b Representative Oil Red staining images depicting different pattern of lipid accumulation in HFD-fed WT and TG2?/? mice; on the right, quantification of lipid droplets (percentage of positive area) using Oil Red O staining. which causes the impairment of autophagy/mitophagy, prospects to worsening of disease progression. Data were confirmed by pharmacological inhibition of TG2 in WT animals. In addition, the analysis of human liver samples from NAFLD individuals validated the enzymes involvement in the liver extra fat disease pathogenesis. Our findings strongly suggest that TG2 activation may present safety in the context of NAFLD, therefore representing a novel restorative target for tackling the NAFLD progression. Introduction Non-alcoholic fatty liver disease (NAFLD) is definitely a pathological switch characterized by the build up of fat, called steatosis, which is found at least in 5% of hepatocytes. NAFLD is an progressively recognized condition that has become the most common liver disorder in developed countries, with prevalence estimations around 24% in Europe1. It is closely associated with features of the metabolic syndrome such as obesity, insulin-resistance, type 2 diabetes, and hyperlipidemia2. In individuals with chronic hepatitis C, steatosis has a prevalence of 40C86% and its rate of recurrence varies with genotype; it is more common in genotype 3 illness, where it happens in 73% of individuals, while the prevalence of steatosis in individuals infected WHI-P 154 with additional genotypes WHI-P 154 is around 50%3. NAFLD is definitely a spectrum of disorders, beginning as simple steatosis which can evolve into non-alcoholic steatohepatitis (NASH) and fibrosis, often resulting in cirrhosis and even hepatocellular carcinoma4. The clinical importance of NAFLD and the current lack of effective medications to limit or reverse disease progression in individuals with NASH have aroused great interest and intense investigation into the fundamental mechanisms involved in the diseases development and progression. Hepatic fat build up results from an imbalance between triglycerides acquisition and IkappaBalpha removal5. Classically, the NAFLD physiopathology and progression has been summarized in the two hits hypothesis, WHI-P 154 with the 1st hit becoming steatosis, and the oxidative stress being involved in the second hit leading to the progression to NASH6. A multiple-hit hypothesis is now identified, in which the timing and combination of genetic, external, and intracellular events, rather than the simple sequence of hepatic insults, result in different pathways, which lead to WHI-P 154 steatosis or NASH, respectively7. The enzyme transglutaminase type 2 (TG2) is definitely a part of the cell response evoked by stress conditions, and its deregulation has been demonstrated to be involved in inflammatory and fibrotic diseases8. TG2 is definitely a ubiquitous member of the TG family. Under pathological conditions it can be located in the extracellular matrix (ECM) or in the cell surface in association with the ECM9 as well as with the cytoplasm, where it is mostly soluble; it is also associated, however, with the inner face of the plasma or nuclear membrane10, and in the mitochondria11. TG2 has been implicated in a variety of cellular processes, such as differentiation, cell death, swelling, cell migration, and wound healing12C14. In addition, we have shown that TG2 is an essential component for the proper maturation of autophagosomes under basal and particularly under stressful cellular conditions15. Considering all these findings we explored whether TG2 could be involved in the development of fatty liver disease. To this end the pathogenesis of NAFLD was analyzed in vivo using a TG2-null mouse model exposed to an experimental nutritional induction. Data acquired showed that TG2 deficiency is a key element to limit NAFLD progression. Results Changes in body and liver excess weight To investigate the effect of TG2 in NAFLD progression, wild-type (WT) and TG2?/? C57BL/6 mice were fed with high-fat diet (HFD) for 16 weeks, starting from 6 weeks of age, and the body excess weight increase was monitored once a week. At the the start of the diet the knockout (KO) mice experienced a slightly lower body excess weight compared to the WT (Fig.?1a), however at the end of the 16 weeks of the treatment they gained excess weight much like WT mice (Fig.?1a). As expected, HFD feeding promoted obesity, with significant weight gain as compared WHI-P 154 to settings for both WT and TG2?/? animals. Interestingly, after 16 weeks of diet, WT mice gained 68% of their unique body weight vs 120% in the case of KO mice (p?