6). We next analysed whether the signalling pathways identified by in-vitro

assays on cell lines also operate in intestinal tissue. Basal TG2 expression was detected in healthy tissue, but levels were significantly higher in samples from untreated CD patients (Fig. 7). Incubation with TNF-α + IFN-γ induced TG2 expression in biopsy samples from both CD patients and control individuals. Therefore, TG2 is expressed physiologically in healthy mucosal tissue and is increased in intestinal mucosa of untreated CD patients, as a consequence of the proinflammatory environment in intestinal mucosa Tamoxifen purchase in active CD, due mainly to abundant IFN-γ, a key player in the pathogenic mechanism of CD. Because IkBα is a key negative modulator of the NF-κB pathway, inactivation of IkBα by cross-linking induced strongly by TG2 activates NF-κB. Consequently, TG2 and NF-κB can enhance each other’s actions amplifying the inflammatory cycle [11]. Interestingly, constitutive NF-κB activation often accompanies increased TG2 expression in inflammatory disease such as inflammatory bowel disease, rheumatoid arthritis and coeliac disease [6,7,26,27]. Similar to NF-κB, other transcription factors that activate TG2 also induce the production

of proinflammatory cytokines characteristically present in the intestinal mucosa of untreated CD patients. Therefore, NF-κB, as a key element together with HDAC inhibitor others transcription factors, may exacerbate a complex vicious circle of inflammation through interactions with TG2. Induction of these inflammatory pathways drives activation and recruitment of effector cells, such as neutrophils, macrophages, dendritic cells and T cells, which cause and amplify the pathogenic mechanisms of different chronic disorders [7,28,29]. Based on our results obtained by qRT–PCR, flow cytometry and Western blotting, we propose a model for the signalling pathways activated by TNF-α and IFN-γ involved in the regulation of TG2 expression. As a consequence of dysregulation

of TG2 expression and activity, a distinct pathogenic process may be initiated (Fig. 8). Therapeutic approaches aimed to modulate the activity of TG2 are being taken into consideration as a way to reduce, or even cancel, ADP ribosylation factor the disease processes in which the enzyme is involved [15,30]. This study on TG2 gene regulation provides useful information for the development of new therapeutic strategies to down-modulate chronic inflammatory disorders. The authors declare no conflict of interest. Fig. S1. Dose–response curve of transglutaminase 2 (TG2) induction by tumour necrosis factor (TNF)-α and interferon (IFN)-γ. Human acute monocytic leukaemia cell line (THP-1) cells were incubated for 24 h with different concentrations of TNF-α and/or IFN-γ as indicated. Grey bars correspond to the cytokine concentration selected for further studies; TNF-α (10 ng/ml) and IFN-γ (200 UI/ml). Fig. S2.