Accumulation Of Lipid Droplets In Hepatocytes Research Articles

Ethanol disrupts hepatocellular lipophagy by altering Rab5-centric LD-lysosome trafficking.

Previous reports suggest that lipid droplets (LDs) in the hepatocyte can be catabolized by a direct engulfment from nearby endolysosomes (microlipophagy). Further, it is likely that this process is compromised by chronic ethanol (EtOH) exposure leading to hepatic steatosis. This study investigates the hepatocellular machinery supporting microlipophagy and EtOH-induced alterations in this process with a focus on the small, endosome-associated, GTPase Rab5. Here we report that this small Ras-related GTPase is a resident component of LDs, and its activity is important for hepatocellular LD-lysosome proximity and physical interactions. We find that Rab5 siRNA knockdown causes an accumulation of LDs in hepatocytes by inhibiting lysosome dependent LD catabolism. Importantly, Rab5 appears to support this process by mediating the recruitment of early endosomal and or multivesicular body compartments to the LD surface before lysosome fusion. Interestingly, while wild-type or a constituently active GTPase form (Q79L) of Rab5 supports LD-lysosome transport, this process is markedly reduced in cells expressing a GTPase dead (S34N) Rab5 protein or in hepatocytes exposed to chronic EtOH. These findings support the novel premise of an early endosomal/multivesicular body intermediate compartment on the LD surface that provides a "docking" site for lysosomal trafficking, not unlike the process that occurs during the hepatocellular degradation of endocytosed ligands that is also known to be compromised by EtOH exposure.

The hepatitis B virus promotes the progression of non-alcoholic fatty liver disease through incomplete autophagy.

Hepatitis B virus (HBV) infection is still a serious public health problem. In recent years, with the increasing incidence of chronic hepatitis B (CHB) combined with nonalcoholic fatty liver disease (NAFLD), a more in-depth exploration of the pathogenesis of CHB combined with NAFLD is required. HBV can induce autophagy and use to increase replication. The removal of fat by autophagy, also known as lipophagy, is also currently considered an alternative pathway for lipid metabolism in liver cells. This degradation of autophagy prevents hepatotoxicity and steatosis. However, it is not known whether there is a correlation between HBV-related autophagy and the progression of NAFLD. We explored how HBV affects disease progression in NAFLD should be " and determined whether it is associated with HBV-associated autophagy. In this study, we constructed HBV-TG mouse high-fat diet (HFD) models and controls, and the results showed that the presence of HBV promoted the occurrence of NAFLD. We also demonstrated that HBV promotes lipid droplet accumulation in hepatocytes using HBV-stable expression cell lines HepG2.2.15 and AML12-HBV. In addition, this study also found that exogenous OA supplementation reduced HBV replication. We further studied the mechanism and found that HBV-related autophagy can promote the absorption of liver cells to lipid droplets. It can reduce the decomposition of lipid droplets by inhibiting the function of autophagolysosome, and eventually lead to the accumulation of lipid droplets in hepatocytes. In a word, HBV promotes the progression of NAFLD by increasing lipid accumulation in hepatocytes through incomplete autophagy.

Hif1α/Dhrs3a Pathway Participates in Lipid Droplet Accumulation via Retinol and Ppar-γ in Fish Hepatocytes.

Excessive hepatic lipid accumulation is a common phenomenon in cultured fish; however, its underlying mechanisms are poorly understood. Lipid droplet (LD)-related proteins play vital roles in LD accumulation. Herein, using a zebrafish liver cell line (ZFL), we show that LD accumulation is accompanied by differential expression of seven LD-annotated genes, among which the expression of dehydrogenase/reductase (SDR family) member 3 a/b (dhrs3a/b) increased synchronously. RNAi-mediated knockdown of dhrs3a delayed LD accumulation and downregulated the mRNA expression of peroxisome proliferator-activated receptor gamma (pparg) in cells incubated with fatty acids. Notably, Dhrs3 catalyzed retinene to retinol, the content of which increased in LD-enriched cells. The addition of exogenous retinyl acetate maintained LD accumulation only in cells incubated in a lipid-rich medium. Correspondingly, exogenous retinyl acetate significantly increased pparg mRNA expression levels and altered the lipidome of the cells by increasing the phosphatidylcholine and triacylglycerol contents and decreasing the cardiolipin, phosphatidylinositol, and phosphatidylserine contents. Administration of LW6, an hypoxia-inducible factor 1α (HIF1α) inhibitor, reduced the size and number of LDs in ZFL cells and attenuated hif1αa, hif1αb, dhrs3a, and pparg mRNA expression levels. We propose that the Hif-1α/Dhrs3a pathway participates in LD accumulation in hepatocytes, which induces retinol formation and the Ppar-γ pathway.

Impaired Function of Solute Carrier Family 19 Leads to Low Folate Levels and Lipid Droplet Accumulation in Hepatocytes.

Low serum folate levels are inversely related to metabolic associated fatty liver disease (MAFLD). The role of the folate transporter gene (SLC19A1) was assessed to clarify its involvement in lipid accumulation during the onset of MAFLD in humans and in liver cells by genomic, transcriptomic, and metabolomic techniques. Genotypes of 3 SNPs in a case-control cohort were initially correlated to clinical and serum MAFLD markers. Subsequently, the expression of 84 key genes in response to the loss of SLC19A1 was evaluated with the aid of an RT2 profiler-array. After shRNA-silencing of SLC19A1 in THLE2 cells, folate and lipid levels were measured by ELISA and staining techniques, respectively. In addition, up to 482 amino acids and lipid metabolites were semi-quantified in SLC19A1-knockdown (KD) cells through ultra-high-performance liquid chromatography coupled with mass spectrometry. SNPs, rs1051266 and rs3788200, were significantly associated with the development of fatty liver for the single-marker allelic test. The minor alleles of these SNPs were associated with a 0.6/-1.67-fold decreased risk of developing MAFLD. When SLC19A1 was KD in THLE2 cells, intracellular folate content was four times lower than in wild-type cells. The lack of functional SLC19A1 provoked significant changes in the regulation of genes associated with lipid droplet accumulation within the cell and the onset of NAFLD. Metabolomic analyses showed a highly altered profile, where most of the species that accumulated in SLC19A1-KD-cells belong to the chemical groups of triacylglycerols, diacylglycerols, polyunsaturated fatty acids, and long chain, highly unsaturated cholesterol esters. In conclusion, the lack of SLC19A1 gene expression in hepatocytes affects the regulation of key genes for normal liver function, reduces intracellular folate levels, and impairs lipid metabolism, which entails lipid droplet accumulation in hepatocytes.

MTORC2 Facilitates Liver Regeneration Through Sphingolipid-Induced PPAR-α-Fatty Acid Oxidation.

During liver regeneration after partial hepatectomy, the function and metabolic pathways governing transient lipid droplet accumulation in hepatocytes remain obscure. Mammalian target of rapamycin 2 (mTORC2) facilitates de novo synthesis of hepatic lipids. Under normal conditions and in tumorigenesis, decreased levels of triglyceride (TG) and fatty acids (FAs) are observed in the mTORC2-deficient liver. However, during liver regeneration, their levels increase in the absence of mTORC2. Rictor liver-specific knockout and control mice underwent partial hepatectomy, followed by measurement of TG and FA contents during liver regeneration. FA metabolism was evaluated by analyzing the expression of FA metabolism-related genes and proteins. Intraperitoneal injection of the peroxisome proliferator-activated receptor α (PPAR-α) agonist, p53 inhibitor, and protein kinase B (AKT) activator was performed to verify the regulatory pathways involved. Lipid mass spectrometry was performed to identify the potential PPAR-α activators. The expression of FA metabolism-related genes and proteins suggested that FAs are mainly transported into hepatocytes during liver regeneration. The PPAR-α pathway is down-regulated significantly in the mTORC2-deficient liver, resulting in the accumulation of TGs. The PPAR-α agonist WY-14643 rescued deficient liver regeneration and survival in mTORC2-deficient mice. Furthermore, lipidomic analysis suggested that mTORC2 deficiency substantially reduced glucosylceramide (GluCer) content. GluCer activated PPAR-α. GluCer treatment in vivo restored the regenerative ability and survival rates in the mTORC2-deficient group. Our data suggest that FAs are mainly transported into hepatocytes during liver regeneration, and their metabolism is facilitated by mTORC2 through the GluCer-PPAR-α pathway, thereby establishing a novel role for mTORC2 in lipid metabolism.

A high-fat diet catalyzes progression to hyperglycemia in mice with selective impairment of insulin action in Glut4-expressing tissues

Insulin resistance impairs postprandial glucose uptake through glucose transporter type 4 (GLUT4) and is the primary defect preceding type 2 diabetes. We previously generated an insulin-resistant mouse model with human GLUT4 promoter-driven insulin receptor knockout (GIRKO) in the muscle, adipose, and neuronal subpopulations. However, the rate of diabetes in GIRKO mice remained low prior to 6 months of age on normal chow diet (NCD), suggesting that additional factors/mechanisms are responsible for adverse metabolic effects driving the ultimate progression of overt diabetes. In this study, we characterized the metabolic phenotypes of the adult GIRKO mice acutely switched to high-fat diet (HFD) feeding in order to identify additional metabolic challenges required for disease progression. Distinct from other diet-induced obesity (DIO) and genetic models (e.g., db/db mice), GIRKO mice remained leaner on HFD feeding, but developed other cardinal features of insulin resistance syndrome. GIRKO mice rapidly developed hyperglycemia despite compensatory increases in β-cell mass and hyperinsulinemia. Furthermore, GIRKO mice also had impaired oral glucose tolerance and a limited glucose-lowering benefit from exendin-4, suggesting that the blunted incretin effect contributed to hyperglycemia. Secondly, GIRKO mice manifested severe dyslipidemia while on HFD due to elevated hepatic lipid secretion, serum triglyceride concentration, and lipid droplet accumulation in hepatocytes. Thirdly, GIRKO mice on HFD had increased inflammatory cues in the gut, which were associated with the HFD-induced microbiome alterations and increased serum lipopolysaccharide (LPS). In conclusion, our studies identified important gene/diet interactions contributing to diabetes progression, which might be leveraged to develop more efficacious therapies.

Free fatty acids induce the demethylation of the fructose 1,6-biphosphatase 2 gene promoter and potentiate its expression in hepatocytes.

Obesity is a serious health issue as it is a social burden and the main risk factor for other metabolic diseases. Increasing evidence indicates that a high-fat diet (HFD) is the key factor for the development of obesity, but the key genes and their associated molecular mechanisms are still not fully understood. In this study, we performed integrated bioinformatic analysis and identified that fructose-1,6 biphosphatase 2 (FBP2) was involved in free fatty acids (FFAs)-induced lipid droplet accumulation in hepatocytes and HFD-induced obesity in mice. Our data showed that palmitate (PA) and oleic acid (OA) induced the expression of FBP2 in time- and dose-dependent manners, and accelerated the development of lipid droplets in LO2 human normal liver cells. In HFD-fed C57BL/6 mice, accompanied by insulin resistance and lipid droplet accumulation, the mRNA and protein levels of FBP2 in the livers also increased significantly. The results from the methylation sequencing PCR (MSP) and bisulfite specific PCR (BSP) indicated that PA/OA induced the demethylation of the FBP2 gene promoter in LO2 cells. Moreover, betaine, a methyl donor, attenuated the expression of the FBP2 gene, the accumulation of lipid droplets, and the expression of perilipin-2, a biomarker of lipid droplets, in LO2 cells. All these findings revealed that FBP2 might be involved in HFD-induced obesity, and it is of interest to investigate the role of FBP2 in the treatment and prevention of obesity and its associated complications.

The protective effects of the \u03b23 adrenergic receptor agonist BRL37344 against liver steatosis and inflammation in a rat model of high-fat diet-induced nonalcoholic fatty liver disease (NAFLD)

BackgroundOur objective was to investigate the efficacy of the beta-3 adrenergic receptor (β3-AR) agonist BRL37344 for the prevention of liver steatosis and inflammation associated with nonalcoholic fatty liver disease (NAFLD).MethodsFour groups were established: a control group (given a standard diet), a high-fat diet (HFD) group, an HFD + β3-AR agonist (β3-AGO) group, and an HFD + β3-AR antagonist (β3-ANT) group. All rats were fed for 12 weeks. The β3-AR agonist BRL37344 and the antagonist L748337 were administered for the last 4 weeks with Alzet micro-osmotic pumps. The rat body weights (g) were measured at the end of the 4th, 8th, and 12th weeks. At the end of the 12th week, the liver weights were measured. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were analyzed with a Hitachi automatic analyzer. The lipid levels of the triglycerides (TGs), total cholesterol (TC), and low-density lipoprotein cholesterol (LDL-C) and the concentrations of free fatty acids (FFAs) were also measured. An oil red O kit was used to detect lipid droplet accumulation in hepatocytes. Steatosis, ballooning degeneration and inflammation were histopathologically determined. The protein and mRNA expression levels of β3-AR, peroxisome proliferator-activated receptor-alpha (PPAR-α), peroxisome proliferator-activated receptor-gamma (PPAR-γ), mitochondrial carnitine palmitoyltransferase-1 (mCPT-1), and fatty acid translocase (FAT)/CD36 were measured by western blot analysis and RT-qPCR, respectively.ResultsAfter treatment with the β3-AR agonist BRL37344 for 4 weeks, the levels of ALT, AST, TGs, TC, LDL-C and FFAs were decreased in the NAFLD model group compared with the HFD group. Body and liver weights, liver index values and lipid droplet accumulation were lower in the HFD + β3-AGO group than in the HFD group. Decreased NAFLD activity scores (NASs) also showed that liver steatosis and inflammation were ameliorated after treatment with BRL37344. Moreover, the β3-AR antagonist L748337 reversed these effects. Additionally, the protein and gene expression levels of β3-AR, PPAR-α, and mCPT-1 were increased in the HFD + β3-AGO group, whereas those of PPAR-γ and FAT/CD36 were decreased.ConclusionThe β3-AR agonist BRL37344 is beneficial for reducing liver fat accumulation and for ameliorating liver steatosis and inflammation in NAFLD. These effects may be associated with PPARs/mCPT-1 and FAT/CD36.

Theaflavin TF3 Relieves Hepatocyte Lipid Deposition through Activating an AMPK Signaling Pathway by targeting Plasma Kallikrein.

Nonalcoholic fatty liver disease (NAFLD) is rapidly becoming the leading cause of chronic liver diseases throughout the world. The deficit of pharmacotherapy for NAFLD calls for an urgent need for a new drug discovery and lifestyle management. Black tea is the most popular and functional drink consumed worldwide. Its main bioactive constituent theaflavin helps to prevent obesity-a major risk factor for NAFLD. To find new targets for the development of effective and safe therapeutic drugs from natural plants for NAFLD, we found a theaflavin monomer theaflavin-3,3'-digallate (TF3), which significantly reduced lipid droplet accumulation in hepatocytes, and directly bound and inhibited the activation of plasma kallikrein (PK), which was further proved to stimulate adenosine monophosphate activated protein kinase (AMPK) and its downstream targets. Taken together, we proposed that the TF3-PK-AMPK regulatory axis is a novel mechanism of lipid deposition mitigation, and PK could be a new target for NAFLD treatment.

Deletion of the Glutaredoxin-2 Gene Protects Mice from Diet-Induced Weight Gain, Which Correlates with Increased Mitochondrial Respiration and Proton Leaks in Skeletal Muscle.

Aims: The aim of this study was to determine whether deleting the gene encoding glutaredoxin-2 (GRX2) could protect mice from diet-induced weight gain. Results: Subjecting wild-type littermates to a high fat diet (HFD) induced a significant increase in overall body mass, white adipose tissue hypertrophy, lipid droplet accumulation in hepatocytes, and higher circulating insulin and triglyceride levels. In contrast, GRX2 heterozygotes (GRX2+/-) fed an HFD had a body mass, white adipose tissue weight, and hepatic and circulating lipid and insulin levels similar to littermates fed a control diet. Examination of the bioenergetics of muscle mitochondria revealed that this protective effect was associated with an increase in respiration and proton leaks. Muscle mitochondria from GRX2+/- mice had a ∼2- to 3-fold increase in state 3 (phosphorylating) respiration when pyruvate/malate or succinate served as substrates and a ∼4-fold increase when palmitoyl-carnitine was being oxidized. Proton leaks were ∼2- to 3-fold higher in GRX2+/- muscle mitochondria. Treatment of mitochondria with either guanosine diphosphate, genipin, or octanoyl-carnitine revealed that the higher rate of O2 consumption under state 4 conditions was associated with increased leaks through uncoupling protein-3 and adenine nucleotide translocase. GRX2+/- mitochondria also had better protection from oxidative distress. Innovation: For the first time, we demonstrate that deleting the Grx2 gene can protect from diet-induced weight gain and the development of obesity-related disorders. Conclusions: Deleting the Grx2 gene protects mice from diet-induced weight gain. This effect was related to an increase in muscle fuel combustion, mitochondrial respiration, proton leaks, and reactive oxygen species handling. Antioxid. Redox Signal. 31, 1272-1288.

Natural Schistosoma mansoni Infection in the Wild Reservoir Nectomys squamipes Leads to Excessive Lipid Droplet Accumulation in Hepatocytes in the Absence of Liver Functional Impairment.

Schistosomiasis is a neglected tropical disease of a significant public health impact. The water rat Nectomys squamipes is one of the most important non-human hosts in the schistosomiasis mansoni transmission in Brazil, being considered a wild reservoir. Cellular mechanisms that contribute to the physiological adaptation of this rodent to the Schistosoma mansoni parasite are poorly understood. Here we identified, for the first time, that a hepatic steatosis, a condition characterized by excessive lipid accumulation with formation of lipid droplets (LDs) within hepatocytes, occurs in response to the natural S. mansoni infection of N. squamipes, captured in an endemic region. Significant increases of LD area in the hepatic tissue and LD numbers/hepatocyte, detected by quantitative histopathological and ultrastructural analyses, were paralleled by increased serum profile (total cholesterol and triglycerides) in infected compared to uninfected animals. Raman spectroscopy showed high content of polyunsaturated fatty acids (PUFAs) in the liver of both groups. MALDI-TOFF mass spectroscopy revealed an amplified pool of omega-6 PUFA arachidonic acid in the liver of infected animals. Assessment of liver functional activity by the levels of hepatic transaminases (ALT and AST) did not detect any alteration during the natural infection. In summary, this work demonstrates that the natural infection of the wild reservoir N. squamipes with S. mansoni elicits hepatic steatosis in the absence of liver functional harm and that accumulation of lipids, markedly PUFAs, coexists with low occurrence of inflammatory granulomatous processes, suggesting that lipid stores may be acting as a protective mechanism for dealing with the infection.

Lipid droplet breakdown requires Dynamin 2 for vesiculation of autolysosomal tubules in hepatocytes

Lipid droplets (LDs) are lipid storage organelles that in hepatocytes may be catabolized by autophagy for use as an energy source, but the membrane-trafficking machinery regulating such a process is poorly characterized. We hypothesized that the large GTPase dynamin 2 (Dyn2), well known for its involvement in membrane deformation and cellular protein trafficking, could orchestrate autophagy-mediated LD breakdown. Accordingly, depletion or pharmacologic inhibition of Dyn2 led to a substantial accumulation of LDs in hepatocytes. Strikingly, the targeted disruption of Dyn2 induced a dramatic four- to fivefold increase in the size of autolysosomes. Chronic or acute Dyn2 inhibition combined with nutrient deprivation stimulated the excessive tubulation of these autolysosomal compartments. Importantly, Dyn2 associated with these tubules along their length, and the tubules vesiculated and fragmented in the presence of functional Dyn2. These findings provide new evidence for the participation of the autolysosome in LD metabolism and demonstrate a novel role for dynamin in the function and maturation of an autophagic compartment.

Lipocalin-2 (LCN2) in experimental liver injury and human liver diseases

Background: LCN2 is a member of the lipocalin family that is expressed under pernicious conditions such as intoxication, infection, inflammation and cellular stress. Sustained LCN2 expression is induced in experimental liver injury. Immunohistochemistry and primary liver cell isolation identified injured hepatocytes as the main source of LCN2. Methods: LCN2-/- mice were used in experimental models (CCl4, BDL, LPS and ConA) for both acute and chronic liver injury. Liver function analysis, FACS-based liver leukocyte characterization, qRT-PCR for chemokine and cytokine expression and Western Blot as well as histology for fibrotic markers was performed. In addition, serum LCN2 from 91 control and 194 liver diseased patients was analyzed in ELISA. Results: Acute single dose CCl4 intoxication showed lipid droplet accumulation in hepatocytes in LCN2-/- mice. Liver leukocyte isolation showed CD45+ and CD11b+F4–80+ to significantly decrease in LCN2-/- mice, while we found no difference after 5 day BDL or during chronic CCl4 application or BDL for 4 weeks. In ConA models however, LCN2-/- mice showed slightly more liver damage and increased values of CD11b+F4–80+, CD45+ and Tregs than wild type animals. Although serum LCN2 levels in diseased humans were higher than in control patients, we found no difference between cirrhotic and non-cirrhotic patients. Patients with Child A and Child C showed higher LCN2 levels compared to control patients that were independent from disease etiology and serum AST and ALT. By contrast, serum LCN2 showed significant correlation with glomerular filtration rate. Conclusion: LCN2 is a fatty acid transport protein that mobilizes fatty acids from hepatocytes. LCN2-/- mice are unable to appropriately respond to acute CCl4 intoxication. LCN2 expression during inflammatory responses in liver depends on the chosen model. It acts as an acute-phase response protein but the expression during chronic liver insult does not correlate to AST and ALT.

Western-diet enhances expression of death receptors and fatty acid transporters in a mouse model of 70% hepatectomy and leads to increased cell proliferation in the liver

Aims: The non-alcoholic fatty liver disease (NAFLD) is associated with excessive lipid-droplet accumulation in hepatocytes, increased death-receptor expression, altered apoptosis-induction and may progress to liver damage and fibrosis. Active transport of free fatty acids (FFA) is thought to participate in liver cell damage and enhanced apoptosis sensitivity. Death Receptors (DR) apart from apoptosis signalling, also initiate cell proliferation. Thus we investigated, whether high fat diet (HFD) has an impact on apoptosis or regeneration after hepatectomy. Methods: C57Bl/6J-mice received standard (SD) or HFD for 6 weeks, underwent 70% partial hepatectomy (Phx) and were sacrificed on postoperative day (POD) 1, 2 or 7. mRNA expression of FABP-1, FATP-5, CD36/FAT, CD95/Fas, TNFR-1, NOXA and cFLIPL were investigated by qrt-PCR. Proliferation after 70% PHx was visualized by Ki-67 immunhistochemistry. Results: HFD enhanced DR-expression after 70% PHx compared to SD-animals (i.e. CD95/Fas: POD2: x5.53±1.63, p=0.03; TNFR-1: POD1: x27.48±3 .77, p<0.001). Postoperative expression of fatty acid transporters were elevated by HFD (FABP-1: x3.8 5±0.8; p=0.002; CD36/FAT POD1: x3.68±0 .72, p=0.01; POD2: x17.65±3.06, p=0.003; FATP-5 POD2: HFD x3.9±0.77; p=0.013). Ki-67-staining showed increased liver cell proliferation in HFD-mice after PHx compared to SD-mice (POD1: 3% vs. 0.3%, p<0.0001; POD2: 52% vs. 34%; p=0.0002). Conclusion: Data of a mouse model of hepatectomy show increased mRNA expression of FFA-transport proteins, DR’s and elevated cell-proliferation after 70% PHx. FFA may thus pose a risk by increasing cellular susceptibility to apoptosis in general, but could have a beneficial effect in regeneration after hepatectomy.

Increased expression of FABP1 and cFLIPL by free fatty acid treatment is associated with reduced death-receptor mediated apoptosis in human primary hepatocytes

Aims: The non-alcoholic fatty liver disease (NAFLD) is associated with excessive accumulation of lipid-droplets in hepatocytes and causes increased death-receptor expression, altered apoptosis-induction, liver damage and fibrosis. Investigation of fatty acid transport protein expression and lipid accumulation in steatotic hepatocytes may elucidate the correlation of FFA transport mechanisms, death-receptor expression and apoptosis. Methods: Human primary hepatocytes were treated with 0.5mM and 1mM free fatty acids (FFA; oleate: palmitate=2: 1) for 24–96h. Expression of the FFA transport proteins FABP-1, FATP-5, CD36/FAT and Caveolin-1 as well as the death-receptors DR4, DR5, CD95/Fas and TNFR-1 and the apoptosis-related proteins PUMA, NOXA, cFLIPL was investigated. Activation of the stress-responsive kinase ERK was determined by Western-blot. Results: Expression of CD36 (0.5±0.1,p<0.01;means±SEM) and Caveolin-1 (0.5±0.1,p<0.01) was reduced in hepatocytes after treatment with FFA for 96h while FABP-1 (2.2±0.2,p<0.01) expression-levels were elevated. FATP-5-expression was not changed significantly under FFA-treatment. FFA-incubation for 96h slightly decreased the expression of DR4 (0.7±0.1,p=0.014) and DR5 (0.6±0.1,p=0.011). Expression-levels of CD95/Fas (2.7±0.1, p=0.04) and TNFR-1 (2.1±0.1,p<0.01) were increased after incubation with 0.5mM FFA. Furthermore the expression of the pro-apoptotic molecule NOXA (0.4±0.1,p<0.01) was repressed. FFA-treatment did not affect apoptosis-induction (M30) significantly but activated the expression of the anti-apoptotic protein cFLIPL (2.9±0.5,p=0.017). Western-blot analyses showed increased phosphorylation of the stress-responsive kinase ERK after FFA-treatment. Conclusions: FFA-treatment induces FABP-1-expression consequently FFA-uptake seems to be an active mechanism. FFA-incubation activates the expression of CD95/Fas and TNFR-1 but does not activate apoptosis, which may be due to increased cFLIPL expression and ERK activation.

Prevention of orotic-acid-induced fatty liver in male rats by dehydroepiandrosterone and/or phenobarbital.

Dehydroepiandrosterone (DHEA) is a steroid hormone which induces the peroxisome proliferation in rodents. The fatty liver induced by orotic acid and a high sucrose diet in male rats was prevented by the administration of DHEA and/or phenobarbital (PB). A significant increase in the liver weight was induced in the DHEA group (relative weight) and the DHEA + PB group (absolute and relative weight). The liver weight increased more conspicuously in the DHEA + PB group than in the DHEA group. The increase in the liver weight was caused by an increase in the cell size and peroxisome number. In addition, the administration of DHEA alone and the combination of DHEA and PB prevented the lipid droplet accumulation in hepatocytes. The administration of PB alone also prevented the accumulation of lipid droplets without any increase in the liver weight.