Interleukin-17 programs liver progenitor cell transformation into cancer stem cells through miR-122 downregulation with increased risk of primary liver cancer initiation.

Chronic inflammation is a key component in the development of virtually all types of primary liver cancers. However, how chronic inflammation potentiates or even may initiate liver parenchymal cell transformation remains unclear. Cancer stem cells (CSCs) represent an exciting target for novel anticancer therapeutic strategies in several types of cancers and were also described in primary liver cancers as tumor initiating cells. Recently, we reported a key role of Interleukin (IL)-17 in Liver Progenitor Cell (LPC) accumulation in preneoplastic cirrhotic livers. In this study, we evidenced in vitro, that long-term stimulation of LPCs with IL-17 led to their transformation into CSCs. Indeed, they acquired CSC-marker expression, and self-renewal properties, showed by their increased capacity to form spheroids. The miRNome analysis revealed that long-term IL-17 treatment of LPCs led to a 90% decrease in miR-122 expression. In a model using immunodeficient mice, ectopic engraftment of LPCs in an IL-17-enriched environment led to tumor occurrence with an aggressive phenotype. Contrastingly, in a murine model of hepatocellular carcinoma induced by a unique injection of diethyl-nitrosamine associated with chronic administration of carbon tetrachloride, IL-17-deficiency or anti-IL-17 therapy protected mice from liver tumor growth. In conclusion, we showed that a chronic exposure of LPCs to IL-17 cytokine promotes their transformation into CSCs. In addition, we demonstrated that IL-17-neutralizing strategies limit CSC occurrence and liver tumor progression through miR-122 restored-expression.

Autofluorescence imaging within the liver: a promising tool for the detection and characterization of primary liver tumors.

OBJECTIVES: To assess the performance of 405 nm-induced autofluorescence for the characterization of primary liver nodules on ex vivo resected specimens. MATERIALS AND METHODS: Forty resected liver specimens bearing 53 primary liver nodules were included in this IRB-approved prospective study. Intratissular spectroscopic measurements were performed using a 25-G fibered-needle on all ex vivo specimens: 5 autofluorescence measurements were performed in both nodules and adjacent parenchyma. The spectra derivatives of the 635 and 670 nm autofluorescence peaks observed in nodules and in adjacent liver parenchyma were compared (Kruskal-Wallis and Mann-Whitney when appropriate). RESULTS: A total of 42 potentially evolutive primary liver nodules-34 hepatocellular carcinomas, 4 intrahepatic cholangiocarcinomas, 4 hepatocellular adenomas-and 11 benign nodules-5 focal nodular hyperplasias, 6 regenerative nodules-were included. Both 635 and 670 nm Δderivatives were significantly higher in benign as compared to potentially evolutive (PEV) nodules (respectively 32.9 ± 4.5 vs 15.3 ± 1.4; p < 0.0001 and 5.7 ± 0.6 vs 2.5 ± 0.1; p < 0.0001) with respective sensitivity and specificity of 78% and 91% for distinguishing PEV from benign nodules. CONCLUSION: 405 nm-induced autofluorescence enables the discrimination of benign from PEV primary liver nodules, suggesting that autofluorescence imaging could be used to optimize US targeted liver biopsies. KEY POINTS: • 405 nm-induced autofluorescence can distinguish liver tumors from the adjacent liver parenchyma. • The analysis of autofluorescence imaging observed within primary liver tumors can discriminate benign tumors from those requiring follow-up or targeted liver biopsy. • In current practice, autofluorescence imaging could be embedded within biopsy needle, to enable, in addition to ultrasound guidance, optimal targeting of liver nodules which could optimize tissue sampling.

Intrahepatic immune changes after hepatitis c virus eradication by direct-acting antiviral therapy.

BACKGROUND & AIMS: The recent approval of direct acting anti-virals (DAA) has dramatically changed the landscape of hepatitis C virus (HCV) therapy. Whether viral clearance could promote liver carcinogenesis is debated. It has been hypothesized that changes in intrahepatic immune surveillance following viral cure could favour tumour growth. This study aimed at characterizing the intrahepatic immune changes induced by HCV cure following DAA therapy. METHODS: Patients with compensated cirrhosis who underwent surgical resection for hepatocellular carcinoma (HCC) after sustained virological response (SVR) to DAA therapy were included. A control group of untreated HCV-infected patients with compensated cirrhosis was selected. RNA was extracted from tumoral and non-tumoral tissues and analysed using the Nanostring Immuno-Oncology-360 panel. Immune cells were quantified by immunohistochemistry. RESULTS: Twenty patients were included: 10 patients with a DAA-induced SVR and 10 untreated controls. All of them had a de novo BCLC 0/A HCC. Non-tumoral tissue profiling showed down-regulation of interferon-related genes (including MX1, ISG15 and IFIT1) after DAA therapy. No other differences in immune profiles/immune cell densities were identified between the two groups. The intra-tumoral immune profiles of HCCs that occurred after DAA therapy were not qualitatively or quantitatively different from those of tumours occurring in untreated patients. CONCLUSION: In conclusion, removal of HCV infection after DAA-based therapy results only in a down-regulation of interferon-stimulated genes in non-tumoral tissues from patients with cirrhosis who develop HCC. These minor changes in the liver immune microenvironment are unlikely to favour HCC occurrence or recurrence after DAA-induced SVR.

Nasal airway epithelial cell IL-6 and FKBP51 gene expression and steroid sensitivity in asthmatic children.

BACKGROUND: Many asthmatic patients exhibit uncontrolled asthma despite high-dose inhaled corticosteroids (ICS). Airway epithelial cells (AEC) have distinct activation profiles that can influence ICS response. OBJECTIVES: A pilot study to identify gene expression markers of AEC dysfunction and markers of corticosteroid sensitivity in asthmatic and non-asthmatic control children, for comparison with published reports in adults. METHODS: AEC were obtained by nasal brushings and primary submerged cultures, and incubated in control conditions or in the presence of 10 ng/ml TNFalpha, 10-8M dexamethasone, or both. RT-PCR-based expression of FKBP51 (a steroid hormone receptor signalling regulator), NF-kB, IL-6, LIF (an IL-6 family neurotrophic cytokine), serpinB2 (which inhibits plasminogen activation and promotes fibrin deposition) and porin (a marker of mitochondrial mass) were determined. RESULTS: 6 patients without asthma (median age 11yr; min-max: 7-13), 8 with controlled asthma (11yr, 7-13; median daily fluticasone dose = 100 µg), and 4 with uncontrolled asthma (12yr, 7-14; 1000 µg fluticasone daily) were included. Baseline expression of LIF mRNA was significantly increased in uncontrolled vs controlled asthmatic children. TNFalpha significantly increased LIF expression in uncontrolled asthma. A similar trend was observed regarding IL-6. Dexamethasone significantly upregulated FKBP51 expression in all groups but the response was blunted in asthmatic children. No significant upregulation was identified regarding NF-kB, serpinB2 and porin. CONCLUSION: LIF and FKBP51 expression in epithelial cells were the most interesting markers of AEC dysfunction/response to corticosteroid treatment.

N-Acetyl-ß-D-glucosaminidase activity in feral Carcinus maenas exposed to cadmium.

Cadmium is a priority hazardous substance, persistent in the aquatic environment, with the capacity to interfere with crustacean moulting. Moulting is a vital process dictating crustacean growth, reproduction and metamorphosis. However, for many organisms, moult disruption is difficult to evaluate in the short term, what limits its inclusion in monitoring programmes. N-acetyl-ß-D-glucosaminidase (NAGase) is an enzyme acting in the final steps of the endocrine-regulated moulting cascade, allowing for the cast off of the old exoskeleton, with potential interest as a biomarker of moult disruption. This study investigated responses to waterborne cadmium of NAGase activity of Carcinus maenas originating from estuaries with different histories of anthropogenic contamination: a low impacted and a moderately polluted one. Crabs from both sites were individually exposed for seven days to cadmium concentrations ranging from 1.3 to 2000 µg/L. At the end of the assays, NAGase activity was assessed in the epidermis and digestive gland. Detoxification, antioxidant, energy production, and oxidative stress biomarkers implicated in cadmium metabolism and tolerance were also assessed to better understand differential NAGase responses: activity of glutathione S-transferases (GST), glutathione peroxidase (GPx) glutathione reductase (GR), levels of total glutathiones (TG), lipid peroxidation (LPO), lactate dehydrogenase (LDH), and NADP(+)-dependent isocitrate dehydrogenase (IDH). Animals from the moderately polluted estuary had lower NAGase activity both in the epidermis and digestive gland than in the low impacted site. NAGase activity in the epidermis and digestive gland of C. maenas from both estuaries was sensitive to cadmium exposure suggesting its usefulness for inclusion in monitoring programmes. However, in the digestive gland NAGase inhibition was found in crabs from the less impacted site but not in those from the moderately contaminated one. Altered glutathione levels were observed in cadmium-treated crabs from the contaminated site possibly conferring enhanced tolerance to these animals through its chelator action. Investigation of enhanced tolerance should thus be accounted for in monitoring programmes employing NAGase as biomarker to avoid data misinterpretation.

Sertraline accumulation and effects in the estuarine decapod Carcinus maenas: importance of the history of exposure to chemical stress.

Sertraline is widely prescribed worldwide and frequently detected in aquatic systems. There is, however, a remarkable gap of information on its potential impact on estuarine and coastal invertebrates. This study investigated sertraline accumulation and effects in Carcinus maenas. Crabs from a moderately contaminated (Lima) and a low-impacted (Minho) estuary were exposed to environmental and high levels of sertraline (0.05, 5, 500 µg L(-1)). A battery of biomarkers related to sertraline mode of action was employed to assess neurotransmission, energy metabolism, biotransformation and oxidative stress pathways. After a seven-day exposure, sertraline accumulation in crabs' soft tissues was found in Lima (5 µg L(-1): 15.3 ng L(-1) ww; 500 µg L(-1): 1010 ng L(-1) ww) and Minho (500 µg L(-1): 605 ng L(-1) ww) animals. Lima crabs were also more sensitive to sertraline than those from Minho, exhibiting decreased acetylcholinesterase activity, indicative of ventilatory and locomotory dysfunction, inhibition of anti-oxidant enzymes and increased oxidative damage at ≥ 0.05 µg L(-1). The Integrated Biomarker Response (IBR) index indicated their low health status. In addition, Minho crabs showed non-monotonic responses of acetylcholinesterase suggestive of hormesis. The results pointed an influence of the exposure history on differential sensitivity to sertraline and the need to perform evaluations with site-specific ecological receptors to increase relevance of risk estimations when extrapolating from laboratory to field conditions.

Integrated biomarker responses of an estuarine invertebrate to high abiotic stress and decreased metal contamination.

An integrated chemical-biological effects monitoring was performed in 2010 and 2012 in two NW Iberian estuaries under different anthropogenic pressure. One is low impacted and the other is contaminated by metals. The aim was to verify the usefulness of a multibiomarker approach, using Carcinus maenas as bioindicator species, to reflect diminishing environmental contamination and improved health status under abiotic variation. Sampling sites were assessed for metal levels in sediments and C. maenas, water abiotic factors and biomarkers (neurotoxicity, energy metabolism, biotransformation, anti-oxidant defences, oxidative damage). High inter-annual and seasonal abiotic variation was observed. Metal levels in sediments and crab tissues were markedly higher in 2010 than in 2012 in the contaminated estuary. Biomarkers indicated differences between the study sites and seasons and an improvement of effects measured in C. maenas from the polluted estuary in 2012. Integrated Biomarker Response (IBR) index depicted sites with higher stress levels whereas Principal Component Analysis (PCA) showed associations between biomarker responses and environmental variables. The multibiomarker approach and integrated assessments proved to be useful to the early diagnosis of remediation measures in impacted sites.

Joint effects of salinity and the antidepressant sertraline on the estuarine decapod Carcinus maenas.

Concurrent exposure of estuarine organisms to man-made and natural stressors has become a common occurrence. Numerous interactions of multiple stressors causing synergistic or antagonistic effects have been described. However, limited information is available on combined effects of emerging pharmaceuticals and natural stressors. This study investigated the joint effects of the antidepressant sertraline and salinity on Carcinus maenas. To improve knowledge about interactive effects and potential vulnerability, experiments were performed with organisms from two estuaries with differing histories of exposure to environmental contamination. Biomarkers related to mode of action of sertraline were employed to assess effects of environmentally realistic concentrations of sertraline at two salinity levels. Synergism and antagonism were identified for biomarkers of cholinergic neurotransmission, energy production, anti-oxidant defences and oxidative damage. Different interactions were found for the two study sites highlighting the need to account for differences in tolerance of local ecological receptors in risk evaluations.