Background & Aims Despite high morbidity and mortality of alcoholic liver disease worldwide, the molecular mechanisms underlying alcohol-induced liver cell death are not fully understood. Transglutaminase 2 (TG2) is a cross-linking enzyme implicated in apoptosis. TG2 levels and activity are increased in association with various types of liver injury. However, how TG2 induces hepatic apoptosis is not known. Methods Human hepatic cells or primary hepatocytes from rats or TG2+/+ and TG2−/− mice were treated with ethanol. Mice were administered anti-Fas antibody or alcohol. Liver sections were prepared from patients with alcoholic steatohepatitis. Changes in TG2 levels, Sp1 cross-linking and its activities, expression of hepatocyte growth factor receptor, c-Met, and hepatic apoptosis were measured. Results Ethanol induced apoptosis in hepatic cells, enhanced activity and nuclear accumulation of TG2 as well as accumulation of cross-linked and inactivated Sp1, and reduced expression of the Sp1-responsive gene, c-Met. These effects were rescued by TG2 knockdown, restoration of functional Sp1, or addition of hepatocyte growth factor, whereas apoptosis was reproduced by Sp1 knockdown or TG2 overexpression. Compared with TG2+/+ mice, TG2−/− mice showed markedly reduced hepatocyte apoptosis and Sp1 cross-linking following ethanol or anti-Fas treatment. Treatment of TG2+/+ mice with the TG2 inhibitors putrescine or cystamine blocked anti-Fas–induced hepatic apoptosis and Sp1 silencing. Moreover, enhanced expression of cross-linked Sp1 and TG2 was evident in hepatocyte nuclei of patients with alcoholic steatohepatitis. Conclusions TG2 induces hepatocyte apoptosis via Sp1 cross-linking and inactivation, with resultant inhibition of the expression of c-Met required for hepatic cell viability.

Non-alcoholic steatohepatitis (NASH), a progressive form of fatty liver, shares histological similarities with alcoholic steatohepatitis (ASH), including accumulated fat, hepatic apoptosis, and fibrous tissues in the liver, but the molecular mechanisms responsible for hepatic apoptosis remain unclear. We previously reported that transglutaminase 2 (TG2), induced in the nuclei of ethanol-treated hepatocytes, crosslinks and inactivates the transcription factor Sp1, leading to hepatic apoptosis. In this study, we investigated whether a similar change is involved in NASH, and if so, how TG2 and crosslinked Sp1 (CLSp1) are induced. Elevated nuclear TG2 and CLSp1 formation was demonstrated in NASH patients, as well as increased activation of apoptosis inducing factor (AIF) and release of cytochrome c. In Hc human normal hepatocytes treated with free fatty acids (FFAs), biochemical analyses revealed that ethanol and FFAs provoked fat accumulation, endoplasmic reticulum (ER) stress, increased nuclear factor kappa B (NFκB), and nuclear TG2. Salubrinal, a selective inhibitor of the ER stress-induced pancreatic ER kinase (PERK) signaling pathway, inhibited NFκB activation, nuclear TG2 expression, and apoptosis only if it was induced by FFAs, but not by ethanol. These results suggest that FFAs could increase ER stress and lead to nuclear NFκB activation and TG2 induction through PERK-dependent pathways, resulting in TG2-mediated apoptosis accompanying crosslinking and inactivation of Sp1, activation of AIF, and release of cytochrome c.

Pancreatic cancer is a highly aggressive and notoriously difficult to treat. As the vast majority of patients are diagnosed at advanced stage of the disease, only a small population is curative by surgical resection. Although gemcitabine-based chemotherapy is typically offered as standard of care, most patients do not survive longer than 6 months. Thus, new therapeutic approaches are needed. Pancreatic cancer cells that develop gemcitabine resistance would still be suitable targets for immunotherapy. Therefore, one promising treatment approach may be immunotherapy that is designed to target pancreatic-cancer-associated antigens. In this paper, we detail recent work in immunotherapy and the advances in concept of combination therapy of immunotherapy and chemotherapy. We offer our perspective on how to increase the clinical efficacy of immunotherapies for pancreatic cancer.

Byproducts of cytokine activation are sometimes useful as surrogate biomarkers for monitoring cytokine generation in patients. Transforming growth factor (TGF)-β plays a pivotal role in pathogenesis of hepatic fibrosis. TGF-β is produced as part of an inactive latent complex, in which the cytokine is trapped by its propeptide, the latency-associated protein (LAP). Therefore, to exert its biological activity, TGF-β must be released from the latent complex. Several proteases activate latent TGF-β by cutting LAP. We previously reported that Camostat Mesilate, a broad spectrum protease inhibitor, which is especially potent at inhibiting plasma kallikrein (PLK), prevented liver fibrosis in the porcine serum-induced liver fibrosis model in rats. We suggested that PLK may work as an activator of latent TGF-β during the pathogenesis of liver diseases in the animal models. However, it remained to be elucidated whether this activation mechanism also functions in fibrotic liver in patients.Here, we report that PLK cleaves LAP between R58 and L59 residues. We have produced monoclonal antibodies against two degradation products of LAP (LAP-DP) by PLK, and we have used these specific antibodies to immunostain LAP-DP in liver tissues from both fibrotic animals and patients.The N-terminal side LAP-DP ending at R58 (R58 LAP-DP) was detected in liver tissues, while the C-terminal side LAP-DP beginning at L59 (L59 LAP-DP) was not detectable. The R58 LAP-DP was seen mostly in α-smooth muscle actin-positive activated stellate cells.These data suggest for the first time that the occurrence of a PLK-dependent TGF-β activation reaction in patients and indicates that the LAP-DP may be useful as a surrogate marker reflecting PLK-dependent TGF-β activation in fibrotic liver both in animal models and in patients.Electronic supplementary materialThe online version of this article (doi:10.1186/2193-1801-3-221) contains supplementary material, which is available to authorized users.