Lipid and lipoprotein metabolism in Hep G2 cells.
ABSTRACT Lipid composition, lipid synthesis and lipoprotein secretion by the Hep G2 cell line have been studied with substrate and insulin supplied under different conditions. The lipid composition of Hep G2 cells was close to that of normal human liver, except for a higher content in sphingomyelin (P less than 0.005) and a lower phosphatidylcholine/sphingomyelin ratio. Most of the [14C]triacylglycerols secreted into the medium were recovered by ultracentrifugation at densities of 1.006 to 1.020 g/ml. The main apolipoproteins secreted were apo B-100 and apo A-I. Hep G2 mRNA synthesized in vitro the pro-apolipoproteins A-I and E. Triacylglycerol secretion was 7.38 +/- 1.04 micrograms/mg cell protein per 20 h with 5.5 mM glucose in the medium and increased linearly with glucose concentration. Oleic acid (1 mM) increased the incorporation of [3H]glycerol into the medium and cell triacylglycerols by 251 and 899%, with a concomitant increment in cell triacylglycerols and cholesterol ester. Insulin (1 mU or 7 pmol/ml) inhibited triacylglycerol secretion and [35S]methionine incorporation into secreted protein by 47 and 28%, respectively, with a corresponding increase in the cells. Preincubation of cells with 2.5-10 mM mevalonolactone decreased the incorporation of [14C]acetate into cholesterol 6.2-fold, indicating an inhibitory effect on HMG-CoA reductase. It is concluded that in spite of some differences between Hep G2 and normal human hepatocytes, this line offers an alternative and reliable model for studies on liver lipid metabolism.
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ABSTRACT: Phospholipid transfer protein (PLTP) is a serum glycoprotein with a central role in high-density lipoprotein metabolism. We created a fusion protein in which enhanced green fluorescent protein (EGFP) was fused to the carboxyl-terminus of PLTP. Stably transfected HepG2 cells, which overexpress this fusion protein, were generated. PLTP-EGFP was translocated into the ER and fluoresced within the biosynthetic pathway, showing a marked concentration in the Golgi complex. The transfected cells secreted into the growth medium phospholipid transfer activity 7-fold higher than that of the mock-transfected controls. The medium of the PLTP-EGFP - expressing cells displayed EGFP fluorescence, demonstrating that both the PLTP and the EGFP moieties had attained a biologically active conformation. However, the specific activity of PLTP-EGFP in the medium was markedly reduced as compared with that of endogenous PLTP. This suggests that the EGFP attached to the carboxyl-terminal tail of PLTP interferes with the interaction of PLTP with its substrates or with the lipid transfer process itself. Fluorescently tagged PLTP is a useful tool for elucidating the intracellular functions of PLTP and the interaction of exogenously added PLTP with cells, and will provide a means of monitoring the distribution of exogenously added PLTP between serum lipoprotein subspecies.Biochemistry and Cell Biology 05/2006; 84(2):117-25. · 2.92 Impact Factor
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ABSTRACT: The nuclear receptor farnesoid X receptor (FXR) regulates pathways in lipid, glucose, and energy metabolism. Activation of FXR in mice significantly improved high-fat diet-induced hepatic steatosis. It has been reported that activation of imidazoline I-1 receptor by rilmenidine increases the expression of FXR in human hepatoma cell line, Hep G2 cell, to regulate the target genes relating to lipid metabolism; activation of FXR by rilmenidine exerts an antihyperlipidemic action. However, signals for this action of rilmenidine are still unknown. In the present study, hepatic steatosis induced in mice by high-fat diet was improved by rilmenidine after intraperitoneal injection at 1 mg/kg daily for 12 weeks. Also, mediation of I-1 receptors was identified using the specific antagonist efaroxan. Moreover, rilmenidine decreased the oleic acid-induced lipid accumulation in Hep G2 cells. Otherwise, rilmenidine increased the phosphorylation of p38 to increase the expression of FXR. Deletion of calcium ions by BAPTA-AM reversed the rilmenidine-induced p38 phosphorylation. In conclusion, we suggest that rilmenidine activates I-1 receptor to increase intracellular calcium ions that may enhance the phosphorylation of p38 to higher the expression of FXR for improvement of hepatic steatosis in both animals and cells.Archiv für Experimentelle Pathologie und Pharmakologie 09/2011; 385(1):51-6. · 2.15 Impact Factor
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ABSTRACT: Mulberries are a traditional edible food used to treat hepatic disease. The anti-obesity and hypolipidemic effects of mulberry water extracts (MWE) have attracted increasing interest. In the present study, MWE were assessed for their hepatic lipid-lowering potential when administered in fatty acid overload conditions in HepG2 cells. We found that MWE significantly reduced lipid accumulation, suppressed fatty acid synthesis, and stimulated fatty acid oxidation. Furthermore, MWE also inhibited acetyl coenzyme A carboxylase activities by stimulating adenosine monophosphate-activated protein kinase (AMPK). MWE attenuated the expression of sterol regulatory element-binding protein-1 (SREBP-1) and its target molecules, such as fatty acid synthase. Similar results were also measured in the expressions of enzymes involved in triglyceride and cholesterol biosyntheses including glycerol-3-phosphate acyltransferase, 3-hydroxy-3-methylglutaryl-CoA reductase, and SREBP-2. In contrast, the lipolytic enzyme expression of peroxisome proliferator activated receptor α and carnitine palmitoyltransferase-1 were increased. Our study suggests that the hypolipidemic effects of MWE occur via phosphorylation of AMPK and inhibition of lipid biosynthesis. Therefore, the mulberry extract may be active in the prevention of fatty liver.Journal of the Science of Food and Agriculture 12/2011; 91(15):2740-8. · 1.76 Impact Factor