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Hyperammonemia is a metabolic condition characterized by elevated levels of ammonia and a common event in acute liver injury/failure and chronic liver disease. Even though hepatic ammonia levels are potential predictive factors of patient outcome, easy and inexpensive methods aiming at the detection of liver ammonia accumulation in the clinical setting remain unavailable. Thus, herein we have developed a morphological method, based on the utilization of Nessler´s reagent, to accurately and precisely detect the accumulation of ammonia in biological tissue. We have validated our method against a commercially available kit in mouse tissue samples and, by using this modified method, we have confirmed the hepatic accumulation of ammonia in clinical and animal models of acute and chronic advanced liver injury as well as in the progression of fatty liver disease. Overall, we propose a morphological method for ammonia detection in liver that correlates well with the degree of liver disease severity and therefore can be potentially used to predict patient outcome.

ObjectiveThe prevalence of psoriatic arthritis (PsA) is very heterogeneous. There are no data on its frequency in the general population in Spain. The aim of EPISER2016 study was to estimate the prevalence of PsA in people aged ≥20 years in Spain.MethodsCross-sectional multicenter population-based study. Subjects from all the autonomous communities in Spain were randomly selected using multistage stratified cluster sampling. Participants in each of the municipalities randomly selected for the study were administered a telephone-based questionnaire to screen for the study diseases. If the participant reported being previously diagnosed, rheumatologists from the participant's reference hospital confirmed the diagnosis based on a review of the clinical history. Subjects not previously diagnosed but whose screening result was positive based on symptoms received a second telephone call from the investigating rheumatologist in order to evaluate the suspicion. If the suspicion remained, an appointment was made at the reference hospital to complete the diagnostic confirmation process according to CASPAR criteria. To calculate the prevalence and its 95% confidence interval (CI), the sample design was taken into account and weighing was calculated considering age, sex and geographic origin.ResultsThe sample comprised 4916 subjects. The prevalence of PsA was 0.58% (95%CI: 0.38-0.87). All but 1 of the 27 cases (96.30%) had been diagnosed prior to EPISER2016.ConclusionThe prevalence of PsA in Spain was among the highest reported to date, only below that reported in Norway (0.67%) and slightly higher than that reported in Italy (0.42%).

Dalbergia ecastaphyllum is a native Brazil plant with importance for beekeeping, and widely used in folk medicine. For the first time, the extracts of this plant were assessed for the presence of hydrophilic and lipophilic antioxidants, as well as inhibition of tyrosinase, free radicals scavenging and sunscreen protection. The antioxidant activity was evaluated by free radical scavenging (DPPH) and β-carotene bleaching assay. The tyrosinase inhibitory activity was evaluated and calculated the EC50. The photoprotective activity was measured using different concentrations of D. ecastaphyllum extracts. The Sun Protection Factor (SPF) of the samples was higher than 6, and the sample from Ilhéus showed the most pronounced photoprotective effect. Sample from Canavieiras presented the highest antioxidant activity by free radical scavenging DPPH and β-carotene bleaching method, with 92.41% and 48.34%, respectively. All samples inhibited the tyrosinase, especially the sample from Prado that was most effective (124.62 μg.mL-1). Significant negative correlation was found between flavonoid contents and inhibition of tyrosinase. The overall results provide relevant information about the Dalbergia ecastaphyllum species, indicating as potential material to cosmetic and pharmaceutical industry.

The formation of acquired drug resistance is a major reason for the failure of anti-cancer therapies after initial response. Here, we introduce a novel model of acquired oxaliplatin resistance, a sub-line of the non-MYCN-amplified neuroblastoma cell line SK-N-AS that was adapted to growth in the presence of 4000 ng/mL oxaliplatin (SK-N-ASrOXALI4000). SK-N-ASrOXALI4000 cells displayed enhanced chromosomal aberrations compared to SK-N-AS, as indicated by 24-chromosome fluorescence in situ hybridisation. Moreover, SK-N-ASrOXALI4000 cells were resistant not only to oxaliplatin but also to the two other commonly used anti-cancer platinum agents cisplatin and carboplatin. SK-N-ASrOXALI4000 cells exhibited a stable resistance phenotype that was not affected by culturing the cells for 10 weeks in the absence of oxaliplatin. Interestingly, SK-N-ASrOXALI4000 cells showed no cross resistance to gemcitabine and increased sensitivity to doxorubicin and UVC radiation, alternative treatments that like platinum drugs target DNA integrity. Notably, UVC-induced DNA damage is thought to be predominantly repaired by nucleotide excision repair and nucleotide excision repair has been described as the main oxaliplatin-induced DNA damage repair system. SK-N-ASrOXALI4000 cells were also more sensitive to lysis by influenza A virus, a candidate for oncolytic therapy, than SK-N-AS cells. In conclusion, we introduce a novel oxaliplatin resistance model. The oxaliplatin resistance mechanisms in SK-N-ASrOXALI4000 cells appear to be complex and not to directly depend on enhanced DNA repair capacity. Models of oxaliplatin resistance are of particular relevance since research on platinum drugs has so far predominantly focused on cisplatin and carboplatin.

Deep brain stimulation (DBS) has become a widely used technique for treating advanced stages of neurological and psychiatric illness. In the case of motor disorders related to basal ganglia (BG) dysfunction, several mechanisms of action for the DBS therapy have been identified which might be involved simultaneously or in sequence. However, the identification of a common key mechanism underlying the clinical relevant DBS configurations has remained elusive due to the inherent complexity related to the interaction between the electrical stimulation and the neural tissue, and the intricate circuital structure of the BG-thalamocortical network. In this work, it is shown that the clinically relevant range for both, the frequency and intensity of the electrical stimulation pattern, is an emergent property of the BG anatomy at the system-level that can be addressed using mean-field descriptive models of the BG network. Moreover, it is shown that the activity resetting mechanism elicited by electrical stimulation provides a natural explanation to the ineffectiveness of irregular (i.e., aperiodic) stimulation patterns, which has been commonly observed in previously reported pathophysiology models of Parkinson's disease. Using analytical and numerical techniques, these results have been reproduced in both cases: 1) a reduced mean-field model that can be thought as an elementary building block capable to capture the underlying fundamentals of the relevant loops constituting the BG-thalamocortical network, and 2) a detailed model constituted by the direct and hyperdirect loops including one-dimensional spatial structure of the BG nuclei. We found that the optimal ranges for the essential parameters of the stimulation patterns can be understood without taking into account biophysical details of the relevant structures.

Hyperammonemia is a metabolic condition characterized by elevated levels of ammonia and a common event in acute liver injury/failure and chronic liver disease. Even though hepatic ammonia levels are potential predictive factors of patient outcome, easy and inexpensive methods aiming at the detection of liver ammonia accumulation in the clinical setting remain unavailable. Thus, herein we have developed a morphological method, based on the utilization of Nessler´s reagent, to accurately and precisely detect the accumulation of ammonia in biological tissue. We have validated our method against a commercially available kit in mouse tissue samples and, by using this modified method, we have confirmed the hepatic accumulation of ammonia in clinical and animal models of acute and chronic advanced liver injury as well as in the progression of fatty liver disease. Overall, we propose a morphological method for ammonia detection in liver that correlates well with the degree of liver disease severity and therefore can be potentially used to predict patient outcome.

Several etiologies result in chronic liver diseases including chronic hepatitis C virus infection (HCV). Despite its high incidence and the severe economic and medical consequences, liver disease is still commonly overlooked due to the lack of efficient non-invasive diagnostic methods. While several techniques have been tested for the detection of fibrosis, the available biomarkers still present severe limitations that preclude their use in clinical diagnostics. Liver diseases have also been the subject of metabolomic analysis. Here, we demonstrate the suitability of 1H NMR spectroscopy for characterizing the metabolism of liver fibrosis induced by HCV. Serum samples from HCV patients without fibrosis or with liver cirrhosis were analyzed by NMR spectroscopy and the results were submitted to multivariate and univariate statistical analysis. PLS-DA test was able to discriminate between advanced fibrotic and non-fibrotic patients and several metabolites were found to be up or downregulated in patients with cirrhosis. The suitability of the most significantly regulated metabolites was validated by ROC analysis. Our study reveals that choline, acetoacetate and low-density lipoproteins are the most informative biomarkers for predicting cirrhosis in HCV patients. Our results demonstrate that statistical analysis of 1H-NMR spectra is able to distinguish between fibrotic and non-fibrotic patients suffering from HCV, representing a novel diagnostic application for NMR spectroscopy.

In general, in composting facilities the active, or intensive, stage of the process is done separately from the maturation stage, using a specific technology and time. The pre-composted material to be matured can contain enough biodegradable substrates to cause microbial proliferation, which in turn can cause temperatures to increase. Therefore, not controlling the maturation period during waste management at an industrial level can result in undesired outcomes. The main hypothesis of this study is that controlling the maturation stage through turning provides one with an optimized process when compared to the static approach. The waste used was sludge from a seafood-processing plant, mixed with shredded wood (1:2, v/v). The composting system consists of an intensive stage in a 600L static reactor, followed by maturation in triplicate in 200L boxes for 112 days. Two tests were carried out with the same process in reactor and different treatments in boxes: static maturation and turning during maturation when the temperature went above 55°C. PLFAs, organic matter, pH, electrical conductivity, forms of nitrogen and carbon, hydrolytic enzymes and respiratory activity were periodically measured. Turning significantly increased the duration of the thermophilic phase and consequently increased the organic-matter degradation. PCA differentiated significantly the two treatments in function of tracking parameters, especially pH, total carbon, forms of nitrogen and C/N ratio. So, stability and maturity optimum values for compost were achieved in less time with turnings. Whereas turning resulted in microbial-group stabilization and a low mono/sat ratio, static treatment produced greater variability in microbial groups and a high mono/sat ratio, the presence of more degradable substrates causes changes in microbial communities and their study during maturation gives an approach of the state of organic-matter degradation. Obtaining quality compost and optimizing the composting process requires using turning as a control mechanism during maturation.

Alport syndrome results from mutations in the COL4A5 (X-linked) or COL4A3/COL4A4 (recessive) genes. This study examined 754 previously- unpublished variants in these genes from individuals referred for genetic testing in 12 accredited diagnostic laboratories worldwide, in addition to all published COL4A5, COL4A3 and COL4A4 variants in the LOVD databases. It also determined genotype-phenotype correlations for variants where clinical data were available. Individuals were referred for genetic testing where Alport syndrome was suspected clinically or on biopsy (renal failure, hearing loss, retinopathy, lamellated glomerular basement membrane), variant pathogenicity was assessed using currently-accepted criteria, and variants were examined for gene location, and age at renal failure onset. Results were compared using Fisher's exact test (DNA Stata). Altogether 754 new DNA variants were identified, an increase of 25%, predominantly in people of European background. Of the 1168 COL4A5 variants, 504 (43%) were missense mutations, 273 (23%) splicing variants, 73 (6%) nonsense mutations, 169 (14%) short deletions and 76 (7%) complex or large deletions. Only 135 of the 432 Gly residues in the collagenous sequence were substituted (31%), which means that fewer than 10% of all possible variants have been identified. Both missense and nonsense mutations in COL4A5 were not randomly distributed but more common at the 70 CpG sequences (pAla substitutions were underrepresented in all three genes (p< 0.0001) probably because of an association with a milder phenotype. The average age at end-stage renal failure was the same for all mutations in COL4A5 (24.4 ±7.8 years), COL4A3 (23.3 ± 9.3) and COL4A4 (25.4 ± 10.3) (COL4A5 and COL4A3, p = 0.45; COL4A5 and COL4A4, p = 0.55; COL4A3 and COL4A4, p = 0.41). For COL4A5, renal failure occurred sooner with non-missense than missense variants (p

IntroductionCushing's syndrome (CS) increases cardiovascular risk (CVR) and adipocytokine imbalance, associated with an increased inflammatory state. Telomere length (TL) shortening is a novel CVR marker, associated with inflammation biomarkers. We hypothesized that inflammatory state and higher CVR in CS might be related to TL shortening, as observed in premature aging.AimTo evaluate relationships between TL, CVR and inflammation markers in CS.MethodsIn a cross-sectional study, 77 patients with CS (14 males, 59 pituitary-, 17 adrenal- and 1 ectopic-origin; 21 active disease) and 77 age-, gender-, smoking-matched controls were included. Total white blood cell TL was measured by TRF-Southern technique. Clinical data and blood samples were collected (lipids, adrenal function, glucose). Adiponectin, interleukin-6 (IL6) and C-reactive protein (CRP) were available in a subgroup of patients (n=32). Correlations between TL and clinical features were examined and multiple linear regression analysis was performed to investigate potential predictors of TL.ResultsDyslipidemic CS had shorter TL than non-dyslipidemic subjects (7328±1274 vs 7957±1137 bp, pConclusionTL is shortened in dyslipidemic CS patients, further worse if hypertension and/or obesity coexist and is negatively correlated with increased inflammation markers. Increased lipids and a "low" grade inflammation may contribute to TL shortening and consequently to premature ageing and increased morbidity in CS.

Nonalcoholic fatty liver disease (NAFLD) is a risk factor for Hepatocellular carcinoma (HCC), but he transition from NAFLD to HCC is poorly understood. Feature selection algorithms in human and genetically modified mice NAFLD and HCC microarray data were applied to generate signatures of NAFLD progression and HCC differential survival. These signatures were used to study the pathogenesis of NAFLD derived HCC and explore which subtypes of cancers that can be investigated using mouse models. Our findings show that: (I) HNF4 is a common potential transcription factor mediating the transcription of NAFLD progression genes (II) mice HCC derived from NAFLD co-cluster with a less aggressive human HCC subtype of differential prognosis and mixed etiology (III) the HCC survival signature is able to correctly classify 95% of the samples and gives Fgf20 and Tgfb1i1 as the most robust genes for prediction (IV) the expression values of genes composing the signature in an independent human HCC dataset revealed different HCC subtypes showing differences in survival time by a Logrank test. In summary, we present marker signatures for NAFLD derived HCC molecular pathogenesis both at the gene and pathway level.

Hyperthermia is a predictor of poor outcome in ischemic (IS) and intracerebral hemorrhagic (ICH) stroke. Our aim was to study the plausible mechanisms involved in the poor outcome associated to hyperthermia in stroke. We conducted a case-control study including patients with IS (n = 100) and ICH (n = 100) within the first 12 hours from symptom onset. Specifically, IS and ICH patients were consecutively included into 2 subgroups, according to the highest body temperature within the first 24 hours: Tmax 2) at 3 months. Serum levels of glutamate and active MMP-9 were measured at admission. Our results showed that Tmax ≥37.5°C within the first 24 hours was independently associated with poor outcome in both IS (OR, 12.43; 95% CI, 3.73-41.48; p

Glutamate excitotoxicity, metabolic rate and inflammatory response have been associated to the deleterious effects of temperature during the acute phase of stroke. So far, the association of temperature with these mechanisms has been studied individually. However, the simultaneous study of the influence of temperature on these mechanisms is necessary to clarify their contributions to temperature-mediated ischemic damage. We used non-invasive Magnetic Resonance Spectroscopy to simultaneously measure temperature, glutamate excitotoxicity and metabolic rate in the brain in animal models of ischemia. The immune response to ischemia was measured through molecular serum markers in peripheral blood. We submitted groups of animals to different experimental conditions (hypothermia at 33°C, normothermia at 37°C and hyperthermia at 39°C), and combined these conditions with pharmacological modulation of glutamate levels in the brain through systemic injections of glutamate and oxaloacetate. We show that pharmacological modulation of glutamate levels can neutralize the deleterious effects of hyperthermia and the beneficial effects of hypothermia, however the analysis of the inflammatory response and metabolic rate, demonstrated that their effects on ischemic damage are less critical than glutamate excitotoxity. We conclude that glutamate excitotoxicity is the key molecular mechanism which is influenced by body temperature during the acute phase of brain stroke.

BACKGROUND: Obesity modulates inflammation and activation of immune pathways which can lead to liver complications. We aimed at identifying expression patterns of inflammatory and immune response genes specifically associated with obesity and NASH in the liver of morbidly obese patients. METHODOLOGY/PRINCIPAL FINDINGS: Expression of 222 genes was evaluated by quantitative RT-PCR in the liver of morbidly obese patients with histologically normal liver (n = 6), or with severe steatosis without (n = 6) or with NASH (n = 6), and in lean controls (n = 5). Hepatic expression of 58 out of 222 inflammatory and immune response genes was upregulated in NASH patients. The most notable changes occurred in genes encoding chemokines and chemokine receptors involved in leukocyte recruitment, CD and cytokines involved in the T cell activation towards a Th1 phenotype, and immune semaphorins. This regulation seems to be specific for the liver since visceral adipose tissue expression and serum levels of MCP1, IP10, TNFα and IL6 were not modified. Importantly, 47 other genes were already upregulated in histologically normal liver (e.g. CRP, Toll-like receptor (TLR) pathway). Interestingly, serum palmitate, known to activate the TLR pathway, was increased with steatosis. CONCLUSION/SIGNIFICANCE: The liver of obese patients without histological abnormalities already displayed a low-grade inflammation and could be more responsive to activators of the TLR pathway. NASH was then characterized by a specific gene signature. These findings help to identify new potential actors of the pathogenesis of NAFLD.

Cannabinoids have deleterious effects on prefrontal cortex (PFC)-mediated functions and multiple evidences link the endogenous cannabinoid (endocannabinoid) system, cannabis use and schizophrenia, a disease in which PFC functions are altered. Nonetheless, the molecular composition and the physiological functions of the endocannabinoid system in the PFC are unknown.Here, using electron microscopy we found that key proteins involved in endocannabinoid signaling are expressed in layers v/vi of the mouse prelimbic area of the PFC: presynaptic cannabinoid CB1 receptors (CB1R) faced postsynaptic mGluR5 while diacylglycerol lipase alpha (DGL-alpha), the enzyme generating the endocannabinoid 2-arachidonoyl-glycerol (2-AG) was expressed in the same dendritic processes as mGluR5. Activation of presynaptic CB1R strongly inhibited evoked excitatory post-synaptic currents. Prolonged synaptic stimulation at 10Hz induced a profound long-term depression (LTD) of layers V/VI excitatory inputs. The endocannabinoid -LTD was presynaptically expressed and depended on the activation of postsynaptic mGluR5, phospholipase C and a rise in postsynaptic Ca(2+) as predicted from the localization of the different components of the endocannabinoid system. Blocking the degradation of 2-AG (with URB 602) but not of anandamide (with URB 597) converted subthreshold tetanus to LTD-inducing ones. Moreover, inhibiting the synthesis of 2-AG with Tetrahydrolipstatin, blocked endocannabinoid-mediated LTD. All together, our data show that 2-AG mediates LTD at these synapses.Our data show that the endocannabinoid -retrograde signaling plays a prominent role in long-term synaptic plasticity at the excitatory synapses of the PFC. Alterations of endocannabinoid -mediated synaptic plasticity may participate to the etiology of PFC-related pathologies.