A Membrane Biophysics Perspective on the Mechanism of Alcohol Toxicity.

Motivations for understanding the underlying mechanisms of alcohol toxicity range from economical to toxicological and clinical. On the one hand, acute alcohol toxicity limits biofuel yields, and on the other hand, acute alcohol toxicity provides a vital defense mechanism to prevent the spread of disease. Herein the role that stored curvature elastic energy (SCE) in biological membranes might play in alcohol toxicity is discussed, for both short and long-chain alcohols. Structure-toxicity relationships for alcohols ranging from methanol to hexadecanol are collated, and estimates of alcohol toxicity per alcohol molecule in the cell membrane are made. The latter reveal a minimum toxicity value per molecule around butanol before alcohol toxicity per molecule increases to a maximum around decanol and subsequently decreases again. The impact of alcohol molecules on the lamellar to inverse hexagonal phase transition temperature (TH) is then presented and used as a metric to assess the impact of alcohol molecules on SCE. This approach suggests the nonmonotonic relationship between alcohol toxicity and chain length is consistent with SCE being a target of alcohol toxicity. Finally, in vivo evidence for SCE-driven adaptations to alcohol toxicity in the literature are discussed.

Establishment of a long-term hypertrophic scar model by injection of anhydrous alcohol: A rabbit model.

The processes of hypertrophic scar formation are extremely complex, and current animal models have limitations in terms of the complete characterization of lesions. An ideal animal model is indispensable for exploring the complex progression of scar formation to elucidate its pathophysiology and to perform therapeutic testing. This study aimed to establish a long-term, consistent and easily testable animal model by injecting anhydrous alcohol into the dorsal trunk dermis of rabbits. The rabbits were injected with different amounts of anhydrous alcohol. Anhydrous alcohol was infiltrated into the subcutaneous and superficial fascia. The optimal amount of anhydrous alcohol was determined by measuring the area and thickness of the scar. The typical model was established by determining the optimum dosage, and then we analysed the histological characteristics and fibrosis-associated protein expression. The dermal scar was generated by treating with 2 ml/kg anhydrous alcohol and displayed histopathologic features that characterize human hypertrophic scarring, including a parallel collagen fibre orientation, dermal and epidermal thickening, broad collagen deposition and the loss of dermal adnexal structures. The expression of fibrotic pan-markers was also enhanced. Moreover, the scar features and duration were compared between the anhydrous alcohol model and the rabbit ear model. Our results show that injecting anhydrous alcohol in the rabbit model thickened the dermal tissue, stimulated dermal fibroproliferation and resulted in hypertrophic scars with protein and histologic features similar to those seen in humans. Taken together, the findings from this study show that our model could be a feasible and useful tool for further research on the pathogenesis of hypertrophic scars.

High Resolution Genomic Scans Reveal Genetic Architecture Controlling Alcohol Preference in Bidirectionally Selected Rat Model.

Investigations on the influence of nature vs. nurture on Alcoholism (Alcohol Use Disorder) in human have yet to provide a clear view on potential genomic etiologies. To address this issue, we sequenced a replicated animal model system bidirectionally-selected for alcohol preference (AP). This model is uniquely suited to map genetic effects with high reproducibility, and resolution. The origin of the rat lines (an 8-way cross) resulted in small haplotype blocks (HB) with a corresponding high level of resolution. We sequenced DNAs from 40 samples (10 per line of each replicate) to determine allele frequencies and HB. We achieved ~46X coverage per line and replicate. Excessive differentiation in the genomic architecture between lines, across replicates, termed signatures of selection (SS), were classified according to gene and region. We identified SS in 930 genes associated with AP. The majority (50%) of the SS were confined to single gene regions, the greatest numbers of which were in promoters (284) and intronic regions (169) with the least in exon's (4), suggesting that differences in AP were primarily due to alterations in regulatory regions. We confirmed previously identified genes and found many new genes associated with AP. Of those newly identified genes, several demonstrated neuronal function involved in synaptic memory and reward behavior, e.g. ion channels (Kcnf1, Kcnn3, Scn5a), excitatory receptors (Grin2a, Gria3, Grip1), neurotransmitters (Pomc), and synapses (Snap29). This study not only reveals the polygenic architecture of AP, but also emphasizes the importance of regulatory elements, consistent with other complex traits.

Therapeutic and Protective Effects of Liposomal Encapsulation of Astaxanthin in Mice with Alcoholic Liver Fibrosis.

Astaxanthin (Asta) has been demonstrated to possess anti-inflammatory, antitumor, and free radical-clearing activities. However, the poor stability and low water solubility of Asta hamper its bioavailability. The objectives of this study were to fabricate Asta-loaded liposomes (Asta-lipo) and investigate the therapeutic effects of Asta-lipo on alcoholic liver fibrosis in mice. The mice were administered with Asta-lipo or liposomes alone prior to a 3-week dose containing 30% alcohol with or without feeding with a second dose of 30% alcohol. The prepared Asta-lipo of 225.0 ± 58.3 nm in diameter, had an encapsulation efficiency of 98%. A slow release profile of 16.2% Asta from Asta-lipo was observed after a 24-h incubation. Restorative actions against alcoholic liver fibrosis were observed after oral administration of Asta-lipo for 4 weeks. Hepatic repair, followed by a second dose of 30% alcohol, suggested that Asta-lipo exerted protective and reparative effects against liver injuries induced by repeated consumption of alcohol. The changes of serum ALT and AST values were principally in consistence with the histopathologic findings. Asta-lipo exerted rapid and direct effects against repeated alcohol-induced liver disease, whereas Asta-lipo given orally could boost recovery from liver injuries obtained due to previous long-term alcohol use. These data demonstrate that Asta-lipo has applicable protective and therapeutic potential to treat alcohol-induced liver diseases.

Tandem repeated irritation test (TRIT) studies and clinical relevance: Post 2006.

Background/Aims: Single or multiple applications of irritants can lead to occupational contact dermatitis, and most commonly irritant contact dermatitis (ICD). Tandem irritation, the sequential application of two irritants to a target skin area, has been studied using the Tandem Repeated Irritation Test (TRIT) to provide a more accurate representation of skin irritation. Objective: Here we present an update to Kartono's review on tandem irritation studies since 2006. Methods: We surveyed the literature available on PubMed, Embase, Google Scholar, and the UCSF Dermatology library databases since 2006. Results and discussion: The studies included discuss the tandem effects of common chemical irritants, organic solvents, occlusion as well as clinical relevance - and enlarge our ability to discern whether multiple chemical exposures are more or less likely to enhance irritation.

Lipopolysaccharide enhances TGF-ß1 signalling pathway and rat pancreatic fibrosis.

Pancreatic stellate cells (PSCs) play a critical role in fibrogenesis during alcoholic chronic pancreatitis (ACP). Transforming growth factor-beta1 (TGF-ß1) is a key regulator of extracellular matrix production and PSC activation. Endotoxin lipopolysaccharide (LPS) has been recognized as a trigger factor in the pathogenesis of ACP. This study aimed to investigate the mechanisms by which LPS modulates TGF-ß1 signalling and pancreatic fibrosis. Sprague-Dawley rats fed with a Lieber-DeCarli alcohol (ALC) liquid diet for 10 weeks with or without LPS challenge during the last 3 weeks. In vitro studies were performed using rat macrophages (Mφs) and PSCs (RP-2 cell line). The results showed that repeated LPS challenge resulted in significantly more collagen production and PSC activation compared to rats fed with ALC alone. LPS administration caused overexpression of pancreatic TLR4 or TGF-ß1 which was paralleled by an increased number of TLR4-positive or TGF-ß1-positive Mφs or PSCs in ALC-fed rats. In vitro, TLR4 or TGF-ß1 production in Mφs or RP-2 cells was up-regulated by LPS. LPS alone or in combination with TGF-ß1 significantly increased type I collagen and α-SMA production and Smad2 and 3 phosphorylation in serum-starved RP-2 cells. TGF-ß pseudoreceptor BAMBI production was repressed by LPS, which was antagonized by Si-TLR4 RNA or by inhibitors of MyD88/NF-kB. Additionally, knockdown of Bambi with Si-Bambi RNA significantly increased TGF-ß1 signalling in RP-2 cells. These findings indicate that LPS increases TGF-ß1 production through paracrine and autocrine mechanisms and that LPS enhances TGF-ß1 signalling in PSCs by repressing BAMBI via TLR4/MyD88/NF-kB activation.

Calcium signalling in the acinar environment of the exocrine pancreas: physiology and pathophysiology.

KEY POINTS: Ca2+ signalling in different cell types in exocrine pancreatic lobules was monitored simultaneously and signalling responses to various stimuli were directly compared. Ca2+ signals evoked by K+ -induced depolarization were recorded from pancreatic nerve cells. Nerve cell stimulation evoked Ca2+ signals in acinar but not in stellate cells. Stellate cells are not electrically excitable as they, like acinar cells, did not generate Ca2+ signals in response to membrane depolarization. The responsiveness of the stellate cells to bradykinin was markedly reduced in experimental alcohol-related acute pancreatitis, but they became sensitive to stimulation with trypsin. Our results provide fresh evidence for an important role of stellate cells in acute pancreatitis. They seem to be a critical element in a vicious circle promoting necrotic acinar cell death. Initial trypsin release from a few dying acinar cells generates Ca2+ signals in the stellate cells, which then in turn damage more acinar cells causing further trypsin liberation. ABSTRACT: Physiological Ca2+ signals in pancreatic acinar cells control fluid and enzyme secretion, whereas excessive Ca2+ signals induced by pathological agents induce destructive processes leading to acute pancreatitis. Ca2+ signals in the peri-acinar stellate cells may also play a role in the development of acute pancreatitis. In this study, we explored Ca2+ signalling in the different cell types in the acinar environment of the pancreatic tissue. We have, for the first time, recorded depolarization-evoked Ca2+ signals in pancreatic nerves and shown that whereas acinar cells receive a functional cholinergic innervation, there is no evidence for functional innervation of the stellate cells. The stellate, like the acinar, cells are not electrically excitable as they do not generate Ca2+ signals in response to membrane depolarization. The principal agent evoking Ca2+ signals in the stellate cells is bradykinin, but in experimental alcohol-related acute pancreatitis, these cells become much less responsive to bradykinin and then acquire sensitivity to trypsin. Our new findings have implications for our understanding of the development of acute pancreatitis and we propose a scheme in which Ca2+ signals in stellate cells provide an amplification loop promoting acinar cell death. Initial release of the proteases kallikrein and trypsin from dying acinar cells can, via bradykinin generation and protease-activated receptors, induce Ca2+ signals in stellate cells which can then, possibly via nitric oxide generation, damage more acinar cells and thereby cause additional release of proteases, generating a vicious circle.

Ethanol and Other Short-Chain Alcohols Inhibit NLRP3 Inflammasome Activation through Protein Tyrosine Phosphatase Stimulation.

Immunosuppression is a major complication of alcoholism that contributes to increased rates of opportunistic infections and sepsis in alcoholics. The NLRP3 inflammasome, a multiprotein intracellular pattern recognition receptor complex that facilitates the cleavage and secretion of the proinflammatory cytokines IL-1ß and IL-18, can be inhibited by ethanol, and we sought to better understand the mechanism through which this occurs and whether chemically similar molecules exert comparable effects. We show that ethanol can specifically inhibit activation of the NLRP3 inflammasome, resulting in attenuated IL-1ß and caspase-1 cleavage and secretion, as well as diminished apoptosis-associated speck-like protein containing a CARD (ASC) speck formation, without affecting potassium efflux, in a mouse macrophage cell line (J774), mouse bone marrow-derived dendritic cells, mouse neutrophils, and human PBMCs. The inhibitory effects on the Nlrp3 inflammasome were independent of γ-aminobutyric acid A receptor activation or N-methyl-d-asparate receptor inhibition but were associated with decreased oxidant production. Ethanol treatment markedly decreased cellular tyrosine phosphorylation, whereas administration of the tyrosine phosphatase inhibitor sodium orthovanadate prior to ethanol restored tyrosine phosphorylation and IL-1ß secretion subsequent to ATP stimulation. Furthermore, sodium orthovanadate-induced phosphorylation of ASC Y144, necessary and sufficient for Nlrp3 inflammasome activation, and secretion of phosphorylated ASC were inhibited by ethanol. Finally, multiple alcohol-containing organic compounds exerted inhibitory effects on the Nlrp3 inflammasome, whereas 2-methylbutane (isopentane), the analogous alkane of the potent inhibitor isoamyl alcohol (isopentanol), did not. Our results demonstrate that ethanol antagonizes the NLRP3 inflammasome at an apical event in its activation through the stimulation of protein tyrosine phosphatases, an effect shared by other short-chain alcohols.

Methods for enhancing cyanobacterial stress tolerance to enable improved production of biofuels and industrially relevant chemicals.

Cyanobacteria are photosynthetic prokaryotes that can fix atmospheric CO2 and can be engineered to produce industrially important compounds such as alcohols, free fatty acids, alkanes used in next-generation biofuels, and commodity chemicals such as ethylene or farnesene. They can be easily genetically manipulated, have minimal nutrient requirements, and are quite tolerant to abiotic stress making them an appealing alternative to other biofuel-producing microbes which require additional carbon sources and plants which compete with food crops for arable land. Many of the compounds produced in cyanobacteria are toxic as titers increase which can slow growth, reduce production, and decrease overall biomass. Additionally, many factors associated with outdoor culturing of cyanobacteria such as UV exposure and fluctuations in temperature can also limit the production potential of cyanobacteria. For cyanobacteria to be utilized successfully as biofactories, tolerance to these stressors must be increased and ameliorating stress responses must be enhanced. Genetic manipulation, directed evolution, and supplementation of culture media with antioxidants are all viable strategies for designing more robust cyanobacterial strains that have the potential to meet industrial production goals.

Toxicological assessment of lower alkyl methacrylate esters by a category approach.

Categories and read-across are essential tools for supplying information for assessments of endpoints without data while minimizing animal testing. This study is based on the guidance of ECHA in its Read-Across Framework (RAAF). A category of C1 - C8 alkyl methacrylate esters (methyl, ethyl, n-butyl, iso-butyl and 2-ethylhexyl) was constructed to fill in missing information for human health endpoints using read-across as a permitted adaptation under EU REACH. The esters form a series with common functional groups, small incremental changes of electrophilicity by molecular weight, and rapid hydrolysis by ester cleavage. Read-across is justified by two common specific modes of action, direct electrophilic reaction of the parent compounds and the potential inherent toxicities of the common metabolites methacrylic acid and the corresponding alcohols. The toxicological profile is very similar for all category members and not driven by the alcohol metabolites. Data gaps can be filled in with high confidence based on the number of studies available, the effects therein observed and the toxicological profiles of the hydrolysis products. The guidance provided by the RAAF enabled data gaps to be filled in a robust manner.

Understanding and engineering alcohol-tolerant bacteria using OMICS technology.

Microbes are capable of producing alcohols, making them an important source of alternative energy that can replace fossil fuels. However, these alcohols can be toxic to the microbes themselves, retaring or inhibiting cell growth and decreasing the production yield. One solution is improving the alcohol tolerance of such alcohol-producing organisms. Advances in omics technologies, including transcriptomic, proteomic, metabolomic, and genomic technologies, have helped us understand the complex mechanisms underlying alcohol toxicity, and such advances could assist in devising strategies for engineering alcohol-tolerant strains. This review highlights these advances and discusses strategies for improving alcohol tolerance using omics analyses.

Formulas for Calculated Osmolarity and Osmolal Gap: A Study of Diagnostic Accuracy.

BACKGROUND: The osmolal gap has been used for decades to screen for exposure to toxic alcohols. However, several issues may affect its reliability. We aimed to develop equations to calculate osmolarity with improved performance when used to screen for intoxication to toxic alcohols. STUDY DESIGN: Retrospective cohort study. SETTING & PARTICIPANTS: 7,525 patients undergoing simultaneous measurements of osmolality, sodium, potassium, urea, glucose, and ethanol or undergoing similar measurements performed within 30 minutes of a measurement of toxic alcohol levels at a single tertiary-care center from April 2001 to June 2016. Patients with detectable toxic alcohols were excluded. INDEX TEST: Equations to calculate osmolarity using multiple linear regression. OUTCOMES: The performance of new equations compared with published equations developed to calculate osmolarity, and to diagnose toxic alcohol intoxications more accurately. RESULTS: We obtained 7,525 measurements, including 100 with undetectable toxic alcohols. Among them, 3,875 had undetectable and 3,650 had detectable ethanol levels. In the entire cohort, the best equation to calculate osmolarity was 2.006×Na + 1.228×Urea + 1.387×Glucose + 1.207×Ethanol (values in mmol/L, R2=0.96). A simplified equation, 2.0×Na + 1.2×Urea + 1.4×Glucose + 1.2×Ethanol, had a similar R2 with 95% of osmolal gap values between -10.9 and 13.8. In patients with undetectable ethanol concentrations, the range of 95% of osmolal gap values was narrower than previous published formulas, and in patients with detectable ethanol concentrations, the range was narrower or similar. We performed a subanalysis of 138 cases for which both the toxic alcohol concentration could be measured and the osmolal gap could be calculated. Our simplified equation had superior diagnostic accuracy for toxic alcohol exposure. LIMITATIONS: Single center, no external validation, limited number of cases with detectable toxic alcohols. CONCLUSIONS: In a large cohort, coefficients from regression analyses estimating the contribution of glucose, urea, and ethanol were higher than 1.0. Our simplified formula to precisely calculate osmolarity yielded improved diagnostic accuracy for suspected toxic alcohol exposures than previously published formulas.

Use of in silico models for prioritization of heat-induced food contaminants in mutagenicity and carcinogenicity testing.

Numerous Maillard reaction and lipid oxidation products are present in processed foods such as heated cereals, roasted meat, refined oils, coffee, and juices. Due to the lack of experimental toxicological data, risk assessment is hardly possible for most of these compounds. In the present study, an in silico approach was employed for the prediction of the toxicological endpoints mutagenicity and carcinogenicity on the basis of the structure of the respective compound, to examine (quantitative) structure-activity relationships for more than 800 compounds. Five software tools for mutagenicity prediction (T.E.S.T., SARpy, CAESAR, Benigni-Bossa, and LAZAR) and three carcinogenicity prediction tools (CAESAR, Benigni-Bossa, and LAZAR) were combined to yield so-called mutagenic or carcinogenic scores for every single substance. Alcohols, ketones, acids, lactones, and esters were predicted to be mutagenic and carcinogenic with low probability, whereas the software tools tended to predict a considerable mutagenic and carcinogenic potential for thiazoles. To verify the in silico predictions for the endpoint mutagenicity experimentally, twelve selected compounds were examined for their mutagenic potential using two different validated in vitro test systems, the bacterial reverse mutation assay (Ames test) and the in vitro micronucleus assay. There was a good correlation between the results of the Ames test and the in silico predictions. However, in the case of the micronucleus assay, at least three substances, 2-amino-6-methylpyridine, 6-heptenoic acid, and 2-methylphenol, were clearly positive although they were predicted to be non-mutagenic. Thus, software tools for mutagenicity prediction are suitable for prioritization among large numbers of substances, but these predictions still need experimental verification.

Economic, human and environmental health benefits of replacing formaldehyde in the preservation of corpses.

Formaldehyde has been prominent in preserving biological tissues since the nineteenth century. Despite being admittedly harmful to health and to the environment, it is still widely used. The Morphology Department of the University of Brasília - Brazil, applied the rethink, reduce, reuse, recycle and responsibility methodology to their activities in an effort to protect the health of laboratory workers and users, save resources and reduce damage to the environment. Here we evaluate the results obtained a decade after the implementation of this proposal (2005-2015). Formaldehyde was replaced by alcohol and glycerol solutions in corpse conservation. Over five thousand dollars in public funds that would have been destined to buying preserving substances were saved annually, and over a hundred thousand liters of water that would have been contaminated and thrown into the sewage system were spared. The environment used to implement the study was improved and anatomical parts kept for study had their lifespan extended. It is noteworthy that such simple adjustments could cause pronounced changes in laboratory activities. We would avoid contaminating billions of liters of water and it would be possible to save millions if similar practices were implemented in all educational institutions having similar routines.

Association between alcohol intake and the risk of pancreatic cancer: a dose-response meta-analysis of cohort studies.

BACKGROUND: Studies examining the association between alcohol intake and the risk of pancreatic cancer have given inconsistent results. The purpose of this study was to summarize and examine the evidence regarding the association between alcohol intake and pancreatic cancer risk based on results from prospective cohort studies. METHODS: We searched electronic databases consisting of PubMed, Ovid, Embase, and the Cochrane Library identifying studies published up to Aug 2015. Only prospective studies that reported effect estimates with 95% confidence intervals (CIs) for the risk of pancreatic cancer, examining different alcohol intake categories compared with a low alcohol intake category were included. Results of individual studies were pooled using a random-effects model. RESULTS: We included 19 prospective studies (21 cohorts) reporting data from 4,211,129 individuals. Low-to-moderate alcohol intake had little or no effect on the risk of pancreatic cancer. High alcohol intake was associated with an increased risk of pancreatic cancer (risk ratio [RR], 1.15; 95% CI: 1.06-1.25). Pooled analysis also showed that high liquor intake was associated with an increased risk of pancreatic cancer (RR, 1.43; 95% CI: 1.17-1.74). Subgroup analyses suggested that high alcohol intake was associated with an increased risk of pancreatic cancer in North America, when the duration of follow-up was greater than 10 years, in studies scored as high quality, and in studies with adjustments for smoking status, body mass index, diabetes mellitus, and energy intake.. CONCLUSIONS: Low-to-moderate alcohol intake was not significantly associated with the risk of pancreatic cancer, whereas high alcohol intake was associated with an increased risk of pancreatic cancer. Furthermore, liquor intake in particular was associated with an increased risk of pancreatic cancer.

Alcohol-related breast cancer in postmenopausal women - effect of CYP19A1, PPARG and PPARGC1A polymorphisms on female sex-hormone levels and interaction with alcohol consumption and NSAID usage in a nested case-control study and a randomised controlled trial.

BACKGROUND: Alcohol consumption is associated with increased risk of breast cancer (BC), and the underlying mechanism is thought to be sex-hormone driven. In vitro and observational studies suggest a mechanism involving peroxisome proliferator-activated receptor gamma (PPARγ) in a complex with peroxisome proliferator-activated receptor gamma coactivator 1-α (PGC-1α) and interaction with aromatase (encoded by CYP19A1). Use of non-steroidal anti-inflammatory drugs (NSAID) may also affect circulating sex-hormone levels by modifying PPARγ activity. METHODS: In the present study we assessed whether genetic variation in CYP19A1 is associated with risk of BC in a case-control study group nested within the Danish "Diet, Cancer and Health" cohort (ncases = 687 and ncontrols = 687) and searched for gene-gene interaction between CYP19A1 and PPARGC1A, and CYP19A1 and PPARG, and gene-alcohol and gene-NSAID interactions. Association between the CYP19A1 polymorphisms and hormone levels was also examined among 339 non-HRT users. Incidence rate ratios were calculated based on Cox' proportional hazards model. Furthermore, we performed a pilot randomised controlled trial to determine the effect of the PPARG Pro(12)Ala polymorphism and the PPARγ stimulator Ibuprofen on sex-hormone levels following alcohol intake in postmenopausal women (n = 25) using linear regression. RESULTS: Genetic variations in CYP19A1 were associated with hormone levels (estrone: P rs11070844 = 0.009, estrone sulphate: P rs11070844 = 0.01, P rs749292 = 0.004, P rs1062033 = 0.007 and P rs10519297 = 0.03, and sex hormone-binding globulin (SHBG): P rs3751591 = 0.03) and interacted with alcohol intake in relation to hormone levels (estrone sulphate: P interaction/rs2008691 = 0.02 and P interaction/rs1062033= 0.03, and SHBG: P interaction/rs11070844 = 0.03). CYP19A1/rs3751591 was both associated with SHBG levels (P = 0.03) and with risk of BC (Incidence Rate Ratio = 2.12; 95 % Confidence Interval: 1.02-4.43) such that homozygous variant allele carriers had increased levels of serum SHBG and were at increased risk of BC. Acute intake of alcohol decreased blood estrone (P = <0.0001), estrone sulphate (P = <0.0001), and SHBG (P = 0.009) levels, whereas Ibuprofen intake and PPARG Pro(12)Ala genotype had no effect on hormone levels. CONCLUSIONS: Our results suggest that genetically determined variation in CYP19A1 is associated with differences in sex hormone levels. However, the genetically determined differences in sex hormone levels were not convincingly associated with BC risk. The results therefore indicate that the genetically determined variation in CYP19A1 contributes little to BC risk and to alcohol-mediated BC risk. TRIAL REGISTRATION: NCT02463383, June 3, 2015.

Alcohol-triggered signs of migraine: An animal model.

We describe an animal model where characteristics of migraine can be triggered by alcohol administration. In rats chronically implanted with a cannula overlying the transverse sinus, we applied potassium chloride (KCl) (or saline) to the meninges to sensitize trigeminovascular afferents. We assessed effects of repeated KCl application on animal behavior using conditioned place avoidance paradigm. In KCl-treated rats we discovered that alcohol injections (0.2 mg/kg), but not saline, resulted in the development of extracephalic allodynia and signs of ongoing pain.

Dietary Nucleotides Supplementation and Liver Injury in Alcohol-Treated Rats: A Metabolomics Investigation.

BACKGROUND: Previous studies suggested that nucleotides were beneficial for liver function, lipid metabolism and so on. The present study aimed to investigate the metabolic response of dietary nucleotides supplementation in alcohol-induced liver injury rats. METHODS: Five groups of male Wistar rats were used: normal control group (basal diet, equivalent distilled water), alcohol control group (basal diet, 50% alcohol (v/v)), dextrose control group (basal diet, isocaloric amount of dextrose), and 0.04% and 0.16% nucleotides groups (basal diet supplemented with 0.4 g and 1.6 g nucleotides kg(-1) respectively, 50% alcohol (v/v)). The liver injury was measured through traditional liver enzymes, expression of oxidative stress markers and histopathological examination. Ultra-performance liquid chromatography quadrupole-time-flight mass spectrometry (UPLC-Q-TOF-MS) was applied to identify liver metabolite profiles. RESULTS: Nucleotides supplementation prevented the progression of hepatocyte steatosis. The levels of total proteins, globulin, alanine aminotransferase, aspartate aminotransferase, total cholesterol triglyceride, as well as the oxidative stress markers altered by alcohol, were improved by nucleotides supplementation. Elevated levels of liver bile acids (glycocholic acid, chenodeoxyglycocholic acid, and taurodeoxycholic acid), as well as lipids (stearic acid, palmitic acid, oleic acid, phosphatidylcholine, and lysophosphatidylethanolamine) in alcohol-treated rats were reversed by nucleotides supplementation. In addition, supplementation with nucleotides could increase the levels of amino acids, including valyl-Leucine, L-leucine, alanyl-leucine and L-phenylalanine. CONCLUSION: These data indicate potential biomarkers and confirm the benefit of dietary nucleotides on alcoholic liver injury.