Identification of specific gene expression signatures characteristic of oncogenic pathways is an important step toward molecular classification of human malignancies. Aberrant activation of the Met signaling pathway is frequently associated with tumor progression and metastasis. In this study, we defined the Met-dependent gene expression signature using global gene expression profiling of WT and Met-deficient primary mouse hepatocytes. Newly identified transcriptional targets of the Met pathway included genes involved in the regulation of oxidative stress responses as well as cell motility, cytoskeletal organization, and angiogenesis. To assess the importance of a Met-regulated gene expression signature, a comparative functional genomic approach was applied to 242 human hepatocellular carcinomas (HCCs) and 7 metastatic liver lesions. Cluster analysis revealed that a subset of human HCCs and all liver metastases shared the Met-induced expression signature. Furthermore, the presence of the Met signature showed significant correlation with increased vascular invasion rate and microvessel density as well as with decreased mean survival time of HCC patients. We conclude that the genetically defined gene expression signatures in combination with comparative functional genomics constitute an attractive paradigm for defining both the function of oncogenic pathways and the clinically relevant subgroups of human cancers.

Background & Aims The cancer stem cells (CSCs) have important therapeutic implications for multi-resistant cancers including hepatocellular carcinoma (HCC). Among the key pathways frequently activated in liver CSCs is NF-kB signaling. Methods We evaluated the CSCs-depleting potential of NF-kB inhibition in liver cancer achieved by the IKK inhibitor curcumin, RNAi and specific peptide SN50. The effects on CSCs were assessed by analysis of Side Population (SP), sphere formation and tumorigenicity. Molecular changes were determined by RT-qPCR, global gene expression microarray, EMSA, and Western blotting. Results HCC cell lines exposed to curcumin exhibited differential responses to curcumin and were classified as sensitive and resistant. In sensitive lines, curcumin-mediated induction of cell death was directly related to the extent of NF-kB inhibition. The treatment also led to a selective CSC-depletion as evidenced by a reduced SP size, decreased sphere formation, down-regulation of CSC markers and suppressed tumorigenicity. Similarly, NF-kB inhibition by SN50 and siRNA against p65 suppressed tumor cell growth. In contrast, curcumin-resistant cells displayed a paradoxical increase in proliferation and expression of CSC markers. Mechanistically, an important component of the CSC-depleting activity of curcumin could be attributed to a NF-kB-mediated HDAC inhibition. Co-administration of the class I/II HDAC inhibitor trichostatine sensitized resistant cells to curcumin. Further, integration of a predictive signature of curcumin sensitivity with human HCC database indicated that HCCs with poor prognosis and progenitor features are most likely to benefit from NF-kB inhibition. Conclusions These results demonstrate that blocking NF-kB can specifically target CSC populations and suggest a potential for combined inhibition of NF-kB and HDAC signaling for treatment of liver cancer patients with poor prognosis.

Hepatocyte growth factor (HGF) has been reported to have both positive and negative effects on carcinogenesis. Here, we show that the loss of c-Met signaling in hepatocytes enhanced rather than suppressed the early stages of chemical hepatocarcinogenesis. c-Met conditional knockout mice (c-met fl/fl , AlbCre +/À ; MetLivKO) treated with N-nitrosodiethylamine developed significantly more and bigger tumors and with a shorter latency compared with control (w/w, AlbCre +/À ; Cre-Ctrl) mice. Accelerated tumor development was associated with increased rate of cell proliferation and prolonged activation of epidermal growth factor receptor (EGFR) signaling. MetLivKO livers treated with N-nitrosodiethylamine also displayed elevated lipid peroxidation, decreased ratio of reduced glutathione to oxidized glutathione, and up-regulation of superoxide dismutase 1 and heat shock protein 70, all consistent with increased oxidative stress. Likewise, gene expression profiling done at 3 and 5 months after N-nitrosodiethylamine treatment revealed up-regulation of genes associated with cell proliferation and stress responses in c-Met mutant livers. The negative effects of c-Met deficiency were reversed by chronic p.o. administration of antioxidant N-acetyl-L-cysteine. N-acetyl-L-cysteine blocked the EGFR activation and reduced the N-nitrosodiethylamineinitiated hepatocarcinogenesis to the levels of Cre-Ctrl mice. These results argue that intact HGF/c-Met signaling is essential for maintaining normal redox homeostasis in the liver and has tumor suppressor effect(s) during the early stages of Nnitrosodiethylamine-induced hepatocarcinogenesis.

Several countries around the world have faced an important obesity challenge for the past four decades as the result of an obesogenic environment. This disease has a multifactorial origin and it is associated with multiple comorbidities including type 2 diabetes, hypertension, osteoarthritis, metabolic syndrome, cancer, and dyslipidemia. With regard to dyslipidemia, hypertriglyceridemia is a well-known activator of the NLRP3 inflammasome, triggering adipokines and cytokines secretion which in addition induce a systemic inflammatory state that provides an adequate scenario for infections, particularly those mediated by viruses such as HIV, H1N1 influenza, and SARS-CoV-2. The SARS-CoV-2 infection causes the coronavirus disease 2019 (COVID-19) and it is responsible for the pandemic that we are currently living. COVID-19 causes an aggressive immune response known as cytokine release syndrome or cytokine storm that causes multiorgan failure and in most cases leads to death. In the present work, we aimed to review the molecular mechanisms by which obesity-associated systemic inflammation could cause a more severe clinical presentation of COVID-19. The SARS-CoV-2 infection could potentiate or accelerate the pre-existing systemic inflammatory state of individuals with obesity, via the NLRP3 inflammasome activation and the release of pro-inflammatory cytokines from cells trough Gasdermin-pores commonly found in cell death by pyroptosis.

Primary liver cancers represent the second leading cause of cancer-related deaths worldwide. Diverse etiological factors include chronic viral hepatitis, aflatoxin and alcohol exposure as well as aberrant liver lipid overload. Cholesterol has been identified as a key inducer of metabolic impairment, oxidative stress and promoter of cellular dysfunction. The aim of this work was to address the oxidative stress-mediated DNA damage induced by cholesterol overload, and its role in the development of hepatocellular carcinoma.C57BL/6 male mice were fed with a high cholesterol diet, followed by a single dose of N-diethylnitrosamine (DEN, 10 μg/g, ip). Reactive oxygen species generation, DNA oxidation, antioxidant and DNA repair proteins were analyzed at different time points. Diet-induced cholesterol overload caused enhanced oxidative DNA damage in the liver and was associated with a decrease in key DNA repair genes as early as 7 days. Interestingly, we found a cell survival response, induced by cholesterol, judged by a decrement in Bax to Bcl2 ratio. Importantly, N-acetyl-cysteine supplementation significantly prevented DNA oxidation damage. Furthermore, at 8 months after DEN administration, tumor growth was significantly enhanced in mice under cholesterol diet in comparison to control animals. Together, these results suggest that cholesterol overload exerts an oxidative stress-mediated effects and promotes the development of liver cancer.

We applied a genome-wide microarray analysis to three transgenic mouse models of liver cancer in which targeted overexpression of c-Myc, E2f1, and a combination of the two was driven by the albumin promoter. Although gene expression profiles in HCC derived in all three transgenic lines were highly similar, oncogene-specific gene expression signatures were identified at an early dysplastic stage of hepatocarcinogenesis. Overexpression of E2f1 was associated with a strong alteration in lipid metabolism, and Srebp1was identified as a candidate transcription factor responsible for lipogenic enzyme induction. The molecular signature of c-Myc overexpression included the induction of more than 60 genes involved in the translational machinery that correlated with an increase in liver mass. In contrast, the combined activity of c-Myc and E2f1 specifically enhanced the expression of genes involved in mitochondrial metabolism-particularly the components of the respiratory chain-and correlated with an increased ATP synthesis. Thus, the results suggest that E2f1, c-Myc, and their combination may promote liver tumor development by distinct mechanisms. In conclusion, determination of tissue-specific oncogene expression signatures might be useful to identify conserved expression modules in human cancers. Supplementary material for this article can be found on the HEPATOLOGY website (http://interscience.wiley.com/jpages/0270-9139/suppmat/index.html).

Non-alcoholic fatty liver disease (NAFLD) encompasses a broad spectrum of histopathological changes ranging from non-inflammatory intracellular fat deposition to non-alcoholic steatohepatitis (NASH), which may progress into hepatic fibrosis, cirrhosis, or hepatocellular carcinoma. Recent data suggest that impaired hepatic cholesterol homeostasis and its accumulation are relevant to the pathogenesis of NAFLD/NASH. Despite a vital physiological function of cholesterol, mitochondrial dysfunction is an important consequence of dietary-induced hypercholesterolemia and was, subsequently, linked to many pathophysiological conditions. The aim in the current study was to evaluate the morphological and molecular changes of cholesterol overload in mouse liver and particularly, in mitochondria, induced by a high-cholesterol (HC) diet for one month. Histopathological studies revealed microvesicular hepatic steatosis and significantly elevated levels of liver cholesterol and triglycerides leading to impaired liver synthesis. Further, high levels of oxidative stress could be determined in liver tissue as well as primary hepatocyte culture.Transcriptomic changes induced by the HC diet involved disruption in key pathways related to cell death and oxidative stress as well as upregulation of genes related to glutathione homeostasis. Impaired liver function could be associated with a decrease in mitochondrial membrane potential and ATP content and significant alterations in mitochondrial dynamics. We demonstrate that cholesterol overload in the liver leads to mitochondrial changes which may render damaged hepatocytes proliferative and resistant to cell death whereby perpetuating liver damage. K E Y W O R D S apoptosis, cholesterol, mitochondrial dynamics, oxidative stress J Cell Physiol. 2019;234:7213-7223. wileyonlinelibrary.com/journal/jcp Additional supporting information may be found online in the Supporting Information section at the end of the article. How to cite this article: Domínguez-Pérez M, Simoni-Nieves A, Rosales P, et al. Cholesterol burden in the liver induces mitochondrial dynamic changes and resistance to apoptosis.

Non-alcoholic fatty liver disease (NAFLD) and progression to non-alcoholic steatohepatitis (NASH) result as a consequence of diverse conditions, mainly unbalanced diets. Particularly, high-fat and cholesterol content, as well as carbohydrates, such as those commonly ingested in Western countries, frequently drive adverse metabolic alterations in the liver and promote NAFLD development. Lipid liver overload is also one of the main risk factors for initiation and progression of hepatocellular carcinoma (HCC), but detailed knowledge on the relevance of high nutritional cholesterol remains elusive. We were aimed to characterize HCC development in mice fed with a Western diet (high in lipids and cholesterol) and to identify molecular alterations that define a subtype of liver cancer induced by lipid overload. Mice under western or high cholesterol diets more frequently developed tumors with a more aggressive phenotype than animals fed with a chow diet. Associated changes involved macrophage infiltration, angiogenesis, and stemness features. RNA-seq revealed a specific gene expression signature (Slc41a; Fabp5; Igdcc4 and Mthfd1l) resembling the adverse phenotypic features and poor clinical outcomes seen in patients with HCC. In conclusion; consumption of lipid enriched diets; particularly cholesterol; could accelerate HCC development with an aggressive phenotype and poor prognosis