Chronic Alcohol Administration Research Articles

Anxiolytic, memory improvement, hepatoprotective, and antioxidant effects of Thymus vulgaris on alcohol withdrawal syndrome in mice

Introduction: Thymus vulgaris is a plant used in Cameroon to alleviate and treat hangover symptoms. This study was conducted to assess the anxiolytic, memory-improving, hepatoprotective, and antioxidant effects of T. vulgaris on alcohol withdrawal syndrome. Methods: In this preclinical study 42 mice were grouped into seven equal sets of normal control, negative control, positive control (piracetam; 200 mg/kg), and test (T. vulgaris; 25, 50, 100, or 200 mg/kg) groups. All mice (except those of the normal control) were administered ethanol by gavage once daily for 28 days. After alcohol withdrawal, behavioral assessments (elevated plus maze, Y-maze, and open field) were performed from day 29 to day 31, one hour following appropriate treatments. Mice were euthanized on day 32 and the brains and the livers were used for histopathological assessment and oxidative stress parameters evaluation. Results: In the elevated plus maze test, T. vulgaris at 100 and 200 mg/kg significantly (P < 0.001) increased the number of entries and the time spent in the open arms. The plant extract (200 mg/kg) significantly (P < 0.001) increased the spontaneous alternation percentage in the Y maze. T. vulgaris administration decreased malondialdehyde (MDA) and nitric oxide (NO) in the brain and liver and increased glutathione (GSH) and catalase (CAT). The plant extract reduced the neuronal degeneration and hepatocyte damage induced by chronic alcohol administration. Conclusion: T. vulgaris had anxiolytic, memory-enhancing, and hepatoprotective properties against alcohol toxicity. These results may justify the use of thyme in traditional pharmacopeia in the management of alcohol use disorders (AUDs).

Potential Targets for the Protective Effect of Astaxanthin on Ethanolinduced Damage in Rat Liver Mitochondria.

Alcohol intoxication leads to multiple degenerative disorders in the structure and function of mitochondria. The mechanisms underlying these disorders, as well as ways to prevent them, are an urgent task in biomedicine. We investigate the mechanism of the positive effect of AX on rat liver mitochondria after chronic alcohol administration and suggest the targets of its effects. In this work, we continued our studies of astaxanthin (AX) as a possible protector of mitochondria from the toxic effects of ethanol. In our experiments, we used the Lieber-DeCarly model of chronic alcohol intoxication, which allows high-dose alcohol intake. Four groups of animals were used in the experiments: group 1 (control), group 2 (treated with AX), group 3 (treated with ethanol), and group 4 (treated with ethanol and AX together). Rat liver mitochondria (RLM) were isolated by the standard method modified in our laboratory. A multifunctional chamber with built-in electrodes was used to determine mitochondrial functions. Electrophoresis followed by Western blot analysis was used to detect mitochondrial proteins. Statistical significance was calculated using t-test Student-Newman- Keuls test. AX has been shown to have a positive effect on the functioning of the mitochondrial permeability transition pore (mPTP), the regulation of signaling pathways, as well as mitochondrial dynamics. It was found that AX is able to suppress the degenerative effect of alcohol on liver mitochondria. Targets for the protective action of AX in rat liver mitochondria (RLM) have been proposed. The discovered protective effect of AX on liver mitochondria during alcohol damage may contribute to the development of new strategies for the treatment of alcohol- induced damage.

Extracellular vesicle miR-206 improves chronic binge alcohol mediated decreased myoblast differentiation in SIV-infected female macaques.

Alcohol misuse in people with HIV (PWH) and chronic binge alcohol (CBA) administration in simian immunodeficiency virus (SIV)-infected macaques are associated with increased physical frailty and impaired functional skeletal muscle mass, respectively. Previous studies by our group demonstrate that muscle-enriched microRNAs (myomiRs) are differentially expressed in skeletal muscle (SKM) from CBA-administered SIV-infected male macaques and their altered expression contributes to impaired differentiation of SKM stem cells, or myoblasts. MicroRNAs can be transported in extracellular vesicles (EVs) to mediate numerous cellular responses through intercellular communication. The current study tested the hypothesis that EV-mediated delivery of miR-206 can ameliorate CBA-mediated decreased myoblast differentiation. Myoblasts were isolated from SKM of female SIV-infected, antiretroviral therapy-treated macaques that received either CBA (2.5g/kg/day, CBA/SIV) or water (VEH/SIV) for 14.5 months. Myotube and myotube derived EV myomiR expression, including miR-206, was lower in the CBA/SIV group. Overexpression of miR-206 decreased histone deacetylase 4 (HDAC4) and paired box 7 (PAX7) expression in myotubes and increased fusion index, a differentiation index, in CBA/SIV-derived myotubes. Similarly, EV-mediated delivery of miR-206 increased both fusion index and myotube density of CBA/SIV-derived myoblasts. These results support the potential therapeutic utility of EVs in delivering myomiRs to improve SKM stem cell differentiation.

Low-dose alcohol exacerbates hyperdynamic circulation and shunting in non-alcoholic cirrhotic rats.

Portal hypertension affects hepatic, splanchnic and portosystemic collateral systems. Although alcohol is a well-known risk factor for liver cirrhosis, it also affects vascular contractility. However, the relevant effects on portal hypertension have not been evaluated in non-alcoholic cirrhosis. The present study aimed to investigate the impacts of low-dose alcohol on portal hypertension-related derangements in non-alcoholic cirrhotic rats. Sprague-Dawley rats received bile duct ligation to induce cirrhosis or sham operation as controls. The chronic or acute effects of low-dose alcohol (2.4 g/kg/day, oral gavage, approximately 1.3 drinks/day in humans) were evaluated. The chronic administration of low-dose alcohol did not precipitate liver fibrosis in the sham or cirrhotic rats; however, it significantly increased splanchnic blood inflow (P=0.034) and portosystemic collaterals (P=0.001). Mesenteric angiogenesis and pro-angiogenic proteins were up-regulated in the alcohol-treated cirrhotic rats, and poorer collateral vasoresponsiveness to vasoconstrictors (P<0.001) was noted. Consistently, acute alcohol administration reduced splenorenal shunt resistance. Collateral vasoresponsiveness to vasoconstrictors also significantly decreased (P=0.003). In non-alcoholic cirrhosis rats, a single dose of alcohol adversely affected portosystemic collateral vessels due to vasodilatation. Long-term alcohol use precipitated splanchnic hyperdynamic circulation, in which mesenteric angiogenesis played a role. Further studies are warranted to evaluate the benefits of avoiding low-dose alcohol consumption in patients with non-alcoholic cirrhosis.

The Effects of Acute and Chronic Alcohol Administration and Withdrawal on Bone Microstructure, Mechanical Strength, and Remodeling Protein Expression and Their Relation to an Antioxidant and FGF23 In Vivo.

Alcohol's detrimental effects on bone health are well established, yet some literature suggests moderate consumption may offer benefits. With alcohol use on the rise, we investigate the impact of acute and chronic alcohol administration, along with withdrawal, on male Wistar rat femurs. We observed a transient cortical thickness increase with acute alcohol (AA) compared to chronic exposure (CA) but no significant changes in trabecular parameters or mechanical properties. High osteocalcin and osteopontin expression levels were noted in AA, alongside elevated RANKL expression. Conversely, CA showed low TRAP levels. FGF23 expression significantly increased during alcohol withdrawal (AW), while GPX decreased after chronic exposure but rose during withdrawal. Although mechanical strength changes were insignificant, biochemical shifts suggest alcohol exposure promotes bone resorption, reduces antioxidant protection, and potentially hampers active vitamin D and phosphate reabsorption via FGF23 upregulation.

Role of ghrelin hormone in the development of alcohol-associated liver disease

Fatty liver is the earliest response of the liver to excessive alcohol consumption. Previously we identified that chronic alcohol administration increases levels of stomach-derived hormone, ghrelin, which by reducing circulating insulin levels, ultimately contributes to the development of alcohol-associated liver disease (ALD). In addition, ghrelin directly promotes fat accumulation in hepatocytes by enhancing de novo lipogenesis. Other than promoting ALD, ghrelin is known to increase alcohol craving and intake. In this study, we used a ghrelin receptor (GHSR) knockout (KO) rat model to characterize the specific contribution of ghrelin in the development of ALD with emphasis on energy homeostasis. Male Wistar wild type (WT) and GHSR-KO rats were pair-fed the Lieber-DeCarli control or ethanol diet for 6 weeks. At the end of the feeding period, glucose tolerance test was conducted, and tissue samples were collected. We observed reduced alcohol intake by GHSR-KOs compared to a previous study where WT rats were fed ethanol diet ad libitum. Further, when the WTs were pair-fed to GHSR-KOs, the KO rats exhibited resistance to develop ALD through improving insulin secretion/sensitivity to reduce adipose lipolysis and hepatic fatty acid uptake/synthesis and increase fatty acid oxidation. Furthermore, proteomic data revealed that ethanol-fed KO exhibit less alcohol-induced mitochondrial dysfunction and oxidative stress than WT rats. Proteomic data also confirmed that the ethanol-fed KOs are insulin sensitive and are resistant to hepatic steatosis development compared to WT rats. Together, these data confirm that inhibiting ghrelin action prevent alcohol-induced liver and adipose dysfunction independent of reducing alcohol intake.

Investigating the Protective Effects of a Tryptophan-Based Diet in Alcoholics: An Experimental Study on Depression and Anxiety

Depression and anxiety are prevalent co-morbidities among individuals with chronic alcohol consumption, posing significant health challenges worldwide. This study aimed to explore the potential protective effects of a tryptophan-based diet against depressive tendencies and anxiety associated with chronic alcohol consumption. Using an experimental design, Wistar rats were subjected to chronic alcohol administration along with varying doses of a tryptophan-based diet over 28 days. Behavioral tests including the Elevated Plus Maze (EPM) and Forced Swim Test (FST) were conducted to evaluate anxiety-like behavior and depressive tendencies, respectively. Results indicated a significant decrease in immobility time in the FST with increasing doses of the tryptophan-based diet, suggesting antidepressant effects. Moreover, the anxiety index decreased while the time spent on open arms increased with higher doses of the diet in the EPM, indicating anxiolytic effects. These findings suggest that a high tryptophan-based diet could serve as a potential intervention for individuals with alcohol-related depression and anxiety. Keywords: Depression, anxiety, alcohol, tryptophan, diet, alcoholics

Alcohol Impairs Bioenergetics and Differentiation Capacity of Myoblasts from Simian Immunodeficiency Virus-Infected Female Macaques.

Alcohol misuse and HIV independently induce myopathy. We previously showed that chronic binge alcohol (CBA) administration, with or without simian immunodeficiency virus (SIV), decreases differentiation capacity of male rhesus macaque myoblasts. We hypothesized that short-term alcohol and CBA/SIV would synergistically decrease differentiation capacity and impair bioenergetic parameters in female macaque myoblasts. Myoblasts from naïve (CBA-/SIV-), vehicle [VEH]/SIV, and CBA/SIV (N = 4-6/group) groups were proliferated (3 days) and differentiated (5 days) with 0 or 50 mM ethanol (short-term). CBA/SIV decreased differentiation and increased non-mitochondrial oxygen consumption rate (OCR) versus naïve and/or VEH/SIV. Short-term alcohol decreased differentiation; increased maximal and non-mitochondrial OCR, mitochondrial reactive oxygen species (ROS) production, and aldolase activity; and decreased glycolytic measures, ATP production, mitochondrial membrane potential (ΔΨm), and pyruvate kinase activity. Mitochondrial ROS production was closely associated with mitochondrial network volume, and differentiation indices were closely associated with key bioenergetic health and function parameters. Results indicate that short-term alcohol and CBA non-synergistically decrease myoblast differentiation capacity. Short-term alcohol impaired myoblast glycolytic function, driving the bioenergetic deficit. Results suggest potentially differing mechanisms underlying decreased differentiation capacity with short-term alcohol and CBA, highlighting the need to elucidate the impact of different alcohol use patterns on myopathy.

Acetyl-L-carnitine attenuates chronic ethanol-induced oxidative stress, ER stress and apoptosis in rat gastric tissue.

Consuming alcohol affects almost all organs. Acetaldehyde, formed as the main product as a result of alcohol metabolism, causes the production of free superoxide radicals when oxidized, and accordingly oxidative and apoptotic processes are triggered. There are studies showing that carnitine has effects on oxidative and apoptotic processes that occur in various conditions. However, the mechanisms showing the effects of L-carnitine on these effects of alcohol have not been fully elucidated. In our study, the effects of acetyl-L-carnitine administration on the molecular mechanisms of oxidative stress, endoplasmic reticulum stress and apoptotic parameters in gastric tissue of rats chronically exposed to alcohol were investigated. Hematoxylin-eosin staining was used for histopathological studies. Endoplasmic reticulum stress markers were detected with immunohistochemical staining and western blotting. Apoptotic index was evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. Total oxidant and antioxidant status were examined by ELISA. Our results showed that chronic alcohol administration caused a significant increase in TOS levels, an indicator of oxidative stress, the levels of ER-stress-associated proteins XBP1, GRP78 and CHOP, and % apoptotic index values in rat gastric tissues. Additionally, it was determined that acetyl-L-carnitine administration caused an improvement in those values. Based on our data, we can conclude that acetyl-L-carnitine has a tissue protective effect by scavenging free oxygen radicals and reducing ER stress-related proteins XBP1, GRP78 and CHOP and apoptosis in chronic ethanol administered rats, and that this natural antioxidant may be beneficial in the treatment of oxidative stress-induced diseases.

Impact of common ALDH2 inactivating mutation and alcohol consumption on Alzheimer's disease.

Aldehyde dehydrogenase 2 (ALDH2) is an enzyme found in the mitochondrial matrix that plays a central role in alcohol and aldehyde metabolism. A common ALDH2 polymorphism in East Asians descent (called ALDH2*2 or E504K missense variant, SNP ID: rs671), present in approximately 8% of the world's population, has been associated with a variety of diseases. Recent meta-analyses support the relationship between this ALDH2 polymorphism and Alzheimer's disease (AD). And AD-like pathology observed in ALDH2-/- null mice and ALDH2*2 overexpressing transgenic mice indicate that ALDH2 deficiency plays an important role in the pathogenesis of AD. Recently, the worldwide increase in alcohol consumption has drawn attention to the relationship between heavy alcohol consumption and AD. Of potential clinical significance, chronic administration of alcohol in ALDH2*2/*2 knock-in mice exacerbates the pathogenesis of AD-like symptoms. Therefore, ALDH2 polymorphism and alcohol consumption likely play an important role in the onset and progression of AD. Here, we review the data on the relationship between ALDH2 polymorphism, alcohol, and AD, and summarize what is currently known about the role of the common ALDH2 inactivating mutation, ALDH2*2, and alcohol in the onset and progression of AD.

Glucagon-like peptide 1 receptor agonist, exendin-4, reduces alcohol-associated fatty liver disease

Fatty liver is the earliest response to excessive ethanol consumption, which increases the susceptibility of the liver to develop advanced stage of liver disease. Our previous studies have revealed that chronic alcohol administration alters metabolic hormone levels and their functions. Of current interest to our laboratory is glucagon-like peptide 1 (GLP-1), a widely studied hormone known to reduce insulin resistance and hepatic fat accumulation in patients with metabolic-associated fatty liver disease. In this study, we examined the beneficial effects of exendin-4 (a GLP-1 receptor agonist) in an experimental rat model of ALD. Male Wistar rats were pair-fed the Lieber-DeCarli control or ethanol diet. After 4 weeks of this feeding regimen, a subset of rats in each group were intraperitoneally injected every other day with either saline or exendin-4 at a dose of 3 nmol/kg/day (total 13 doses) while still being fed their respective diet. At the end of the treatment, rats were fasted for 6 h and glucose tolerance test was conducted. The following day, the rats were euthanized, and the blood and tissue samples collected for subsequent analysis. We found that exendin-4 treatment had no significant effect on body weight gain among the experimental groups. Exendin-4-treated ethanol rats exhibited improved alcohol-induced alterations in liver/body weight and adipose/body weight ratio, serum ALT, NEFA, insulin, adiponectin and hepatic triglyceride levels. Reduction in indices of hepatic steatosis in exendin-4 treated ethanol-fed rats was attributed to improved insulin signaling and fat metabolism. These results strongly suggest that exendin-4 mitigates alcohol-associated hepatic steatosis by regulating fat metabolism.

Chronic binge alcohol-induced hippocampal plasticity in SIV-infected female rhesus macaques

Human immunodeficiency virus (HIV) infection is detrimental to central nervous system (CNS) function and may result in HIV-associated neurocognitive disorder (HAND) over time. HIV-associated cognitive deficits are further exacerbated by chronic alcohol use. With the rising prevalence of alcohol use disorder (AUD) in females and people living with HIV (PLWH), understanding the neurobiological impact of chronic alcohol exposure in these populations is increasingly important. As the hippocampus plays a prominent role in cognition, examining neuroadaptations in this brain region in the context of comorbid HIV infection and alcohol exposure is critical for understanding the mechanisms of neurocognitive impairment. We hypothesized that simian immunodeficiency virus (SIV)-infected female rhesus macaques with a history of chronic binge alcohol (CBA) administration (n=7-8 per group) would exhibit increased phosphorylation of mitogen-activated protein kinase (MAPK) family members, indicative of increased hippocampal activity and potential hyperexcitability. We also measured cognitive performance in macaques via the novel object recognition (NOR) task and further hypothesized a manifestation of cognitive deficits in the CBA-treated group. Finally, we incorporated additional groups of ovariectomized (OVX) females in the design as we hypothesized that the loss of ovarian hormones would reduce cognitive performance. Data were analyzed via two-way ANOVA with CBA treatment and OVX as factors. Western blot analyses demonstrated that CBA significantly increased phosphorylation of MAPK-family members extracellular signal-regulated kinase (ERK; F=8.278, p=0.0093) and p38 mitogen-activated protein kinase (p38 MAPK; F=4.414, p=0.0485). Furthermore, CBA significantly increased both phosphorylated and total levels of Jun N-terminal kinase (JNK; F=5.989, p=0.0233; F=7.158, p=0.0145). While analysis of the NOR task revealed a significant reduction in latency to approach the novel versus familiar object on test day across all groups (F = 11.97, p=0.0013), no significant difference was seen between CBA and control groups. Interestingly, no main effects of OVX or interactive CBA/OVX effects were observed for either molecular or behavioral outcomes. We are currently considering alternative cognitive tasks that may better reveal CBA-associated cognitive deficits, as well as neuroadaptations in nuclear signaling downstream of MAPK activity. Together, these data suggest that chronic alcohol use results in profound changes in hippocampal plasticity, potentially facilitating cognitive decline and other CNS symptoms observed in PLWH. This research was supported by grant # T32AA007577 and #P60AA009803 from the NIAAA at the National Institutes of Health. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

MicroRNA-206 improves alcohol-mediated decreased myoblast differentiation

Background: With recent advances in antiretroviral therapy (ART), people living with HIV (PLWH) have a near-normal life expectancy but have higher risk for aging-related comorbidities, such as metabolic disorders and frailty earlier in life than the general population. At-risk alcohol use is twice as likely in PLWH compared to the general population and alcohol-related myopathy occurs in 40- 60% of people with alcohol use disorder. Previous studies demonstrated that in vivo chronic binge alcohol (CBA) administration in simian immunodeficiency virus (SIV)-infected male rhesus macaques decreased myoblast differentiation and downregulated microRNA-206 (miR-206) expression in skeletal muscle. We hypothesized that increasing miR-206 expression would increase differentiation in CBA-derived primary macaque myoblasts. Methods: Eight SIV-infected female rhesus macaques received either water (VEH) or CBA (13- 14 g/kg/week) for 14.5 months. Three months into either VEH or CBA administration, animals were vaginally infected with SIVmac251, and ART was started 2.5 months later. Animals were sacrificed after 9 months, and primary myoblasts isolated from vastus lateralis muscle 24 hours after the last dose of alcohol (blood alcohol=0 mM). Myoblasts at passage 4 (VEH &amp; CBA) were proliferated for three days. Cells were then transfected with either miR-206 mimic or control (miR scramble). Four experimental groups were studied at day 5 of differentiation: VEH+scramble, CBA+scramble, VEH+miR-206, and CBA+miR-206. Expression of miR-206 was determined by qPCR. Myoblast differentiation was determined by myotube density, calculated as the average number of myotubes per frame, and fusion index calculated as the percentage of myonuclei that fused into myotubes. Results: In non-transfected CBA-derived cells, myotube density and miR-206 expression was lower compared to VEH-derived myotubes (p&lt;0.001 and p&lt;0.05, respectively). miR-206 transfection resulted in over 150-fold higher miR-206 expression compared to VEH-scramble (p&lt;0.05) in CBA- and VEH-derived cells. miR-206 overexpression significantly increased myotube fusion index (p&lt;0.05) compared to myoblasts transfected with miR scramble in CBA-derived cells. Conclusions: Data show significant improvement of alcohol-mediated decreased myoblast differentiation with miR-206 overexpression. These findings suggest that increasing muscle miR-206 can potentially be used as a therapeutic strategy to improve regenerative capacity in response to muscle injury or atrophy. (Supported by: T35AA021097, F30AA029358, P60AA009803, K01AA024494) This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Chronic alcohol-induced mesenteric lymphatic leakage is associated with local and systemic metabolic dysregulation

Introduction: Chronic alcohol administration produces significant metabolic dysfunction as determined in pre-clinical and clinical studies. Our studies using a rodent model of chronic alcohol feeding showed that alcohol induces mesenteric lymphatic leakage and perilymphatic adipose tissue (PLAT) immunometabolic dysregulation. Specifically, chronic alcohol induced localized PLAT inflammation and decreased glucose uptake. Whether chronic alcohol-induced lymphatic leakage is also associated with altered systemic glucose/insulin dynamics remained to be determined. We hypothesized that chronic alcohol feeding impairs the endocrine response to a glucose tolerance test. Methods: Male Fischer 344 rats were randomized to a Lieber-DeCarli liquid diet containing 36% of calories from alcohol or pair-fed an isocaloric diet for 10 weeks. Animals consumed approximately 75 mL of diet per day. Body weights were not significantly different between groups; the average weights were 275.7 grams for control animals and 275.9 grams for alcohol-fed animals. Weights in both groups remained stable for the duration of the study. At 10 weeks, animals were fasted for 16 hours prior to intraperitoneal (IP) injections of 20% glucose solution (2g/kg). Blood samples were collected from the rat tail prior to and at 15-minute intervals following initial glucose injection for 2 hours. Blood glucose was tested at each interval using the AlphaTRAK glucometer. 2-way ANOVA and paired T-test were used to assess significant differences between groups. Results and Conclusion: Prior to IP glucose injection, blood glucose levels averaged 77 mg/dL for control animals and 71.5 mg/dL for alcohol-fed animals. Glucose levels in alcohol-fed animals averaged 38% higher at 15 minutes and 37% higher 30 minutes after IP glucose bolus than that of control animals. Total area under the curve (AUC) calculated from the glucose concentrations measured following IP glucose injection was significantly higher in alcohol-fed animals compared to control animals; average AUC for alcohol-fed animals was 26.8% higher than AUC for control animals. These results suggest that chronic alcohol-induced mesenteric lymphatic leakage and localized PLAT inflammation and altered glucose metabolism are also associated with disturbances in systemic glucose dysregulation. Future studies will examine the potential link between alcohol-induced PLAT immunometabolic dysregulation and alterations in systemic glucose metabolism. Supported by T32 (PM), K01 (FS). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Chronic alcohol administration alters metabolomic profile of murine bone marrow.

People with hazardous alcohol use are more susceptible to viral, bacterial, and fungal infections due to the effect of alcohol on immune system cell function. Metabolized ethanol reduces NAD+ to NADH, affecting critical metabolic pathways. Here, our aim was to investigate whether alcohol is metabolized by bone marrow cells and if it impacts the metabolic pathways of leukocyte progenitor cells. This is said to lead to a qualitative and quantitative alteration of key metabolites which may be related to the immune response. We addressed this aim by using C57BL/6 mice under chronic ethanol administration and evaluating the metabolomic profile of bone marrow total cells by gas chromatography-coupled mass spectrometry (GC-MS). We identified 19 metabolites. Our data demonstrated that chronic ethanol administration alters the metabolomic profile in the bone marrow, resulting in a statistically diminished abundance of five metabolites in ethanol-treated animals: uracil, succinate, proline, nicotinamide, and tyrosine. Our results demonstrate for the first time in the literature the effects of alcohol consumption on the metabolome content of hematopoietic tissue and open a wide range of further studies to investigate mechanisms by which alcohol compromises the cellular function of the immune system.

Differential Regulation of Tachykinin and Opioid System Gene Expression in Brain and Immune Cells of Chronic Binge Alcohol-Treated Simian Immunodeficiency Virus-Infected Macaques.

People living with HIV have a high likelihood of at-risk alcohol use and are at increased risk for neurocognitive decline. The underlying mechanisms involved in HIV-associated neurocognitive disorder (HAND) are not completely understood. Previously, we showed that chronic binge alcohol (CBA) administration produced behavioral deficits in non antiretroviral therapy (ART)-treated simian immunodeficiency virus (SIV)-infected macaques. Moreover, we observed that CBA/SIV enhanced neuroinflammatory gene expression and attenuated growth factor signaling in the frontal cortex (FC) and basal ganglia, effects that were partially ameliorated by ART. We hypothesized that the neuroinflammatory and growth factor changes observed could be associated with alterations in opioid, tachykinin, and endocannabinoid gene expression. Furthermore, we proposed that gene expression patterns in peripheral blood mononuclear cells (PBMCs) could serve as an indicator of expression changes in the brain (FC). We examined gene expression patterns of opioid, tachykinin, and endocannabinoid systems in FC and PBMCs isolated from CBA/SIV macaques. Expression of targeted genes as determined by reverse transcription-quantitative polymerase chain reaction was analyzed in relation to CBA, ART, plasma, and brain viral loads (PVL and BVL, respectively) and compared with baseline (PBMC) or FC from SIV- controls. FC expression of ORM1, POMC, and TACR1 was negatively associated with PVL (p = .03, .002, .05 respectively). FC expression of TAC1 was positively associated with CBA exposure (p = .05). PBMC expression of DAGLA was positively associated with CBA exposure; but negatively associated with combined CBA/ART exposure (p = .03). Our findings reflect the complex interactions of SIV, CBA, and ART in modulating opioid and tachykinin system gene expression. Contrary to our prediction, results did not reveal parallel changes (in magnitude or direction) in PBMC and FC gene expression. Further studies are warranted to determine the relevance of these transcriptional changes in modulating HAND-related behaviors resulting from at-risk alcohol use and HIV/SIV exposure.

RAGE promotes dysregulation of iron and lipid metabolism in alcoholic liver disease

Alcoholic liver disease (ALD) is associated with hepatic inflammatory activation and iron overload. The receptor for advanced glycation end products (RAGE) is an important metabolic mediator during the development of ALD. The aim of this study was to determine the effect of RAGE on iron homeostasis in ALD. We found increased circulating transferrin, hepcidin and ferritin in ALD patients and positively correlated with RAGE level. RAGE knockout (RAGE−/−) and wild-type mice were subjected to chronic alcoholic feeding for 6 weeks to induce ALD, and RAGE inhibitor, iron chelator or lipid peroxidation inhibitor were administered. We showed that chronic alcohol administration triggered hepatic steatosis, inflammation, and oxidative stress, which were eliminated by deficiency or inhibition of RAGE. Surprisingly, pathways of hepatic iron metabolism were significantly altered, including increased iron uptake (Tf/TfR) and storage (Ferritin), as well as decreased iron export (FPN1/Hepcidin). In vitro experiments confirmed that RAGE had different effects on the mechanism of iron metabolism of hepatocytes and macrophages respectively. In conclusion, our data revealed preclinical evidence for RAGE inhibition as an effective intervention for alleviating alcohol-induced liver injury.

Chronic Binge Alcohol Differentially Affects Extracellular Matrix Phenotype in Omental and Subcutaneous Adipose Tissue of SIV‐infected Macaques

The incidence of at‐risk alcohol use among people living with HIV (PLWH) is two times greater than in the general population. Though antiretroviral therapy (ART) has significantly reduced HIV‐associated mortality, increased survival of PLWH on ART is associated with increased incidence of metabolic comorbidities. Adipose tissue maladaptation is central to metabolic alterations in PLWH and at‐risk alcohol use. This involves adipokine disruption, adipocyte dysfunction, inflammation, and dysregulation of the extracellular milieu. Using a relevant preclinical model of HIV‐infection, the objective of our study was to investigate the effects of chronic binge alcohol (CBA) administration on omental adipose tissue (OmAT) and subcutaneous adipose tissue (SAT) extracellular matrix (ECM) phenotype. Rhesus macaques were administered CBA or isovolumetric water (VEH) through an intragastric catheter for 30 min, 5 days a week, with blood alcohol concentrations reaching 50‐60 mM. Three months after initiation of CBA or VEH administration, macaques were inoculated with SIVMac251, and 2.5 months later animals were initiated on ART. CBA or VEH administration was continued throughout the duration of the study (14.5 mo). Animals were euthanized at 11.5 mo of SIV infection, and adipose (OmAT and SAT) samples excised. Collagen content, adipocyte diameter and adipocyte numbers were measured by Picrosirius Red Staining (PSR). A custom PCR Array was used to assess differential expression of 41 ECM‐related genes. CBA administration significantly increased collagen expression and decreased adipocyte diameter in the OmAT without altering adipocyte numbers. There was a strong negative correlation of OmAT adipocyte diameter and collagen expression. CBA administration did not result in significant differences in collagen expression, adipocyte diameter or number in SAT. CBA administration significantly upregulated expression of transforming growth factor beta‐1 (TGFB1), platelet‐derived growth factor receptor alpha (PDGFRA) and secreted protein acidic and rich in cysteine (SPARC), mediators of a pro‐fibrotic milieu in OmAT. In contrast, CBA did not significantly alter expression of ECM‐related genes in the SAT. These data suggest that the maladaptive adipose changes associated with CBA in SIV infection are depot specific. We speculate that changes in the ECM contribute to altered gene expression and impaired adipocyte metabolic capacity.

Ethanol‐Induced Lymphatic Endothelial Cell Permeability: Role of Prox‐1 and Vascular Endothelial Growth Factors

Acute and chronic alcohol (Ethanol, EtOH) administration to rodents promote mesenteric lymphatic vessel permeability leading to immune cell leakage into perilymphatic adipose tissue. We speculate this contributes to impaired immune dialog between the gut and adjacent lymphoid tissue. Mechanistic in vitro studies show that EtOH‐induced lymphatic endothelial cell (LEC) permeability is not associated with alterations in tight and adherens junction expression but is partially rescued by p38 MAP kinase inhibition. Transcription factor Prox‐1 and Vascular endothelial growth factor receptor‐3 (VEGFR‐3) play essential roles in maintaining the integrity of mesenteric lymphatic vessels and of lymphatic vascular functions. The lack of Prox‐1 and VEGFR‐3 expression has been shown to contribute to excessive lymph and vascular leakage, respectively, promoting increased lymphatic permeability. Prox‐1 and VEGFR‐3 are negatively regulated by notch signaling. Notch activation suppresses Prox‐1 and notch target genes, Hey 1 and Hey 2, inhibit VEGFR‐3. We hypothesize that EtOH‐induced LEC permeability is mediated by decreased Prox‐1 and VEGFR‐3 expression via notch signaling activation. We used an in vitro model of rat LEC to characterize Prox‐1 and VEGFR‐3 expression in EtOH‐induced LEC permeability. Cells were seeded in T‐75 flask and cultured with rat endothelial cell medium supplemented with EtOH (0, 25mM, and 50mM) for 30 minutes in an EtOH water bath to limit EtOH evaporation loss from culture media. Gene expression of Prox‐1, VEGFR‐3, Notch 1, Notch 4, Hey 1, and Hey 2 was measured using Quantitative Polymerase Chain Reaction. EtOH at 50mM concentration for 30 minutes significantly decreased gene expression of Prox‐1 and VEGFR‐3 and increased gene expression of Notch 1 and Hey 2. No significant changes were observed in LEC exposed to 25mM EtOH. These findings support our hypothesis that decreases in Prox‐1 and VEGFR‐3 expression involving notch signaling activation may be implicated in EtOH induced LEC permeability leading to mesenteric lymphatic dysfunction.

Changes in Brain Dopamine Extracellular Concentration after Ethanol Administration; Rat Microdialysis Studies.

The purpose of this review is to evaluate microdialysis studies where alterations in the dopaminergic system have been evaluated after different intoxication states, in animals showing preference or not for alcohol, as well as during alcohol withdrawal. Ethanol administration induces varying alterations in dopamine microdialysate concentrations, thereby modulating the functional output of the dopaminergic system. Administration of low doses of ethanol, intraperitoneally, intravenously, orally or directly into the nucleus accumbens, NAc, increases mesolimbic dopamine, transmission, as shown by increases in dopamine content. Chronic alcohol administration to rats, which show alcohol-dependent behaviour, induced little change in basal dopamine microdialysis content. In contrast, reduced basal dopamine content occurred after ethanol withdrawal, which might be the stimulus to induce alcohol cravings and consumption. Intermittent alcohol consumption did not identify any consistent changes in dopamine transmission. Animals which have been selectively or genetically bred for alcohol preference did not show consistent changes in basal dopamine content although, exhibited a significant ethanol-evoked dopamine response by comparison to non-preference animals. Microdialysis has provided valuable information about ethanol-evoked dopamine release in the different animal models of alcohol abuse. Acute ethanol administration increases dopamine transmission in the rat NAc whereas chronic ethanol consumption shows variable results which might reflect whether the rat is prior to or experiencing ethanol withdrawal. Ethanol withdrawal significantly decreases the extracellular dopamine content. Such changes in dopamine surges will contribute to both drug dependence, e.g. susceptibility to drug withdrawal, and addiction, by compromising the ability to react to normal dopamine fluctuations.