Targeting liver and adipose tissue in obese mice: Effects of a N-acylethanolamine mixture on insulin resistance and adipocyte reprogramming.

N-acylethanolamines (NAEs) are endogenous lipid-signalling molecules involved in inflammation and energy metabolism. The potential pharmacological effect of NAE association in managing inflammation-based metabolic disorders is unexplored. To date, targeting liver-adipose axis can be considered a therapeutic approach for the treatment of obesity and related dysfunctions. Here, we investigated the metabolic effect of OLALIAMID® (OLA), an olive oil-derived NAE mixture, in limiting liver and adipose tissue (AT) dysfunction of high-fat diet (HFD)-fed mice. OLA reduced body weight and fat mass in obese mice, decreasing insulin resistance (IR), as shown by homeostasis model assessment index, and leptin/adiponectin ratio, a marker of adipocyte dysfunction. OLA improved serum lipid and hepatic profile and the immune/inflammatory pattern of metainflammation. In liver of HFD mice, OLA treatment counteracted glucose and lipid dysmetabolism, restoring insulin signalling (phosphorylation of AKT and AMPK), and reducing mRNAs of key markers of fatty acid accumulation. Furthermore, OLA positively affected AT function deeply altered by HFD by reprogramming of genes involved in thermogenesis of interscapular brown AT (iBAT) and subcutaneous white AT (scWAT), and inducing the beigeing of scWAT. Notably, the NAE mixture reduced inflammation in iBAT and promoted M1-to-M2 macrophage shift in scWAT of obese mice. The tissue and systemic anti-inflammatory effects of OLA and the increased expression of glucose transporter 4 in scWAT contributed to the improvement of gluco-lipid toxicity and insulin sensitivity. In conclusion, we demonstrated that this olive oil-derived NAE mixture is a valid nutritional strategy to counteract IR and obesity acting on liver-AT crosstalk, restoring both hepatic and AT function and metabolism.

Bisphenol A exacerbates anxiety-like behavior and neuroinflammation in prefrontal cortex of adult obese mice.

AIMS: Bisphenol A (BPA) is an endocrine-disrupting chemical inducing several damages such as neurotoxicity, immunotoxicity, and metabolic disorders. Obesity is the main risk factor for the increased occurrence of metabolic alterations as well as mood disorders. Here, we investigated in obese mice the effects of BPA on anxiety-like behavior, associated with neuroinflammation and immune activation. MAIN METHODS: Male C57Bl/6J mice were divided into 4 groups: control group (STD) receiving chow diet and BPA vehicle; STD group treated with BPA (50 µg/kg/die); high-fat diet (HFD) group receiving BPA vehicle; HFD group treated with BPA. BPA treatment started 12 weeks after HFD feeding and lasted 3 weeks. KEY FINDINGS: The open field and elevated plus-maze tests showed in HFD + BPA group the worsening of HFD-induced anxiety-like behavior. The anxiogenic effects of BPA also emerged from hyperactivation of the hypothalamus-pituitary-adrenal gland axis, determined by the increased transcription of Crh and its receptor in the prefrontal cortex (PFC). Furthermore, BPA activated NLRP3 inflammasome and exacerbated the neuroinflammation induced by HFD, increasing IL-1ß, TNF-α and monocyte chemoattractant protein (MCP)-1 in PFC. Furthermore, it induced inflammation and monocyte recruitment in hypothalamus and amygdala. Contextually, BPA significantly amplified the immune activation caused by lipid overload as evidenced by the increased expression of TLR-4 and MCP-1 in the PFC and triggered mastocytosis in the hypothalamus rather than STD mice. SIGNIFICANCE: All these data show that sub-chronic BPA exposure represents an additional risk factor for mood disorders strictly related to obesity, enhancing neuroinflammation and immune activation triggered by HFD feeding.

Levels of polychlorinated biphenyls and organochlorine pesticides in donkey milk: Correlation with the infection level by intestinal strongyles.

AIM: The study aimed at evaluating the concentration levels of organochlorine pollutants in donkey milk and their modulation on the intestinal strongyle infection. Risk evaluation for consumer health was also investigated. METHODS: We analyzed milk of grazing donkeys living in areas of Southern of Italy affected by organochlorine compounds environmental pollution and parasite infection. The presence of pollutants was assessed through summary statistics; regression analysis of intestinal strongyle on pollutant concentration was performed to investigate the relationship between the two variables. RESULTS: PCB concentrations (mainly non-dioxin-like (ndl)-PCBs) were higher than OCP ones. Mean values of ndl-PCBs across areas ranged from 93.13 to 263.64 ng g-1. In all sample units we detected the six indicator PCBs with the prevalence of the PCB 153, followed by the PCB 28 and the PCB 101. Among the dioxin-like (dl)-PCBs, non-ortho PCB 169, 77 and 126 were assessed in some milk samples; in all areas we detected the mono-ortho PCB 118 and PCB 105. Positive correlation between infection level and six indicator PCBs as well as between the former and HCB, on WW and LW, were observed (at least statistically significant at 5 percent). In some cases, Dl-PCB concentrations emerged as dangerous given the EU maximum residue limit for PCDD/Fs and dl-PCBs. CONCLUSION: Evidence supports the hypothesis of an immunosuppressive role of organochlorine pollutants; risk evaluation reveals the potential health impact of dl-PCB intake, particularly for major donkey milk consumers such as infants, children with cow milk and multiple food intolerance, and elders.

Food contamination by PCBs and waste disposal crisis: Evidence from goat milk in Campania (Italy).

AIM: The study aims at investigating whether, and if so, to what extent the strong presence of urban and industrial waste in a territory may cause PCB contamination in goat milk produced therein. METHODS: We compared PCB concentrations in goat milk from three different locations in the Campania region (Italy). One of the three locations, together with its surrounding area, has long suffered from illegal waste disposal and burning mainly by the so-called Ecomafia. The other locations, not involved in these illegal activities, allowed us to create a control group of goats with characteristics very similar to those of main interest. RESULTS: In milk from the waste contaminated area we identified high PCB concentrations (six indicator PCBs amounted to 170 ng g-1 on lipid weight, on average), whereas there was an almost total absence of such pollutants in milk from the control group. Concentrations of the six indicator PCBs were above the current European maximum residue limit fixed by the EU. At the same time, we found a lower average value of lipid content and a negative relationship between lipid content and PCB concentrations. CONCLUSION: Evidence indicates the potential health risk for consumers living in areas involved in illegal dumping of waste.

Cadmium accumulation and antioxidant responses in Sparus aurata exposed to waterborne cadmium.

Cadmium (Cd), a nonessential trace element, is rapidly accumulated by most living organisms and subsequently exerts its toxicity at different molecular levels. This study exposed gilthead sea bream (Sparus aurata) to waterborne 0.1 mg/l Cd for 11 days and investigated the Cd accumulation pattern, lipid oxidation, and response of antioxidant defences. At the end of the experiment, mean Cd concentrations in gills and liver, the organs most prone to metal accumulation, were 209.4 and 371.7 ng/g ww, respectively. Muscle did not show any Cd retention during the 11 days of exposure. In liver, the cytosolic fraction of the metal was chelated into the nontoxic form by metallothionein (MT), a specific Cd-inducible protein. Zn and Cu concentrations were not influenced by Cd exposure. Glutathione (GSH) concentrations and the antioxidant enzyme activities of GSH reductase and GSH peroxidase showed an overall decreasing trend. In addition, lipid and aqueous hydroperoxide levels did not show any significant variation. Oxidative stress indirectly generated by Cd seems to be compensated for by the different biochemical systems tailored to decrease cellular damage. In particular, the negative effects of Cd accumulation in tissues were probably counteracted by the induction of MT.

Effects of non-dioxin-like polychlorinated biphenyl congeners (PCB 101, PCB 153 and PCB 180) alone or mixed on J774A.1 macrophage cell line: modification of apoptotic pathway.

Non-dioxin-like polychlorinated biphenyls (PCBs) are stable and lipophilic chemicals that persist in the environment and tend to bioaccumulate in the food chains. In the present study, we have investigated the effect of PCBs 101, 153, and 180 on macrophage J774A.1 by assessing cell viability and apoptotic cell death. We have combined morphological techniques and biochemical ones to establish the relevance of apoptosis in macrophage cell death induced by PCBs, alone or in combination. Treatment with the examined PCBs caused the loss of cell viability and accelerated apoptosis in a concentration-dependent manner. Moreover, a synergistic effect on cell death and apoptosis was evidenced for all PCBs at concentrations which were inactive alone. The apoptosis induced by PCBs involved the increase of caspase-3 activity. Also, Bcl-2 and Bax proteins were assessed to elucidate the apoptosis machinery induced in macrophage cultures by PCBs. Our results indicate that the increase in PCB-induced apoptosis correlates with a reduction in the expression of antiapoptotic Bcl-2 and an increase in the expression of proapoptotic Bax. Interestingly, concentrations of PCBs inactive by themselves induce apoptosis when PCBs are combined. In conclusion, our findings suggest that, although less toxic than dioxin like congeners, the examined non-dioxin-like PCBs are equally dangerous as immunotoxic pollutants, also considering their presence as mixtures at higher levels than dioxin-like PCBs in biotic and abiotic matrices.

Differential modification of inflammatory enzymes in J774A.1 macrophages by ochratoxin A alone or in combination with lipopolysaccharide.

Ochratoxin A (OTA) is a fungal metabolite with controversial immunomodulatory effects. A prolonged in vivo exposure to the mycotoxin may result in impaired immunity and decreased resistance to infections. In the present study, OTA modulation of lipopolysaccharide (LPS)-induced inflammatory process is described in the macrophagic cell line, J774A.1 in order to better understand the mechanisms underlying OTA immunotoxicity. OTA (30 nM-100 microM) induces a time and concentration dependent cytotoxic effect, increased when cells were co-stimulated with LPS (100 ng/ml), a concentration that alone did not modify the cellular viability. Moreover, OTA (3 microM) alone induces a significant increase in cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expression, while at the highest concentration (10 microM) a reduced expression of both enzymes was shown, consistently with the mycotoxin cytotoxic profile. The role of nuclear factor-kB (NF-kB) in the mycotoxin effect was also demonstrated. Conversely, when cells were co-stimulated with LPS, OTA showed a concentration-dependent reduction of COX-2 and iNOS expression and their respective metabolites (PGE(2) and NO). These results confirm the pro-inflammatory role of OTA by itself, and demonstrate the impaired capability of OTA-treated macrophages to respond properly to noxious stimuli, such as LPS, mimicking the environmental co-exposure to both compounds.

Persistent organochlorine pollutants in liver of birds of different trophic levels from coastal areas of Campania, Italy.

Liver samples of 12 species of birds of different trophic levels, collected during the period 1998-2000 from coastal areas of the Campania region, Southern Italy, were analyzed for organochlorine pesticides (OCs), such as dichlorodiphenyltrichloroethane (DDT) and its metabolites, hexachlorobenzene (HCB), gamma-hexachlorocycloexane (gamma-HCH), aldrin, dieldrin, endrin, and the seven PCB "target" congeners, IUPAC Nos. 28, 52, 101, 118, 138, 153 and 180. p,p'-DDE was present in all the samples analyzed, at concentrations ranging from 4 to 4504 ng/g wet wt, which were much higher than those found for HCB, dieldrin, and p,p'-DDD. The concentrations of the others OCs were below the detection limit in all the samples. PCBs were found in all the bird species at levels ranging between 6 and 8431 ng/g wet wt. The hepta-, hexa-, and penta-chlorinated congeners 180, 153, 138, and 118 were predominant since, in almost all the species, they contributed to more than 98% of the total seven determined PCBs. No significant differences in mean concentrations of organochlorine pesticides are detected between single species or between species grouped according to their feeding habits (p > 0.05). However, p,p'-DDE levels were higher in carnivorous species than in omnivorous and insectivorous ones (carnivorous > omnivorous > insectivorous). Concentrations of total PCBs were significantly higher in omnivorous birds than in carnivorous (p < 0.01) and insectivorous ones (p < 0.001), whereas carnivorous birds exhibited significantly higher total PCB levels than insectivorous ones (p < 0.01). Marked differences in total PCB concentrations were found also between single species (from p < 0.05 to p < 0.001). Levels of OCs and PCBs were generally below the thresholds known to affect reproduction, however, mean hepatic concentrations of total PCBs in the yellow-legged herring gull (Larus cachinnans), black-headed gull (Larus ridibundus), and kestrel (Falcus tinnunculus) were far higher than those estimated to elicit immunosuppressive effects and possibly increase susceptibility to parasitoses.

Coding criteria of bladder cancer: effects on estimating survival.

The aim of this study is to evaluate the consistency between routine methods for coding urinary bladder tumours in eight Italian cancer registries and the European Network of cancer registries (ENCR) criteria. Furthermore, it aims to evaluating the impact of the discordance on survival data. Eight cancer registries took part in the study: Ferrara, Florence, Macerata, Ragusa, Romagna, Sassari, Turin and Varese. The first 100 cases of neoplasm of the urinary bladder incident in the years 1993-1994 were identified from the files of each registry. The original pathology reports were made available. A working group considered eligible to the study 699 cases of microscopically confirmed transitional carcinoma (ICD-O morphology code 812-813). Using the ENCR criteria, each of these was classified according to morphology code (8120 vs. 8130) and behaviour (1/ uncertain, /2 non-invasive, 3/ invasive). Information of tumour behaviour was classified as follows: (i) present, when expressly stated in the original report, (ii) deducible, when not expressly stated but suggested by the pathologist's description, and (iii) absent, when impossible to determine on the basis of the original pathology report. The working group classification of tumour behaviour and the classification of the registry of origin were compared. There was a full concordance in the case of complete agreement on the morphology code, and partial concordance when only the invasive or non-invasive behaviour code was agreed upon. As much as 92.5% cases were microscopically confirmed. Tumour behaviour was expressly stated in the original report of 69.2% cases, not stated but suggested by the pathologist's description in 21.2% cases, and impossible to determine in 9.6%. Agreement between the panel and the registry of origin was complete in 71.2% cases and partial in 12.3% while there was a complete discordance in 16.5% cases. The panel interpreted as non-invasive 111 cases coded as invasive by the registry of origin. Conversely, it was estimated that 24% cases included in incidence data were non-invasive. This article discusses the impact of misclassification on survival data.

Effect of fumonisin B1 on inducible nitric oxide synthase and cyclooxygenase-2 in LPS-stimulated J774A.1 cells.

Fumonisin B1 (FB1) is a water-soluble fungal metabolite that elicits a wide spectrum of toxicological effects. Cellular targets of FB1 include immune cells and in particular macrophages. In the present study the cytotoxic effect of FB1 (1-100 microM) was evaluated using a murine macrophage cell line (J774A.1) as model system. The effect of FB1 on nitric oxide (NO) and prostaglandin E2 (PGE2) production induced by lipopolysaccharide (LPS, 10 and 100 ng/ml) was also investigated. Macrophages were pretreated with FB1 for 72 h and then stimulated with LPS for 24 h. The increase of LPS-induced production of these inflammatory mediators was observed at increasing concentrations of FB1 (0.1-10 microM) and was found to be concentration dependent. By western blot analysis we demonstrated that the observed increase of NO and PGE2 production by FB1 was related to an enhancement of iNOS and COX-2 expression.