Transglutaminases: new target molecules for inflammatory bowel disease?

T he family of transglutaminases (TG) includes the plasma form factor XIIIa as well as the tissue transglutaminase (tTG) and keratinocyte transglutaminase (TGk). In particular, tTG reminds every gastroenterologist primarily of coeliac disease where tTG represents the key autoantigen. Disease induction is confined to tTG, a ubiquitous enzyme which is released from fibroblasts, endothelial, and inflammatory cells during mechanical irritation or inflammation. At acidic pH, which occurs with inflammation, tTG can, apart from its physiological function described in more detail below, also simply deamidate some of the glutamine residues of the gluten peptides. In coeliac disease, deamidation introduces a negative charge into the gluten peptides which can increase the binding affinity to HLA-DQ2 or HLA-DQ8, the primary HLA association in coeliac disease. Binding of gluten peptide to either HLA-DQ2 or HLA-DQ8 results in an increase in their capacity to stimulate T cells, thus inducing intestinal inflammation. 3 As indicated in this issue of Gut by D’Argenio and colleagues, TG may be involved in intestinal inflammation in a completely different way and may even represent a therapeutic target (see page 496). But how can the physiological function of TG be defined? TG are enzymes catalysing the covalent cross linking between proteins by forming e(c-glutamyl)-lysine isopeptide bonds. These bonds are formed between glutaminase and lysine residues thus stabilising intraand extracellular proteins, a process which is required for a variety of essential physiological purposes such as barrier function in epithelia, apoptosis, and extracellular matrix formation. Substrates include intracellular (actin cytoskeleton, histones) as well as extracellular (collagen, vitronectin, fibronectin) proteins many of which are found in the matrix of the colonic mucosa. The balance of cross linking alters the susceptibility of the matrix to proteolysis and thus affects the rate of its formation and remodelling during mucosal damage and repair. In the work presented in this issue of Gut, D’Argenio and colleagues hypothesise that due to impaired healing in ulcerative colitis, an imbalance of TG may participate in the inflammatory process. To approach this question, peripheral blood as well as colonic biopsies from patients with active and inactive ulcerative colitis as well as noninflamed control samples were examined for the three members of the TG family, tTG, factor XIIIa, and TGk. Endoscopic and histological disease indices were applied to determine disease activity. Factor XIIIa activity was evaluated in plasma. In addition, tTG, TGk, and factor XIIIa protein as well as RNA content in colonic tissue was analysed by western blot and reverse transcription-polymerase chain reaction, respectively. An elegant tool in this study proved to be the immunohistochemical studies visualising colonic localisation of TG and their reaction products, the e(c-glutamyl)-lysine isopeptide bonds. With the results obtained, a scheme can be suggested as to how TG act in active ulcerative colitis. Primarily, factor XIIIa activity in plasma is significantly reduced in active disease compared with inactive ulcerative colitis or healthy controls. Immunohistochemical analysis indicated that tTG and factor XIIIa colocalise in damaged areas with the isopeptide bonds, which may explain the decrease in factor XIIIa in plasma. The TGk protein as well as RNA expression is significantly reduced in active disease in colonic tissue. tTG RNA is upregulated while tTG protein expression remains unaltered. Interestingly, in active disease, tTG appeared with two bands in the western blot analysis representing degradation probably due to endogenous proteolysis by calpain, an enzyme that inactivates tTG, known to be increased in active ulcerative colitis. Importantly, TGk was detected for the first time in colonic tissue where it was localised at the upper part of the crypts and it was shown that TGk was significantly downregulated in active disease. Based on these results the authors conclude that the abnormal pattern of TG contributes to the course of ulcerative colitis. Whether or not these changes are specific for ulcerative colitis or whether they also occur in other intestinal inflammations, for example, infectious colitis, is unknown. Similar data have been obtained for Crohn’s disease. 8 Here, in active disease, factor XIIIa was equally decreased in serum, and immunhistochemical staining also showed colocalisation of factor XIIIa and tTG to the extracellular matrix. Interestingly, the western blot from colonic tissue from patients with active Crohn’s disease showed an identical double band in active disease, an observation that was not discussed further by the authors. These data indicate that independent of the type of chronic inflammatory bowel disease, a similar pattern of TG changes can be seen. These data as well as data from the literature indicate a protective role of TG in Crohn’s disease referred to above. This protective function is already evident given the functional properties of TG: TGk is abundantly expressed in epithelial cells cross linking a series of defined structural proteins, thus exerting barrier function. Consequently, the decrease in TGk is associated with a barrier defect, a well described phenomenon in intestinal inflammation. tTG serves as G protein in most mammalian cells; when Ca levels rise this protein becomes active in cross linking TG reactions. The potential protective effect is supported by the observation that inhibition of calpain, which can inactivate tTG, reduced colonic injury in experimental colitis. Thus wound healing or tissue remodelling is an active process consisting of a destructive part, including extracellular proteinases such as metalloproteinases or calpain, and a constructive part involving processing enzymes such as TG. During acute inflammation the presence of extracellular matrix proteins increases, thus inactivating at least part of the TG, as indicated in the present study as well as in the study referred to above in Crohn’s disease patients. Additional experimental data support this theory. Na-butyrate enemas have been demonstrated to ameliorate experimental colitis. Nabutyrate is known to upregulate tTG expression. Furthermore, a case report including three patients demonstrated a beneficial effect for recombinant factor XIII concentrate in patients with inflammatory bowel disease. 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