EtOH Group Research Articles

Proteomic Profiling of Liver and Plasma in Chronic Ethanol Feeding Model of Hepatic Alcohol Dehydrogenase-Deficient Deer Mice.

Chronic alcohol abuse, a major risk factor for such diseases as hepatitis and cirrhosis, impairs hepatic alcohol dehydrogenase (ADH; key ethanol [EtOH]-metabolizing enzyme). Therefore, differentially altered hepatic and plasma proteomes were identified in chronic EtOH feeding model of hepatic ADH-deficient (ADH- ) deer mice to understand the metabolic basis of alcoholic liver disease (ALD). ADH- deer mice were fed 3.5g% EtOH via Lieber-DeCarli liquid diet daily for 3months and histology of the liver assessed. Liver and plasma proteins were separated by 2-dimensional gel electrophoresis. The proteins differentially expressed were identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry. Histology of the liver showed panlobular steatosis and infiltration of T lymphocytes. Using the criteria of ≥1.5 for fold change (p-value ≤0.05) with expectation value (E ≤10-3 ) and protein score (≥64), 18 proteins in the livers and 5 in the plasma of EtOH-fed mice were differentially expressed and identified. Prolyl 4-hydroxylase, cytochrome b-5, endo A cytokeratin, ATP synthase, heat-shock 70kD proteins, enoyl CoA hydratase, stress-70 protein, peroxiredoxin 1, and ornithine carbamoyl transferase were up-regulated in the livers. However, carbonic anhydrase 3, mitochondrial ATP synthase, aldolase 2, actin γ, laminin receptor, and carbamoyl phosphate synthase were down-regulated. Contrary to the increased expression of creatine kinase M-type, a decreased expression of serine protease inhibitor A3A precursor, sulfated glycoprotein-2 (clusterin), and apolipoprotein E isoforms were found in the plasma of EtOH group. Chronic EtOH feeding in ADH- deer mice causes steatosis and infiltration of T lymphocytes in the livers along with increased expression of proteins involved in endoplasmic reticulum (ER) stress, fibrosis, fatty acid β oxidation and biogenesis, and decreased expression of proteins involved in ATP synthesis, carbohydrate metabolism, in cell regulation and architecture. Reduced expression of various carrier proteins as found in the plasma of EtOH group has a biomarker potential.

Acute Alcohol Binge Deteriorates Metabolic and Respiratory Compensation Capability After Blunt Chest Trauma Followed by Hemorrhagic Shock-A New Research Model.

The clinical relevance of blunt (thoracic) chest trauma (TxT) and hemorrhagic shock is indisputable due to the high prevalence of this injury type, as well as its close association with mortality and/or preventable deaths. Furthermore, there is an ongoing discussion about the influence of alcohol in trauma patients. Thus, we established a model of TxT followed by hemorrhagic shock with resuscitation (H/R) in alcohol-intoxicated rats. Depending on group allocation, 12 (subacute) or 2 (acute) hours before experimentation, the animals received a single oral dose of alcohol (ethanol [EtOH]) or saline (NaCl) followed by TxT, hemorrhagic shock (35±3mm Hg), and resuscitation (TxT+H/R). Arterial blood gas analyses and continuous monitoring of blood pressure were performed during the experimentation period. Survival during the experimentation procedure was determined. Subacute and acute EtOH group exhibited lower baseline mean arterial blood pressure values compared with the corresponding NaCl group, respectively. Both EtOH groups showed lower maximal bleed-out volume, which was necessary to induce hemorrhagic shock compared to NaCl groups, and the recovery during the resuscitation period was attenuated. During the experimentation in all groups, a trend to acidic pH was observed. Acute EtOH group showed lowest pH values compared to all other groups. Higher pCO2 values were observed in both EtOH groups. All groups developed negative base excess and decreasing HCO3- values until the end of hemorrhagic shock and showed increasing base excess and HCO3- values during resuscitation. Significantly higher mortality rate was found in the acute EtOH group. This study indicates that alcohol limits the metabolic and respiratory compensation capability, thereby promoting mortality.

Repeated ethanol exposure alters social behavior and oxidative stress parameters of zebrafish

Repeated ethanol (EtOH) consumption induces neurological disorders in humans and is considered an important public health problem. The physiological effects of EtOH are dose- and time-dependent, causing relevant changes in the social behavior. In addition, alcohol-induced oxidative stress has been proposed as a key mechanism involved in EtOH neurotoxicity. Here we investigate for the first time whether repeated EtOH exposure (REE) alters the social behavior of zebrafish and influences brain oxidation processes. Animals were exposed to water (control group) or 1% (v/v) EtOH (EtOH group) for 8 consecutive days (20min per day). EtOH was added directly to the tank water. At day 9, the social behavior and biochemical parameters were assessed. REE increased shoal cohesion by reducing inter-fish and farthest neighbor distances. SOD and CAT activities, as well as NPSH levels decreased in brain tissue. Moreover, REE increased lipid peroxidation suggesting oxidative damage. In summary, changes in oxidation processes may play a role in the CNS effects of EtOH, influencing the social behavior of zebrafish. Furthermore, in a translational neuroscience perspective, our data reinforces the utility of zebrafish to clarify the biochemical and behavioral effects of intermittent EtOH administration.

Effect of a heme oxygenase-1 inducer on NADPH oxidase expression in alcohol-induced liver injury in male Wistar rats

Purpose: To investigated the effect of hemin, a heme oxygenase-1 (HO-1) inducer, on nicotinamide adenine dinucleotide phosphate oxidase (NOX) expression in rats with alcohol-induced liver injury. Methods: Male Wistar rats were randomly divided into four groups consisting of the control group, the ethanol (EtOH) group, the EtOH + zinc protoporphyrin IX (ZnPP-IX) group and EtOH + hemin group. Hepatic NOX gene expression and immunohistochemistry of hepatic NOX1 and NOX4 were investigated in week 4. Results: EtOH significantly increased levels of NOX. An immunohistochemical study demonstrated a high number of immunopositive hepatocytes for NOX1 in the EtOH group and EtOH + ZnPP-IX group compared with the control group. Hemin administration downregulated NOX gene expression and lowered the number of immunopositive hepatocytes for NOX1. In contrast, ZnPP-IX (HO-1 inhibitor) administration caused upregulation of NOX gene expression and increased the number of immunopositive hepatocytes for NOX1. Conclusion: HO-1 inducer, hemin, alleviates oxidative stress-induced alcoholic liver injury by reducing NOX, especially NOX1. Keywords: NADPH oxidase, Immunohistochemistry, Heme oxygenase-1, Hemin, Reactive oxygen species, Alcohol-induced liver disease

Acanthoic Acid Can Partially Prevent Alcohol Exposure-Induced Liver Lipid Deposition and Inflammation.

Aims: The present study aims to detect the effect of acanthoic acid (AA) on alcohol exposure-induced liver lipid deposition and inflammation, and to explore the mechanisms.Methods: C57BL/6 mice were pretreated with single dose of AA (20 and 40 mg/kg) by oral gavage or equal volume of saline, and then exposed to three doses of ethanol (5 g/kg body weight, 25%, w/v) by gavage within 24 h. The mice were sacrificed at 6 h after the last ethanol dosing. Serum and hepatic indexes were detected by western blot, RT-PCR, and histopathological assay. AML-12 cells were pretreated with AA (5, 10, 20 μM), or AICAR (500 μM), GW3965 (1 μM), SRT1720 (6 μM), Nicotinamide (20 mM) for 2 h, respectively, and then following treated with EtOH (200 mM) and lipopolysaccharide (LPS) (10 ng/ml) for additional 48 h. Cell protein and mRNA were collected for western blot and RT-PCR. Cytokines interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) release were detected by ELISA assay.Results: It was found that AA significantly decreased acute ethanol-induced increasing of the serum ALT/AST, LDH, ALP levels, and hepatic and serum triglyceride levels, and reduced fat droplets accumulation in mice liver. AA significantly suppressed the levels of sterol regulatory element binding protein 1 (SREBP-1), cytochrome P4502E1 (CYP2E1), IL-1β, and caspase-1 induced by ethanol. Furthermore, a significant decline of sirtuin 1 (Sirt1) and liver X receptors (LXRs) levels was observed in EtOH group, compared with normal group mice. And AA pretreatment increased the Sirt1 and LXRs levels, and also ameliorated phosphorylation of liver kinase B-1 (LKB-1), adenosine monophosphate-activated protein kinase (AMPK), acetyl CoA carboxylase (ACC) proteins, compared with EtOH group. However, the levels of peroxisome proliferator activated receptor -α or -γ (PPAR-α or PPAR-γ) induced by acute ethanol were reversed by AA. In EtOH/LPS cultivated AML-12 cells, AA decreased IL-1β and TNF-α levels, lipid droplets, and SREBP-1 and CYP2E1 expressions, compared with EtOH/LPS treatment. AA also significantly increased protein expressions of Sirt1, p-LKB1, p-ACC, PPARα, and decreased protein expression of PPARγ, compared with EtOH/LPS treatment.Conclusion: Acanthoic acid can partially prevent alcohol exposure-induced liver lipid deposition and inflammation via regulation of LKB1/Sirt1/AMPK/ACC and LXRs pathways.

Effects of Alcohol on Postoperative Adhesion Formation in Ischemic Myocardium and Pericardium

Postoperative formation of adhesions increases risk of complications during cardiac reoperations. We previously demonstrated that swine supplemented with vodka had a significant reduction in adhesions at sternotomy after previous thoracotomy. This follow-up study was conducted to determine reproducibility and the mechanism for adhesion reduction in swine supplemented with ethanol. An ameroid constrictor was placed in the left circumflex in 14 male Yorkshire swine to induce chronic myocardial ischemia through left minithoracotomy. Animals were supplemented postoperatively with ethanol (45 g ETOH, n= 7) or sucrose (80 g SUC, n= 7) for 7 weeks, followed by a reoperative median sternotomy. The ETOH group had significantly fewer adhesions, thinner pericardial thickness, decreased intramyocardial fibrosis, and decreased myocardial collagen deposition compared with the SUC group. In the myocardium, ETOH animals had decreased expression of proadhesion proteins focal adhesion kinase, paxillin, integrin-β1, transforming growth factor-β1 and phosphorylated SMAD and increased expression of adhesion breakdown proteins matrix metalloproteinase (MMP) 1, MMP2, MMP3, and MMP9 compared with SUC animals. However in the pericardium, ETOH animals had increased expression of proadhesion proteins focal adhesion kinase, paxillin, phosphorylated paxillin, vinculin, integrin-β1, tumor necrosis factor-α, transforming growth factor-β, and phosphorylated SMAD3, and decreased expression of adhesion breakdown proteins MMP1, MMP3, MMP9, and plasmin compared with SUC animals. Alcohol supplementation substantially reduced postoperative pericardial adhesion formation, attenuated pericardial thickening, and reduced myocardial fibrosis in response to chronic ischemia. Alcohol supplementation modulates adhesion protein and MMP/tissue inhibitor of metalloproteinase expression, favoring a profile associated with reduced pericardial adhesions. These results suggest that the myocardium is the driving factor in reducing pericardial adhesions and mediating the postoperative healing process.

Rutin ameliorates oxidative stress and preserves hepatic and renal functions following exposure to cadmium and ethanol

Context: Rutin (RUT) is an antioxidant flavonoid with well-known metal chelating potentials.Objective: This study was designed to evaluate the protective effects of RUT against cadmium (Cd) + ethanol (EtOH)-induced hepatic and renal toxicity in rats.Materials and methods: Wistar rats were treated with Cd (50 mg/kg) alone or in combination with EtOH (5 mg/kg) and RUT (25, 50 and 100 mg/kg) for 15 days. After treatment, the liver, kidney and serum were removed for biochemical assays by spectrophotometric methods.Results: Serum, hepatic and renal malondialdehyde (MDA) levels were highest in the Cd + EtOH group and lowest in Cd + EtOH animals co-treated with the highest dose of RUT (2.98 ± 0.34, 10.08 ± 2.32, 4.99 ± 1.21 vs. 1.69 ± 0.33, 6.13 ± 0.28, 3.66 ± 1.12 μmol MDA/mg protein, respectively). The serum level of Cd was increased in the Cd + EtOH treated animals compared to Cd + EtOH animals co-treated with 100 mg/kg RUT (2.54 ± 0.08 vs. 1.28 ± 0.04 ppm). Furthermore, RUT at the highest dose protected against Cd + EtOH-induced elevation of bilirubin and uric acid levels as well as activities of lactate dehydrogenase and γ-glutamyl transferase (62.86 ± 2.74 vs. 122.52 ± 6.35 µmol/L; 1.77 ± 0.35 vs. 3.23 ± 0.55 mmol/L; 9.56 ± 1.22 vs. 16.21 ± 1.64 U/L; 288.92 ± 40.12 vs. 159.8 ± 18.01 U/L). The histo-pathological changes in the liver and kidney were also reduced in the Cd + EtOH animals co-treated with RUT in a dose-dependent manner.Discussion and conclusion: RUT protected against the combined effects of Cd + EtOH on hepatic and renal functions and improved the antioxidant defence system in the blood.

Hepatoprotective role of Solenostemma argel growing in Egypt on ethanol induced oxidative damage in rats

Abstract Objective The objective of the current study is to investigate the protective effect of Solenostemma argel (S. argel) treatment on ethanol (EtOH)-induced hepatotoxicity in rat liver. Methods Forty adult male albino rats were divided into four groups as control, S. argel (100 mg/kg), EtOH (3 g/kg) and S. argel + EtOH groups. S. argel was given 1 h prior to EtOH by oral gavage for 28 days. Results The results showed that, administration of EtOH caused a significant decrease (p<0.05) in serum total protein and albumin, whereas ALT and AST and lipid peroxidation (LPO) were increased following EtOH treatment. S. argel treatment significantly (p<0.05) ameliorated the previous parameters. Protein carbonyl, reduced glutathione and LPO levels were significantly (p<0.05) increased along with glutathione-S-transferase activity was decreased (as markers of oxidative stress) in EtOH treated rat liver. Previous oxidative stress was attenuated significantly by S. argel treatment. Moreover, in EtOH group, tumor necrosis factor-α (TNF-α) and nitric oxide (NO) contents and cytochrome P450 2E1 activity were significantly increased in liver tissues showing oxidative organ damage. Co-administration of S. argel with EtOH significantly reversed the inflammation in rat livers. Conclusion S. argel had a hepatoprotective role against EtOH-induce oxidative stress and inflammation in rat liver.

Pontine Changes in Metabolites and Axonal Fibres of Rats Following Four-week Alcohol Exposure: In Vivo Diffusion Tensor Imaging and 1h-magnetic Resonance Spectroscopy Study at 7.0 T.

Aims We characterized the metabolite content and axonal fibre changes in the rat pontine, in vivo , using 1H-magnetic resonance spectroscopy (1H-MRS) and diffusion tensor imaging (DTI) with a 7.0 T magnetic resonance imaging (MRI) scanner following chronic alcohol administration to explore new indicators for the early diagnosis and treatment of chronic alcoholism. Methods Forty male Sprague-Dawley rats were randomly divided into the EtOH and control groups (water group). To establish a chronic alcohol administration rat model, rats were intragastrically administered edible wine (56% vol/vol) twice daily for four weeks. Quantitation of the changes in metabolite concentrations and the apparent diffusion coefficient (ADC), as well as the fractional anisotropy (FA) values, within the pontine of brain stem were detected using MRS and DTI at 7.0 T. Differences of the pontine metabolite content, ADC and FA values between the two groups were compared. Results The 7.0 T MRIs showed no abnormal T2 phase pontine signals in either the EtOH or control groups. A 1H-MRS scan showed that the values for NAA, Cr and Cho in the EtOH group (4.54 ± 0.69, 4.69 ± 0.70 and 1.32 ± 0.18) were lower than those in the controls (6.78 ± 0.76, 8.50 ± 0.83 and 1.89 ± 0.24) ( P 0.05). DTI showed that the FA value in chronic alcohol exposure rats (0.41 ± 0.12) was lower than the control rats (0.53 ± 0.08) ( P 0.05). Conclusions After chronic alcohol exposure for four weeks, the metabolite content and axonal fibre changes in the rat pontine can be detected using MRS and DTI at 7.0 T earlier than conventional MRI, which provides information for the early diagnosis and treatment of chronic alcoholism. Short Summary In the present study, the authors showed that the metabolite content and axonal fibre changes in the pontine of chronic ethanol-treated rats can be detected using MRS and DTI at 7.0 T earlier than conventional MRI, which provide objective indicators for the early diagnosis and treatment of chronic alcoholism.

Role of resistance physical exercise in preventing testicular damage caused by chronic ethanol consumption in UChB rats.

Ethanol consumption is associated with spermatogenesis damage and testosterone level alterations. Alcohol remains the most commonly used substance among athletes and sports enthusiasts. This study evaluated whether resistance physical exercise can reduce the testicular damage caused by ethanol exposure. A total of 36 ethanol drinking (UChB) rats were divided into four groups: C (control rats), ETOH (ethanol consumption), ETOH + T (ethanol consumption + physical training), and T (group physical training). The physical training component of the T and ETOH + T groups was based on a resistance training model consisting of four sets of 10 jumps, with an increasing overload of 50-70% of the body weight attached to the chest three times per week. Rats in the ETOH and ETOH +T groups received 10% ethanol. At postnatal day 90, the rats were sacrificed. Blood sample was collected for hormonal analysis, and the testicles were weighed and processed for histopathological, morphometric, and immunohistochemical analyses. The ETOH group showed an increase in testosterone levels. The immunohistochemical of androgen receptor and the absolute weight of the testes were higher in the ETOH and ETOH + T groups, while the ETOH animals showed a decreased weight gain index. The number of abnormal seminiferous tubules increased in the ETOH and T groups compared to those in the control group (C); however, the association with treatment (ETOH + T group) prevented this effect and decreased caspase-3 production. In conclusion, these findings show that the combination of ethanol consumption and resistance physical exercise can prevent testicular damage in adult UChB rats.

Moderate-dose Regular Lifelong Alcohol Intake Changes the Intestinal Flora, Protects against Aging, and Keeps Spatial Memory in the Senescence-accelerated Mouse Prone 8 (SAMP8) Model.

Heavy and long-term alcohol consumption increase the risk of alcohol-related diseases. Epidemiological studies show moderate drinking reduces the risk of mortality, cardiovascular diseases, and brain infarction in the J-shaped or U-shaped curve effect. However, why moderate drinkers may be healthy and non-drinkers may be ill in diverse populations remains controversial. Herein, we examined the relationship between moderate/lifelong alcohol intake and aging, especially aging-related cognitive functions in senescence-accelerated mouse prone 8 (SAMP8) model. Methods: SAMP8 model (5-week-old, male, n = 36), a model of age-related cognitive deficit, were group-housed (n = 6/cage) and provided free access to water (water group, n = 18) or 1% ethanol (EtOH group, n = 18, intake started when mice were 9 weeks old). The object recognition test (ORT) and object location test (OLT) were used to evaluate cognitive functions. The intestinal flora at the age of 87 weeks was analyzed by terminal restriction fragment length polymorphism (T-RFLP). The lifespan of the EtOH-group mice was about 4 weeks longer than that of the water-group mice. In the EtOH group, spatial recognition impairment, assessed by OLT, was observed later (age, 73 weeks) than that in the water group (age, 52 weeks). The spinal curvature and skin conditions progressed significantly slower in the EtOH group than in the water group. Moreover, diarrhea symptoms only appeared in the water group, at the age of 82 weeks. The T-RFLP analysis of the intestinal flora indicated higher Lactobacillales order and lower Clostridium cluster XI in the EtOH group than in the water group, although those were extremely high in some mice close to death in both groups. Water-group mice with diarrhea presented significantly higher Clostridium cluster XI than did those without diarrhea (P = 0.017). Moderate alcohol intake changes intestinal flora and positively affects aging of SAMP8 model.

Protective effect of Pyropiayezoensis glycoprotein on chronic ethanol consumption-induced hepatotoxicity in rats.

The present study investigated the protective effect of Pyropiayezoensis glycoprotein (PYGP) against chronic ethanol consumption‑mediated hepatotoxicity in rats. Male Sprague-Dawley rats (n=20; 6weeks old) were randomly divided into four groups. The rats in each group were treated for 30days with the following: i)CON group, distilled water only; ii)EtOH group, 20% ethanol 3.7g/kg/BW; iii)EtOH+150group, 20%ethanol 3.7g/kg/BW+PYGP 150mg/kg/BW; iv)EtOH+300 group, 20% ethanol 3.7g/kg/BW+PYGP 300mg/kg/BW. EtOH, PYGP and water were orally administered. The rats were sacrificed after 30days, and blood and liver samples were collected for analysis. Treatment with ethanol caused significant elevation of serum levels of glutamic oxaloacetic transaminase (GOT) and glutamic pyruvic transaminase (GPT). Furthermore, inhibition of the antioxidant defense system in the liver, including glutathione (GSH), glutathione peroxidase (GSH‑px) and catalase (CAT) was observed. However, co‑administration with PYGP recovered the antioxidant defense system, and the serum levels of GOT and GPT. PYGP was shown to attenuate ethanol toxicity via the inactivation of mitogen‑activated protein kinases (MAKPs). PYGP suppressed the overexpression of cytochrome P4502E1 (CYP2E1), inducible nitric oxide synthase and cyclooxygenase‑2. These results suggested that the protective effect of PYGP was associated with antioxidant activities, MAPKs and the CYP2E1 signaling pathway.

The effects of repeated forced ethanol consumption during adolescence on reproductive behaviors in male rats.

Adolescence is a sensitive period of brain development when changes in hormone levels may have long-lasting effects on synaptic connections and behavior. In humans, alcohol consumption frequently begins during this critical period, although the impact of early exposure has not been fully examined. The current study was designed to investigate short- and long-term effects of repeated forced ethanol consumption during adolescence on emerging reproductive behaviors. Twenty-six young male Long-Evans rats were assigned to ethanol (Young EtOH, n=12) or water (Young Control, n=14) groups at postnatal day (P) 32, receiving a modified binge protocol of 3g/kg of solution via gavage twice per week from P32 to P80. For comparison, another cohort of rats received a similar treatment paradigm in adulthood from P75-P133 (Adult EtOH, n=8; Adult Control, n=10). Reproductive behavior was assessed with tests for copulation, partner preference, and 50-kHz vocalizations during forced consumption (intoxication) and again after a 4-5 week period of abstinence. During forced consumption, the Young EtOH group showed significantly longer latencies on copulation tests than Young Controls, but these differences did not persist after abstinence. Different patterns were observed in Adult animals, who only showed significant, delayed impairments in the post-ejaculatory interval. Preference for sexually receptive females increased with sexual experience in both adolescent and adult rats, regardless of treatment during the forced consumption phase. However, after abstinence, the Young EtOH group showed a significantly reduced partner preference compared to the Young Control group, which may indicate long-term effects on sexual motivation. Additionally, during forced consumption the Young EtOH group tended to emit fewer ultrasonic vocalizations, perhaps reflecting impairments in sexual communication. Adult groups showed no differences in partner preference or vocalization tests at any time. Taken together, these findings indicate that repeated, intermittent ethanol exposure may have moderate effects on reproductive behavior that vary as a function of age. After abstinence, differences were only observed in the younger group, suggesting that the adolescent brain and behavior are more sensitive to ethanol exposure than the adult brain for sexual motivation and performance.

Medial Prefrontal Cortical Dopamine Responses During Operant Self-Administration of Sweetened Ethanol.

Medial prefrontal cortex (mPFC) dysfunction is present in heavy alcohol consumers. Dopamine signaling in mPFC is associated with executive functioning and affects drinking behavior; however, direct measurement of extracellular mPFC dopamine during appetitive and consummatory ethanol (EtOH) self-administration behavior has not been reported. We used invivo microdialysis in freely behaving, adult, male, Long Evans rats to determine extracellular dopamine concentration in the mPFC during operant self-administration of an EtOH-plus-sucrose or sucrose solution. The model separated appetitive/seeking from consummatory phases of the operant session. Dopamine was also monitored in an untrained handling control group, and dialysate EtOH was measured in the EtOH-drinking group. Home cage baseline dopamine was lower in rats that experienced a week of drinking sweetened EtOH compared with sucrose-drinking and handling controls. Transfer into the operant chamber and the initiation of consumption stimulated a relatively higher change in dopamine over baseline in the sweetened EtOH group compared with sucrose and handling controls. However, all groups show a dopamine response during transfer into the operant chamber, and the sucrose group had a relatively higher change in dopamine over baseline during initiation of consumption compared with handling controls. The time courses of dopamine and EtOH in the mPFC differ in the EtOH-consuming rats. Differences in extracellular mPFC dopamine between EtOH drinkers compared with control groups suggest that mPFC dopamine is involved in the mechanism of operant self-administration of sweetened EtOH and sucrose. Furthermore, the increase in dopamine during consumption is consistent with a role of mPFC dopamine in reward prediction.

Protective Effects of Selenium, N-Acetylcysteine and Vitamin E Against Acute Ethanol Intoxication in Rats

The aim of this study was to determine possible protective influences of selenium (Se), N-acetylcysteine (NAC), and vitamin E (Vit E) against acute ethanol (EtOH) intoxication. Thirty-six rats were divided into six groups: I (control), II (EtOH), III (EtOH + Se), IV (EtOH + Vit E), V (EtOH + NAC), and VI (EtOH + mix). Except group I, EtOH was given the other pretreated (groups III, IV, V, and VI) and untreated groups (group II). Compared with the EtOH group, serum aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, lactate dehydrogenase, creatine kinase, and creatine kinase-MB levels were significantly decreased in all pretreated groups, whereas slightly diminished amylase and lipase were observed. Compared with the control group, a remarkably lower total antioxidant status (TAS), but higher total oxidant status (TOS), and oxidative stress index (OSI) were seen in brain, liver, and kidney tissues. The values of these parameters were less affected from EtOH-exposed brain tissue of EtOH + NAC and liver of EtOH + mix groups. Both significant decrease of catalase activity and marked increases of adenosine deaminase and myeloperoxidase were determined only in liver tissue of the EtOH group. Activities of these enzymes were restored in almost all pretreated groups. Moreover, an increase of xanthine oxidase activity was prevented in brain tissue of pretreated groups. In histopathological examination of the liver, hydropic degeneration, sinusoidal dilatation, mononuclear cell infiltration, and marked congestion, which were seen in the EtOH group, were prevented in all pretreated groups. Relative protection against acute EtOH toxicity, in both single and combined pretreatments of Se, NAC, and Vit E supplementation, was probably through antioxidant and free radical-neutralizing effects of foregoing materials.

Soy Protein Isolate Protects Against Ethanol-Mediated Tumor Progression in Diethylnitrosamine-Treated Male Mice.

In this study, diethylnitrosamine-treated male mice were assigned to three groups: (i) a 35% high fat ethanol liquid diet (EtOH) with casein as the protein source, (ii) the same EtOH liquid diet with soy protein isolate as the sole protein source (EtOH/SPI), (iii) and a chow group. EtOH feeding continued for 16 weeks. As expected, EtOH increased the incidence and multiplicity of basophilic lesions and adenomas compared with the chow group, P < 0.05. Soy protein replacement of casein in the EtOH diet significantly reduced adenoma progression when compared with the EtOH and EtOH/SPI group (P < 0.05). Tumor reduction in the EtOH/SPI group corresponded to reduced liver injury associated with decreased hepatic Tnfα and Cd14 antigen (Cd14) expression and decreased nuclear accumulation of NF-κB1 protein compared with the EtOH group (P < 0.05). Detection of sphingolipids using high-resolution matrix-assisted laser desorption/ionization-Fourier transform ion cyclotron resonance (MALDI-FTICR) imaging mass spectrometry revealed increased accumulation of long acyl chain ceramide species, and sphingosine-1-phosphate (S1P) in the EtOH group that were significantly reduced in the EtOH/SPI group. Chronic EtOH feeding also increased mRNA expression of β-catenin transcriptional targets, including cyclin D1 (Ccnd1), matrix metallopeptidase 7 (Mmp7), and glutamine synthetase (Glns), which were reduced in the EtOH/SPI group (P < 0.05). We conclude that soy prevents tumorigenesis by reducing proinflammatory and oxidative environment resulting from EtOH-induced hepatic injury, and by reducing hepatocyte proliferation through inhibition of β-catenin signaling. These mechanisms may involve changes in sphingolipid signaling. Cancer Prev Res; 9(6); 466-75. ©2016 AACR.

Chronic plus binge ethanol feeding induces myocardial oxidative stress, mitochondrial and cardiovascular dysfunction, and steatosis.

Alcoholic cardiomyopathy in humans develops in response to chronic excessive alcohol consumption; however, good models of alcohol-induced cardiomyopathy in mice are lacking. Herein we describe mouse models of alcoholic cardiomyopathies induced by chronic and binge ethanol (EtOH) feeding and characterize detailed hemodynamic alterations, mitochondrial function, and redox signaling in these models. Mice were fed a liquid diet containing 5% EtOH for 10, 20, and 40 days (d) combined with single or multiple EtOH binges (5 g/kg body wt). Isocalorically pair-fed mice served as controls. Left ventricular (LV) function and morphology were assessed by invasive pressure-volume conductance approach and by echocardiography. Mitochondrial complex (I, II, IV) activities, 3-nitrotyrosine (3-NT) levels, gene expression of markers of oxidative stress (gp91phox, p47phox), mitochondrial biogenesis (PGC1α, peroxisome proliferator-activated receptor α), and fibrosis were examined. Cardiac steatosis and fibrosis were investigated by histological/immunohistochemical methods. Chronic and binge EtOH feeding (already in 10 days EtOH plus single binge group) was characterized by contractile dysfunction (decreased slope of end-systolic pressure-volume relationship and preload recruitable stroke work), impaired relaxation (decreased time constant of LV pressure decay and maximal slope of systolic pressure decrement), and vascular dysfunction (impaired arterial elastance and lower total peripheral resistance). This was accompanied by enhanced myocardial oxidative/nitrative stress (3-NT; gp91phox; p47phox; angiotensin II receptor, type 1a) and deterioration of mitochondrial complex I, II, IV activities and mitochondrial biogenesis, excessive cardiac steatosis, and higher mortality. Collectively, chronic plus binge EtOH feeding in mice leads to alcohol-induced cardiomyopathies (National Institute on Alcohol Abuse and Alcoholism models) characterized by increased myocardial oxidative/nitrative stress, impaired mitochondrial function and biogenesis, and enhanced cardiac steatosis.

Synthesis of layered PLGA-PLA microparticles for tunable delayed protein release

Event Abstract Back to Event Synthesis of layered PLGA-PLA microparticles for tunable delayed protein release Dipankar Dutta1, Chase Fauer1, Mariama Salifu1 and Sarah E. Stabenfeldt1 1 Arizona State University, School of Biological and Health Systems Engineering, United States Introduction: Layered microparticles (MPs) formulated using poly(L-lactic acid) (PLA) and poly(D,L-lactic-co-glycolic acid) (PLGA) have been widely investigated for delayed release of small molecule drugs. However, the extension to protein delivery is limited even though temporal presentation/activity of proteins is critical for cellular signalling. Additionally, most reports rely on complex tools (i.e. tricapillary electrosprayer) to successfully fabricate layered MPs[1]. Here, we report a straightforward protocol for: 1) encapsulating the model protein, bovine serum albumin (BSA) in core-shell MPs, and 2) modulating the protein release profile using ethanol (EtOH). Materials and Methods: Protein-loaded PLGA-PLA particles were formulated by adapting a previously described technique to encapsulate BSA[2]. Both PLGA and PLA were dissolved in dichloromethane where EtOH was added (0, 2.5 & 5%; v/v) to the PLA solution. The aqueous protein phase was mixed with the PLGA solution, transferred to the PLA solution and emulsified in a 4x volume excess of 1.0% (w/v) poly-vinyl alcohol (PVA) using a table-top vortexer. After solvent evaporation in 15x excess of 0.5% PVA, the MPs were recovered by subsequent washes and lyophilization. Particle size and architecture were evaluated through scanning electron microscopy (SEM). Release profiles were determined from protein loading and release assays. Protein localization (fluorophore-conjugated insulin) within particles was quantified using confocal microscopy and MATLAB. Results: Average particle diameters ranged between 32-44µm and showed an increasing trend with the addition of EtOH. SEM images of freshly prepared and DMSO exposed particles verified a core-shell architecture with a PLGA-rich core and a PLA-rich shell (Figures 1 & 2). Protein encapsulation efficiencies were between 65-73% and release profiles indicate an initial burst release between 12-20% of cumulative cargo. Marked differences are seen in cumulative release over 40days: 32%, 43%, and 100% for 0%, 2.5% & 5% (v/v) EtOH groups, respectively (Figure 3A). Confocal images indicate protein localization within the core of PLGA-PLA MPs. Upon addition of EtOH, protein distribution was more diffuse across MPs (Figure 3B). Discussion: To our knowledge, we are the first to demonstrate that adding EtOH in the shell phase modulates release profiles in core-shell (PLGA-PLA) MPs. EtOH likely influences: 1) protein mobility within the oil phase, and 2) infiltration of aqueous continuous phase into the dispersed oil phase during solvent evaporation. The result is a more diffuse protein distribution (Figure 3B) within MPs and a relatively porous surface morphology (as seen from SEM). Release profiles indicate EtOH can modulate protein release rate and delay period. Conclusion: BSA was encapsulated in core-shell PLGA-PLA particles using a straightforward protocol and we report ethanol as a novel means for tuning their release profile. Dr. Rachael Sirianni, Barrow Neurological Institute, Phoenix, AZ; John M. Cowley Center for High Resolution Electron Microscopy, Tempe, AZ

Ethanol and Acetaldehyde After Intraperitoneal Administration to Aldh2-Knockout Mice-Reflection in Blood and Brain Levels.

This paper reports, for the first time, on the analysis of ethanol (EtOH) and acetaldehyde (AcH) concentrations in the blood and brains of Aldh2-knockout (Aldh2-KO) and C57B6/6J (WT) mice. Animals were administrated EtOH (1.0, 2.0 or 4.0g/kg) or 4-methylpyrazole (4-MP, 82mg/kg) plus AcH (50, 100 or 200mg/kg) intraperitoneally. During the blood tests, samples from the orbital sinus of the eye were collected. During the brain tests, dialysates were collected every 5min (equal to a 15µl sample) from the striatum using in vivo brain microdialysis. Samples were collected at 5, 10, 15, 20, 25, 30 and 60min intervals post-EtOH and -AcH injection, and then analyzed by head-space GC. In the EtOH groups, high AcH levels were found in the blood and brains of Aldh2-KO mice, while only small traces of AcH were seen in the blood and brains of WT mice. No significant differences in EtOH levels were observed between the WT and the Aldh2-KO mice for either the EtOH dose. EtOH concentrations in the brain were comparable to the EtOH concentrations in the blood, but the AcH concentrations in the brain were four to five times lower compared to the AcH concentrations in the blood. In the AcH groups, high AcH levels were found in both WT and Aldh2-KO mice. Levels reached a sharp peak at 5min and then quickly declined for 60min. Brain AcH concentrations were almost equal to the concentrations found in the blood, where the AcH concentrations were approximately two times higher in the Aldh2-KO mice than in the WT mice, both in the blood and the brain. Our results suggest that systemic EtOH and AcH administration can cause a greater increase in AcH accumulation in the blood and brains of Aldh2-KO mice, where EtOH concentrations in the Aldh2-KO mice were comparable to the EtOH concentrations in the WT mice. Furthermore, detection of EtOH and AcH in the blood and brain was found to be dose-dependent in both genotypes.

Heme oxygenase-1 alleviates alcoholic liver steatosis: histopathological study.

Excessive alcohol consumption is one of the most important causes of hepatic steatosis, which involves oxidative stress. In particular, increased oxidative stress has been strongly linked to stimulation of the expression of heme oxygenase-1 (HO-1). This study aimed to investigate whether HO-1 could alleviates alcoholic steatosis in rats. Male Wistar rats were randomly divided into 4 groups: 1) the control group, 2) the EtOH group, 3) the EtOH + ZnPP-IX group and 4) the EtOH + Hemin group. Liver histopathology was investigated in weeks 1 and 4 after the start of the treatment period. Alcohol treatment significantly increased the hepatic malondialdehyde (MDA) levels, an oxidative stress marker. In addition, it increased the triglyceride, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels in both weeks. Gross examination demonstrated a yellowish and slightly enlarged liver in the alcohol-treated rats. Hematoxylin and eosin (H&E) and Oil Red O staining indicated hepatic steatosis, which was characterized by diffuse, extensive fatty accumulation and discrete lipid droplets of variable size in hepatocytes of the alcohol-treated rats. Administration of the HO-1 inducer hemin resulted in upregulation of hepatic HO-1 gene expression, reduced the MDA, triglyceride, ALT and AST levels and alleviated alcoholic hepatic steatosis, whereas administration of the HO-1 inhibitor zinc protoporphyrin IX (ZnPP-IX) resulted in downregulation of hepatic HO-1 gene expression and could not alleviate alcoholic hepatic steatosis either week. In conclusion, HO-1 could alleviate alcoholic hepatic steatosis in male Wistar rats and may be useful in development of a new therapeutic approach.