A Panel of Protein Kinase Chemosensors Distinguishes Different Types of Fatty Liver Disease

The worldwide incidence of fatty liver disease continues to rise, which may account for concurrent increases in the frequencies of more aggressive liver ailments. Given the existence of histologically identical fatty liver disease subtypes, there is a critical need for the identification of methods that can classify disease and potentially predict progression. Herein, we show that a panel of protein kinase chemosensors can distinguish fatty liver disease subtypes. These direct activity measurements highlight distinct differences between histologically identical fatty liver diseases arising from diets rich in fat versus alcohol and identify a previously unreported decrease in p38α activity associated with a high-fat diet. In addition, we have profiled kinase activities in both benign (dietinduced) and progressive (STAM) disease models. These experiments provide temporal insights into kinase activity during disease development and progression. Altogether, this work provides the basis for the future development of clinical diagnostics and potential treatment strategies

Chronic alcohol exposure alters circulating insulin and ghrelin levels: role of ghrelin in hepatic steatosis

Fatty liver is the earliest response of the liver to excessive ethanol consumption. Central in the development of alcoholic steatosis is increased mobilization of nonesterified free fatty acids (NEFAs) to the liver from the adipose tissue. In this study, we hypothesized that ethanol-induced increase in ghrelin by impairing insulin secretion, could be responsible for the altered lipid metabolism observed in adipose and liver tissue. Male Wistar rats were fed for 5–8 wk with control or ethanol Lieber-DeCarli diet, followed by biochemical analyses in serum and liver tissues. In addition, in vitro studies were conducted on pancreatic islets isolated from experimental rats. We found that ethanol increased serum ghrelin and decreased serum insulin levels in both fed and fasting conditions. These results were corroborated by our observations of a significant accumulation of insulin in pancreatic islets of ethanol-fed rats, indicating that its secretion was impaired. Furthermore, ethanol-induced reduction in circulating insulin was associated with lower adipose weight and increased NEFA levels observed in these rats. Additionally, we found that increased concentration of serum ghrelin was due to increased synthesis and maturation in the stomach of the ethanol-fed rats. We also report that in addition to its effect on the pancreas, ghrelin can also directly act on hepatocytes via the ghrelin receptors and promote fat accumulation. In conclusion, alcohol-induced elevation of circulating ghrelin levels impairs insulin secretion. Consequently, reduced circulating insulin levels likely contribute to increased free fatty acid mobilization from adipose tissue to liver, thereby contributing to hepatic steatosis

Elevated S-Adenosylhomocysteine Induces Adipocyte Dysfunction to Promote Alcohol-Associated Liver Steatosis

It has been previously shown that chronic ethanol administration-induced increase in adipose tissue lipolysis and reduction in the secretion of protective adipokines collectively contribute to alcohol-associated liver disease (ALD) pathogenesis. Further studies have revealed that increased adipose S-adenosylhomocysteine (SAH) levels generate methylation defects that promote lipolysis. Here, we hypothesized that increased intracellular SAH alone causes additional related pathological changes in adipose tissue as seen with alcohol administration. To test this, we used 3-deazaadenosine (DZA), which selectively elevates intracellular SAH levels by blocking its hydrolysis. Fully differentiated 3T3-L1 adipocytes were treated in vitro for 48 h with DZA and analysed for lipolysis, adipokine release and differentiation status. DZA treatment enhanced adipocyte lipolysis, as judged by lower levels of intracellular triglycerides, reduced lipid droplet sizes and higher levels of glycerol and free fatty acids released into the culture medium. These findings coincided with activation of both adipose triglyceride lipase and hormone sensitive lipase. DZA treatment also significantly reduced adipocyte differentiation factors, impaired adiponectin and leptin secretion but increased release of pro-inflammatory cytokines, IL-6, TNF and MCP-1. Together, our results demonstrate that elevation of intracellular SAH alone by DZA treatment of 3T3-L1 adipocytes induces lipolysis and dysregulates adipokine secretion. Selective elevation of intracellular SAH by DZA treatment mimics ethanol\u27s effects and induces adipose dysfunction. We conclude that alcohol-induced elevations in adipose SAH levels contribute to the pathogenesis and progression of ALD

Chronic alcohol exposure alters circulating insulin and ghrelin levels: role of ghrelin in hepatic steatosis

Fatty liver is the earliest response of the liver to excessive ethanol consumption. Central in the development of alcoholic steatosis is increased mobilization of nonesterified free fatty acids (NEFAs) to the liver from the adipose tissue. In this study, we hypothesized that ethanol-induced increase in ghrelin by impairing insulin secretion, could be responsible for the altered lipid metabolism observed in adipose and liver tissue. Male Wistar rats were fed for 5–8 wk with control or ethanol Lieber-DeCarli diet, followed by biochemical analyses in serum and liver tissues. In addition, in vitro studies were conducted on pancreatic islets isolated from experimental rats. We found that ethanol increased serum ghrelin and decreased serum insulin levels in both fed and fasting conditions. These results were corroborated by our observations of a significant accumulation of insulin in pancreatic islets of ethanol-fed rats, indicating that its secretion was impaired. Furthermore, ethanol-induced reduction in circulating insulin was associated with lower adipose weight and increased NEFA levels observed in these rats. Additionally, we found that increased concentration of serum ghrelin was due to increased synthesis and maturation in the stomach of the ethanol-fed rats. We also report that in addition to its effect on the pancreas, ghrelin can also directly act on hepatocytes via the ghrelin receptors and promote fat accumulation. In conclusion, alcohol-induced elevation of circulating ghrelin levels impairs insulin secretion. Consequently, reduced circulating insulin levels likely contribute to increased free fatty acid mobilization from adipose tissue to liver, thereby contributing to hepatic steatosis

The Effects of Age and Alcohol on Lipid Metabolism in the Liver

Background: Alcohol-associated liver disease (ALD) encompasses the liver manifestation of chronic alcohol abuse, characterized by different stages of liver damage that progresses from fat accumulation to steatohepatitis, fibrosis and eventually cirrhosis. The severity of liver damage is influenced by age, which is also a predictor for ALD-related mortality. Thus, the purpose of this study was to investigate how aging and alcohol affect lipid metabolism in the liver. Methods: Rats aged 4 months, 8 months, 12 months, and 22 months-old were pair-fed Lieber-DeCarli control or ethanol diet for 6 weeks. Serum and liver were collected for analyses when rats were euthanized. Analyses included histopathology, measurements of non-esterified fatty acid content and hepatic triglyceride content, and gene expression.https://digitalcommons.unmc.edu/surp2022/1034/thumbnail.jp

Acute Ethanol-Induced Liver Injury is Prevented by Betaine Administration

Binge drinking is the most common form of excessive alcohol use. Repeated episodes of binge drinking cause multiple organ injuries, including liver damage. We previously demonstrated that chronic ethanol administration causes a decline in the intrahepatic ratio of S-adenosylmethionine (SAM) to S-adenosylhomocysteine (SAH). This decline causes impairments in essential methylation reactions that result in alcohol-induced fatty liver (steatosis) and other features of alcohol-associated liver disease (ALD). Co-treatment with betaine during chronic ethanol feeding, normalizes hepatocellular SAM:SAH ratio and alleviates many features of liver damage including steatosis. Here, we sought to examine whether betaine treatment similarly protects against liver injury in an alcohol binge-drinking model. We hypothesized that ethanol binge with prior or simultaneous betaine administration would prevent or attenuate acute alcohol-induced liver damage. Male C57Bl/6 mice were gavaged twice, 12 h apart, with either 6 g ethanol/kg BW or with an equal volume/kg BW of 0.9% NaCl. Two separate groups of mice (n = 5/group) were gavaged with 4 g betaine/kg BW, either 2 h before or simultaneously with the ethanol or saline gavages. All mice were sacrificed 8 h after the last gavage and serum and liver parameters were quantified. Ethanol binges caused a 50% decrease in hepatic SAM:SAH ratio and a \u3e3-fold rise in liver triglycerides (p ≤ 0.05). These latter changes were accompanied by elevated serum AST and ALT activities and blood alcohol concentrations (BAC) that were ∼three-times higher than the legal limit of intoxication in humans. Mice that were treated with betaine 2 h before or simultaneously with the ethanol binges exhibited similar BAC as in mice given ethanol-alone. Both betaine treatments significantly elevated hepatic SAM levels thereby normalizing the SAM:SAH ratio and attenuating hepatic steatosis and other injury parameters, compared with mice given ethanol alone. Simultaneous betaine co-administration with ethanol was more effective in preventing or attenuating liver injury than betaine given before ethanol gavage. Our findings confirm the potential therapeutic value of betaine administration in preventing liver injury after binge drinking in an animal model

Alcoholic vs non-alcoholic fatty liver in rats: distinct differences in endocytosis and vesicle trafficking despite similar pathology

Background: Non-alcoholic and alcoholic fatty liver disease (NAFLD and AFLD, respectively) are major health problems, as patients with either condition can progress to hepatitis, fibrosis, and cirrhosis. Although histologically similar, key differences likely exist in these two models. For example, altered content of several vesicle trafficking proteins have been identified in AFLD, but their content in NAFLD is unknown. In this study, we compared select parameters in NAFLD and AFLD in a rat model. Methods: We fed either Lieber- DeCarli liquid control or alcohol-containing (35 % as calories) diet (AFLD model) or lean or high-fat (12 or 60 % derived from fat, respectively) pellets (NAFLD model) for 8–10 weeks, n = 8 in each model. Serum, hepatocytes and liver tissue were analyzed. Liver injury markers were measured in serum, triglyceride content and endocytosis (binding and internalization of 125I- asialoorosomucoid) was measured in isolated hepatocytes, and content of selected trafficking proteins (Rab3D, Rab7 and Rab18) were determined in whole liver tissue. Results: Although liver injury markers and triglyceride content were similar in both models, binding and internalization of 125I- asialoorosomucoid was significantly impaired in the hepatocytes from AFLD, but not NAFLD, animals. In addition, protein content of the asialoglycoprotein receptor (ASGPR) and three trafficking proteins, Rab3D, Rab7and Rab18, were significantly decreased after alcohol, but not high-fat feeding. Levels of protein carbonylation, amount of glutathione stores, and lipid peroxidation were similar irrespective of the insult to the livers that resulted in fatty liver. Conclusion: Impairments in protein trafficking in AFLD are likely a direct result of alcohol administration, and not a function of fatty liver

\u3ci\u3eIn Vivo\u3c/i\u3e Selection To Identify Bacterial Strains with Enhanced Ecological Performance in Synbiotic Applications

One strategy for enhancing the establishment of probiotic bacteria in the human intestinal tract is via the parallel administration of a prebiotic, which is referred to as a synbiotic. Here we present a novel method that allows a rational selection of putative probiotic strains to be used in synbiotic applications: in vivo selection (IVS). This method consists of isolating candidate probiotic strains from fecal samples following enrichment with the respective prebiotic. To test the potential of IVS, we isolated bifidobacteria from human subjects who consumed increasing doses of galactooligosaccharides (GOS) for 9 weeks. A retrospective analysis of the fecal microbiota of one subject revealed an 8-fold enrichment in Bifidobacterium adolescentis strain IVS-1 during GOS administration. The functionality of GOS to support the establishment of IVS-1 in the gastrointestinal tract was then evaluated in rats administered the bacterial strain alone, the prebiotic alone, or the synbiotic combination. Strain-specific quantitative real-time PCR showed that the addition of GOS increased B. adolescentis IVS-1 abundance in the distal intestine by nearly 2 logs compared to rats receiving only the probiotic. Illumina 16S rRNA sequencing not only confirmed the increased establishment of IVS-1 in the intestine but also revealed that the strain was able to outcompete the resident Bifidobacterium population when provided with GOS. In conclusion, this study demonstrated that IVS can be used to successfully formulate a synergistic synbiotic that can substantially enhance the establishment and competitiveness of a putative probiotic strain in the gastrointestinal tract

\u3ci\u3eIn Vivo\u3c/i\u3e Selection To Identify Bacterial Strains with Enhanced Ecological Performance in Synbiotic Applications

One strategy for enhancing the establishment of probiotic bacteria in the human intestinal tract is via the parallel administration of a prebiotic, which is referred to as a synbiotic. Here we present a novel method that allows a rational selection of putative probiotic strains to be used in synbiotic applications: in vivo selection (IVS). This method consists of isolating candidate probiotic strains from fecal samples following enrichment with the respective prebiotic. To test the potential of IVS, we isolated bifidobacteria from human subjects who consumed increasing doses of galactooligosaccharides (GOS) for 9 weeks. A retrospective analysis of the fecal microbiota of one subject revealed an 8-fold enrichment in Bifidobacterium adolescentis strain IVS-1 during GOS administration. The functionality of GOS to support the establishment of IVS-1 in the gastrointestinal tract was then evaluated in rats administered the bacterial strain alone, the prebiotic alone, or the synbiotic combination. Strain-specific quantitative real-time PCR showed that the addition of GOS increased B. adolescentis IVS-1 abundance in the distal intestine by nearly 2 logs compared to rats receiving only the probiotic. Illumina 16S rRNA sequencing not only confirmed the increased establishment of IVS-1 in the intestine but also revealed that the strain was able to outcompete the resident Bifidobacterium population when provided with GOS. In conclusion, this study demonstrated that IVS can be used to successfully formulate a synergistic synbiotic that can substantially enhance the establishment and competitiveness of a putative probiotic strain in the gastrointestinal tract

Natural Recovery by the Liver and Other Organs after Chronic Alcohol Use

Chronic, heavy alcohol consumption disrupts normal organ function and causes structural damage in virtually every tissue of the body. Current diagnostic terminology states that a person who drinks alcohol excessively has alcohol use disorder. The liver is especially susceptible to alcohol-induced damage. This review summarizes and describes the effects of chronic alcohol use not only on the liver, but also on other selected organs and systems affected by continual heavy drinking-including the gastrointestinal tract, pancreas, heart, and bone. Most significantly, the recovery process after cessation of alcohol consumption (abstinence) is explored. Depending on the organ and whether there is relapse, functional recovery is possible. Even after years of heavy alcohol use, the liver has a remarkable regenerative capacity and, following alcohol removal, can recover a significant portion of its original mass and function. Other organs show recovery after abstinence as well. Data on studies of both heavy alcohol use among humans and animal models of chronic ethanol feeding are discussed. This review describes how (or whether) each organ/tissue metabolizes ethanol, as metabolism influences the organ\u27s degree of injury. Damage sustained by the organ/tissue is reviewed, and evidence for recovery during abstinence is presented