Alcohol Metabolism, Functional Consequences, Apoptosis Signaling Pathways, and Organ Damage

National Institute On Alcohol Abuse And Alcoholism

Investigators

Linked publications, trials & patents

Abstract

My lab has continued studying the opposing roles of ethanol-inducible cytochrome P450-2E1 (CYP2E1), a pro-oxidant enzyme, and mitochondrial aldehyde dehydrogenase-2 (ALDH2), an antioxidant enzyme for the removal of toxic acetaldehyde and lipid peroxides, in tissue injury caused by alcohol and other potentially toxic substances. Alcohol-induced oxidative and nitrative (nitro-oxidative) stress can inhibit the activities of ALDH2 and other mitochondrial proteins, resulting in accumulation of toxic aldehydes and mitochondrial dysfunction, contributing to cell death. In addition, CYP2E1-mediated nitro-oxidative stress can stimulate different types of post-translational protein modification (PTM), contributing to endoplasmic reticulum (ER) stress, mitochondrial dysfunction, and tissue/organ damage. These PTMs include oxidation, S-nitrosylation, nitration, phosphorylation, acetylation, adduct formation, etc. All these PTMs generally occur shortly after exposure to alcohol and nonalcohol-containing substances or under pathological conditions. In the past, we demonstrated a causal role of CYP2E1 in promoting tissue injury by evaluating the time-dependent changes between PTMs and actual cellular damage in wild-type (WT) compared to the age- and sex-matched Cyp2e1-null mice in the presence or absence of a protective agent. We also showed that these oxidative PTMs and functional alterations of covalently modified proteins are observed in the liver and other tissues such as the gut and brain. In fact, we recently reported the critical role of CYP2E1 in binge alcohol-mediated gut barrier dysfunction (leaky gut), endotoxemia, and inflammatory liver injury in rats and WT mice by studying the pattern of various PTMs of the gut tight and adherens junction (TJ/AJ) proteins and their decreased levels. Exposure to binge alcohol or nonalcohol-containing substances significantly decreased the levels of gut TJ/AJ proteins while it increased the levels of intestinal CYP2E1, iNOS, nitrated proteins, apoptosis-related marker proteins, serum endotoxin, and fecal albumin contents, suggesting elevated gut leakiness and endotoxemia compared to the corresponding Cyp2e1-null mice. These results demonstrate a direct or indirect, permissive role of CYP2E1 in promoting gut leakiness, endotoxemia, and inflammatory or fibrotic liver disease. In addition, we reported that small amounts of alcohol (administered as single or multiple doses) can cause acute tissue injury in young Aldh2-KO mice (on a C57BL/6J background), while no or little damage was observed in age- and sex-matched WT mice. In these models, elevated levels of various oxidative PTMs were observed in the gut, liver, and brain of alcohol-exposed Aldh2-KO mice compared to the corresponding WT mice, suggesting a contributing role of PTMs in alcohol-mediated gut leakiness and liver or brain injury through the gut-liver-brain axis. These results also indicate the protective role of ALDH2 against binge alcohol-induced tissue injury, including neurodegeneration. In contrast, our unpublished results showed that binge alcohol significantly elevated the levels of oxidative PTMs and neurodegeneration, assessed by confocal images of Fluoro-Jade-C (FJC)-stained damaged neurons and neurobehavioral tests, in WT mice compared to the corresponding Cyp2e1-KO mice. These results indicate the contributing role of CYP2E1 in alcohol-mediated brain injury. Based on the opposing roles of CYP2E1 and ALDH2, suppression of CYP2E1 and activation of ALDH2 would be an important concept to develop a therapeutic agent for preventing or treating alcohol-mediated multiple organ damage. We also studied the molecular mechanisms of acetaminophen (APAP)- and thioacetamide (TAA)-mediated acute liver injury (ALI) by investigating the contributing or causal role of oxidative PTMs in promoting gut leakiness, endotoxemia, and ALI. In this study, we particularly focused on the early changes (e.g., 1, 2, 4, or 7 h) in oxidative PTMs and gut injury after drug challenges. Our results showed that a single injection of APAP or TAA increased the oxidative PTMs at early time points, leading to decreased intestinal TJ/AJ proteins, elevated gut leakiness, and endotoxemia prior to ALI observed at 24 h after APAP or TAA exposure. However, ALI was virtually not observed in the APAP- or TAA-exposed Cyp2e1-null mice, demonstrating the direct role of CYP2E1 in APAP or TAA metabolism and subsequent oxidative PTMs, gut leakiness, and ALI. Our novel results about the contributing role of oxidative PTMs in promoting gut leakiness in ALI are clearly different from the earlier study on the changes in gut microbiota and transcriptomic regulation, which reflect the consequences but not the causal factors, since the earlier report analyzed the liver and gut tissues collected at 20 hours after drug challenges. More importantly, our results emphasize the clinical importance of early changes in many other tissues, including the gut, in APAP-mediated multiple organ damage, as we reported recently. Furthermore, our approaches can also be applied to the understanding of pathological mechanisms of other acute tissue injury models caused by viral infections and many drugs, including antibiotics and chemotherapeutic agents. In collaboration with a former lab member, Dr. Young-Eun Cho, we continued to study the beneficial effects of dietary antioxidant compounds or extracellular nanovesicles (EVs) prepared from plants against alcohol- or nonalcohol-induced tissue injury. For instance, we investigated the molecular mechanisms by which exosome-like nanoparticles from ginseng or pomegranate (POM) prevented Western-style high-fat diet (HFD) or binge alcohol-induced gut leakiness and liver fibrosis or brain injury. Histology and biochemical analyses showed that HFD feeding caused gut, liver, and brain injury through the gut-liver-brain axis. Immunoblot analyses showed the increased levels of oxidative PTMs and cell death marker proteins with decreased levels of many gut TJ/AJ proteins that were nitrated and degraded by ubiquitin-dependent proteolysis in HFD-fed mice. The exosome-like EV nanoparticles from POM also protected against binge alcohol-mediated oxidative PTMs, gut leakiness, and liver or brain injury via similar mechanisms. Furthermore, these plant-derived EVs significantly restored the composition and abundance of beneficial gut microbiomes that were negatively altered by binge alcohol or Western-style HFD. Our results also showed that the nanosized EVs from ginseng or POM significantly alleviated the severities of gut and liver injury, suggesting the utilities of plant-derived EVs as non-toxic antioxidant nanocarriers in preventing alcohol- and HFD-mediated multiple organ damage. Two reports on these new results have been recently published. All these experimental rodent and cell studies not only represent examples of our mechanistic and translational research with safe dietary supplements or plant-derived EVs to prevent oxidative stress-mediated PTMs, intestinal damage, and liver fibrosis caused by alcohol or HFD, but also suggest a conserved mechanism of gut and liver injury in different disease models through the gut-liver axis. Furthermore, we also published a few review articles on the important roles of various PTMs in mitochondria and nuclear proteins, contributing to increased mitochondrial dysfunction, altered gene expression (epigenetics), and multiple tissue injury through the gut-liver-brain axis. In these review articles, we described the opposing roles of CYP2E1 and ALDH2 in promoting mitochondrial dysfunction, autophagy impairments, and organ damage, as well as the protective mechanisms of POM, ellagic acid, and other naturally occurring antioxidants against gut leakiness liver and brain injury after exposure to binge alcohol or other potentially toxic agents, including HFD.