Publications

BACKGROUND: Chronic alcohol consumption is associated with neuroinflammation, neuronal damage, and behavioral alterations including addiction. Alcohol-induced neuroinflammation is characterized by increased expression of proinflammatory cytokines (including TNFα, IL-1β, and CCL2) and microglial activation. We hypothesized chronic alcohol consumption results in peripheral immune cell infiltration to the CNS. Since chemotaxis through the CCL2-CCR2 signaling axis is critical for macrophage recruitment peripherally and centrally, we further hypothesized that blockade of CCL2 signaling using the dual CCR2/5 inhibitor cenicriviroc (CVC) would prevent alcohol-induced CNS infiltration of peripheral macrophages and alter the neuroinflammatory state in the brain after chronic alcohol consumption.

METHODS: C57BL/6J female mice were fed an isocaloric or 5% (v/v) ethanol Lieber DeCarli diet for 6 weeks. Some mice received daily injections of CVC. Microglia and infiltrating macrophages were characterized and quantified by flow cytometry and visualized using CX3CR1eGFP/+ CCR2RFP/+ reporter mice. The effect of ethanol and CVC treatment on the expression of inflammatory genes was evaluated in various regions of the brain, using a Nanostring nCounter inflammation panel. Microglia activation was analyzed by immunofluorescence. CVC-treated and untreated mice were presented with the two-bottle choice test.

RESULTS: Chronic alcohol consumption induced microglia activation and peripheral macrophage infiltration in the CNS, particularly in the hippocampus. Treatment with CVC abrogated ethanol-induced recruitment of peripheral macrophages and partially reversed microglia activation. Furthermore, the expression of proinflammatory markers was upregulated by chronic alcohol consumption in various regions of the brain, including the cortex, hippocampus, and cerebellum. Inhibition of CCR2/5 decreased alcohol-mediated expression of inflammatory markers. Finally, microglia function was impaired by chronic alcohol consumption and restored by CVC treatment. CVC treatment did not change the ethanol consumption or preference of mice in the two-bottle choice test.

CONCLUSIONS: Together, our data establish that chronic alcohol consumption promotes the recruitment of peripheral macrophages into the CNS and microglia alterations through the CCR2/5 axis. Therefore, further exploration of the CCR2/5 axis as a modulator of neuroinflammation may offer a potential therapeutic approach for the treatment of alcohol-associated neuroinflammation.

Acute liver injury (ALI) is associated with multiple cellular events such as necrosis, apoptosis, oxidative stress and inflammation, which can lead to liver failure. In this study, we demonstrate a new role of microRNA (miR)-208a in ALI. ALI was induced in wild-type (WT) and miR-208a knockout (KO) mice by CCl4 administration. Increased alanine aminotransferase and decreased hepatic miR-208a levels were found in WT mice after acute CCl4 treatment. Histopathological evaluations revealed increased necrosis and decreased inflammation in miR-208a KO compared with WT mice after CCl4 treatment. CCl4 treatment induced a higher alanine aminotransferase elevation and increased numbers of circulating extracellular vesicles (exosomes and microvesicles) in miR-208a KO compared with WT mice. We found increased CCl4-induced nuclear factor kappa B activation and tumor necrosis factor-α induction and decreased monocyte chemoattractant protein 1 levels in miR-208a KO compared with WT mice. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling assay indicated aggravated hepatic apoptosis and necrosis in CCl4 -treated miR-208a KO compared with WT mice. CCl4 treatment induced a greater increase in cleaved caspase-8, p18, and caspase-3 in miR-208a KO compared with WT mice. p53 is involved in various cell death pathways, including necrosis and apoptosis. Our in silico analysis revealed p53 as a predicted miR-208a target, and we found enhanced p53 and cyclophilin D protein expressions in miR-208a KO mice after CCl4 treatment. Increased liver injury in miR-208a KO mice was further associated with increased Bax (B cell lymphoma 2-associated X protein) and p21 expression. Our in vitro results indicated a role of miR-208a in cell death. We found that CCl4-induced cytotoxicity was partially rescued by miR-208a overexpression in RAW macrophages. Altogether, our results revealed a role of miR-208a in ALI in mice and suggest a role for miR-208a in regulating cell death.

Severe alcoholic hepatitis (AH) is an acute and often devastating form of alcohol-associated liver disease. Clinically, AH is characterized by elevated bilirubin, model for end stage liver disease scores >20, and nonspecific symptoms that are caused by underlying inflammation, hepatocyte injury, and impaired intestinal barrier function. Compromised immune defense in AH contributes to infections, sepsis and organ failure. To date, corticosteroids are the only recommended treatment for severe AH, however it does not provide survival benefits beyond 1 month. Recent preclinical and early clinical studies in AH aided understanding of the disease and presented opportunities for new therapeutic options targeting inflammation, oxidative stress, liver regeneration and modification of intestinal microbiota. In this comprehensive review, we discuss promising preclinical results and ongoing clinical trials evaluating novel therapeutic agents for the treatment of severe AH.

BACKGROUND/AIMS: Despite high mortality of alcohol-associated hepatitis, there has been limited advancement in treatment strategies. Defeat Alcoholic Steatohepatitis (DASH) is a multicenter, randomized, double-blind controlled trial whose primary objective was to evaluate the safety and efficacy of a novel combination of 3 drugs targeting different perturbations in AH.

METHODS: Severe AH was diagnosed by liver biopsy or clinical and biochemical criteria and model for end stage liver disease (MELD) score ≥ 20 stratified by MELD scores (20-25 and ≥ 26) and randomized to a combination of an interleukin receptor 1 antagonist, Anakinra(100 mg daily for 14 days) to suppress acute inflammation, pentoxifylline (400 mg three times a day for 28 days) to prevent hepatorenal syndrome, and zinc sulfate (220 mg orally once daily for 6 months) or the standard of care therapy including methylprednisolone 32 mg orally once daily for 28 days. The primary efficacy outcome was the unadjusted log-rank test of the Kaplan-Meier survival estimates for the two treatment groups at 180 days.

RESULTS: Between July 2012 to March 2018, 500 subjects with severe AH were screened of which 104 subjects were enrolled with MELD score of 25.6 ± 3.2 (20.0-35.0) in the investigational arm and 25.8 ± 4.5 (20.0-40.0) in the standard of care arm. Causes of screen failures included not meeting eligibility criteria (n = 347), declining to participate (n = 39), and other reasons (n = 10).

CONCLUSIONS: Data from the DASH consortium studies will determine if a combination of drugs targeting multiple mechanisms of injury in the severe AH will improve clinical outcomes.

This review provides a summary of the symposium titled "Role of Non-Genetic Risk Factors in Exacerbating Alcohol-Related Organ Damage", which was held at the 42nd Annual Meeting of the Research Society on Alcoholism. The goals of the symposium were to provide newer insights into the role of non-genetic factors, including specific external factors, notably infectious agents or lifestyle factors, that synergistically act to exacerbate alcohol pathogenicity to generate more dramatic downstream biological defects. This summary of the symposium will benefit junior/senior basic scientists and clinicians currently investigating/treating alcohol-induced organ pathology, as well as undergraduate, graduate, and post-graduate students and fellows.

INTRODUCTION: Excessive alcohol consumption has steadily risen to become the third leading cause of preventable death in the USA. One consequence of heavy alcohol use recently under considerable investigation is alcoholic hepatitis. Although many risk factors for developing alcoholic hepatitis have been documented, our aim in this study was to examine the potential association between sarcopenia and severity, mortality, 30 days readmission rate, complication, infections and length of hospital stay in alcoholic hepatitis patients.

METHODS: A retrospective analysis was performed at a large, academic hospital in 194 alcoholic hepatitis patients aged 18-60 who had cross-sectional computed tomography imaging and met our clinical definition of alcoholic hepatitis. The fifth percentile of the psoas muscle index was used as a cutoff for sarcopenia.

RESULTS: One hundred ninety-four patients met the criteria for alcoholic hepatitis and had cross-sectional imaging. Higher Model for End-Stage Liver disease score was found in the sarcopenia group when compared to the non-sarcopenia group (mean Model for End-Stage Liver disease 21.5 and 24.2, respectively, P = 0.03). Sarcopenia also correlated with significantly longer hospital stay; the average length of stay in the sarcopenia group was 17.2 days while the non-sarcopenia patients had an average of 12.4 days. We found higher risk of developing pneumonia, sepsis and hepatic encephalopathy in sarcopenic patients.

CONCLUSION: Alcoholic hepatitis patients with sarcopenia have significantly worse outcomes when compared with the patients without sarcopenia, including a severe form of alcoholic hepatitis, longer hospital stays, higher risk of developing pneumonia, sepsis and hepatic encephalopathy.

Alcohol-related liver disease (ALD), which includes a range of disorders of different severity and is one of the most prevalent types of liver disease worldwide, has recently regained increased attention. Among other reasons, the realisation that any alcohol intake, regardless of type of beverage represents a health risk, and the new therapeutic strategies tested in recently published or undergoing clinical trials spur scientific interest in this area.In April 2019, Gut convened a round table panel of experts during the European Association for the Study of the Liver International Liver Congress in Vienna to discuss critical and up-to-date issues and clinical trial data regarding ALD, its epidemiology, diagnosis, management, pathomechanisms, possible future treatments and prevention. This paper summarises the discussion and its conclusions.

BACKGROUND: Alcohol use disorder is a significant societal and medical burden that is associated with both organ pathology and addiction. Excessive alcohol use results in neuroinflammation characterized by activation of the inflammasome, a multiprotein complex, and IL-1β increase in the brain. Recent studies suggest that inflammation could contribute to alcohol addiction. Here, we targeted components of the NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome cascade, which senses and responds to immunologic stimuli, to determine whether NLRP3 inhibition modulates alcohol consumption.

METHODS: C57BL/6J male and female mice were provided a 2-bottle choice of alcohol at increasing concentrations (3, 6, 9, and 12%, 4 days each) or water, and some were treated with daily injections of an NLRP3 inhibitor (MCC950), a caspase-1 inhibitor (VX765), IL-1 receptor antagonist (IL-1ra; anakinra), or vehicle injection.

RESULTS: Treatment with VX765, MCC950, and IL-1ra significantly reduced alcohol consumption and preference in female mice (p < 0.05). Treatment with MCC950 and IL-1ra reduced alcohol consumption, while IL-1ra reduced alcohol preference in male mice (p < 0.05). VX765 did not affect alcohol consumption or preference in male mice.

CONCLUSIONS: These findings highlight gender differences in alcohol preference and demonstrate that inhibition of different steps in inflammasome signaling can reduce alcohol consumption in females. Inhibition of NLRP3 inflammasome activation and the inflammasome-IL-1β cascade opens novel insights into the development of new therapies to address alcohol use disorder in an era of targeted and precision medicine.