CPT I overexpression protects L6E9 muscle cells from fatty acid-induced insulin resistance

Abstract

In this study, we tested the hypothesis that overexpression of carnitine palmitoyltransferase I (CPT I) could protect myotubes from fatty acid-induced insulin resistance by reducing lipid accumulation in the muscle cell. Incubation of L6E9 myotubes with palmitate caused accumulation of triglycerides, diacylgycerol, and ceramide, produced an activation of PKC and PKC, and blocked insulin-stimulated glucose metabolism, reducing insulin-stimulated PKB activity by 60%. Transduction of L6E9 myotubes with adenoviruses encoding for liver CPT I (LCPT I) wild-type (WT), or a mutant form of LCPT I (LCPT I M593S), which is insensitive to malonyl-CoA, produced a twofold increase in palmitate oxidation when LCPT I activity was increased threefold. LCPT I WT and LCPT I M593S-overexpressing L6E9 myotubes showed normal insulin-stimulated glucose metabolism and an improvement in PKB activity when pretreated with palmitate. Moreover, LCPT I WTand LCPT I M593S-transduced L6E9 myotubes were protected against the palmitate-induced accumulation of diacylglycerol and ceramide and PKC and -activation. These results suggest that LCPT I overexpression protects L6E9 myotubes from fatty acid-induced insulin resistance by inhibiting both the accumulation of lipid metabolites and the activation of PKC and PKC.carnitine palmitoyltransferase I; diacylglycerol; ceramides; triacylglycerides; protein kinase C SKELETAL MUSCLE is responsible for 70 -80% of whole body insulin-stimulated glucose uptake (13) and is therefore generally considered the most important site of insulin resistance. Insulin resistance in skeletal muscle plays a major role in the pathogenesis of type 2 diabetes (51), although the mechanism responsible remains unclear. Intramuscular lipid accumulation is evident in a wide set of experimental models, including insulin resistance induced acutely by lipid infusion in both humans (5) and rodents (11), genetic forms of obesity such as Zucker rats (29), high-fat-fed rats (12, 41), and muscle cells in culture incubated with high concentrations of fatty acids (35,44,55). These observations have promoted the so-called lipotoxic model of skeletal muscle insulin resistance, which proposes that high plasma fatty acid levels observed in the obese or insulin-resistant state lead to muscle lipid accumulation, which in turn causes a predisposition toward decreased insulin sensitivity and worsening of the disease (57).Although the association between increased intramyocellular lipid and the development of insulin resistance is compelling, the molecular mechanisms linking free fatty acids (FFAs) to the inhibition of insulin action remain unclear. Recent studies in either humans (6), rodent models of obesity and/or insulin resistance (60), or cultured cells (44,50,55) indicate that fatty acids disrupt one or more of the early steps in insulin signal transduction. The mechanism for this disruption involves the accumulation of fatty acids in lipid-derived metabolites such as long-chain acyl-CoA (LCACoA) species (16), diacylglycerol (DAG) (23, 49), ...

Overexpression of Carnitine Palmitoyltransferase-1 in Skeletal Muscle Is Sufficient to Enhance Fatty Acid Oxidation and Improve High-Fat Diet–Induced Insulin Resistance

“…Having performed hyperinsulinemic-euglycemic clamps with 2-deoxyglucose tracers, we were able to demonstrate that CPT1 overexpression improves insulin action in diet-induced obese models. Our results are entirely consistent with previous work in vitro, where it has been reported that a two-to threefold overexpression of CPT1 in L6 myotubes increased oxidation of the long-chain fatty acids palmitate and oleate and increased insulin stimulation of [ 14 C]glucose incorporation into glycogen by 60% (21,22).…”

“…Carnitine palmitoyltransferase 1 (CPT1) is a mitochondrial transmembrane enzyme thought to be rate limiting for long-chain fatty acid entry into the mitochondria for ␤-oxidation (16,19). Inhibition of CPT1 with the chemical etomoxir increases lipid deposition and exacerbates insu-lin resistance when animals are placed on a high-fat diet (20), whereas overexpression of CPT1 protects myotubes against lipid-induced insulin resistance (21,22), arguing that alterations in fatty acid flux into the mitochondria are critical in regulating lipid effects on insulin sensitivity. Thus, to test whether increasing the capacity for fatty acid flux into the mitochondria is, in itself, sufficient to increase fat oxidation and alter insulin action, we used an approach in which we selectively overexpressed the muscle isoform of CPT1 in skeletal muscle in vivo.…”

Enhanced fatty acid oxidation in adipocytes and macrophages reduces lipid-induced triglyceride accumulation and inflammation

“…Despite their excess fat, obese individuals have reduced visceral WAT CPT1 mRNA and protein levels (20). This prompted our group and others to overexpress CPT1 in liver (26,29,45), muscle (3,33,40), and white adipocytes (9), which led to a decrease in triglyceride (TG) content and an improvement in insulin sensitivity.…”

Myocardial Insulin Resistance: An Overview of Its Causes, Effects, and Potential Therapy

“…Accelerating fatty acid oxidation rates potentially prevents fatty acid, acetyl-CoA and subsequent DAG accumulation and may improve insulin sensitivity. This proposal was recently supported by a study showing that carnitine palmitoyltransferase I (CPT-1) over-expression in L6E9 myotubes increases mitochondrial fatty acid uptake, decreased intracellular DAG concentrations and protects against elevated fatty acid induced insulin resistance [82].…”