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Originally published In Press as doi:10.1194/jlr.M400505-JLR200 on February 16, 2005

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Journal of Lipid Research, Vol. 46, 988-993, May 2005
Copyright © 2005 by American Society for Biochemistry and Molecular Biology

Acute inhibition of hepatic ß-oxidation in APOE*3Leiden mice does not affect hepatic VLDL secretion or insulin sensitivity

Ilse Duivenvoorden*,{dagger}, Bas Teusink1,*, Patrick C. N. Rensen*,{dagger}, Folkert Kuipers§, Johannes A. Romijn**, Louis M. Havekes*,{dagger},{dagger}{dagger} and Peter J. Voshol2,*,**

* Netherlands Organization for Applied Scientific Research-Quality of Life, Gaubius Laboratory, Leiden, The Netherlands
{dagger} Department of General Internal Medicine, Leiden University Medical Center, Leiden, The Netherlands
** Department of Endocrinology and Metabolic Diseases, Leiden University Medical Center, Leiden, The Netherlands
{dagger}{dagger} Department of Cardiology, Leiden University Medical Center, Leiden, The Netherlands
§ Center for Liver, Digestive, and Metabolic Diseases, Laboratory of Pediatrics, University Hospital Groningen, Groningen, The Netherlands

Published, JLR Papers in Press, February 16, 2005. DOI 10.1194/jlr.M400505-JLR200

1 Present address of B. Teusink: NIZO Food Research, Ede, The Netherlands. Back

2 To whom correspondence should be addressed. e-mail: p.j.voshol{at}lumc.nl


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Hepatic VLDL and glucose production is enhanced in type 2 diabetes and associated with hepatic steatosis. Whether the derangements in hepatic metabolism are attributable to steatosis or to the increased availability of FA metabolites is not known. We used methyl palmoxirate (MP), an inhibitor of carnitine palmitoyl transferase I, to acutely inhibit hepatic FA oxidation and investigated whether the FAs were rerouted into VLDL secretion and whether this would affect hepatic glucose production. After an overnight fast, male APOE3*Leiden transgenic mice received an oral dose of 10 mg/kg MP. Administration of MP led to an 83% reduction in plasma ß-hydroxybutyrate (ketone body) levels compared with vehicle-treated mice (0.47 ± 0.07 vs. 2.81 ± 0.16 mmol/l, respectively; P < 0.01), indicative of impaired ketogenesis. Plasma FFA levels were increased by 32% and cholesterol and insulin levels were decreased by 17% and 50%, respectively, in MP-treated mice compared with controls. MP treatment led to a 30% increase in liver triglyceride (TG) content. Surprisingly, no effect on hepatic VLDL-TG production was observed between the groups at 8 h after MP administration. In addition, the capacity of insulin to suppress endogenous glucose production was unaffected in MP-treated mice compared with controls.

In conclusion, acute inhibition of FA oxidation increases hepatic lipid content but does not stimulate hepatic VLDL secretion or reduce insulin sensitivity.

Abbreviations: acc1, acetyl-coenzyme A carboxylase 1; apoB, apolipoprotein B; CPT1, carnitine palmitoyl transferase 1; dgat, acyl:diacylglycerol transferase; fas, fatty acid synthase; ß-HB, ß-hydroxybutyrate; hmgs, 3-hydroxy-3-methylglutaryl-coenzyme A synthase; mcad, medium-chain acyl coenzyme A; MP, methyl palmoxirate; MTP, microsomal triglyceride transfer protein; mttp, microsomal triglyceride transfer protein; ppar{alpha}, peroxisome proliferator-activated receptor {alpha}; srebp1c, sterol regulatory element binding protein 1c; TG, triglyceride

Supplementary key words triglycerides • fatty acids • steatosis • glucose metabolism • very low density lipoprotein


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Hepatic VLDL-triglyceride (TG) secretion is an important determinant of plasma lipid levels. The rate of VLDL secretion is generally believed to be substrate-driven (i.e., the hepatic content of TG and FA determines VLDL assembly and the rate of VLDL secretion) (1–3). Indeed, addition of FA to hepatocytes in vitro leads to increased hepatic TG and eventually to enhanced VLDL secretion (4). However, the hypothesis that VLDL secretion is substrate-driven in vivo has not been proven.

Hepatic TG content is the result of uptake and synthesis on the one hand and ß-oxidation and VLDL-TG secretion on the other. The importance of ß-oxidation is illustrated by the severe phenotype of humans and mice with impaired ß-oxidation. Deficiency in hepatic ß-oxidation enzymes results in severe plasma hypoketosis and fatty liver (5, 6). In ß-oxidation, the rate-limiting enzyme is carnitine palmitoyl transferase 1 (CPT1), which couples long-chain FAs to carnitine for transportation into the mitochondria (7, 8). Methyl palmoxirate (MP) is a specific and irreversible inhibitor of CPT1 and thereby inhibits ß-oxidation (9, 10).

In this study, we tested the effect of acute inhibition of ß-oxidation in vivo by MP on hepatic VLDL production in fasted hyperlipidemic APOE3*Leiden transgenic mice. APOE3*Leiden mice are characterized by a human-like lipoprotein profile (11, 12). We and others have shown that these mice provide a suitable model in which to study hepatic VLDL metabolism, because they have decreased VLDL-TG production and fatty liver comparable to human conditions (12–14). We observed that acute inhibition of fatty acid ß-oxidation in the liver leads to increased hepatic TG content but neither increases hepatic VLDL secretion nor induces hepatic insulin resistance.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Animals and diet
Male 3 to 4 month old APOE3*Leiden transgenic mice were fed a Western-type diet containing 15% cacao butter and 0.25% cholesterol (Hope Farms, Woerden, The Netherlands) for 8 weeks before the experiments. The animals were allowed free access to food and water and were kept on a normal diurnal rhythm under standard conditions. After an overnight fast, the animals were matched for body weight and received a dose of 10 mg/kg MP (McN-3716: methyl 2-tetradecyloxiranecarboxylate; kindly provided by Dr. Hegardt, Barcelona, Spain) by gavage in 0.05% methyl cellulose solution or methyl cellulose solution alone (vehicle) as described previously (9). Blood samples were taken by tail-tip bleeding every 2 h after MP or vehicle administration. After 8 h, the animals were killed and liver samples were taken and snap-frozen in liquid nitrogen for lipid content and mRNA expression analysis. Parallel groups of mice were used either to study VLDL-TG production or to perform hyperinsulinemic clamp analyses (see below). The animal care committee of Netherlands Oganization for Applied Scientific Research (TNO) approved all experiments.

Plasma parameters
Blood samples were taken from the tail vein into chilled paraoxon-coated capillaries to prevent ex vivo lipolysis (15). Plasma was collected via centrifugation and plasma cholesterol (Roche Diagnostics GmbH, Mannheim, Germany), TG (without free glycerol; Triglyceride GPO-Trinder, Sigma Diagnostics, Inc., St. Louis, MO), glucose (glucose trinder 500, Sigma Diagnostics), ß-hydroxybutyrate (ß-HB) (Sigma Diagnostics), and FFA levels (NEFA-C, Wako Chemicals GmbH, Neuss, Germany) were determined using standard commercial kits, according to the manufacturer's instructions. Plasma insulin was measured by a radioimmunoassay using rat insulin standards, which show 100% cross-reaction with mouse insulin (Sensitive Rat Insulin RIA Kit; Linco Research, Inc., St. Charles, MO).

Liver lipid levels
Liver samples taken from mice 8 h after administration of MP or vehicle were homogenized in phosphate-buffered saline (10% wet weight per volume), and samples were taken to measure protein content by the Lowry assay (16). Lipid content was determined by extraction of lipids using the Bligh and Dyer method (17), followed by lipid separation using high performance thin-layer chromatography on silica gel plates as described previously (18) and subsequent analysis by TINA2.09 software (19) (Raytest Isotopen Meßgeräte GmbH, Straubenhardt, Germany).

Hepatic VLDL-TG production
Hepatic VLDL-TG production, de novo apolipoprotein B (apoB) secretion, and VLDL composition were determined in overnight-fasted APOE3*Leiden mice given a gavage of MP (10 mg/kg) or vehicle. After 6 h of MP or vehicle administration, the animals were anesthetized [0.5 ml/kg Hypnorm (Janssen Pharmaceutica, Beerse, Belgium) and 12.5 mg/g midazolam (Roche, Mijdrecht, The Netherlands)] and injected intravenously with 0.1 ml of phosphate-buffered saline containing 100 µCi of Tran35S-labelTM (ICN Biomedicals, Inc., Irvine, CA) to measure de novo total apoB synthesis. After 30 min, the animals received a 15% (by volume) Triton WR1339 injection (500 mg/kg body weight; Tyloxapol; Sigma Chemicals, Steinheim, Germany) to prevent systemic lipolysis of newly secreted hepatic VLDL-TG (20). Blood samples were drawn at 0, 15, 30, 60, and 90 min after Triton WR1339 injection, and TG concentrations were determined in the plasma as described above. At 90 min, the animals were killed and blood was collected by cardiac puncture for isolation of VLDL.

VLDL composition analyses
VLDL particles (d < 1.019) were separated from other lipoproteins in plasma by density gradient ultracentrifugation as described (21). Protein content of the VLDL fraction was determined by the Lowry assay (16). TG and total cholesterol were measured as in plasma (see above). Phospholipids and free cholesterol were determined using standard commercial kits according to the manufacturer's instructions (Wako Chemicals GmbH). 35S-labeled total apoB content was measured in the VLDL fraction after precipitation with isopropanol as described previously (22, 23).

Hyperinsulinemic clamp analysis
Insulin sensitivity was determined in a group of APOE3* Leiden mice, fed a Western-type diet, that were fasted overnight, body weight-matched, and given either MP or vehicle. Basal and insulin-mediated suppression of endogenous (hepatic) glucose production was studied by hyperinsulinemic euglycemic clamp analysis using [3H]D-glucose as a tracer. The clamp analysis and calculations were performed as described previously (24, 25).

Hepatic mRNA expression
Livers were immediately removed from the mice and snap-frozen in liquid nitrogen. Total RNA was isolated as described by Chomczynski and Sacchi (26) by use of RNA-BeeTM (Campro Scientific, Berlin, Germany). cDNA synthesis was done according to Bloks et al. (27). Real-time quantitative PCR (28) was performed using an Applied Biosystems 7700 Sequence Detector according to the manufacturer's instructions. Primers were obtained from Invitrogen (Paisley, UK), and fluorogenic probes, labeled with 6-carboxyfluorescein and 6-carboxytetramethylrhodamine, were supplied by Eurogentec (Seraing, Belgium). Primers and probes used in this experiment were described previously (29–31). All expression data were subsequently standardized for cyclophilin RNA, which was analyzed in a separate run.

Statistical analysis
The Mann-Whitney U test was used to determine differences between MP- and vehicle-treated mice. The level of significance was set at P < 0.05. All data are presented as means ± SD. Analyses were performed using SPSS11.0 (SPSS, Inc., Chicago, IL).


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
MP decreases ß-HB and increases FFA levels in plasma
Male APOE3*Leiden mice, fed a Western-type diet for 8 weeks, were fasted overnight and treated with 10 mg/kg MP or vehicle. Plasma ß-HB (one of the ketone bodies produced by the liver) was decreased by 83% at 8 h after MP administration, confirming the inhibition of ß-oxidation. A strong decrease in plasma ß-HB was already apparent after 2 h, and levels remained constant up to at least 8 h, whereas the solvent had no effect (Fig. 1) . Table 1 summarizes plasma parameters measured at 8 h after MP or vehicle administration. Plasma FFA levels were increased by 32% upon MP treatment. Glucose and TG did not show significant differences, whereas plasma insulin and cholesterol levels were both decreased significantly, by 50% and 17%, respectively, in the MP-treated group versus controls.



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  		Fig. 1. Time course effect of methyl palmoxirate (MP) on ß-hydroxybutyrate (ß-HB) levels. Plasma samples from overnight-fasted APOE*3 Leiden mice, treated with MP (closed circles) or vehicle (open circles), were taken every 2 h to determine plasma ß-HB levels. Values represent means ± SD of five mice per group. ** P < 0.01.

 

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  		TABLE 1. Plasma parameters determined in overnight-fasted APOE*3Leiden mice at 8 h after MP administration (10 mg/kg body weight)

 
Inhibition of ß-oxidation increases lipid storage in liver without affecting VLDL production
Liver lipid analysis showed that MP-treated mice had 30% increased TG content in liver compared with vehicle-treated animals (Fig. 2A) . To analyze whether the increased hepatic TG content was associated with altered hepatic VLDL-TG production, mice were injected with Triton WR1339 at 6 h after MP or vehicle gavage, and the accumulation of endogenous VLDL-TG in plasma was determined over time. Plasma ß-HB levels in this subset of mice were decreased similarly, as shown in Fig. 1. As is evident from Fig. 2B, the VLDL-TG production rate, as determined from the slope of the curve, was unchanged in MP-treated animals compared with controls (2.12 ± 0.58 vs. 2.51 ± 0.53 mmol/l TG/h, respectively). Furthermore, the composition of the VLDL particles (Table 2) as well as the rate of de novo total apoB production of newly synthesized VLDL particles did not differ between MP- and vehicle-treated mice (35 ± 3 vs. 36 ± 4 x 104 dpm/ml plasma/mg protein, respectively).



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  		Fig. 2. Effect of MP on hepatic lipid levels and VLDL production. A: Liver samples were taken 8 h after MP or vehicle treatment. Lipids were extracted from liver homogenates and subsequently separated on thin-layer silica gel plates as described. Liver free cholesterol, cholesteryl ester, and triglyceride levels are depicted for MP-treated (closed bars) and vehicle-treated (open bars) mice. B: The increase in plasma triglyceride (TG) levels in MP-treated (closed circles) and vehicle-treated (open circles) mice was measured in time after Triton WR1339 injection. Values represent means ± SD for five mice per group. * P < 0.05.

 

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  		TABLE 2. Composition of VLDL obtained from Triton WR1339- injected mice at 8 h after treatment with MP or vehicle

 
Acute inhibition of ß-oxidation does not lead to hepatic insulin resistance
Increased hepatic TG content is negatively associated with insulin sensitivity (32). Because plasma insulin levels decreased in the MP-treated mice, we analyzed the insulin-mediated suppression of endogenous (hepatic) glucose production during a hyperinsulinemic clamp procedure. Although hepatic TG content was increased in the MP-treated mice, endogenous (hepatic) glucose production was equally suppressed by insulin in both groups. The glucose production decreased from 32.5 ± 6.5 to 19.1 ± 10.1 µmol/min/kg body weight (–40%) in MP-treated mice and from 28.7 ± 7.1 to 14.7 ± 8.8 µmol/min/kg body weight (–46%) in vehicle-treated mice. These results show that the livers of MP-treated mice displayed normal hepatic insulin sensitivity.

Hepatic mRNA levels
Hepatic expression of several genes was studied in mice at 8 h after treatment with MP or vehicle using RT-PCR. The expression of genes involved in ß-oxidation [peroxisome proliferator-activated receptor {alpha} (ppar{alpha}), cpt1a, medium-chain acyl-CoA (mcad), and HMG-CoA synthase (hmgs)] were all 40–56% higher in MP-treated livers compared with controls, albeit statistical significance was achieved for only ppar{alpha} and cpt1a (Table 3). Regarding genes involved in VLDL production, microsomal TG transfer protein (mttp) expression was increased in MP-treated mice, whereas no differences were observed in acyl:diacylglycerol transferase 1 (dgat1), dgat2, apob, and apoe expression. Expression of sterol regulatory element binding protein 1c (srebp1c), which is involved in the activation of genes involved in the uptake and synthesis of FA, TG, and cholesterol, was strongly decreased in MP-treated mice. In FA synthesis, gene expression levels of fatty acid synthase (fas) and acetyl-CoA carboxylase 1 (acc1) were not changed between the two groups. Also, no changes were observed in gene expression levels of pyruvate kinase (pk) and phosphoenolpyruvate carboxykinase (pepck), which are involved in hepatic glucose metabolism.


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  		TABLE 3. Hepatic mRNA expression levels in control and MP-treated mice as determined by RT-PCR and related to cyclophilin

 

   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
Hepatic FA and TG metabolism encompasses a number of interacting and dynamic processes, including uptake of plasma FFA and TG from lipoprotein remnants, storage in the form of TG, ß-oxidation, and VLDL-TG formation/secretion. The rate of VLDL-TG secretion has long been thought to be substrate-driven (1–3). We questioned whether an acute inhibition of hepatic ß-oxidation would lead to a redirection of FA toward hepatic TG synthesis specifically directed to VLDL secretion. MP was used to irreversibly inhibit the crucial enzyme in ß-oxidation, CPT1A (9, 10). Indeed, within 2 h after oral dosing of MP, plasma ketone bodies (i.e., ß-HB) decreased and remained low for up to 8 h after gavage (Fig. 1). Because plasma ketone bodies are derived solely from hepatic ß-oxidation, we concluded that hepatic ß-oxidation of long-chain FAs was almost completely inhibited by the applied dose of MP. It is known that at low doses, MP inhibits mostly CPT1A (present in liver and heart), whereas higher doses also inhibit CPT1B (present in skeletal muscle and heart) in rat (7, 9).

Several studies have demonstrated that oxirane carboxylates (such as etomoxir and MP) are effective at decreasing both ketone body and glucose levels in rodents, dogs, and humans (9, 33–36). However, in our overnight-fasted mice, plasma glucose levels were similar between MP-treated and control mice, suggesting that muscle ß-oxidation was not completely inhibited, in contrast to liver ß-oxidation, as judged from the extremely low ß-HB levels. The residual level of ß-HB in MP-treated animals may be derived from short-chain FA oxidation that does not depend on CPT1A for transport into mitochondria.

We observed that the hepatic expression of genes involved in ß-oxidation (i.e., ppar{alpha}, cpt1a, mcad, and hmgs) was higher in MP-treated mice (Table 3), albeit statistically significant differences were observed only for ppar{alpha} and cpt1a, as a result of the moderate sample sizes in combination with relatively large standard deviations in the control group. This increase of genes involved in ß-oxidation may be an attempt by the liver to compensate for the strongly decreased hepatic ß-oxidation.

Plasma FFA levels increased significantly at 8 h after MP treatment (Table 2). This increase was most likely caused by the decreased hepatic FA oxidation. Interestingly, we observed that acute inhibition of ß-oxidation and/or the decrease of plasma ketone bodies was associated with strongly decreased plasma insulin levels (Table 2). In agreement with this, Boden and Chen (37) showed that in humans there is a positive correlation between plasma ß-HB concentrations and insulin secretion capacity.

As expected, inhibition of hepatic ß-oxidation led to a significant accumulation of TG within the liver (Fig. 2A). Hepatic TG accumulation is in agreement with the symptoms seen in patients and animal models with ß-oxidation disorders (5, 6). Microscopic analysis of liver slices revealed a mixed micro/macrovesicular accumulation of lipid droplets, also called micro/macrovesicular steatosis (data not shown). This observation of microvesicular steatosis seems to be in agreement with the recent finding with cyclopropane carboxylic acid (CPCA), another ß-oxidation inhibitor, in rats (38). Although one might expect to observe increased liver mRNA levels of TG-synthesizing enzymes such as DGAT1, this was not observed in our experimental setting.

Interestingly, the increased TG accumulation in the liver was not associated with increased hepatic VLDL-TG production and/or changes in VLDL composition (Fig. 2B, Table 2). We did observe an increase in gene mRNA expression of mttp in the livers of MP-treated mice. We did not analyze microsomal triglyceride transfer protein (MTP) activity, so we can only conclude that this increased mttp expression did not (yet) lead to increased MTP protein levels or activity. We cannot exclude the possibility that chronic, long-term inhibition of hepatic ß-oxidation might induce hepatic VLDL-TG production.

Hepatic srebp1c expression was strongly decreased in MP-treated mice. Because insulin increases hepatic srebp1c expression (39), it is possible that the low plasma insulin levels caused this downregulation of liver srebp1c expression. However, the low srebp1c expression did not seem to affect RNA expression levels of enzymes involved in de novo lipogenesis, such as fas or acc1, at least at the examined time point.

Because TG accumulation is known to be associated with insulin resistance in the liver (32), we performed hyperinsulinemic clamp procedures to assess the insulin-mediated suppression of endogenous glucose production. Under normal conditions, the major part of endogenous glucose production is derived from liver (37). Our results clearly show that there was no significant difference in insulin-mediated suppression of endogenous (hepatic) glucose production between treated and untreated mice. Gene expression of enzymes involved in hepatic glucose production also did not show any differences between the two groups. Whole body insulin-mediated glucose uptake was not significantly different between MP-treated and control mice (data not shown). This confirms, in part, that muscle FA ß-oxidation must still be functional at the applied low dose of MP. It has been shown by Dobbins et al. (40) that chronic suppression of hepatic ß-oxidation leads to insulin resistance in both liver and muscle in rats. We clearly show that acute inhibition of FA ß-oxidation does not affect either hepatic or muscle insulin sensitivity in vivo in mice.

We conclude that acute inhibition of hepatic FA ß-oxidation leads to hepatic micro/macrovesicular steatosis but does not affect either hepatic VLDL secretion or hepatic insulin sensitivity.


   ACKNOWLEDGMENTS
 
The authors are grateful to Fjodor van der Sluijs and Anita van Nieuwkoop for excellent technical assistance. This study was conducted in the framework of the Leiden Center for Cardiovascular Research, Leiden University Medical Centre-Netherlands Organization for Applied Scientific Research and supported by the Netherlands Organization for Scientific Research (NWO Grant 903-39-179 and NWO VIDI Grant 917.36.351 to P.C.N.R.; NWO VENI Grant 916.36.071 to P.J.V.; and Program Grant 903-39-291 to L.M.H.) and the Leiden University Medical Centre (Gisela Thier Fellowship to P.C.N.R.).

Manuscript received December 20, 2004 and in revised form January 31, 2005.


   REFERENCES
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
  1. Lewis, G. F. 1997. Fatty acid regulation of very low density lipoprotein production. Curr. Opin. Lipidol. 8: 146–153.[Medline]

  2. Randle, P. J., P. B. Garland, C. N. Hales, and E. A. Newsholme. 1963. The glucose fatty-acid cycle. Its role in insulin sensitivity and the metabolic disturbances of diabetes mellitus. Lancet. 1: 785–789.[Medline]

  3. Shelness, G. S., and J. A. Sellers. 2001. Very-low-density lipoprotein assembly and secretion. Curr. Opin. Lipidol. 12: 151–157.[CrossRef][Medline]

  4. Kuipers, F., M. C. Jong, Y. Lin, M. Eck, R. Havinga, V. Bloks, H. J. Verkade, M. H. Hofker, H. Moshage, T. J. Berkel, et al. 1997. Impaired secretion of very low density lipoprotein-triglycerides by apolipoprotein E-deficient mouse hepatocytes. J. Clin. Invest. 100: 2915–2922.[Medline]

  5. Roe, C. R., and P. M. Coates. 1997. Mitochondrial fatty acid oxidation disorders. In The Metabolic and Molecular Bases of Inherited Disease. C. R. Scriver, A. L. Beaudet, W. S. Sly, and D. Valle, editors. McGraw-Hill, New York. 1501–1533.

  6. Schuler, A. M., and P. A. Wood. 2002. Mouse models for disorders of mitochondrial fatty acid beta-oxidation. ILAR J. 43: 57–65.[Medline]

  7. Eaton, S. 2002. Control of mitochondrial beta-oxidation flux. Prog. Lipid Res. 41: 197–239.[CrossRef][Medline]

  8. McGarry, J. D., and N. F. Brown. 1997. The mitochondrial carnitine palmitoyltransferase system. From concept to molecular analysis. Eur. J. Biochem. 244: 1–14.[Medline]

  9. Friedman, M. I., R. B. Harris, H. Ji, I. Ramirez, and M. G. Tordoff. 1999. Fatty acid oxidation affects food intake by altering hepatic energy status. Am. J. Physiol. 276: R1046–R1053.

  10. Kiorpes, T. C., D. Hoerr, W. Ho, L. E. Weaner, M. G. Inman, and G. F. Tutwiler. 1984. Identification of 2-tetradecylglycidyl coenzyme A as the active form of methyl 2-tetradecylglycidate (methyl palmoxirate) and its characterization as an irreversible, active site-directed inhibitor of carnitine palmitoyltransferase A in isolated rat liver mitochondria. J. Biol. Chem. 259: 9750–9755.[Abstract/Free Full Text]

  11. van Vlijmen, B. J., A. M. van den Maagdenberg, M. J. Gijbels, H. van der Boom, H. HogenEsch, R. R. Frants, M. H. Hofker, and L. M. Havekes. 1994. Diet-induced hyperlipoproteinemia and atherosclerosis in apolipoprotein E3-Leiden transgenic mice. J. Clin. Invest. 93: 1403–1410.

  12. van Vlijmen, B. J., H. B. van't Hof, M. J. Mol, H. van der Boom, A. van der Zee, R. R. Frants, M. H. Hofker, and L. M. Havekes. 1996. Modulation of very low density lipoprotein production and clearance contributes to age- and gender-dependent hyperlipoproteinemia in apolipoprotein E3-Leiden transgenic mice. J. Clin. Invest. 97: 1184–1192.[Medline]

  13. Mensenkamp, A. R., M. J. van Luyn, H. van Goor, V. Bloks, F. Apostel, J. Greeve, M. H. Hofker, M. C. Jong, B. J. van Vlijmen, L. M. Havekes, et al. 2000. Hepatic lipid accumulation, altered very low density lipoprotein formation and apolipoprotein E deposition in apolipoprotein E3-Leiden transgenic mice. J. Hepatol. 33: 189–198.[CrossRef][Medline]

  14. Mensenkamp, A. R., B. Teusink, J. F. Baller, H. Wolters, R. Havinga, K. W. van Dijk, L. M. Havekes, and F. Kuipers. 2001. Mice expressing only the mutant APOE3Leiden gene show impaired VLDL secretion. Arterioscler. Thromb. Vasc. Biol. 21: 1366–1372.[Abstract/Free Full Text]

  15. Zambon, A., S. I. Hashimoto, and J. D. Brunzell. 1993. Analysis of techniques to obtain plasma for measurement of levels of free fatty acids. J. Lipid Res. 34: 1021–1028.[Abstract]

  16. Lowry, O. H., N. J. Rosebrough, A. L. Farr, and R. J. Randall. 1951. Protein measurement with the Folin phenol reagent. J. Biol. Chem. 193: 265–275.[Free Full Text]

  17. Bligh, E. G., and W. J. Dyer. 1959. A rapid method of total lipid extraction and purification. Can. J. Med. Sci. 37: 911–917.

  18. Havekes, L. M., E. C. de Wit, and H. M. Princen. 1987. Cellular free cholesterol in Hep G2 cells is only partially available for down-regulation of low-density-lipoprotein receptor activity. Biochem. J. 247: 739–746.[Medline]

  19. Post, S. M., E. C. de Wit, and H. M. Princen. 1997. Cafestol, the cholesterol-raising factor in boiled coffee, suppresses bile acid synthesis by downregulation of cholesterol 7 alpha-hydroxylase and sterol 27-hydroxylase in rat hepatocytes. Arterioscler. Thromb. Vasc. Biol. 17: 3064–3070.[Abstract/Free Full Text]

  20. Aalto-Setala, K., E. A. Fisher, X. Chen, T. Chajek-Shaul, T. Hayek, R. Zechner, A. Walsh, R. Ramakrishnan, H. N. Ginsberg, and J. L. Breslow. 1992. Mechanism of hypertriglyceridemia in human apolipoprotein (apo) CIII transgenic mice. Diminished very low density lipoprotein fractional catabolic rate associated with increased apo CIII and reduced apo E on the particles. J. Clin. Invest. 90: 1889–1900.

  21. Jong, M. C., V. E. Dahlmans, P. J. van Gorp, M. L. Breuer, M. J. Mol, A. van der Zee, R. R. Frants, M. H. Hofker, and L. M. Havekes. 1996. Both lipolysis and hepatic uptake of VLDL are impaired in transgenic mice coexpressing human apolipoprotein E*3Leiden and human apolipoprotein C1. Arterioscler. Thromb. Vasc. Biol. 16: 934–940.[Abstract/Free Full Text]

  22. Li, X., F. Catalina, S. M. Grundy, and S. Patel. 1996. Method to measure apolipoprotein B-48 and B-100 secretion rates in an individual mouse: evidence for a very rapid turnover of VLDL and preferential removal of B-48- relative to B-100-containing lipoproteins. J. Lipid Res. 37: 210–220.[Abstract]

  23. Pietzsch, J., S. Subat, S. Nitzsche, W. Leonhardt, K. U. Schentke, and M. Hanefeld. 1995. Very fast ultracentrifugation of serum lipoproteins: influence on lipoprotein separation and composition. Biochim. Biophys. Acta. 1254: 77–88.[Medline]

  24. Koopmans, S. J., M. C. Jong, I. Que, V. E. Dahlmans, H. Pijl, J. K. Radder, M. Frolich, and L. M. Havekes. 2001. Hyperlipidaemia is associated with increased insulin-mediated glucose metabolism, reduced fatty acid metabolism and normal blood pressure in transgenic mice overexpressing human apolipoprotein C1. Diabetologia. 44: 437–443.[CrossRef][Medline]

  25. Voshol, P. J., M. C. Jong, V. E. Dahlmans, D. Kratky, S. Levak-Frank, R. Zechner, J. A. Romijn, and L. M. Havekes. 2001. In muscle-specific lipoprotein lipase-overexpressing mice, muscle triglyceride content is increased without inhibition of insulin-stimulated whole-body and muscle-specific glucose uptake. Diabetes. 50: 2585–2590.[Abstract/Free Full Text]

  26. Chomczynski, P., and N. Sacchi. 1987. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal. Biochem. 162: 156–159.[Medline]

  27. Bloks, V. W., T. Plosch, H. van Goor, H. Roelofsen, J. Baller, R. Havinga, H. J. Verkade, A. van Tol, P. L. Jansen, and F. Kuipers. 2001. Hyperlipidemia and atherosclerosis associated with liver disease in ferrochelatase-deficient mice. J. Lipid Res. 42: 41–50.[Abstract/Free Full Text]

  28. Heid, C. A., J. Stevens, K. J. Livak, and P. M. Williams. 1996. Real time quantitative PCR. Genome Res. 6: 986–994.[Abstract/Free Full Text]

  29. Bandsma, R. H., T. H. Van Dijk, A. A. Harmsel, T. Kok, D. J. Reijngoud, B. Staels, and F. Kuipers. 2004. Hepatic de novo synthesis of glucose 6-phosphate is not affected in peroxisome proliferator-activated receptor alpha-deficient mice but is preferentially directed toward hepatic glycogen stores after a short term fast. J. Biol. Chem. 279: 8930–8937.[Abstract/Free Full Text]

  30. Post, S. M., M. Groenendijk, K. Solaas, P. C. Rensen, and H. M. Princen. 2004. Cholesterol 7alpha-hydroxylase deficiency in mice on an APOE*3-Leiden background impairs very-low-density lipoprotein production. Arterioscler. Thromb. Vasc. Biol. 24: 768–774.[Abstract/Free Full Text]

  31. Heijboer, A. C., E. Donga, P. J. Voshol, Z. C. Dang, L. M. Havekes, J. A. Romijn, and E. P. Corssmit. 2005. Sixteen hours fasting differentially affects hepatic and muscle insulin sensitivity in mice. J. Lipid Res. 46: 582–588.[Abstract/Free Full Text]

  32. den Boer, M., P. J. Voshol, F. Kuipers, L. M. Havekes, and J. A. Romijn. 2004. Hepatic steatosis: a mediator of the metabolic syndrome. Lessons from animal models. Arterioscler. Thromb. Vasc. Biol. 24: 644–649.[Abstract/Free Full Text]

  33. Foley, J. E. 1992. Rationale and application of fatty acid oxidation inhibitors in treatment of diabetes mellitus. Diabetes Care. 15: 773–784.[Abstract]

  34. Gonzalez-Manchon, C., M. S. Ayuso, and R. Parrilla. 1990. On the mechanism of sodium 2-5-4 chlorophenylpentyloxirane-2-carboxylate (POCA) inhibition of hepatic gluconeogenesis. Biochem. Pharmacol. 40: 1695–1699.[CrossRef][Medline]

  35. Mandarino, L., E. Tsalikian, S. Bartold, H. Marsh, A. Carney, E. Buerklin, G. Tutwiler, M. Haymond, B. Handwerger, and R. Rizza. 1984. Mechanism of hyperglycemia and response to treatment with an inhibitor of fatty acid oxidation in a patient with insulin resistance due to antiinsulin receptor antibodies. J. Clin. Endocrinol. Metab. 59: 658–664.[Abstract]

  36. Tuman, R. W., G. F. Tutwiler, J. M. Joseph, and N. H. Wallace. 1988. Hypoglycaemic and hypoketonaemic effects of single and repeated oral doses of methyl palmoxirate (methyl 2-tetradecylglycidate) in streptozotocin/alloxan-induced diabetic dogs. Br. J. Pharmacol. 94: 130–136.[Medline]

  37. Boden, G., and X. Chen. 1999. Effects of fatty acids and ketone bodies on basal insulin secretion in type 2 diabetes. Diabetes. 48: 577–583.[Abstract]

  38. Jolly, R. A., R. Ciurlionis, D. Morfitt, M. Helgren, R. Patterson, R. G. Ulrich, and J. F. Waring. 2004. Microvesicular steatosis induced by a short chain fatty acid: effects on mitochondrial function and correlation with gene expression. Toxicol. Pathol. 32 (Suppl. 2): 19–25.

  39. Horton, J. D., and I. Shimomura. 1999. Sterol regulatory element-binding proteins: activators of cholesterol and fatty acid biosynthesis. Curr. Opin. Lipidol. 10: 143–150.[CrossRef][Medline]

  40. Dobbins, R. L., L. S. Szczepaniak, B. Bentley, V. Esser, J. Myhill, and J. D. McGarry. 2001. Prolonged inhibition of muscle carnitine palmitoyltransferase-1 promotes intramyocellular lipid accumulation and insulin resistance in rats. Diabetes. 50: 123–130.[Abstract/Free Full Text]


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M. A. M. den Boer, P. J. Voshol, F. Kuipers, J. A. Romijn, and L. M. Havekes
Hepatic glucose production is more sensitive to insulin-mediated inhibition than hepatic VLDL-triglyceride production
Am J Physiol Endocrinol Metab, December 1, 2006; 291(6): E1360 - E1364.
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