The role of ethanol metabolism in development of alcoholic steatohepatitis in the rat

The importance of ethanol metabolism in the development of alcoholic liver disease remains controversial.

The present study examined the effects of selective inhibition of the cytochrome P450 enzyme CYP2E1 compared with the inhibition of overall ethanol metabolism on the development of alcoholic steatohepatitis.

Adult male Sprague–Dawley rats were fed via total enteral nutrition for 45 days with or without 10–12g/kg/d ethanol. Some groups were given 200mg/kg/d of the CYP2E1 inhibitor diallyl sulfide (DAS). Other groups were treated with 164mg/kg/d of the alcohol dehydrogenase (ADH) inhibitor 4-methylpyrazole (4-MP) and dosed at 2–3g/kg/d ethanol to maintain similar average urine ethanol concentrations.

Liver pathology scores and levels of apoptosis were elevated by ethanol (P.05) but did not differ significantly on cotreatment with DAS or 4-MP.

However, liver triglycerides were lower when ethanol-fed rats were treated with DAS or 4-MP (P.05). Serum alanine aminotransferase values were significantly lower in ethanol-fed 4-MP–treated rats indicating reduced necrosis.

Hepatic oxidative stress and the endoplasmic reticulum (ER) stress marker tribbles-related protein 3 were increased after ethanol (P.05); further increased by DAS but partly attenuated by 4-MP.

Both DAS and 4-MP reversed ethanol increases in the cytokine, tumor necrosis factor-alpha (TNF-α), and the chemokine CXCL-2 (P.05). However, neither inhibitors prevented ethanol suppression of interleukins IL-4 or IL-12. Moreover, neither inhibitors prevented ethanol increases in tumor growth factor-beta mRNA.

Ethanol and DAS additively induced hepatic hyperplasia (P.05).

These data suggest that a significant proportion of hepatic injury after ethanol exposure is independent of alcohol metabolism. Ethanol metabolism by CYP2E1 may be linked in part to triglyceride accumulation, to induction of TNF-α, and to chemokine production. Ethanol metabolism by ADH may be linked in part to oxidative and ER stress and necrotic injury.

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Alcohol consumption among patients with hepatitis B infection in northern Portugal considering gender and hepatitis B virus genotype differences

Alcohol abuse is an important public health problem. In Portugal with a population of 10 millions of inhabitants, there are around 10% of alcoholics or excessive alcohol drinkers and 1% of chronically infected patients with hepatitis B virus (HBV).

To examine the characteristics of patients with higher levels of alcohol consumption and to investigate the association between alcohol consumption and liver damage a total of 298 chronically infected individuals, with HBV genotyped and submitted to liver biopsy, were classified with Child's grading and separated by habits of alcohol intake, less and greater than 20g/day.

No significant differences were observed about genotype but genotypes A and D were predominant in both of them. A higher percentage of males (P.001) were observed in the group with alcohol intake above 20g/day, as well a lower proportion of patients with HBeAg negativity (P≤.035). In this group, biochemistry parameters, such as alanine aminotransferase (P=.006), aspartate aminotransferase (P=.001), gamma-glutamyl transferase (P.001) were elevated in a significantly higher proportion than in the other group.

The analysis of hematological parameters showed significantly lower values of platelets (P=.042) and mean corpuscular volume (P.001) and significantly higher values of prothrombin time (P.001) in the group with higher levels of alcohol consumption.

The characteristics of biopsy (P.001) and Child–Phug's classification (P=.002) revealed more severe results in this group.

Logistic regression showed a positive association between liver damage and alcohol intake, increasing with age. In female patients, a strong positive association between alcohol intake and liver damage was also found (odds ratio: 9.379; 95% confidence interval: 0.859–468.422; P = .037); however, the most severe cases were only observed in women older than 45 years.

In patients with HBV infection, alcohol is associated with a more severe liver disease.

No evidence was found concerning association with HBV genotype.
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Breast pumping and lactational state exert differential effects on ethanol pharmacokinetics

Prior research revealed that breast stimulation altered the way the lactating body handles alcohol. Its effects depended upon when it occurred relative to drinking.

The goal of the present study was to determine whether breast pumping works independently of the physiological and metabolic changes that accompany lactation.

To this end, we tested 12 women when they were exclusively breastfeeding 3–5-month-old infants and then again several months after lactation had ceased. Subjects were randomly assigned to one of two groups that differed in the timing of breast pumping relative to drinking a 0.4g/kg dose of alcohol: one group breast pumped 0.6h after drinking (pumped after group) and the other pumped 1h before drinking (pumped before group). For each reproductive stage, subjects were tested on 2 separate days, consuming a standardized meal 1 h before drinking during 1 test day and remaining fasted during the other. Breath alcohol concentrations (BrAC) and temperature readings were obtained before and at fixed intervals after drinking.

Pumping before drinking significantly decreased BrAC during both reproductive stages, whereas pumping after drinking resulted in different BrAC time curves during lactation when compared with after lactation. That is, levels were significantly lower during the descending phase of the time curve during than after lactation. The interactions between pumping and reproductive stage were most apparent during fed condition. Furthermore, women were more sensitive to hypothermic effects of both fasting and drinking alcohol during lactation.

These findings add to the growing literature that lactating women metabolize alcohol differently, in part, due to the frequent breast stimulation during breastfeeding and the pronounced physiological changes that accompany one of the most energetically costly mammalian activities.

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Request Reprint E-Mail: mennella@monell.org
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Behind Bars II: Substance Abuse and America’s Prison Population

Of the 2.3 million inmates crowding our nations prisons and jails, 1.5 million meet the DSM IV medical criteria for substance abuse or addiction, and another 458,000, while not meeting the strict DSM IV criteria, had histories of substance abuse; were under the influence of alcohol or other drugs at the time of their crime; committed their offense to get money to buy drugs; were incarcerated for an alcohol or drug law violation; or shared some combination of these characteristics, according to "Behind Bars II: Substance Abuse and America’s Prison Population".

Combined these two groups constitute 85 percent of the U.S. prison population.

The new 144-page report released by The National Center on Addiction and Substance Abuse (CASA) at Columbia University also reveals that alcohol and other drugs are significant factors in all crime.

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The UN tackles road safety

WHO welcomes the proclamation by the UN General Assembly of the first "Decade of Action for Road Safety 2011-2020" which seeks to halt the increasing trends in road traffic deaths and injuries worldwide.

"This Decade of Action for Road Safety is long overdue," says WHO Assistant Director-General Dr Ala Alwan. "It will help us increase action to address what will otherwise become the fifth leading cause of death by 2030."

Road traffic injuries are a major public health problem, killing nearly 1.3 million people each year and injuring as many as 50 million. They are the leading cause of death for children and young people aged 5–29 years. Almost half of the world's road traffic fatalities are among pedestrians, cyclists and motorcyclists and more than 90% occur in developing countries.

While road traffic death rates in many high-income countries have stabilized or declined in recent decades, research suggests road deaths are increasing in most regions of the world. If trends continue unabated deaths will rise to an estimated 2.4 million a year by 2030. . . . . .

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Long-Evans Rats Acquire Operant Self-Administration of 20% Ethanol Without Sucrose Fading

A major obstacle in the development of new medications for the treatment of alcohol use disorders (AUDs) has been the lack of preclinical, oral ethanol consumption paradigms that elicit high consumption.

We have previously shown that rats exposed to 20 ethanol intermittently in a two-bottle choice paradigm will consume two times more ethanol than those given continuous access without the use of water deprivation or sucrose fading (5–6% g/kg every 24 h vs 2–3 g/kg every 24 h, respectively).

In this study, we have adapted the model to an operant self-administration paradigm. Long-Evans rats were given access to 20% ethanol in overnight sessions on one of two schedules: (1) intermittent (Monday, Wednesday, and Friday) or (2) daily (Monday through Friday).

With the progression of the overnight sessions, both groups showed a steady escalation in drinking (3–6 g/kg every 14 h) without the use of a sucrose-fading procedure.

Following the acquisition phase, the 20% ethanol groups consumed significantly more ethanol than did animals trained to consume 10% ethanol with a sucrose fade (1.5 vs 0.7 g/kg every 30 min) and reached significantly higher blood ethanol concentrations.

In addition, training history (20% ethanol vs 10% ethanol with sucrose fade) had a significant effect on the subsequent self-administration of higher concentrations of ethanol.

Administration of the pharmacological stressor yohimbine following extinction caused a significant reinstatement of ethanol-seeking behavior.

Both 20% ethanol models show promise and are amenable to the study of maintenance, motivation, and reinstatement. Furthermore, training animals to lever press for ethanol without the use of sucrose fading removes a potential confound from self-administration studies.

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Request Reprint E-Mail: selenab@gallo.ucsf.edu
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Early onset alcohol dependence with high density of family history is not "male limited".

Based on classical adoption studies, early onset type II alcoholism was originally described as “male limited.”

We examined the possible expression of this subtype in present day alcohol-dependent women. Detailed systematic assessment was obtained from 200 treatment-seeking alcohol-dependent women and 189 healthy population controls.

Women fulfilling type II alcoholism criteria had higher alcoholism severity as measured by The Alcohol Use Disorders Identification Test and markedly higher use of illicit drugs.

Both alcoholism subtypes scored higher than normal on anxiety and impulsivity traits, but type II women scored markedly higher on aggression subscales than either of the other groups. Importantly, density of family history was markedly higher in type II women, suggesting a higher heritability.

Despite its original description as male limited, early onset alcoholism with high density of family history is likely to be a valid construct in women. Its recognition has important implications for diagnosis, treatment, and research.

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Request Reprint E-Mail: markus.heilig@mail.nih.gov
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Analysis of the alcohol biomarker phosphatidylethanol by NACE with on-line ESI-MS

Phosphatidylethanol (Peth), a group of aberrant phospholipids formed in cell membranes in the presence of ethanol, has been recently proposed as biomarker of chronic alcohol abuse.

The aim of this study was to develop a new analytical method, based on NACE online coupled with a mass spectrometer for the analysis of Peth in blood.

For this purpose an ion-trap mass spectrometer equipped with an orthogonal ESI source operating in negative ion mode was used. An alkaline solution of ammonium acetate 5 mM (pH 9) in water/methanol (MeOH) (80:20 v/v) was delivered as coaxial sheath liquid. All experiments were performed using an uncoated fused-silica capillary (90 cm×75 m id).

The effects of variable percentages of ACN, MeOH, 2-propanol, dichloromethane, along with variable concentrations of ammonium acetate were investigated for the separation of Peth. Collectively, a separation medium composed of ACN (45% v/v), 2-propanol (20% v/v), dichloromethane (20% v/v), MeOH (10% v/v), water (5% v/v), and ammonium acetate (25 mM) was chosen.

The estimated LOD was 0.1 M, while LOQ was 0.4 M. Within-run (intra-day) and between-run (inter-day) precision was always lower than 15%.

The method proved to be robust and reliable. The MS detector allowed the simultaneous identification of several Peth homologues, and the use of an internal standard (phosphatidylbutanol) with similar electrophoretic properties of that of Peth increased quantitation effectiveness.

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Request Reprint E-Mail: santodavide.ferrara@unipd.it
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