Together, these results suggest a role for endogenous endocannabinoid signalling in regulation of endocrine secretion in the human pancreas.

Interleukin-6 (IL-6) has emerged as an important mediator of fatty acid metabolism with paradoxical effects in the liver. Administration of IL-6 has been reported to confer protection against steatosis, but plasma and tissue IL-6 concentrations are elevated in chronic liver diseases, including fatty liver diseases associated with obesity and alcoholic ingestion. In this study, we further investigated the role of IL-6 on steatosis induced through a high-fat diet (HFD) in wild-type (WT) and IL-6-deficient (IL-6−/−) mice. Additionally, HFD-fed IL-6−/− mice were also chronically treated with recombinant IL-6 (rIL-6). Obesity in WT mice fed a HFD associated with elevated serum IL-6 levels, fatty liver, upregulation of carnitine palmitoyltransferase 1 (CPT1) and signal transducer and activator of transcription-3 (STAT3), increased AMP kinase phosphorylation (p-AMPK), and downregulation of the hepatic lipogenic enzymes fatty acid synthase (FAS) and stearoyl-CoA desaturase 1 (SCD1). The HFD-fed IL-6−/− mice showed severe steatosis, no changes in CPT1 levels or AMPK activity, no increase in STAT3 amounts, inactivated STAT3, and marked downregulation of the expression of acetyl-CoA carboxylase (ACCα/β), FAS and SCD1. The IL-6 chronic replacement in HFD-fed IL-6−/− mice restored hepatic STAT3 and AMPK activation but also increased the expression of the lipogenic enzymes ACCα/β, FAS and SCD1. Furthermore, rIL-6 administration was associated with aggravated steatosis and elevated fat content in the liver. We conclude that, in the context of HFD-induced obesity, the administration of rIL-6 might contribute to the aggravation of fatty liver disease through increasing lipogenesis.

We have examined the effects of inactivation of the p53 tumor suppressor gene on the incidence of apoptotic cell death in two stages of the adenoma-to-carcinoma progression in the intestine: in early adenomas where p53 mutations are rare and in highly dysplastic adenomas where loss of p53 occurs frequently. Homozygosity for an inactivating germline mutation of p53 had no effect on the incidence or the rate of progression of Apc Min͞؉ -induced adenomas in mice and also did not affect the frequency of apoptosis in the cells of these adenomas. To examine the effect of p53 loss on apoptosis in late-stage adenomas, we compared the incidence of apoptotic cell death before and after the appearance of highly dysplastic cells in human colonic adenomas. The appearance of highly dysplastic cells, which usually coincides during colon tumor progression with loss of heterozygosity at the p53 locus, did not correlate with a reduction in the incidence of apoptosis. These studies suggest that p53 is only one of the genes that determine the incidence of apoptotic in colon carcinomas and that wild-type p53 retards the progression of many benign colonic adenoma to malignant carcinomas by mechanism(s) other than the promotion of apoptosis.

The majority of colorectal cancers develop from adenomatous polyps under the influence of factors that are still poorly understood. Tumourigenesis is generally considered a multistep process in which multiple genetic alterations occur, eventually reflected in abnormalities of the cellular DNA content. Macroscopical features such as tumour size and tumour architecture (tubular, tubulovillous, or villous) are correlated wit the chance of malignancy in the lesion. Grade of dysplasia can be considered an indicator for the level of progression of the adenoma towards invasive carcinoma. These characteristics were correlated with the presence or absence of K‐ras mutations and the DNA ploidy in a prospective study performed on 46 large sporadic colorectal adenomas resected by endoscopy. DNA ploidy and K‐ras mutations were analysed in two samples taken at distant sites in the adenomas. Aneuploidy was present in 12 adenomas (26 per cent) and K‐ras mutations occurred in 26 (57 per cent). A highly significant correlation was found between aneuploidy and adenoma size, architecture, and grade of dysplasia. The presence of K‐ras mutations was significantly correlated only with the size of the adenomas. The proportion of adenomas with aneuploidy and/or a K‐ras mutation increased when two samples were analysed instead of one. This observation suggests that the prevalence of genetic mutations and of aneuploidy is probably underestimated, as generally only one sample is investigated. No correlation was observed between K‐ras mutations and ploidy. This study demonstrates the presence of genetic heterogeneity in colorectal adenomas and supports the notion that K‐ras mutation is an early event, while aneuploidy is a late event in the adenoma–carcinoma sequence. © 1997 John Wiley & Sons, Ltd.

Protective mechanisms against drug-induced liver injury are actively being searched to identify new therapeutic targets. Among them, the anti-inflammatory N-acyl ethanolamide (NAE)-peroxisome proliferators activated receptor alpha (PPARα) system has gained much interest after the identification of its protective role in steatohepatitis and liver fibrosis. An overdose of paracetamol (APAP), a commonly used analgesic/antipyretic drug, causes hepatotoxicity, and it is being used as a liver model. In the present study, we have analyzed the impact of APAP on the liver NAE-PPARα system. A dose-response (0.5–5–10–20 mM) and time-course (2–6–24 h) study in human HepG2 cells showed a biphasic response, with a decreased PPARα expression after 6-h APAP incubation followed by a generalized increase of NAE-PPARα system-related components (PPARα, NAPE-PLD, and FAAH), including the NAEs oleoyl ethanolamide (OEA) and docosahexaenoyl ethanolamide, after a 24-h exposure to APAP. These results were partially confirmed in a time-course study of mice exposed to an acute dose of APAP (750 mg/kg). The gene expression levels of Pparα and Faah were decreased after 6 h of treatment and, after 24 h, the gene expression levels of Nape-pld and Faah, as well as the liver levels of OEA and palmitoyl ethanolamide, were increased. Repeated APAP administration (750 mg/kg/day) up to 4 days also decreased the expression levels of PPARα and FAAH, and increased the liver levels of NAEs. A resting period of 15 days completely restored these impairments. Liver immunohistochemistry in a well-characterized human case of APAP hepatotoxicity confirmed PPARα and FAAH decrements. Histopathological and hepatic damage (Cyp2e1, Caspase3, αSma, Tnfα, and Mcp1)-related alterations observed after repeated APAP administration were aggravated in the liver of Pparα-deficient mice. Our results demonstrate that the anti-inflammatory NAE-PPARα signaling system is implicated in liver toxicity after exposure to APAP overdose, and may contribute to its recovery through a long-term time-dependent response.

High-fat diet-induced obesity (DIO) is associated with fatty liver and elevated IL-6 circulating levels. IL-6 administration in rodents has yielded contradictory results regarding its effects on steatosis progression. In some models of fatty liver disease, high doses of human IL-6 ameliorate the liver steatosis, whereas restoration of IL-6 in DIO IL-6-/- mice up-regulates hepatic lipogenic enzymes and aggravates steatosis. We further examined the effects of chronic low doses of murine IL-6 on hepatic lipid metabolism in WT mice in DIO. IL-6 was delivered twice daily in C57BL/6J DIO mice for 15 days. The status and expression of IL-6-signalling mediators and targets were investigated in relation to the steatosis and lipid content in blood and in liver. IL-6 administration in DIO mice markedly raised circulating levels of lipids, glucose and leptin, elevated fat liver content and aggravated steatosis. Under IL-6 treatment there was hepatic Stat3 activation and increased gene expression of Socs3 and Tnf-alpha whereas the gene expression of endogenous IL-6, IL-6-receptor, Stat3, Cpt1 and the enzymes involved in lipogenesis was suppressed. These data further implicate IL-6 in fatty liver disease modulation in the context of DIO, and indicate that continuous stimulation with IL-6 attenuates the IL-6-receptor response, which is associated with high serum levels of leptin, glucose and lipids, the lowering levels of lipogenic and Cpt1 hepatic enzymes and with increased Tnf-alpha hepatic expression, a scenario evoking that observed in IL-6-/- mice exposed to DIO and in obese Zucker rats.

De novo lipogenesis and hypercaloric diets are thought to contribute to increased fat mass, particularly in abdominal fat depots. CB1 is highly expressed in adipose tissue, and CB1-mediated signalling is associated with stimulation of lipogenesis and diet-induced obesity, though its contribution to increasing fat deposition in adipose tissue is controversial. Lipogenesis is regulated by transcription factors such as liver X receptor (LXR), sterol-response element binding protein (SREBP) and carbohydrate-responsive-element-binding protein (ChREBP). We evaluated the role of CB1 in the gene expression of these factors and their target genes in relation to lipogenesis in the perirenal adipose tissue (PrAT) of rats fed a high-carbohydrate diet (HCHD) or a high-fat diet (HFD). Both obesity models showed an up-regulated gene expression of CB1 and Lxrα in this adipose pad. The Srebf-1 and ChREBP gene expressions were down-regulated in HFD but not in HCHD. The expression of their target genes encoding for lipogenic enzymes showed a decrease in diet-induced obesity and was particularly dramatic in HFD. In HCHD, CB1 blockade by AM251 reduced the Srebf-1 and ChREBP expression and totally abrogated the remnant gene expression of their target lipogenic enzymes. The phosphorylated form of the extracellular signal-regulated kinase (ERK-p), which participates in the CB1-mediated signalling pathway, was markedly present in the PrAT of obese rats. ERK-p was drastically repressed by AM251 indicating that CB1 is actually functional in PrAT of obese animals, though its activation loses the ability to stimulate lipogenesis in PrAT of obese rats. Even so, the remnant expression levels of lipogenic transcription factors found in HCHD-fed rats are still dependent on CB1 activity. Hence, in HCHD-induced obesity, CB1 blockade may help to further potentiate the reduction of lipogenesis in PrAT by means of inducing down-regulation of the ChREBP and Srebf-1 gene expression, and consequently in the expression of lipogenic enzymes.

Lipogenesis is intimately controlled by hormones and cytokines as well as nutritional conditions. IL-6 participates in the regulation of fatty acid metabolism in the liver. We investigated the role of IL-6 in mediating fasting/re-feeding changes in the expression of hepatic lipogenic enzymes. EXPERIMENTAL APPROACHGene and protein expression of lipogenic enzymes were examined in livers of wild-type (WT) and IL-6-deficient (IL-6 À/À ) mice during fasting and re-feeding conditions. Effects of exogenous IL-6 administration on gene expression of these enzymes were evaluated in vivo. The involvement of STAT3 in mediating these IL-6 responses was investigated by using siRNA in human HepG2 cells. KEY RESULTSDuring feeding, the up-regulation in the hepatic expression of lipogenic genes presented similar time kinetics in WT and IL-6 À/À mice. During fasting, expression of lipogenic genes decreased gradually over time in both strains, although the initial drop was more marked in IL-6 À/À mice. Protein levels of hepatic lipogenic enzymes were lower in IL-6 À/À than in WT mice at the end of the fasting period. In WT, circulating IL-6 levels paralleled gene expression of hepatic lipogenic enzymes. IL-6 administration in vivo and in vitro showed that IL-6-mediated signalling was associated with the up-regulation of hepatic lipogenic enzyme genes. Moreover, silencing STAT3 in HepG2 cells attenuated IL-6 mediated up-regulation of lipogenic gene transcription levels. CONCLUSIONS AND IMPLICATIONSIL-6 sustains levels of hepatic lipogenic enzymes during fasting through activation of STAT3. Our findings indicate that clinical use of STAT3-associated signalling cytokines, particularly against steatosis, should be undertaken with caution. AbbreviationsACC, (Accα/β, Acaca/b) acetyl-CoA carboxylase; FAS, (Fasn) fatty acid synthase; rIL-6, recombinant IL-6; SCD1, (Scd1, Scd1) stearoyl-CoA desaturase-1; STAT3, (Stat3, Stat3) signal transducer and activator of transcription factor 3; Srebp-1c, (Srebp-1c) sterol regulatory element-binding proteins-1c