metallothionein and Alcoholism

Alcohol-associated liver disease is a spectrum of liver disorders with histopathological changes ranging from simple steatosis to steatohepatitis, cirrhosis, and hepatocellular carcinoma. Recent data suggest that chronic-plus-binge ethanol intake induces steatohepatitis by promoting release by hepatocytes of proinflammatory mitochondrial DNA-enriched (mtDNA-enriched) extracellular vesicles (EVs). The aim of the present study was to investigate the role of the stress kinase apoptosis signal-regulating kinase 1 (ASK1) and p38 mitogen-activated protein kinase (p38) in chronic-plus-binge ethanol-induced steatohepatitis and mtDNA-enriched EV release. Microarray analysis revealed the greatest hepatic upregulation of metallothionein 1 and 2 (Mt1/2), which encode 2 of the most potent antioxidant proteins. Genetic deletion of the Mt1 and Mt2 genes aggravated ethanol-induced liver injury, as evidenced by elevation of serum ALT, neutrophil infiltration, oxidative stress, and ASK1/p38 activation in the liver. Inhibition or genetic deletion of Ask1 or p38 ameliorated ethanol-induced liver injury, inflammation, ROS levels, and expression of phagocytic oxidase and ER stress markers in the liver. In addition, inhibition of ASK1 or p38 also attenuated ethanol-induced mtDNA-enriched EV secretion from hepatocytes. Taken together, these findings indicate that induction of hepatic mtDNA-enriched EVs by ethanol is dependent on ASK1 and p38, thereby promoting alcoholic steatohepatitis.

Topics: Alcoholism; Alcohols; Animals; Binge Drinking; Disease Models, Animal; DNA, Mitochondrial; Extracellular Vesicles; Fatty Liver, Alcoholic; Hepatocytes; Humans; Inflammation; Liver; MAP Kinase Kinase Kinase 5; Matrix Metalloproteinase 14; Metallothionein; Mice; p38 Mitogen-Activated Protein Kinases; Signal Transduction

Chronic alcohol abuse has been linked to the disruption of executive function and allostatic conditioning of reward response dysregulation in the mesocorticolimbic pathway (MCL). Here, we analyzed genome-wide mRNA and miRNA expression from matched cases with alcohol dependence (AD) and controls (n = 35) via gene network analysis to identify unique and shared biological processes dysregulated in the prefrontal cortex (PFC) and nucleus accumbens (NAc). We further investigated potential mRNA/miRNA interactions at the network and individual gene expression levels to identify the neurobiological mechanisms underlying AD in the brain. By using genotyped and imputed SNP data, we identified expression quantitative trait loci (eQTL) uncovering potential genetic regulatory elements for gene networks associated with AD. At a Bonferroni corrected p≤0.05, we identified significant mRNA (NAc = 6; PFC = 3) and miRNA (NAc = 3; PFC = 2) AD modules. The gene-set enrichment analyses revealed modules preserved between PFC and NAc to be enriched for immune response processes, whereas genes involved in cellular morphogenesis/localization and cilia-based cell projection were enriched in NAc modules only. At a Bonferroni corrected p≤0.05, we identified significant mRNA/miRNA network module correlations (NAc = 6; PFC = 4), which at an individual transcript level implicated miR-449a/b as potential regulators for cellular morphogenesis/localization in NAc. Finally, we identified eQTLs (NAc: mRNA = 37, miRNA = 9; PFC: mRNA = 17, miRNA = 16) which potentially mediate alcohol's effect in a brain region-specific manner. Our study highlights the neurotoxic effects of chronic alcohol abuse as well as brain region specific molecular changes that may impact the development of alcohol addiction.

Topics: Alcoholism; Chronic Disease; Cluster Analysis; Gene Expression Profiling; Gene Expression Regulation; Gene Regulatory Networks; Humans; Metallothionein; MicroRNAs; Nucleus Accumbens; Prefrontal Cortex; Quantitative Trait Loci

Oxidative stress is implicated in pathogenesis of alcoholic liver disease (ALD). This study investigated the possible correlation among the erythrocyte indices of oxidative stress, the leukocyte panels of antioxidant proteins (metallothioneins), the serum biochemical parameters and the liver steatosis grade.. A total of 118 cases including 60 alcoholic subjects and 58 controls were enrolled. All the alcoholic subjects were screened for body mass index (BMI), liver steatosis, and blood chemistry and serology. The level of oxidative stress and oxidative stress-related parameters were measured in the blood and correlated with clinical findings.. Alcoholic subjects showed higher BMI, moderate/severe hepatic steatosis, increase in the levels of triglycerides, cholesterol, glucose, γ-glutamyl-transpeptidase (GGT), alanine aminotransferase (ALT), bilirubin, alpha 1 and beta 2 globulins, iron and a decrease in the levels of aspartate aminotransferase (AST) and beta 1 globulin with respect to the reference values. Moreover, alcoholic subjects showed: (i) an increase in Thiobarbituric Acid Reactive Substance (TBARS) content representing a good estimation of global oxidative stress; (ii) a stimulation of the activities of the antioxidant enzymes catalase and SOD; (iii) a modulation of expression of metallothioneins, with a down-regulation of MT-1A and an up-regulation of MT-1E isoforms.. Our data suggest that alcoholism is strongly associated with altered pattern of blood metallothioneins; this parameter combined with the score calculated by an ad hoc implemented algorithm (HePaTest) could offer a non-invasive alternative approach for evaluating alcohol-related damages and could be used in follow-up of alcoholic patients.

Topics: Adolescent; Adult; Alcoholism; Antioxidants; Case-Control Studies; Erythrocyte Indices; Fatty Liver; Humans; Leukocytes; Lipid Peroxidation; Liver Diseases, Alcoholic; Male; Metallothionein; Middle Aged; Oxidative Stress; Ultrasonography; Up-Regulation; Young Adult

Chronic alcohol abuse impairs both alveolar epithelial and macrophage function, and renders individuals susceptible to acute lung injury, pneumonia, and other serious lung diseases. Zinc deficiency, which is known to impact both epithelial and immune cell functions, is also associated with alcohol abuse. In this study, chronic alcohol ingestion (6 wk) in rats altered expression of key zinc transporters and storage proteins in the small intestine and the lung, and decreased zinc levels in the alveolar compartment. Zinc supplementation of alveolar epithelial monolayers derived from alcohol-fed rats in vitro, or of the diets of alcohol-fed rats in vivo, restored alveolar epithelial barrier function, and these improvements were associated with salutary changes in tight junction protein expression and membrane localization. In parallel, dietary zinc supplementation increased intracellular zinc levels, GM-CSF receptor expression, and bacterial phagocytic capacity in the alveolar macrophages of alcohol-fed rats. Together, these studies implicate zinc deficiency as a novel mechanism mediating alcohol-induced alveolar epithelial and macrophage dysfunction. Importantly, these findings argue that dietary supplementation can overcome alcohol-induced zinc deficiency and restore alveolar epithelial and macrophage function, and therefore could be an effective treatment for the susceptible alcoholic lung phenotype.

Topics: Alcoholism; Animals; Bronchoalveolar Lavage Fluid; Carrier Proteins; Cation Transport Proteins; Cell Line; Dietary Supplements; Epithelial Cells; Ethanol; Gene Expression Regulation; Humans; Macrophages, Alveolar; Male; Membrane Proteins; Membrane Transport Proteins; Metallothionein; Occludin; Phosphoproteins; Pulmonary Alveoli; Rats; Rats, Sprague-Dawley; Receptors, Granulocyte-Macrophage Colony-Stimulating Factor; Zinc; Zonula Occludens-1 Protein

We have previously demonstrated that ethanol teratogenicity is associated with metallothionein-induced fetal zinc (Zn) deficiency, and that maternal subcutaneous Zn treatment given with ethanol in early pregnancy prevents fetal abnormalities and spatial memory impairments in mice. Here we investigated whether dietary Zn supplementation throughout pregnancy can also prevent ethanol-related dysmorphology.. Pregnant mice were injected with saline or 25% ethanol (0.015 ml/g intraperitoneally at 0 and 4 hours) on gestational day (GD) 8 and fed either a control (35 mg Zn/kg) or a Zn-supplemented diet (200 mg Zn/kg) from GD 0 to 18. Fetuses from the saline, saline + Zn, ethanol and ethanol + Zn groups were assessed for external birth abnormalities on GD 18. In a separate cohort of mice, postnatal growth and survival of offspring from these treatment groups were examined from birth until postnatal day 60.. Fetuses from dams treated with ethanol alone in early pregnancy had a significantly greater incidence of physical abnormalities (26%) compared to those from the saline (10%), saline + Zn (9%), or ethanol + Zn (12%) groups. The incidence of abnormalities in ethanol + Zn-supplemented fetuses was not different from saline-treated fetuses. While ethanol exposure did not affect the number of fetal resorptions or pre- or postnatal weight, there were more stillbirths with ethanol alone, and cumulative postnatal mortality was significantly higher in offspring exposed to ethanol alone (35% deaths) compared to all other treatment groups (13.5 to 20.5% deaths). Mice supplemented with Zn throughout pregnancy had higher plasma Zn concentrations than those in un-supplemented groups.. These findings demonstrate that dietary Zn supplementation throughout pregnancy ameliorates dysmorphology and postnatal mortality caused by ethanol exposure in early pregnancy.

Topics: Abnormalities, Drug-Induced; Alcoholism; Animals; Central Nervous System Depressants; Dietary Supplements; Disease Models, Animal; Embryonic Development; Ethanol; Female; Liver; Male; Metallothionein; Mice; Mice, Inbred C57BL; Pregnancy; Pregnancy Outcome; Pregnancy, Animal; Prenatal Exposure Delayed Effects; Sodium Chloride; Zinc

Metallothioneins (MTs) are ubiquitously expressed intracellular proteins that bind heavy metals and are involved in cytoprotection against several types of stress agents including chemicals, hormones, and oxidants. We have previously reported 1 isoform, MT-II, as a possible candidate gene for ethanol (EtOH) preference (EP) determination in mice.. Semiquantitative RT-PCR was used to determine brain mRNA levels of MT-I and MT-III in 4 inbred mouse strains with variable EP. Following this, cosegregation of MT-II brain expression with EP was analyzed in F2 mice from 2 intercrosses (C57BL/6J x BALB/cJ and C57BL/6J x DBA/2J). Studies on MT-I/MT-II knockout (KO) mice were also undertaken to further explore this relationship.. Our results suggest that MT-I is responsive to EtOH, with no evidence of basal-level differences between strains. Conversely, MT-III shows no EtOH response, yet indicates a possible strain-specific feature with C57BL/6J having the lowest levels of brain MT-III. Metallothionein-II expression cosegregates with EP in F2 mice from a C57BL/6J (preferring) and DBA/2J (avoiding) intercross. Although F2 mice from a cross with C57BL/6J and BALB/cJ (avoiding) strains follow a similar pattern, the results are not statistically significant. Metallothionein-I/MT-II knockout (MT-KO) mice appear to have smaller litter sizes as well as higher weight compared with controls (129S1/SvImJ) and also show a slight increase in EP.. Metallothionein-II remains the primary candidate of the mouse MT gene family for involvement in EP. Its effect on EP appears to be dependent on the genetic background. Such conclusions are based on results from C57BL/6J, BALB/cJ, DBA/2J, and 129 inbred mouse strains. Evidence also points to shared neural pathways involved in weight gain and obesity. The complex interactions between MT-II, EP, and weight gain/obesity remain to be studied.

Topics: Alcoholism; Animals; Brain; Chromosome Segregation; Crosses, Genetic; Metallothionein; Metallothionein 3; Mice; Mice, Inbred Strains; Mice, Knockout; Nerve Tissue Proteins; Protein Isoforms; RNA, Messenger; Species Specificity

Hepatic lipid peroxidation, metallothioneins, collagen and proline hydroxylase activity were investigated in 16 ethanol-fed rats and in 16 control animals. The rats were further divided into three groups to receive either a standard diet, a zinc-deficient diet or a zinc-supplemented diet. The animals were sacrificed at week 12 of the experiment for histological and biochemical assessments. Hepatic tissue examination indicated that oral zinc supplementation was associated with a decrease in lipid peroxidation, collagen deposition and proline hydroxylase activity together with an increase in metallothionein concentration in alcoholic rats. There were no significant differences in lipid peroxidation in the control group in relation to the diet. Zinc supplementation was associated with increased concentrations of hepatic metallothioneins together with decreased concentrations of proline-hydroxylase and collagen but to a lesser degree than in alcoholic animals. These results indicate that zinc is an efficient hepato-protective agent against lipid peroxidation in alcoholic rats and its effect may be, in part, mediated by the activation of metallothionein synthesis. Also, lipid peroxidation may be related to changes in hepatic collagen synthesis.

Topics: Alcoholism; Animals; Collagen; Disease Models, Animal; Ethanol; Food, Fortified; Lipid Peroxidation; Liver; Male; Metallothionein; Procollagen-Proline Dioxygenase; Rats; Rats, Wistar; Zinc

The content of metallothionein (MT) was determined in the livers (MT-L) and the kidneys (MT-K) of 145 deceased persons. The individual values showed marked variations; average levels (arithmetic means +/- S.D.) were 154.9 +/- 151.4 mg/kg liver wet wt. and 160.5 +/- 150.4 mg/kg total kidney wet wt. In contrast to MT-L, MT-K increased with age up to a maximum around mid-life and decreased at higher ages. Neither the MT-L nor the MT-K depend on sex. The MT content significantly correlates with the macroscopic and histopathologic status of the liver. As compared to normal tissue, livers with fatty degeneration, cirrhosis and brown atrophy show MT-L values of 40%, 25% and 233% respectively. Low MT-L of 40% of the control values are also observed in individuals with a history of alcohol abuse. This decline preceded visible macroscopic pathological findings. MT-L also responds to the cause of death. At suicidal overdose of drugs the MT-L is lowered to 62% and at "mechanical" suicides it is increased to 137%. Whereas MT-K is essentially independent of the kidney status, it is significantly increased to approximately 190% in male smokers.

Topics: Adolescent; Adult; Age Factors; Aged; Aged, 80 and over; Alcoholism; Cause of Death; Child; Humans; Kidney; Liver; Metallothionein; Middle Aged; Sex Characteristics; Smoking

In mice 4-12 weeks ingestion of ethanol decreased the concentration of a metallothionein-like protein fraction in the liver but not in the kidneys. The zinc and copper concentration also tended to decrease in the liver and remained unaffected in the kidneys.

Topics: Alcoholism; Animals; Copper; Female; Kidney; Liver; Metallothionein; Mice; Organ Specificity; Zinc