member of our group (Dr. T. Kayademir), was characterized. Since morphological analysis (done by collaborating group of Prof G.Stamp) showed no particular difference in regard to normal wild-type mice we decided to look for differences on level of gene expression. Results of these experiments are presented in the manuscript “ Molecular changes due to Tff2 deficiency” . We compared expression of 12.000 genes in mixture of pyloric antrum/Brunner’ s glands by using gene expression microarrays. Differential expression of several interesting genes was confirmed by quantitative qPCR, Northern blot or by immuno-histochemistry. Tff2 deficient mice showed up-regulation of different genes crucial for innate and adaptive immunity. Our data, connecting Tff2 deficiency with differential expression of immune specific genes, prove indirect interplay of TFF2 with immune response. Results of these experiments are presented in the manuscript:” Genetic responses to Tff2 deficiency in murine digestive tract” .

TFF peptides; regulation of gene expression; TFF2 knock out mice

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