Different Types of Dietary Fat and Fructose Interactions Result in Distinct Metabolic Phenotypes in Male Mice.

Fat and fructose are the two major components over-represented in the Western diet. The aim of this study was to determine the combined effects of different types of dietary fat and fructose on the development of nonalcoholic fatty liver disease (NAFLD) in a murine model. Eight-week-old male C57BL/6J mice were fed with high-fat diet enriched with saturated fat (HSF), or omega-6 polyunsaturated fat (n6HUSF), or omega-3 polyunsaturated fat (n3HUSF) with 42% of calories derived from the fat. Fructose supplementation was given via 10% fructose (w/v) in the drinking water ad libitum for 20 weeks. While both HSF and n6HUSF fed mice developed obesity, HSF fed mice exhibited severe hepatic steatosis associated with hepatomegaly and liver injury. Fructose feeding promotes the development of liver fibrosis in HSF fed mice. n6HUSF fed mice were characterized with moderate hepatic steatosis, accompanied with hypertriglyceridemia and hyperlipidemia. Notably, fructose supplementation led to remarkable glucose intolerance in n6HUSF fed mice compared to controls. Hepatic lipidomic analysis revealed that the total saturated fatty acids and total monounsaturated fatty acids were significantly increased by fructose in the free fatty acid pool in HSF fed mice. Moreover, fructose supplementation increased hepatic and plasma cholesterol levels in the HSF fed mice. Our data suggest that excess energy from HSF intake results in fat storage in the liver, likely due to impaired triglyceride secretion; whereas excess energy from n6HUSF diet is stored in the periphery. Both effects are exacerbated by fructose supplementation. n3HUSF is beneficial, even consumed with fructose.

Plasma Metabolomics Analysis of Polyvinyl Chloride Workers Identifies Altered Processes and Candidate Biomarkers for Hepatic Hemangiosarcoma and Its Development.

BACKGROUND: High-level occupational vinyl chloride (VC) exposures have been associated with hepatic hemangiosarcoma, which typically develops following a long latency period. Although VC is genotoxic, a more comprehensive mode of action has not been determined and diagnostic biomarkers have not been established. The purpose of this study is to address these knowledge gaps through plasma metabolomics. METHODS: Plasma samples from polyvinyl chloride polymerization workers who developed hemangiosarcoma (cases, n = 15) and VC exposure-matched controls (n = 17) underwent metabolomic analysis. Random forest and bioinformatic analyses were performed. RESULTS: Cases and controls had similar demographics and routine liver biochemistries. Mass spectroscopy identified 606 known metabolites. Random forest analysis had an 82% predictive accuracy for group classification. 60 metabolites were significantly increased and 44 were decreased vs. controls. Taurocholate, bradykinin and fibrin degradation product 2 were up-regulated by greater than 80-fold. The naturally occurring anti-angiogenic phenol, 4-hydroxybenzyl alcohol, was down-regulated 5-fold. Top affected ontologies involved: (i) metabolism of bile acids, taurine, cholesterol, fatty acids and amino acids; (ii) inflammation and oxidative stress; and (iii) nicotinic cholinergic signaling. CONCLUSIONS: The plasma metabolome was differentially regulated in polyvinyl chloride workers who developed hepatic hemangiosarcoma. Ontologies potentially involved in hemangiosarcoma pathogenesis and candidate biomarkers were identified.

Effect of Epidermal Growth Factor Treatment and Polychlorinated Biphenyl Exposure in a Dietary-Exposure Mouse Model of Steatohepatitis.

BACKGROUND: Polychlorinated biphenyls (PCBs) are signaling disrupting chemicals that exacerbate nonalcoholic steatohepatitis (NASH) in mice. They are epidermal growth factor receptor (EGFR) inhibitors that enhance hepatic inflammation and fibrosis in mice. OBJECTIVES: This study tested the hypothesis that epidermal growth factor (EGF) administration can attenuate PCB-related NASH by increasing hepatic EGFR signaling in a mouse model. METHODS: C57BL/6 male mice were fed a 42% milk fat diet and exposed to Aroclor 1260 (20mg/kg) or vehicle for 12 wk. EGF (0.2µg/g) or vehicle were administered daily for 10 d starting at study week 10. Liver and metabolic phenotyping were performed. The EGF dose was selected based on results of an acute dose-finding study (30 min treatment of EGF at 0.2, 0.02, 0.002µg/g of via intraperitoneal injection). Hepatic phosphoproteomic analysis was performed using liver tissue from this acute study to understand EGFR's role in liver physiology. RESULTS: Markers of EGFR signaling were higher in EGF-treated mice. EGF+PCB-exposed mice had lower hepatic free fatty acids, inflammation, and fibrosis relative to PCB-only exposed mice. EGF-treated mice had higher plasma lipids, with no improvement in hepatic steatosis, and an association with higher LXR target gene expression and de novo lipogenesis. EGF-treated mice showed more severe hyperglycemia associated with lower adiponectin levels and insulin sensitivity. EGF-treated mice had higher hepatic HNF4α, NRF2, and AhR target gene expression but lower constitutive androstane receptor and farnesoid X receptor target gene expression. The hepatic EGF-sensitive phosphoproteome demonstrated a role for EGFR signaling in liver homeostasis. DISCUSSION: These results validated EGFR inhibition as a causal mode of action for PCB-related hepatic inflammation and fibrosis in a mouse model of NASH. However, observed adverse effects may limit the clinical translation of EGF therapy. More data are required to better understand EGFR's underinvestigated roles in liver and environmental health. https://doi.org/10.1289/EHP8222.

Analysis of sex differences in dietary copper-fructose interaction-induced alterations of gut microbial activity in relation to hepatic steatosis.

BACKGROUND: Inadequate copper intake and increased fructose consumption represent two important nutritional problems in the USA. Dietary copper-fructose interactions alter gut microbial activity and contribute to the development of nonalcoholic fatty liver disease (NAFLD). The aim of this study is to determine whether dietary copper-fructose interactions alter gut microbial activity in a sex-differential manner and whether sex differences in gut microbial activity are associated with sex differences in hepatic steatosis. METHODS: Male and female weanling Sprague-Dawley (SD) rats were fed ad libitum with an AIN-93G purified rodent diet with defined copper content for 8 weeks. The copper content is 6 mg/kg and 1.5 mg/kg in adequate copper diet (CuA) and marginal copper diet (CuM), respectively. Animals had free access to either deionized water or deionized water containing 10% fructose (F) (w/v) as the only drink during the experiment. Body weight, calorie intake, plasma alanine aminotransferase, aspartate aminotransferase, and liver histology as well as liver triglyceride were evaluated. Fecal microbial contents were analyzed by 16S ribosomal RNA (16S rRNA) sequencing. Fecal and cecal short-chain fatty acids (SCFAs) were determined by gas chromatography-mass spectrometry (GC-MS). RESULTS: Male and female rats exhibit similar trends of changes in the body weight gain and calorie intake in response to dietary copper and fructose, with a generally higher level in male rats. Several female rats in the CuAF group developed mild steatosis, while no obvious steatosis was observed in male rats fed with CuAF or CuMF diets. Fecal 16S rRNA sequencing analysis revealed distinct alterations of the gut microbiome in male and female rats. Linear discriminant analysis (LDA) effect size (LEfSe) identified sex-specific abundant taxa in different groups. Further, total SCFAs, as well as, butyrate were decreased in a more pronounced manner in female CuMF rats than in male rats. Of note, the decreased SCFAs are concomitant with the reduced SCFA producers, but not correlated to hepatic steatosis. CONCLUSIONS: Our data demonstrated sex differences in the alterations of gut microbial abundance, activities, and hepatic steatosis in response to dietary copper-fructose interaction in rats. The correlation between sex differences in metabolic phenotypes and alterations of gut microbial activities remains elusive.

Proteomics and metabolic phenotyping define principal roles for the aryl hydrocarbon receptor in mouse liver.

Dioxin-like molecules have been associated with endocrine disruption and liver disease. To better understand aryl hydrocarbon receptor (AHR) biology, metabolic phenotyping and liver proteomics were performed in mice following ligand-activation or whole-body genetic ablation of this receptor. Male wild type (WT) and Ahr -/- mice (Taconic) were fed a control diet and exposed to 3,3',4,4',5-pentachlorobiphenyl (PCB126) (61 nmol/kg by gavage) or vehicle for two weeks. PCB126 increased expression of canonical AHR targets (Cyp1a1 and Cyp1a2) in WT but not Ahr -/-. Knockouts had increased adiposity with decreased glucose tolerance; smaller livers with increased steatosis and perilipin-2; and paradoxically decreased blood lipids. PCB126 was associated with increased hepatic triglycerides in Ahr -/-. The liver proteome was impacted more so by Ahr -/- genotype than ligand-activation, but top gene ontology (GO) processes were similar. The PCB126-associated liver proteome was Ahr-dependent. Ahr principally regulated liver metabolism (e.g., lipids, xenobiotics, organic acids) and bioenergetics, but it also impacted liver endocrine response (e.g., the insulin receptor) and function, including the production of steroids, hepatokines, and pheromone binding proteins. These effects could have been indirectly mediated by interacting transcription factors or microRNAs. The biologic roles of the AHR and its ligands warrant more research in liver metabolic health and disease.

Dioxin-like and non-dioxin-like PCBs differentially regulate the hepatic proteome and modify diet-induced nonalcoholic fatty liver disease severity.

Polychlorinated biphenyls (PCBs) are persistent organic pollutants associated with metabolic disruption and non-alcoholic fatty liver disease (NAFLD). Based on their ability to activate the aryl hydrocarbon receptor (AhR), PCBs are subdivided into two classes: dioxin-like (DL) and non-dioxin-like (NDL) PCBs. Previously, we demonstrated that NDL PCBs compromised the liver to promote more severe diet-induced NAFLD. Here, the hepatic effects and potential mechanisms (by untargeted liver proteomics) of DL PCBs, NDL PCBs or co-exposure to both in diet-induced NAFLD are investigated. Male C57Bl/6 mice were fed a 42% fat diet and exposed to vehicle control; Aroclor1260 (20 mg/kg, NDL PCB mixture); PCB126 (20 µg/kg, DL PCB congener); or a mixture of Aroclor1260 (20 mg/kg)+PCB126 (20 µg/kg) for 12 weeks. Each exposure was associated with a distinct hepatic proteome. Phenotypic and proteomic analyses revealed increased hepatic inflammation and phosphoprotein signaling disruption by Aroclor1260. PCB126 decreased hepatic inflammation and fibrosis at the molecular level; while altering cytoskeletal remodeling, metal homeostasis, and intermediary/xenobiotic metabolism. PCB126 attenuated Aroclor1260-induced hepatic inflammation but increased hepatic free fatty acids in the co-exposure group. Aroclor1260+PCB126 exposure was strongly associated with multiple epigenetic processes, and these could potentially explain the observed non-additive effects of the exposures on the hepatic proteome. Taken together, the results demonstrated that PCB exposures differentially regulated the hepatic proteome and the histologic severity of diet-induced NAFLD. Future research is warranted to determine the AhR-dependence of the observed effects including metal homeostasis and the epigenetic regulation of gene expression.

Hepatic Injury Caused by the Environmental Toxicant Vinyl Chloride is Sex-Dependent in Mice.

Vinyl chloride (VC), a common industrial chemical, has been associated with hemangiosarcoma and toxicant-associated steatohepatitis (TASH) in men working at rubber-production plants. Our group previously demonstrated that chronic VC inhalation at environmentally relevant levels (< 1 ppm) in male mice exacerbated hepatic injury caused by high-fat diet (HFD) feeding. Because VC studies on TASH have only been performed in male models, the objective of this study is to examine VC inhalation in female mice in the context of TASH mechanisms. Male and female C57Bl/6 mice were fed either a low-fat diet or HFD and exposed to VC or room air using an inhalation chamber, for 12 weeks (6 h, 5 days/week); and plasma and liver samples were collected after euthanasia. Compared with males, females were less susceptible to HFD+VC-induced obesogenic effects demonstrated by lower body weight and fat composition. Histological analysis revealed that whereas VC exacerbated HFD-induced steatosis in males, this effect was absent in females. In addition, females were more resistant to VC-induced hepatic inflammation whereas males had increased liver weights and higher hepatic Tnfα mRNA levels. Systemic markers of hepatic injury, namely alanine aminotransaminase and thrombin/antithrombin levels were increased by HFD+VC co-exposures only in males. In addition, females did not show significant cell death as previously reported in males. Taken together, the results suggested that VC inhalation led to sex-dependent liver and metabolic toxicity. This study implicated the importance of assessing sex differences in environmental basic science and epidemiologic studies to better identify at-risk populations in both men and women.

Mechanisms of Environmental Contributions to Fatty Liver Disease.

PURPOSE: Fatty liver disease (FLD) affects over 25% of the global population and may lead to liver-related mortality due to cirrhosis and liver cancer. FLD caused by occupational and environmental chemical exposures is termed "toxicant-associated steatohepatitis" (TASH). The current review addresses the scientific progress made in the mechanistic understanding of TASH since its initial description in 2010. RECENT FINDINGS: Recently discovered modes of actions for volatile organic compounds and persistent organic pollutants include the following: (i) the endocrine-, metabolism-, and signaling-disrupting chemical hypotheses; (ii) chemical-nutrient interactions and the "two-hit" hypothesis. These key hypotheses were then reviewed in the context of the steatosis adverse outcome pathway (AOP) proposed by the US Environmental Protection Agency. The conceptual understanding of the contribution of environmental exposures to FLD has progressed significantly. However, because this is a new research area, more studies including mechanistic human data are required to address current knowledge gaps.

Identifying sex differences arising from polychlorinated biphenyl exposures in toxicant-associated liver disease.

Exposures to persistent environmental pollutants like polychlorinated biphenyls (PCBs) has been associated with liver diseases such as toxicant-associated steatohepatitis (TASH). However, previously published PCB hepatotoxicity studies evaluated mostly male animal models. Moreover, epidemiologic studies on PCB-exposed cohorts evaluating sex differences are scarce. Therefore, the objective of this study was to examine hepato-toxicological responses of PCB exposures in the context of sex-dependent outcomes. Male and female C57Bl/6 mice were exposed to Aroclor 1260 (20 mg/kg), and PCB126 (20 µg/kg), by gavage for two weeks. Female mice appeared to be more sensitive to PCB-induced hepatotoxic effects as manifested by increased liver injury markers, namely, hepatic Serpine1 expression. Additionally, compared to their male counterparts, PCB-exposed females exhibited dysregulated hepatic gene expression favoring lipid accumulation rather than lipid breakdown; accompanied by dyslipidemia. Sex differences were also observed in the expression and activation of PCB targets such as the epidermal growth factor receptor (EGFR) while PCB-induced pancreatic toxicity was similar in both sexes. Importantly, PCB exposure appeared to cause pro-androgenic, anti-estrogenic along with sex-dependent thyroid hormone effects. The overall findings demonstrated that the observed PCB-mediated hepatotoxicity was sex-dependent; confirming the existence of sex differences in environmental exposure-induced markers of TASH and warrants further investigation.

Proteomic Analysis Reveals Novel Mechanisms by Which Polychlorinated Biphenyls Compromise the Liver Promoting Diet-Induced Steatohepatitis.

Environmental pollution contributes to fatty liver disease pathogenesis. Polychlorinated biphenyl (PCB) exposures have been associated with liver enzyme elevation and suspected steatohepatitis in cohort studies. Male mice treated with the commercial PCB mixture, Aroclor 1260 (20 mg/kg), and fed high fat diet (HFD) for 12 weeks developed steatohepatitis. Receptor-based modes of action including inhibition of the epidermal growth factor (EGF) receptor were previously proposed, but other mechanisms likely exist. Objectives were to identify and validate the pathways, transcription factors, and mechanisms responsible for the steatohepatitis associated with PCB and HFD coexposures. Comparative proteomics analysis was performed in archived mouse liver samples from the aforementioned chronic exposure study. Pathway and transcription factor analysis (TFA) was performed, and selected results were validated. Liver proteomics detected 1103 unique proteins. Aroclor 1260 upregulated 154 and downregulated 93 of these. Aroclor 1260 + HFD coexposures affected 55 pathways including glutathione metabolism, intermediary metabolism, and cytoskeletal remodeling. TFA of Aroclor 1260 treatment demonstrated alterations in the function of 42 transcription factors including downregulation of NRF2 and key nuclear receptors previously demonstrated to protect against steatohepatitis (e.g., HNF4α, FXR, PPARα/δ/γ, etc.). Validation studies demonstrated that Aroclor 1260 significantly reduced HNF4α protein levels, while Aroclor 1260 + HFD reduced expression of the HNF4α target gene, albumin, in vivo. Aroclor 1260 attenuated EGF-dependent HNF4α phosphorylation and target gene activation in vitro. Aroclor 1260 reduced levels of NRF2, its target genes, and glutathione in vivo. Aroclor 1260 attenuated EGF-dependent NRF2 upregulation, in vitro. Aroclor 1260 indirectly activated hepatic stellate cells in vitro via induction of hepatocyte-derived TGFß. PCB exposures adversely impacted transcription factors regulating liver protection, function, and fibrosis. PCBs, thus, compromised the liver by reducing its protective responses against nutritional stress to promote diet-induced steatohepatitis. The identified mechanisms by which environmental pollutants influence fatty liver disease pathogenesis require confirmation in humans.

Polychlorinated biphenyl exposures differentially regulate hepatic metabolism and pancreatic function: Implications for nonalcoholic steatohepatitis and diabetes.

The endocrine disrupting chemicals, polychlorinated biphenyls (PCBs), have been associated with nonalcoholic steatohepatitis (NASH) and diabetes. However, an integrative analysis of the effects of PCBs on the liver and pancreas has never been performed for the two major PCB subtypes, dioxin-like (DL) and nondioxin-like (NDL), and a mixture of NDL/DL PCBs. Therefore, male C57BL/6 J mice fed a control synthetic diet were treated with either a NDL PCB mixture, Aroclor 1260 (20 mg/kg); a single DL PCB congener, PCB 126 (20 µg/kg); a NDL/DL mixture, Aroclor 1260 plus PCB 126; or vehicle control for 2 weeks. PCB126 had the greatest impact on hepatic lipid metabolism. It caused steatosis due to increased hepatic lipid import with associated hypolipidemia. However, all PCB exposures impacted expression of hepatic lipid metabolism genes in different manners. The 'NASH gene', Pnpla3, was elevated by Aroclor 1260, but decreased by all other exposures. The expression of hepatokines implicated in metabolic syndrome (Fgf21, Igf1, and betatrophin) were differentially regulated. The NDL/DL PCB mixture had the greatest effects on pancreatic histology, including acinar cell atrophy, mild steatosis, and fibrosis without ductal changes or immune cell infiltration. It decreased expression of insulin and altered the expression of genes regulating islet identity. None of these exposures was associated with altered HOMA-IR or HOMA-B. In summary, PCB exposures differentially regulated liver and pancreas structure and function. Novel mechanisms for PCB-induced endocrine/metabolic disruption included altered hepatokines and Pnpla3 as well as 'PCB pancreatopathy' that was associated with altered expression of pancreatic islet identity factors. More research is required to understand fully these findings in the context of human NASH and diabetes.

Hepatic signalling disruption by pollutant Polychlorinated biphenyls in steatohepatitis.

BACKGROUND: Polychlorinated biphenyl-mediated steatohepatitis has been shown to be due in part to inhibition of epidermal growth factor receptor (EGFR) signalling. EGFR signalling regulates many facets of hepatocyte function, but it is unclear which other kinases and pathways are involved in the development of toxicant-associated steatohepatitis (TASH). METHODS: Comparative hepatic phosphoproteomic analysis was used to identify which kinases were affected by either PCB exposure (Aroclor 1260 mixture), high fat diet (HFD), or their interaction in a chronic exposure model of TASH. Cellular assays and western blot analysis were used to validate the phosphoproteomic findings. RESULTS: 1760 unique phosphorylated peptides were identified and of those 588 were significantly different. PCB exposure and dietary interaction promoted a near 25% reduction of hepatic phospho-peptides. Leptin and insulin signalling were pathways highly affected by PCB exposure and liver necrosis was a pathologic ontology over represented due to interaction between PCBs and a HFD. Casein kinase 2 (CK2), Extracellular regulated kinase (ERK), Protein kinase B (AKT), and Cyclin dependent kinase (CDK) activity were demonstrated to be downregulated after PCB exposure and this downregulation was exacerbated with a HFD. PCB exposure led to a loss of hepatic CK2 subunit expression limiting CK2 kinase activity and negatively regulating caspase-3 (CASP3). PCBs promoted secondary necrosis in vitro validating the latter observation. The loss of hepatic phosphoprotein signalling appeared to be due to decreased signal transduction rather than phosphatase upregulation. CONCLUSIONS: PCBs are signal disrupting chemicals that promote secondary necrosis through affecting a myriad of liver processes including metabolism and cellular maintenance. PCB exposure, particularly with interaction with a HFD greatly down-regulates the hepatic kinome. More data are needed on signalling disruption and its impact on liver health.

Ethanol and unsaturated dietary fat induce unique patterns of hepatic ω-6 and ω-3 PUFA oxylipins in a mouse model of alcoholic liver disease.

Alcoholic liver disease (ALD), a significant health problem, progresses through the course of several pathologies including steatosis, steatohepatitis, fibrosis, and cirrhosis. There are no effective FDA-approved medications to prevent or treat any stages of ALD, and the mechanisms involved in ALD pathogenesis are not well understood. Bioactive lipid metabolites play a crucial role in numerous pathological conditions, as well as in the induction and resolution of inflammation. Herein, a hepatic lipidomic analysis was performed on a mouse model of ALD with the objective of identifying novel metabolic pathways and lipid mediators associated with alcoholic steatohepatitis, which might be potential novel biomarkers and therapeutic targets for the disease. We found that ethanol and dietary unsaturated, but not saturated, fat caused elevated plasma ALT levels, hepatic steatosis and inflammation. These pathologies were associated with increased levels of bioactive lipid metabolites generally involved in pro-inflammatory responses, including 13-hydroxy-octadecadienoic acid, 9,10- and 12,13-dihydroxy-octadecenoic acids, 5-, 8-, 9-, 11-, 15-hydroxy-eicosatetraenoic acids, and 8,9- and 11,12-dihydroxy-eicosatrienoic acids, in parallel with an increase in pro-resolving mediators, such as lipoxin A4, 18-hydroxy-eicosapentaenoic acid, and 10S,17S-dihydroxy-docosahexaenoic acid. Elucidation of alterations in these lipid metabolites may shed new light into the molecular mechanisms underlying ALD development/progression, and be potential novel therapeutic targets.

Dehydroepiandrosterone Research: Past, Current, and Future.

The discovery of "oestrus-producing" hormones was a major research breakthrough in biochemistry and pharmacology during the early part of the 20th century. The elucidation of the molecular weight and chemical structure of major oxidative metabolites of dehydroepiandrosterone (DHEA) led to the award of the Nobel Prize in 1939 to Adolf Frederick Johann Butenandt and Leopold Ruzicka. Considered a bulk androgen in the circulation, DHEA and its sulfated metabolite DHEA-S can be taken up by most tissues where the sterols are metabolized to active androgenic and estrogenic compounds needed for growth and development. Butenandt's interactions with the German pharmaceutical company Schering led to production of gram quantities of these steroids and other chemically modified compounds of this class. Sharing chemical expertise allowed Butenandt's laboratory at the Kaiser Wilhelm Institute to isolate and synthesize many steroid compounds in the elucidation of the pathway leading from cholesterol to testosterone and estrogen derivatives. As a major pharmaceutical company worldwide, Schering AG sought these new biological sterols as pharmacological agents for endocrine-related diseases, and the European medical community tested these compounds in women for conditions such as postmenopausal depression, and in men for increasing muscle mass. Since it was noted that circulating DHEA-S levels decline as a function of age, experimental pathology experiments in animals were performed to determine how DHEA may protect against cancer, diabetes, aging, obesity, immune function, bone density, depression, adrenal insufficiency, inflammatory bowel disease, diminished sexual function/libido, AIDS/HIV, chronic obstructive pulmonary disease, coronary artery disease, chronic fatigue syndrome, and metabolic syndrome. While the mechanisms by which DHEA ameliorates these conditions in animal models have been elusive to define, even less is known about its role in human disease, other than as a precursor to other sterols, e.g., testosterone and estradiol. Our groups have shown that DHEA and many of its oxidative metabolites serve as a low-affinity ligands for hepatic nuclear receptors, such as the pregnane X receptor, the constitutive androstane receptor, and estrogen receptors α/ß (ERα/ERß) as well as G protein-coupled ER (GPER1). This chapter highlights the founding research on DHEA from a historical perspective, provides an overview of DHEA biosynthesis and metabolism, briefly summarizes the early work on the beneficial effects attributed to DHEA in animals, and summarizes the human trials addressing the action of DHEA as a therapeutic agent. In general, most human studies involve weak correlations of circulating levels of DHEA and disease outcomes. Some support for DHEA as a therapeutic compound has been demonstrated for postmenopausal women, in vitro fertilization, and several autoimmune disorders, and adverse health effects, such as, acne, embryo virilization during pregnancy, and possible endocrine-dependent cancers.

Mechanisms of Action of Dehydroepiandrosterone.

Dehydroepiandrosterone (3ß-hydroxy-5-androsten-17-one, DHEA) and its sulfated metabolite DHEA-S are the most abundant steroids in circulation and decline with age. Rodent studies have shown that DHEA has a wide variety of effects on liver, kidney, adipose, reproductive tissues, and central nervous system/neuronal function. The mechanisms by which DHEA and DHEA-S impart their physiological effects may be direct actions on plasma membrane receptors, including a DHEA-specific, G-protein-coupled receptor in endothelial cells; various neuroreceptors, e.g., aminobutyric-acid-type A, N-methyl-d-aspartate (NMDA), and sigma-1 (S1R) receptors; by binding steroid receptors: androgen and estrogen receptors (ARs, ERα, or ERß); or by their metabolism to more potent sex steroid hormones, e.g., testosterone, dihydrotestosterone, and estradiol, which bind with higher affinity to ARs and ERs. DHEA inhibits voltage-gated T-type calcium channels. DHEA activates peroxisome proliferator-activated receptor (PPARα) and CAR by a mechanism apparently involving PP2A, a protein phosphatase dephosphorylating PPARα and CAR to activate their transcriptional activity. We review our recent study showing DHEA activated GPER1 (G-protein-coupled estrogen receptor 1) in HepG2 cells to stimulate miR-21 transcription. This chapter reviews some of the physiological, biochemical, and molecular mechanisms of DHEA and DHEA-S activity.

Liver Disease in a Residential Cohort With Elevated Polychlorinated Biphenyl Exposures.

Endocrine and metabolism disrupting chemicals (EDCs/MDCs) have been associated with environmental liver diseases including toxicant-associated steatohepatitis (TASH). TASH has previously been characterized by hepatocellular necrosis, disrupted intermediary metabolism, and liver inflammation. Polychlorinated biphenyls (PCBs) are environmental EDCs/MDCs associated with the genesis and progression of steatohepatitis in animal models and human liver injury in epidemiology studies. The cross-sectional Anniston Community Health Survey (ACHS) investigates ortho-substituted PCB exposures and health effects near a former PCB manufacturing complex. The rates of obesity, diabetes, and dyslipidemia were previously determined to be high in ACHS. In this study, 738 ACHS participants were categorized by liver disease status using the serum cytokeratin 18 biomarker. Associations between PCB exposures and mechanistic biomarkers of intermediary metabolism, inflammation, and hepatocyte death were determined. The liver disease prevalence was high (60.2%), and 80.7% of these individuals were categorized as having TASH. Sex and race/ethnicity differences were noted. TASH was associated with increased exposures to specific PCB congeners, insulin resistance, dyslipidemia, proinflammatory cytokines, and liver necrosis. These findings are consistent with PCB-related steatohepatitis. ΣPCBs was inversely associated with insulin resistance/production, leptin, and hepatocyte apoptosis, while other adipocytokines were increased. This is possibly the largest environmental liver disease study applying mechanistic biomarkers ever performed and the most comprehensive analysis of PCBs and adipocytokines. It provides insight into the mechanisms of PCB-related endocrine and metabolic disruption in liver disease and diabetes. In the future, associations between additional exposures and liver disease biomarkers will be evaluated in the ACHS and follow-up ACHS-II studies.

Epidermal Growth Factor Receptor Signaling Disruption by Endocrine and Metabolic Disrupting Chemicals.

The purpose of this study is to identify an environmentally relevant shared receptor target for endocrine and metabolism disrupting chemical pollutants. A feature of the tested chemicals was that they induced Cyp2b10 in vivo implicating activation of the constitutive androstane receptor (CAR). Recent studies suggest that these compounds could be indirect CAR activators via epidermal growth factor receptor (EGFR) inhibition. Assays included a CAR activity reporter assay, EGF endocytosis assay, and EGFR phosphorylation assay. Docking simulations were used to identify putative binding sites for environmental chemicals on the EGFR. Whole-weight and lipid-adjusted serum mean pollutant exposures were determined using data from the National Health and Examination Survey (NHANES) and compared with the IC50 values determined in vitro. Chlordane, trans-nonachlor, PCB-126, PCB-153, and atrazine were the most potent EGFR inhibitors tested. PCB-126, PCB-153, and trans-nonachlor appeared to be competitive EGFR antagonists as they displaced bound EGF from EGFR. However, atrazine acted through a different mechanism and could be an EGFR tyrosine kinase inhibitor. EGFR inhibition relative effect potencies were determined for these compounds. In NHANES, serum concentrations of trans-nonachlor, PCB-126, and PCB-153 greatly exceeded their calculated IC50 values. A common mechanism of action through EGFR inhibition for three diverse classes of metabolic disrupting chemicals was characterized by measuring inhibition of EGFR phosphorylation and EGF-EGFR endocytosis. Based on NHANES data, EGFR inhibition may be an environmentally relevant mode of action for some PCBs, pesticides, and herbicides.

Dietary copper-fructose interactions alter gut microbial activity in male rats.

Dietary copper-fructose interactions contribute to the development of nonalcoholic fatty liver disease (NAFLD). Gut microbiota play critical roles in the pathogenesis of NAFLD. The aim of this study was to determine the effect of different dietary doses of copper and their interactions with high fructose on gut microbiome. Male weanling Sprague-Dawley rats were fed diets with adequate copper (6 ppm CuA), marginal copper (1.5 ppm CuM) (low copper), or supplemented copper (20 ppm CuS) (high copper) for 4 wk. Deionized water or deionized water containing 30% fructose (wt/vol) was given ad libitum. Copper status, liver enzymes, gut barrier function, and gut microbiome were evaluated. Both low- and high-copper diets led to liver injury in high-fructose-fed rats, and this was associated with gut barrier dysfunction, as shown by the markedly decreased tight junction proteins and increased gut permeability. 16S rDNA sequencing analysis revealed distinct alterations of the gut microbiome associated with dietary low- and high-copper/high-fructose feeding. The common features of the alterations of the gut microbiome were the increased abundance of Firmicutes and the depletion of Akkermansia. However, they differed mainly within the phylum Firmicutes. Our data demonstrated that a complex interplay among host, microbes, and dietary copper-fructose interaction regulates gut microbial metabolic activity, which may contribute to the development of liver injury and hepatic steatosis. The distinct alterations of gut microbial activity, which were associated with the different dietary doses of copper and fructose, imply that separate mechanism(s) may be involved. NEW & NOTEWORTHY First, dietary low- and high-copper/high-fructose-induced liver injury are associated with distinct alterations of gut microbiome. Second, dietary copper level plays a critical role in maintaining the gut barrier integrity, likely by acting on the intestinal tight junction proteins and the protective commensal bacteria Akkermansia. Third, the alterations of gut microbiome induced by dietary low and high copper with or without fructose differ mainly within the phylum Firmicutes.