Overexpression of acyl-CoA synthetase-1 increases lipid deposition in hepatic (HepG2) cells and rodent liver in vivo

Abstract

(TG) requires the activation of fatty acids to long-chain acyl-CoAs (LC-CoA) by the enzyme acyl-CoA synthetase (ACSL). There are five known isoforms of ACSL (ACSL1, -3, -4, -5, -6), which vary in their tissue specificity and affinity for fatty acid substrates. To investigate the role of ACSL1 in the regulation of lipid metabolism, we used adenoviral-mediated gene transfer to overexpress ACSL1 in the human hepatoma cell-line HepG2 and in liver of rodents. Infection of HepG2 cells with the adenoviral construct AdACSL1 increased ACSL activity Ͼ10-fold compared with controls after 24 h. HepG2 cells overexpressing ACSL1 had a 40% higher triglyceride (TG) content (93 Ϯ 3 vs. 67 Ϯ 2 nmol/mg protein in controls, P Ͻ 0.05) after 24-h exposure to 1 mM oleate. Furthermore, ACSL1 overexpression produced a 60% increase in cellular LCA-CoA content (160 Ϯ 6 vs. 100 Ϯ 6 nmol/g protein in controls, P Ͻ 0.05) and increased [ 14 C]oleate incorporation into TG without significantly altering fatty acid oxidation. In mice, AdACSL1 administration increased ACSL1 mRNA and protein more than fivefold over controls at 4 days postinfection. ACSL1 overexpression caused a twofold increase in TG content in mouse liver (39 Ϯ 4 vs. 20 Ϯ 2 mol/g wet wt in controls, P Ͻ 0.05), and overexpression in rat liver increased [1-14 C]palmitate clearance into liver TG. These in vitro and in vivo results suggest a pivotal role for ACSL1 in regulating TG synthesis in liver.acyl-coenzyme A synthetase; hepatic triglyceride synthesis; adenovirus; fatty acid metabolism OBESITY AND ITS ASSOCIATED COMPLICATIONS of insulin resistance, cardiovascular disease, hypertension, hepatic steatosis, and type 2 diabetes have become a major health problem worldwide (50). All of these metabolic disorders exhibit altered regulation of lipid metabolism, which results in the intracellular accumulation of lipid in nonadipose tissues, but the intracellular mechanisms that control partitioning of fatty acids into different metabolic pathways are still being elucidated.Fatty acids can be metabolized in the pathways of ␤-oxidation, triglyceride synthesis, phospholipid synthesis, cholesterol ester synthesis, fatty acid elongation, and protein acylation and can also serve as signaling molecules (7,10,40). It is believed that the partitioning of intracellular fatty acids between storage pathways and ␤-oxidation is controlled by regulation of the mitochondrial acyl-CoA transporter carnitine palmitoyltransferase-1 (32), by regulation of metabolic gene expression (39), and by hormones including insulin. A number of recent studies suggest that partitioning might also involve the different isoforms of acyl-CoA synthetase (ACSL) (7,20,49).ACSL catalyzes the ATP-dependent acylation of fatty acids into long-chain acyl CoAs (LCA-CoAs) and is the first step in lipid metabolism after fatty acid entry into the cell. The LCA-CoAs can then enter the ␤-oxidation pathway for energy production or undergo further esterification for production of phospholipids, cholesterol esters, and triglycerides. Th...

Long-Chain Acyl-Coa Synthetases And Fatty Acid Channeling

“…However in this study, the lack of carnitine in the medium may have limited CPT-1 activity and FA β-oxidation, while the marked increase in ACS activity may have overwhelmed other pathways. Adenoviral-mediated overexpression of ACSL1 in rodent liver in vivo also increased hepatic TAG, but did not affect FA clearance from the blood; cholesterol and phospholipid metabolism were not measured [40]. Taken as a whole, these overexpression studies suggest that, not only do ACSL1 and ACSL5 channel FA towards different lipid pathways, but that the direction of channeling may vary in different cell types.…”

“…Unlike ACSL5, overexpressed ACSL1 increases the incorporation of de novo synthesized FA into glycerolipids [14]. In somewhat contrasting experiments in HepG2 cells, adenovirus-mediated overexpression of ACSL1 resulted in a 20-fold increase in ACS activity, increases in cell acyl-CoA content, and enhanced oleic acid partitioning into cellular TAG [40]. However in this study, the lack of carnitine in the medium may have limited CPT-1 activity and FA β-oxidation, while the marked increase in ACS activity may have overwhelmed other pathways.…”

Reduction of serum free fatty acids and triglycerides by liver-targeted expression of long chain acyl-CoA synthetase 3

“…Furthermore, overexpression of AcSL1 in rat primary hepatocytes increased FA reacylation and channeled FA toward diacylglyceride and phospholipid synthesis and away from cholesterol ester synthesis (15). consistent with in vitro findings, ACSL1 overexpression via adenovirus infection elevated TG in mouse liver and increased palmitate clearance into liver TG in rats (14). Moreover, transgenic mice with heart-specific expression of AcSL1 exhibited impaired cardiac lipid homeostasis with marked accumulation of triglyceride and phospholipids in the heart (16).…”

“…In recent years, the functional involvement of some AcSLs, like AcSL1 (14,15,17) and AcSL5 (13), as regulators for fatty acid partitioning has emerged. However, there is still lack of studies directly addressing the role of the other AcSL isoforms in lipid metabolism in vivo.…”

“…Relative to AcSL4 and 5 (4,13), more studies have been carried out to characterize the function of AcSL1 in the liver using liverderived cell lines, primary hepatocytes and rodents. In vitro, adenoviral-mediated overexpression of rat AcSL1 in HepG2 cells increased triglyceride (TG) content without significantly altering fatty acid oxidation (14). Furthermore, overexpression of AcSL1 in rat primary hepatocytes increased FA reacylation and channeled FA toward diacylglyceride and phospholipid synthesis and away from cholesterol ester synthesis (15).…”