Insulin resistance in the hereditary hypertriglyceridemic rat is associated with an impairment of delta-6 desaturase expression in liver.

Our previous studies have shown that insulin resistance (IR) in the hereditary hypertriglyceridemic (hHTg) rat is accompanied by a specific fatty acid (FA) profile in insulin target tissues, possibly due to a defect in the desaturation pathway. Increased dietary intake of n-3 polyunsaturated fatty acids (PUFAs) was shown to shape FA composition and to improve insulin sensitivity in this animal strain. Thus, the aim of this study is twofold: (1) to evaluate a defect in the FA desaturation by direct measurement of enzyme activity and gene expression for Delta-6 desaturase (Delta-6 D) in liver of hHTg rats and (2) to investigate the effect of dietary n-3 PUFAs on hepatic Delta-6 D in relation to tissue FA composition. Male Wistar or hHTg rats were fed ad libitum for 21 days either the basal or fish oil (FO)-supplemented diets. Triglyceride (Tg) levels in serum and tissue lipid extracts were measured with the aid of a commercially available enzymatic set. Hepatic activity of the Delta-6 D was determined radiometrically in a microsomal fraction using 1-(14)C-linoleic acid as a substrate. The Delta-6 D mRNA levels were measured using the Northern blot technique. Tissue FA composition was determined by gas chromatography in the total phospholipid fraction after TLC separation. Increased levels of Tg in hHTg rat circulation were accompanied by raised accumulation of Tg in skeletal muscles. FO feeding lowered the concentration of Tg in serum and prevented their accumulation in skeletal muscles of hHTg rats. A pronounced decrease in the hepatic Delta-6 D activity in hHTg rats (by about 80%) was not further diminished by FO feeding. On the other hand, the activity of Delta-6 D in liver of control rats was reduced by about 40% after FO supplementation. These changes were paralleled by a decrease in the Delta-6 D index as calculated from the liver phospholipid FA profile. In particular, an increase in the amount of 18:2 n-6 and a decrease in arachidonic acid and PUFA n-6 metabolites were found. The results indicate that a decrease of insulin action in hHTg rats is accompanied by an impairment of the hepatic Delta-6 D activity already at the gene level, which is not further affected by n-3 PUFA supplementation.

An increase in peroxisomal fatty acid oxidation is not sufficient to prevent tissue lipid accumulation in hHTg rats.

We observed earlier that increased skeletal muscle lipid content in the hereditary hypertriglyceridemic (hHTg) rat is accompanied by a decline in plasma leptin. Leptin has recently been shown to enhance peripheral insulin sensitivity by decreasing the tissue triglyceride accumulation, possibly through regulation of fatty acid oxidation and lipogenesis. Thus, to test the hypothesis that insulin resistance and increased skeletal muscle lipid accumulation in hHTg rats are due to a defect in lipid catabolism, we measured mitochondrial and peroxisomal fatty acid oxidation and malonyl-CoA and acetyl-CoA carboxylase-2 content in skeletal muscles of these animals. In addition, we investigated possible molecular mechanisms responsible for the lower leptin levels in hHTg rats by measuring leptin and leptin-receptor (Ob-Ra) mRNA levels. We found the following: (1) in spite of a higher skeletal muscle malonyl-CoA content and an increased sensitivity of carnitine palmitoyltransferase-1 to malonyl-CoA, carnitine palmitoyltransferase-1 activity in muscle of hHTg rats was normal; (2) increased peroxisomal fatty acid oxidation did not seem to be sufficient to prevent the tissue lipid accumulation in these animals; (3) both lower leptin production by white adipose tissue and increased leptin uptake seem to be responsible for lower circulating leptin levels and therefore lower fatty acid catabolism.

The hypotriglyceridemic effect of dietary n-3 FA is associated with increased beta-oxidation and reduced leptin expression.

To study the mechanisms responsible for the hypotriglyceridemic effect of marine oils, we monitored the effects of high dietary intake of n-3 PUFA on hepatic and muscular beta-oxidation, plasma leptin concentration, leptin receptor gene expression, and in vivo insulin action. Two groups of male Wistar rats were fed either a high-fat diet [28% (w/w) of saturated fat] or a high-fat diet containing 10% n-3 PUFA and 18% saturated fat for 3 wk. The hypotriglyceridemic effect of n-3 PUFA was accompanied by increased hepatic oxidation of palmitoyl-CoA (125%, P < 0.005) and palmitoyl-L-carnitine (480%, P < 0.005). These findings were corroborated by raised carnitine palmitoyltransferase-2 activity (154%, P < 0.001) and mRNA levels (91%, P < 0.01) as well as by simultaneous elevation of hepatic peroxisomal acyl-CoA oxidase activity (144%, P < 0.01) and mRNA content (82%, P < 0.05). In contrast, hepatic carnitine palmitoyltransferase-1 activity remained unchanged despite a twofold increased mRNA level after n-3 PUFA feeding. Skeletal muscle FA oxidation was less affected by dietary n-3 PUFA, and the stimulatory effect was found only in peroxisomes. Dietary intake of n-3 PUFA was followed by increased acyl-CoA oxidase activity (48%, P < 0.05) and mRNA level (83%, P < 0.05) in skeletal muscle. The increased FA oxidation after n-3 PUFA supplementation of the high-fat diet was accompanied by lower plasma leptin concentration (-38%, P < 0.05) and leptin mRNA expression (-66%, P < 0.05) in retroperitoneal adipose tissue, and elevated hepatic mRNA level for the leptin receptor Ob-Ra (140%, P < 0.05). Supplementation of the high-fat diet with n-3 PUFA enhanced in vivo insulin sensitivity, as shown by normalization of the glucose infusion rate during euglycemic hyperinsulinemic clamp. Our results indicate that the hypotriglyceridemic effect of dietary n-3 PUFA is associated with stimulation of FA oxidation in the liver and to a smaller extent in skeletal muscle. This may ameliorate dyslipidemia, tissue lipid accumulation, and insulin action, in spite of decreased plasma leptin level and leptin mRNA in adipose tissue.

Delta-6 desaturase activity and gene expression, tissue fatty acid profile and glucose turnover rate in hereditary hypertriglyceridemic rats.

OBJECTIVE: We have shown previously that the impaired insulin action in hereditary hypertriglyceridemic (hHTg) rat is accompanied by a specific fatty acid (FA) profile in the insulin target tissues, possibly due to a desaturation defect. Thus, the aim of this study was to measure the enzymatic activity and gene expression of delta-6 desaturase in liver of hHTg rats and the tissue FA composition in relation to insulin action. METHODS: Glucose, triglycerides and insulin in plasma were measured using commercially available enzymatic sets. The hepatic delta-6 desaturase activity in hHTg rats was determined radiometrically in a microsomal fraction using the 1-14C-linoleic acid as substrate. delta-6 Desaturase gene expression was measured by the Northern blot technique using a specific cDNA probe. Tissue FA profile was determined by gas chromatography in the total lipid fraction extracted to chloroform. The glucose turnover rate was measured in conscious freely moving animals with the aid of euglycemic hyperinsulinic clamp method. RESULTS: Tissue triglycerides showed a high accumulation in skeletal muscle of hHTg rats. In the liver of these animals, a defect in delta-6 desaturase enzymatic activity was found, while the gene expression for delta-6 desaturase was not changed. Such decreased delta-6 desaturase activity in the liver was linked to a decrease of delta-6 desaturase index as calculated from the liver FA composition. Also the concentration of arachidonic acid (a final metabolite in the biosynthesis of polyunsaturated fatty acids of the n-6 series) was significantly decreased in hHTg rat liver. These changes in FA metabolism were accompanied by a decreased glucose infusion rate (a measure of in vivo insulin action) required to maintain euglycemia at hyperinsulinemia in hHTg rats, and correlated with the hepatic delta-6 desaturase activity. CONCLUSIONS: 1. hHTg rats showed a reduced activity of the delta-6 desaturase in liver without any changes in gene expression for this enzyme; 2. such impairment is accompanied by a lower delta-6 desaturase index (18:2n-6/18:3n-6) found in the liver of these animals and by specific FA profiles in the tissues, particularly regarding the amount of long-chain PUFAs and 18:2n-6 metabolites; and (4) these alterations seem to be related to the impaired insulin action of hHTg rats.