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ObjectiveAlcohol-associated liver disease (ALD) is the leading cause of liver-related mortality worldwide. Current strategies to manage ALD largely focus on advanced stage disease, however, metabolic changes such as glucose intolerance are apparent at the earliest stage of alcoholic steatosis and increase the risk of disease progression. Ceramides impair insulin signaling and accumulate in ALD, and metabolic pathways involving ceramide synthase 6 (CerS6) are perturbed in ALD during hepatic steatosis. In this study, we aimed to investigate the role of CerS6 in ALD development.MethodsC57BL/6 WT and CerS6 KO mice of both sexes were fed either a Lieber-DeCarli control (CON) or 15% ethanol (EtOH) diet for 6 weeks.In vivometabolic tests including glucose and insulin tolerance tests (GTT and ITT) were performed. The mice were euthanized, and liver histology and lipid levels in serum and liver were measured. Forin vitrostudies, CerS6 was deleted in human hepatocytes and were incubated with EtOH and/or C16:0-ceramides. RNAseq analysis was performed in mice and in liver from patients with different stages of ALD and diseased controls.ResultsAfter six weeks on an EtOH diet, CerS6 KO mice had reduced body weight, food intake, and %fat mass compared to WT mice. Male (but not female) EtOH-fed KO mice showed significantly higher O2consumption, CO2production, respiratory exchange ratio, and energy expenditure (PIn vitro, EtOH stimulation significantly increased PLIN2 protein expression in VL-17A cells while CerS6 deletion inhibited EtOH-mediated PLIN2 upregulation. C16:0-ceramide treatment significantly increased PLIN2 protein expression compared to CON. Importantly, progression of ALD in humans was associated with increased CerS6 hepatic expression.ConclusionsOur findings demonstrate that CerS6 deletion attenuates EtOH-induced weight gain and hepatic steatosis and improves glucose homeostasis in mice fed an EtOH diet. Notably, we unveil that CerS6 plays a major role as a regulator of lipid droplet biogenesis in alcoholic intra-hepatic lipid droplet formation. Together, our data suggest that CerS6 may be targeted for treatment for early stage ALD.

Chronic endurance exercise is a therapeutic strategy in the treatment of many chronic diseases in humans, including the prevention and treatment of metabolic diseases such as diabetes mellitus. Metabolic, cardiorespiratory, and endocrine pathways targeted by chronic endurance exercise have been identified. In the liver, however, the cellular and molecular pathways that are modified by exercise and have preventive or therapeutic relevance to metabolic disease need to be elucidated. The mouse model used in the current study allows for the quantification of a human-relevant exercise “dosage”. In this study we show hepatic gene expression differences between sedentary female and sedentary male mice and that chronic exercise modifies the transcription of hepatic genes related to metabolic disease and steatosis in both male and female mice. Chronic exercise induces molecular pathways involved in glucose tolerance, glycolysis, and gluconeogenesis while producing a decrease in pathways related to insulin resistance, steatosis, fibrosis, and inflammation. Given these findings, this mouse exercise model has potential to dissect the cellular and molecular hepatic changes following chronic exercise with application to understanding the role that chronic exercise plays in preventing human diseases. Novelty: Exercise modifies the hepatic gene expression and hepatic pathways related to metabolic disease in male and female mice. Sex differences were seen in hepatic gene expression between sedentary and exercised mice. The mouse exercise model used in this study allows for application and evaluation of exercise effects in human disease.

Persistent organic pollutants (POPs) are a group of organic chemicals, as defined by the Stockholm Convention that are persistent in the environment and accumulate in organisms including humans. Most POPs are restricted in use or no longer manufactured because of this persistence, bioaccumulation, and toxicity. In addition, an association between POP exposure and human disease has been suggested. Regarding toxicological effects and particularly cancer, most POPs have been shown to induce hepatic tumors when chronically administered to rodents. Understanding the mechanisms by which POPs induce liver cancer is important in developing meaningful human cancer risk evaluation to these compounds. In this article, we have reviewed the probable mechanisms of hepatic cancer induced by POPs mode of action and molecular initiating events and the relationship of these mechanisms to potential human cancer risk.

Intrahepatic neutrophil infiltration has been implicated in severe alcoholic hepatitis (SAH) pathogenesis; however, the mechanism underlying neutrophil-induced injury in SAH remains obscure. This translational study aims to describe the patterns of intrahepatic neutrophil infiltration and its involvement in SAH pathogenesis. Immunohistochemistry analyses of explanted livers identified two SAH phenotypes despite a similar clinical presentation, one with high intrahepatic neutrophils (Neuhi), but low levels of CD8+ T cells, and vice versa. RNA-Seq analyses demonstrated that neutrophil cytosolic factor 1 (NCF1), a key factor in controlling neutrophilic ROS production, was upregulated and correlated with hepatic inflammation and disease progression. To study specifically the mechanisms related to Neuhi in AH patients and liver injury, we used the mouse model of chronic-plus-binge ethanol feeding and found that myeloid-specific deletion of the Ncf1 gene abolished ethanol-induced hepatic inflammation and steatosis. RNA-Seq analysis and the data from experimental models revealed that neutrophilic NCF1-dependent ROS promoted alcoholic hepatitis (AH) by inhibiting AMP-activated protein kinase (a key regulator of lipid metabolism) and microRNA-223 (a key antiinflammatory and antifibrotic microRNA). In conclusion, two distinct histopathological phenotypes based on liver immune phenotyping are observed in SAH patients, suggesting a separate mechanism driving liver injury and/or failure in these patients.

Since the toxicology field was established, women have played a critical role in it. This article is written to celebrate the 20-year anniversary of the Special Interest Group for Women in Toxicology, affiliated with the Society of Toxicology. Six female pioneers in modern Toxicology from different social classes and education backgrounds are featured. Despite these differences, they overcame similar obstacles in gender, politics, and scientific barriers to disseminate their research. This discussion will start with Ellen Swallow Richards, who, besides being the pioneer in sanitary engineering, founded the home economics movement that applied science to the home. The discussion will continue with Alice Hamilton, a contributor to occupational health, a pioneer in the field of industrial toxicology, and an example of generosity to social movements and those in need. Subsequently, the most famous woman we discuss in this paper is Rachel Carson, whose fundamental work in environmental Toxicology is evidenced in her important book

2,4,6-Tribromophenol (2,4,6-TBP) is a brominated flame retardant that accumulates in human tissues and is a potential toxicant. Previous studies found 2,4,6-TBP levels in human tissues were significantly higher than those of brominated flame retardants measured in the same samples. In contrast, the levels of 2,4,6-TBP in the environment and foodstuff are not elevated, suggesting a low potential for direct intake through environmental exposure or diet. Here, we hypothesized that high levels of 2,4,6-TBP in human tissues are partially from the indirect exposure sources, such as biotransformation of highly brominated substances. We conducted in vitro assays utilizing human and rat liver microsomes to compare the biotransformation rates of four highly brominated flame retardants, which could potentially transform to 2,4,6-TBP, including decabromodiphenyl ethane (DBDPE), 2,4,6-tris-(2,4,6-tribromophenoxy)-1,3,5-triazine (TTBP-TAZ), 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE), and tetrabromobisphenol A (TBBPA). Our results show that TTBP-TAZ rapidly metabolizes in both human and rat liver microsomes with a half-life of 1.1 and 2.2 h, respectively, suggesting that TTBP-TAZ is a potential precursor of 2,4,6-TBP. In contrast, 2,4,6-TBP was not formed as a result of biotransformation of TBBPA, BTBPE, and DBDPE in both human and rat liver microsomes. We applied suspect and target screening to explore the metabolic pathways of TTBP-TAZ and identified 2,4,6-TBP as a major metabolite of TTBP-TAZ accounting for 87% of all formed metabolites. These in vitro results were further tested by an in vivo experiment in which 2,4,6-TBP was detected in the rat blood and liver at concentrations of 270 ± 110 and 50 ± 14 μg/g lipid weight, respectively, after being exposed to 250 mg/kg body weight/day of TTBP-TAZ for a week. The hepatic mRNA expression demonstrated that TTBP-TAZ significantly activates the aryl hydrocarbon receptor (AhR) and promotes fatty degeneration (18 and 28-fold change compared to control, respectively) in rats.

Given the known clinical utility of cardiorespiratory fitness, measurement of physiological responses to submaximal exercise may be a feasible approach well suited for diagnostic and prognostic purposes in non-athletes and the chronically ill. Lactate levels and watt output during a short submaximal exercise and a subsequent relaxation period yield an aerobic score that is consistent with cardiorespiratory fitness grades in non-athletes and that may be used as a marker in such approaches. In this study, 28 females (23 ± 3 years) were submitted to a 15 min submaximal recumbent bike session, and their capillary and saliva lactate concentrations were recorded and plotted against time. An individual aerobic score was calculated from this curve, using watt output during equal relative percentiles of maximum heart-rate benchmarks. The scores were compared with respective results in a V̇O2max test and with a similar scoring system in 14 older (51 ± 9 years) females; they correlated with the 6 categories of the V̇O2max test results and classified into 3 categories of V̇O2max grades (very poor/poor; fair/good; excellent/superior) with a combined accuracy of 80.95%. More studies are required to validate the potential utility of this submaximal test as an additional risk factor for diagnostic purposes in non-athletes. Novelty points: A novel method for estimating cardiorespiratory fitness during submaximal exercise consistent with V̇O2max performance. The method yields an aerobic score that may be used as a marker for diagnostic and prognostic purposes.

We point at several challenges that current exercise oncology rodent models face, which call their human-relevance into question: the vast majority of pre-clinical studies in exercise oncology treat “physical exercise” as a primitive concept without further analysis or qualification, and their results are based on dosages that no human can endure. The lack of analysis and qualification together with the dosage mismatch conceal the fact that rodents do not run like humans. Consequently, while these pre-clinical studies may yield insights into potential biological mechanisms underlying the systemic effects of physical exercise on cancer, the applicability of this knowledge to preventive interventions in healthy humans and the ability to translate it to practical therapies in the critically ill remain limited. We propose an alternative exercise rodent model that has better chances of meeting these challenges.

Ionic Liquids (ILs) has been attracting attention, both from academia and industry, given the numerous applications of these systems. ILs are salts, usually composed by an organic ion, and a counterion which could be organic or inorganic, and, interestingly they are found at liquid state at room temperature. Our interest in studying ILs comes from its low toxicity. Some recent studies have shown that the toxicity of the ILs ishigher than believed, in particular for biologically relevant systems. The main goal of this research is to study the influence of the ionic liquid 1-tetradecyl-3-methylimidazolium chloride ([C14MIM][Cl]) with membrane systems. To do so, we made use of different lipids: POPC, Sphingomyelin, Cholesterol, POPG, DPPC, DPPG and DMPC. For each of these systems, the influence of ILs concentration were elucidated by means of a systematic study through different experimental techniques: Small Angle X-ray scattering (SAXS), dynamic light scattering (DLS), fluorescence anisotropy, optical microscopy and z-potential. Since [C14MIM][Cl] has a positive charge on the imidazolium group, the superficial charge of all vesicles increased. For zwitterionic vesicles no significant change in size and melting temperature were noticed. The imidazolium-based ionic liquid diminished the gel-fluid transition temperature for negatively charged lipids. For DPPC:DPPG (1:1), for instance, the transition temperature decreased from 42.50±0.13oC to 25.27±0.33oC and for DPPG from 46.12±0.22 oC to 36.6±0.38 oC. For DPPG, the vesicle hydrodynamic diameter increased from 84±0.1nm to 176±0.1nm, whereas for DPPC:DPPG it increased from 95±0.1nm to 196±0.1nm. The electronic density profile, obtained by SAXS, supported the penetration of the [C14MIM][Cl] into the negative bilayer structure. 15 mol% of [C14MIM][Cl] increased the polar head thickness of DPPC vesicles from 11.1±0.6 Å to 18.0±0.7 Å, without alter significantly the inner region of the membrane.Qualitative results obtained with optical microscopy showed that the IL incorporation destabilize the membrane asymmetry (between the leaflets) leading to the formation of pores (evidenced by optical contrast lost) and the presence of buds. We believe that this work could improve the understanding of the effects of ILs in the presence of biological relevant systems Os líquidos iônicos (LI) tem atraído grande atenção, tanto da academia quanto da indústria, devido às suas numerosas aplicações. LI são sais, normalmente compostos por um íon orgânico, e um contra-íon que pode ser orgânico ou inorgânico, mas que tem como característica ser encontrado no estado líquido à temperaturas próximas a ambiente. Nosso interesse em estudar LIs vem de sua baixa toxicidade, atribuída a sua baixa volatilidade. Entretanto, alguns estudos recentes mostraram que a toxicidade dos LI é maior do que se acreditava, em particular com sistemas de relevância biológica.O objetivo principal desta dissertação é estudar a influência do líquido iônico 1-tetradecil-3-metilimidazólio cloreto ([C14MIM][Cl]) com sistemas de membrana. Para isso, utilizamos diferentes lipídios, como o POPC, esfingomielina, colesterol, POPG, DPPC,DPPG e o DMPC. Para cada um destes sistemas, a influência da concentração de LI foi elucidada por meio de um estudo sistemático através de diferentes técnicas experimentais, tais como: espalhamento de raio-X a baixos ângulos (SAXS), espalhamento dinâmico de luz (DLS), anisotropia de fluorescência, microscopia óptica e potencial-z. Uma vez que o componente iônico de [C14MIM][Cl] tem uma carga positiva no grupo imidazólio, a carga superficial de todas as vesículas estudadas aqui aumentou. Entretanto, para asvesículas compostas pelos lipídeos zwitteriónicos, não tenha sido observada qualquer alteração significativa no tamanho e na temperatura de transição de fase gel-fluido. O [C14MIM][Cl] altera a organização interna entre as moléculas de lipídio com carga negativa. Consequentemente, à medida que a quantidade de LI aumenta, a temperatura de transição de fase diminui e o tamanho médio das vesículas aumenta. Para o sistema DPPC:DPPG (1:1) a temperatura de transição de fase caiu de 42.50 ± 0.13 oC para 25.27 ± 0.33 oC e para as vesículas de DPPG de 46.12±0.22 oC para 36.6±0.38 oC. Quanto ao diâmetro hidrodinâmico médio, no caso do DPPG este valor aumentou de 84±0.1 nm para 176±0.1 nm, enquanto que para a mistura DPPC:DPPG (1:1) ele passou de 95±0.1nm para 196±0.1nm. Indicando assim que o LI incorpora na bicamada lipídica negativamente carregada. O perfil de densidade eletrônica, obtido por SAXS, confirma a penetração do [C14MIM][Cl] na bicamada lipídica. Diferentemente, para a membrana lipídica zwitteriónica o LI tende a se situar perto da região da cabeça polar sem afetar significativamente a região do interior da bicamada lipídica. Por outro lado, a presença de15 mol% de [C14MIM][Cl] aumenta a espessura da região polar das bicamadas das vesículas de DPPC de ~ 11.1±0.6 Å para ~ 18.0±0.7 Å. Os resultados qualitativos da microscopia óptica mostraram que a incorporação da LI desestabiliza a assimetria da membrana entre as camadas interna e externa, além de sugerir o aparecimento de poros (evidenciado pela perda do contraste ótico das vesículas) e estruturas chamadas de buds. Esperamos que este trabalho melhore a compreensão dos efeitos do LI na presença de organismos biológicos.