Very Low Density Lipoprotein Triglyceride Secretion Research Articles

Leptin increases hepatic triglyceride export via a vagal mechanism in humans

Recombinant human leptin (metreleptin) reduces hepatic lipid content in patients with lipodystrophy and overweight patients with non-alcoholic fatty liver disease and relative hypoleptinemia independent of its anorexic action. In rodents, leptin signaling in the brain increases very-low-density lipoprotein triglyceride (VLDL-TG) secretion and reduces hepatic lipid content via the vagus nerve. In this randomized, placebo-controlled crossover trial (EudraCT Nr. 2017-003014-22), we tested whether a comparable mechanism regulates hepatic lipid metabolism in humans. A single metreleptin injection stimulated hepatic VLDL-TG secretion (primary outcome) and reduced hepatic lipid content in fasted, lean men (n= 13, age range 20-38 years) but failed to do so in metabolically healthy liver transplant recipients (n= 9, age range 26-62 years) who represent a model for hepatic denervation. In an independent cohort of lean men (n= 10, age range 23-31 years), vagal stimulation by modified sham feeding replicated the effects of metreleptin on VLDL-TG secretion. Therefore, we propose that leptin has anti-steatotic properties that are independent of food intake by stimulating hepatic VLDL-TG export via a brain-vagus-liver axis.

276-OR: Impaired Glucagon-Mediated Suppression of VLDL-TG Secretion in Individuals with MAFLD

Background: Individuals with Metabolic Dysfunction-Associated Fatty Liver Disease (MAFLD) have both elevated triglyceride and glucagon levels, although glucagon suppresses the very-low-density lipoprotein-triglyceride (VLDL-TG) secretion in lean individuals. Therefore, we hypothesized that the glucagon-mediated suppression of VLDL-TG secretion is impaired in MAFLD, with greater impairment in individuals with steatohepatitis (MASH) than individuals with simple steatosis (MAFL) . Methods: 28 participants were included in the study (seven overweight control subjects (CON) , patients with MAFL, and with MASH) . We performed a somatostatin-clamp with replacement doses of growth hormone, insulin (0.2 mU/kg/min) and glucagon to suppress endogenous hormone production. Participants were studied during low-dose glucagon infusion (0.65 ng/kg/min, t=0─270 min, LowG) followed by a high-dose glucagon infusion (1.5 ng/kg/min, t=270─450 min, HighG) . We used 14C-labeled VLDL-TG and 3H-labeled glucose tracers to evaluate VLDL-TG secretion and endogenous glucose production (EGP) . Results: HighG suppressed VLDL-TG secretion compared to LowG in all groups, but with a greater reduction in VLDL-TG secretion in CON than in the MAFLD groups (CON 36% (±SD 19%) ; MAFL 10% (±SD 8%) ; MASH 13% (±SD 16%) , p<0.01) . Thus, the suppression was similarly mitigated in both MAFLD groups. This led to insignificant reductions in plasma VLDL-TG, TG and VLDL-TG oxidation rates in the CON group, in contrast to stable or increasing values in the MAFLD groups (CON vs. MAFLD delta-values, p<0.01, p=0.02 and p=0.04) , respectively) . EGP was similar in all groups throughout the LowG period and rose similarly when increasing glucagon infusion rate. Conclusions: Patients with MAFLD, MAFL as well as MASH, have an impaired glucagon-mediated suppression of VLDL-TG secretion which is a probable contributing factor to the dyslipidemia and the increased cardiovascular risk observed in individuals with MAFLD. NCT04042142 Disclosure S.Heebøll: None. J.R.Risikesan: None. H.Gronbaek: Other Relationship; ARLA, Research Support; AbbVie Inc., Intercept Pharmaceuticals, Inc., Speaker's Bureau; AstraZeneca, Novartis AG. E.Søndergaard: None. S.Nielsen: None. Funding Independent Research Fund Denmark (9039-00177B and -00278B) and Aase og Ejnar Danielsens Fond

Liver-Specific Deletion of Mouse Tm6sf2 Promotes Steatosis, Fibrosis, and Hepatocellular Cancer.

Human transmembrane 6 superfamily 2 (TM6SF2) variant rs58542926 is associated with NAFLD and HCC. However, conflicting reports in germline Tm6sf2 knockout mice suggest no change or decreased very low density lipoprotein (VLDL) secretion and either unchanged or increased hepatic steatosis, with no increased fibrosis. We generated liver-specific Tm6Sf2 knockout mice (Tm6 LKO) to study VLDL secretion and the impact on development and progression of NAFLD. Two independent lines of Tm6 LKO mice exhibited spontaneous hepatic steatosis. Targeted lipidomic analyses showed increased triglyceride species whose distribution and abundance phenocopied findings in mice with liver-specific deletion of microsomal triglyceride transfer protein. The VLDL triglyceride secretion was reduced with small, underlipidated particles and unchanged or increased apolipoprotein B. Liver-specific adeno-associated viral, serotype 8 (AAV8) rescue using either wild-type or mutant E167K-Tm6 reduced hepatic steatosis and improved VLDL secretion. The Tm6 LKO mice fed a high milk-fat diet for 3weeks exhibited increased steatosis and fibrosis, and those phenotypes were further exacerbated when mice were fed fibrogenic, high fat/fructose diets for 20weeks. In two models of HCC, either neonatal mice injected with streptozotocin (NASH/STAM) and high-fat fed or with diethylnitrosamine injection plus fibrogenic diet feeding, Tm6 LKO mice exhibited increased steatosis, greater tumor burden, and increased tumor area versus Tm6 flox controls. Additionally, diethylnitrosamine-injected and fibrogenic diet-fed Tm6 LKO mice administered wild-type Tm6 or E167K-mutant Tm6 AAV8 revealed significant tumor attenuation, with tumor burden inversely correlated with Tm6 protein levels. Liver-specific Tm6sf2 deletion impairs VLDL secretion, promoting hepatic steatosis, fibrosis, and accelerated development of HCC, which was mitigated with AAV8- mediated rescue.

No effect of resveratrol on VLDL-TG kinetics and insulin sensitivity in obese men with nonalcoholic fatty liver disease.

The present study (NCT01446276, ClinicalTrials.gov) assessed long-term effects of high-dose Resveratrol (RSV) on basal and insulin-mediated very low-density lipoprotein triglyceride (VLDL-TG), palmitate and glucose kinetics, and liver fat content in men with nonalcoholic fatty liver disease (NAFLD). Participants (n = 16) were non-diabetic, upper-body obese (BMI > 28 kg/m2 , WHR > 0.9) men with NAFLD who were randomized (1:1) in a double-blinded, placebo-controlled clinical trial to either RSV or placebo (500 mg 3 times daily) for 6 months. Magnetic resonance (MR) spectroscopy, dual-X-ray absorptiometry and MR imaging assessed liver fat content and body composition, respectively. 14 C-labeled VLDL-TG and 3 H-labeled glucose and palmitate tracers, in combination with indirect calorimetry and breath samples, were used to assess kinetics and substrate oxidations during basal and hyperinsulinaemic euglycaemic clamp conditions. RSV did not improve either basal or insulin-mediated VLDL-TG secretion, oxidation or clearance rates, nor did it affect palmitate or glucose turnover. Likewise, no changes in body composition or liver fat content occurred following RSV compared with placebo treatment. Therefore, RSV cannot be recommended for treatment of metabolic abnormalities in NAFLD.

Activation of Constitutive Androstane Receptor Ameliorates Renal Ischemia-Reperfusion-Induced Kidney and Liver Injury.

Acute kidney injury (AKI) is associate with high mortality. Despite evidence of AKI-induced distant organ injury, a relationship between AKI and liver injury has not been clearly established. The goal of this study is to investigate whether renal ischemia-reperfusion (IR) can affect liver pathophysiology. We showed that renal IR in mice induced fatty liver and compromised liver function through the downregulation of constitutive androstane receptor (CAR; -90.4%) and inhibition of hepatic very-low-density lipoprotein triglyceride (VLDL-TG) secretion (-28.4%). Treatment of mice with the CAR agonist 1,4-bis[2-(3,5 dichloropyridyloxy)] benzene (TCPOBOP) prevented the development of AKI-induced fatty liver and liver injury, which was associated with the attenuation of AKI-induced inhibition of VLDL-TG secretion. The hepatoprotective effect of TCPOBOP was abolished in CAR-/- mice. Interestingly, alleviation of fatty liver by TCPOBOP also improved the kidney function, whereas CAR ablation sensitized mice to AKI-induced kidney injury and lethality. The serum concentrations of interleukin-6 (IL-6) were elevated by 27-fold after renal IR, but were normalized in TCPOBOP-treated AKI mice, suggesting that the increased release of IL-6 from the kidney may have mediated the AKI responsive liver injury. Taken together, our results revealed an interesting kidney-liver organ cross-talk in response to AKI. Given the importance of CAR in the pathogenesis of renal IR-induced fatty liver and impaired kidney function, fatty liver can be considered as an important risk factor for kidney injury, and a timely management of hepatic steatosis by CAR activation may help to restore kidney function in patients with AKI or kidney transplant.

Impaired Insulin Suppression of VLDL-Triglyceride Kinetics in Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD) is associated with glucose and lipid metabolic abnormalities. However, insulin suppression of very low-density lipoprotein-triglyceride (VLDL-TG) kinetics is not fully understood. The objective of the study was to determine VLDL-TG, glucose, and palmitate kinetics during fasting and hyperinsulinemia in men with (NAFLD+) and without NAFLD (NAFLD−). Twenty-seven nondiabetic, upper-body obese (waist to hip ratio > 0.9, body mass index > 28 kg/m2) men, 18 NAFLD+, and nine NAFLD− determined by magnetic resonance spectroscopy were enrolled.14C-labeled VLDL-TG and 3H-labeled glucose and palmitate tracers were applied in combination with indirect calorimetry and breath samples to assess kinetics and substrate oxidations postabsorptively and during a hyperinsulinemic-euglycemic clamp. Dual-X-ray absorptiometry and magnetic resonance imaging assessed body composition. Liver fat content was greater in NAFLD+ than NAFLD− men (21.0% vs 3.7%), even though body composition, metabolites (except triglycerides), and insulin were similar in the groups. Insulin suppression of VLDL-TG secretion (P = .0001), oxidation (P = .0003), and concentration (P= .008) as well as percentage decreases were lower in NAFLD+ than NAFLD− men (secretion: 31.9% ± 17.2% vs 64.7% ± 19.9%; oxidation: −9.0% ± 24.7% vs 46.5% ± 36.6%; concentration: 11.9% ± 20.7% vs 56.2% ± 22.9%, all P < .001). Likewise, lower insulin suppression of very low-density lipoprotein particle size was present in NAFLD+ than NAFLD− men (P = .0002). Conversely, insulin suppression of endogenous glucose production was similar in the groups. Compared with endogenous glucose production, the inability of NAFLD+ men to suppress VLDL-TG kinetics to compensate for the increased liver fat content seems to be an early pathophysiological manifestation of male NAFLD+. These data suggest therapeutic targets reducing liver fat content may ameliorate metabolic abnormalities associated with NAFLD and presumably diabetes.

Glucose-induced lipid deposition in goose primary hepatocytes is dependent on the PI3K-Akt-mTOR signaling pathway

Previously we showed that fatty liver formation in overfed geese was accompanied by PI3K-Akt-mTOR pathway activation and changes in plasma glucose concentrations. Here, we show that glucose acts in goose hepatocellular lipid metabolism through the PI3K-Akt-mTOR signaling pathway. We observed that glucose increased lipogenesis, decreased fatty acid oxidation and increased very low density lipoprotein triglyceride (VLDL-TG) assembly and secretion. Co-treatment with glucose and inhibitors of the PI3K-Akt-mTOR pathway (LY294002, rapamycin, NVP-BEZ235) decreased the levels of factors involved in lipogenesis and increased the levels of factors involved in fatty acid oxidation and VLDL-TG assembly and secretion. These findings show that inhibition of the PI3K-Akt-mTOR pathway decreased glucose-induced lipogenesis, inhibited the downregulation of fatty acid oxidation by glucose and increased the upregulation of VLDL-TG assembly and secretion by glucose. The results presented herein provide further support for the role of the PI3K-Akt-mTOR pathway in lipid metabolism as we showed that in goose primary hepatocytes, glucose acts through the PI3K-Akt-mTOR-dependent pathway to stimulate lipid deposition by increasing lipogenesis and decreasing fatty acid oxidation and VLDL-TG assembly and secretion.

Hepatic denervation and dyslipidemia in obese Zucker (fa/fa) rats.

Human and animal studies increasingly point toward a neural pathogenesis of the metabolic syndrome, involving hypothalamic and autonomic nervous system dysfunction. We hypothesized that increased very-low-density lipoprotein-triglyceride (VLDL-TG) secretion by the liver in a rat model for dyslipidemia, that is, the obese Zucker (fa/fa) rat, is due to relative hyperactivity of sympathetic, and/or hypoactivity of parasympathetic hepatic innervation. To test the involvement of the autonomic nervous system, we surgically denervated the sympathetic or parasympathetic hepatic nerve in obese Zucker rats. Our results show that cutting the sympathetic hepatic nerve lowers VLDL-TG secretion in obese rats, finally resulting in lower plasma TG concentrations after 6 weeks. In contrast, a parasympathetic denervation results in increased plasma total cholesterol concentrations. The effect of a sympathetic or parasympathetic denervation of the liver was independent of changes in humoral factors or changes in body weight or food intake. In conclusion, a sympathetic denervation improves the lipid profile in obese Zucker rats, whereas a parasympathetic denervation increases total cholesterol levels. We believe this is a novel treatment target, which should be further investigated.

Lean body mass, not FFA, predicts VLDL-TG secretion rate in healthy men

Triglyceride is a risk factor for cardiovascular disease. However, the impact of body composition and free fatty acid (FFA) levels on very-low-density-lipoprotein triglyceride (VLDL-TG) secretion remains controversial. The aim was to identify predictors of VLDL-TG secretion in a data set compiled from seven previously published studies. VLDL-TG kinetics was studied in 96 healthy men covering a wide span in body composition. A primed-constant infusion of ex vivo labeled [1-(14)C]-triolein VLDL-TG was used. Body composition was determined by dual X-ray absorptiometry and computed tomography scanning. Energy expenditure was measured by indirect calorimetry. Palmitate flux was measured by a [9,10-(3)H]-palmitate infusion. VLDL-TG secretion rate correlated significantly with body mass index (BMI), lean body mass (LBM), total fat mass, resting energy expenditure (REE), and insulin. A trend toward an inverse relationship between VLDL-TG secretion rate and FFA concentration was observed. In mixed model linear regression analysis, VLDL-TG secretion rate was positively associated with LBM (P = 0.03), and VLDL-TG clearance rate was inversely related to total fat mass (P < 0.01). LBM is a predictor of VLDL-TG secretion in healthy men, whereas FFA availability is not associated with VLDL-TG secretion. The work suggests reporting VLDL-TG secretion rates normalized for LBM when comparing subjects with differences in body composition.

Abstract 127: Effects of miR-33 Antagonism on Glucose and Triglyceride Metabolism in Nonhuman Primates.

Elevated plasma low-density lipoprotein (LDL) and decreased high-density lipoprotein (HDL) cholesterol levels increase the risk of cardiovascular disease (CVD). While strategies to lower LDL cholesterol have been successful in reducing CVD-related mortality, there is still an unmet need for developing therapies to reduce the residual risk of atherosclerotic CVD. Plasma HDL cholesterol levels are inversely correlated with CVD risk and thus novel therapies to increase plasma HDL cholesterol levels have garnered much attention in recent years. MicroRNA-33a and b (miR-33a/b) are intronically-encoded microRNAs residing in the sterol response element binding protein genes SREBF2 and SREBF1 and suppress the expression of the genes involved in cholesterol efflux and fatty acid oxidation. Recent studies show that antagonism of miR-33 results in increased ABCA1expression and elevated plasma HDL levels in both mice and nonhuman primates. However, findings in mice suggest that long term miR-33 antagonism may lead to hepatic steatosis and increased production of very low-density lipoprotein (VLDL), the precursor to proatherogenic LDL. In the current study, cynomolgus monkeys were fed a high carbohydrate diet plus Kool Aid prior to administration of antisense oligonucleotides (ASOs) specific for miR-33a/b. As expected, miR-33 antagonism resulted in elevated plasma total cholesterol levels, predominantly due to increased HDL. No differences in plasma LDL or triglycerides were observed between treatment groups. MiR-33a/b antagonism had no effect on VLDL triglyceride secretion rates. Anti-miR-33 treatment did not promote hepatic lipid accumulation or adversely affect liver transaminase levels. Glucose tolerance tests revealed no differences in either plasma glucose or insulin levels in response to miR-33a/b antagonism. These findings suggest that anti-miR-33 therapy raises HDL cholesterol without negatively impacting either triglyceride or glucose homeostasis in nonhuman primates.

Effect of Acute Negative and Positive Energy Balance on Basal Very-Low Density Lipoprotein Triglyceride Metabolism in Women

BackgroundAcute reduction in dietary energy intake reduces very low-density lipoprotein triglyceride (VLDL-TG) concentration. Although chronic dietary energy surplus and obesity are associated with hypertriglyceridemia, the effect of acute overfeeding on VLDL-TG metabolism is not known.ObjectiveThe aim of the present study was to investigate the effects of acute negative and positive energy balance on VLDL-TG metabolism in healthy women.DesignTen healthy women (age: 22.0±2.9 years, BMI: 21.2±1.3 kg/m2) underwent a stable isotopically labeled tracer infusion study to determine basal VLDL-TG kinetics after performing, in random order, three experimental trials on the previous day: i) isocaloric feeding (control) ii) hypocaloric feeding with a dietary energy restriction of 2.89±0.42 MJ and iii) hypercaloric feeding with a dietary energy surplus of 2.91±0.32 MJ. The three diets had the same macronutrient composition.ResultsFasting plasma VLDL-TG concentrations decreased by ∼26% after hypocaloric feeding relative to the control trial (P = 0.037), owing to decreased hepatic VLDL-TG secretion rate (by 21%, P = 0.023) and increased VLDL-TG plasma clearance rate (by ∼12%, P = 0.016). Hypercaloric feeding increased plasma glucose concentration (P = 0.042) but had no effect on VLDL-TG concentration and kinetics compared to the control trial.ConclusionAcute dietary energy deficit (∼3MJ) leads to hypotriglyceridemia via a combination of decreased hepatic VLDL-TG secretion and increased VLDL-TG clearance. On the other hand, acute dietary energy surplus (∼3MJ) does not affect basal VLDL-TG metabolism but disrupts glucose homeostasis in healthy women.

Hepatic VLDL-TG Production and MTP Gene Expression are Decreased in Ovariectomized Rats: Effects of Exercise Training

The present study was designed to investigate the effects of estrogen withdrawal and exercise training on hepatic very low density lipoprotein-triglyceride (VLDL-TG) production and on expression of genes involved in hepatic VLDL synthesis in response to lipid infusion. Female Sprague-Dawley rats underwent ovariectomy (Ovx), sham surgery (Sham), and Ovx with 17β-estradiol supplementation (OvxE2) before being subdivided into sedentary (Sed) and trained (Tr) groups for 8 weeks. Exercise training consisted of continuous running on a rodent treadmill 5 times/wk. At the end of the 8-week period, all rats in the fasted state were intravenously infused with a 20% solution of Intralipid for 3-h followed by an injection of Triton WR-1339 to block lipoprotein lipase activity. Plasma TG accumulation was subsequently measured during 90 min to estimate VLDL-TG production. An additional control group consisting of Sham-Sed rats was infused with saline (0.9% NaCl). Estrogen withdrawal resulted in higher (p<0.01) liver fat accumulation concomitantly with lower (p<0.01) VLDL-TG production and lower mRNA and protein content of hepatic microsomal triglyceride transfer protein (MTP). All of these effects in Ovx rats were corrected with estrogen supplementation. Training in Ovx rats reduced (p<0.01) liver fat accumulation and further reduced (p<0.01) hepatic VLDL-TG production along with gene expression of MTP and diacylglycerol acyltransferase-2 (DGAT-2). It is concluded that VLDL-TG synthesis and/or secretion is decreased in Ovx rats probably via MTP regulation and that this decrease may constitute one of the factors involved in hepatic fat accumulation. The training effect on reducing VLDL production was independent of the estrogenic status.

Peroxisome Proliferator-activated Receptor-γ Coactivator-1α (PGC-1α) Stimulates VLDL Assembly through Activation of Cell Death-inducing DFFA-like Effector B (CideB)

Abnormalities in very low density lipoprotein (VLDL) assembly and secretion impact intrahepatic lipid homeostasis, plasma lipoprotein profile, and energy metabolism of distal peripheral tissues. We have evaluated the role of the transcriptional coactivator, the peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha), in VLDL assembly and secretion. PGC-1alpha overexpression in HepG2 cells led to diminished rates of triglyceride (TG) synthesis but strongly stimulated VLDL-TG secretion, markedly increasing the efficiency of secretion of newly synthesized TG. PGC-1alpha overexpression increased the rate of secretion of apoB100 and promoted secretion of larger, less dense VLDL particles. PGC-1alpha overexpression in intact mouse liver also stimulated rates of VLDL TG secretion and attenuated hepatic TG accumulation resulting from high fat diet feeding. To determine the molecular mechanisms mediating the effect of PGC-1alpha on VLDL assembly, we evaluated the expression of several candidate mediators known to be involved in VLDL assembly or hepatic lipid homeostasis. Cell death-inducing DFFA-like effector B (CideB) expression was greatly induced by PGC-1alpha, and siRNA against CideB reversed the effects of PGC-1alpha on the secretion of TG and VLDL-sized particles by HepG2 cells, indicating that CideB is a critical mediator of stimulatory effects of PGC-1alpha on VLDL secretion. Collectively, these data suggest that PGC-1alpha plays an important role in partitioning cytoplasmic TG toward the VLDL secretory compartments and promoting VLDL secretion via transcriptional induction of CideB.

Decrease in hepatic very‐low‐density lipoprotein–triglyceride secretion after weight loss is inversely associated with changes in circulating leptin

Although weight loss usually decreases very-low-density lipoprotein-triglyceride (VLDL-TG) secretion rate, the change in VLDL-TG kinetics is not directly related to the change in body weight. Circulating leptin also declines with weight loss and can affect hepatic lipid metabolism. The aim of this study was to determine whether circulating leptin is associated with weight loss-induced changes in VLDL-TG secretion. Ten extremely obese subjects were studied. VLDL-TG secretion rate and the contribution of systemic (derived from lipolysis of subcutaneous adipose tissue TG) and non-systemic fatty acids (derived primarily from lipolysis of intrahepatic and intraperitoneal TG, and de novo lipogenesis) to VLDL-TG production were determined by using stable isotopically labelled tracer methods before and 1 year after gastric bypass surgery. Subjects lost 33 +/- 12% of body weight, and VLDL-TG secretion rate decreased by 46 +/- 23% (p = 0.001), primarily because of a decrease in the secretion of VLDL-TG from non-systemic fatty acids (p = 0.002). Changes in VLDL-TG secretion rates were not significantly related to reductions in body weight, body mass index, plasma palmitate flux, free fatty acid or insulin concentrations. The change in VLDL-TG secretion was inversely correlated with the change in plasma leptin concentration (r = -0.72, p = 0.013), because of a negative association between changes in leptin and VLDL-TG secretion from non-systemic fatty acids (r = -0.95, p < 0.001). Weight loss-induced changes in plasma leptin concentration are inversely associated with changes in VLDL-TG secretion rate. Additional studies are needed to determine whether the correlation between circulating leptin and VLDL-TG secretion represents a cause-and-effect relationship.

Intrahepatic fat, not visceral fat, is linked with metabolic complications of obesity

Visceral adipose tissue (VAT) is an important risk factor for obesity-related metabolic disorders. Therefore, a reduction in VAT has become a key goal in obesity management. However, VAT is correlated with intrahepatic triglyceride (IHTG) content, so it is possible that IHTG, not VAT, is a better marker of metabolic disease. We determined the independent association of IHTG and VAT to metabolic function, by evaluating groups of obese subjects, who differed in IHTG content (high or normal) but matched on VAT volume or differed in VAT volume (high or low) but matched on IHTG content. Stable isotope tracer techniques and the euglycemic-hyperinsulinemic clamp procedure were used to assess insulin sensitivity and very-low-density lipoprotein-triglyceride (VLDL-TG) secretion rate. Tissue biopsies were obtained to evaluate cellular factors involved in ectopic triglyceride accumulation. Hepatic, adipose tissue and muscle insulin sensitivity were 41, 13, and 36% lower (P < 0.01), whereas VLDL-triglyceride secretion rate was almost double (P < 0.001), in subjects with higher than normal IHTG content, matched on VAT. No differences in insulin sensitivity or VLDL-TG secretion were observed between subjects with different VAT volumes, matched on IHTG content. Adipose tissue CD36 expression was lower (P < 0.05), whereas skeletal muscle CD36 expression was higher (P < 0.05), in subjects with higher than normal IHTG. These data demonstrate that IHTG, not VAT, is a better marker of the metabolic derangements associated with obesity. Furthermore, alterations in tissue fatty acid transport could be involved in the pathogenesis of ectopic triglyceride accumulation by redirecting plasma fatty acid uptake from adipose tissue toward other tissues.

Estimated liver weight is directly related to hepatic very low‐density lipoprotein–triglyceride secretion rate in men

Animal studies suggest that liver weight is directly related to hepatic very low-density lipoprotein-triglyceride (VLDL-TG) secretion, independently of body size. This relationship has never been examined in humans. We measured VLDL-TG secretion rate by using stable isotope-labelled tracers in 21 healthy, non-obese men (age: 25 +/- 3 years; body mass index: 24.8 +/- 1.6 kg m(-2)), and evaluated the relationship between VLDL-TG secretion and indices of total and regional adiposity (body mass index, total body fat, trunk fat), metabolic parameters (free fatty acid, glucose, and insulin concentrations, homeostasis model assessment index of insulin resistance, resting energy expenditure), and estimated liver weight. Correlation analysis showed that estimated liver weight was positively associated with total VLDL-TG secretion rate (r = 0.722, P < 0.001), VLDL-TG secretion rate per liter of plasma (r = 0.562, P = 0.008), VLDL-TG secretion rate per kilogram of body weight (r = 0.555, P = 0.009), and VLDL-TG secretion rate per kilogram of liver weight (r = 0.620, P = 0.003). In multiple regression analysis, estimated liver weight was the only significant predictor of VLDL-TG secretion rate regardless of units of expression, explaining 31-52% of total variance; none of the metabolic parameters and indices of body fatness entered the regression models. We conclude that estimated liver weight is directly related to hepatic VLDL-TG secretion rate in healthy non-obese men; this relationship is likely not mediated by interindividual variation in body size.

Upregulation of Apolipoprotein B Secretion, but Not Lipid, by Tumor Necrosis Factor‐α in Rat Hepatocyte Cultures in the Absence of Extracellular Fatty Acids

Tumor necrosis factor-alpha (TNF-alpha) plays a pivotal role in the host response to infection. Rapidly liberated to the bloodstream, TNF-alpha triggers the production of other cytokines and the acute-phase response. Hypertriglyceridemia is a sepsis hallmark associated with high plasma levels of very low-density lipoprotein (VLDL) particles, partly ascribed to increased hepatic production. The kinetics of the hepatocyte response, the cytokine/s responsible, and the underlying mechanisms are not fully elucidated. VLDL biogenesis is a complex, time-consuming process that depends on lipid availability and microsomal triglyceride transfer protein (MTP) activity for correct apolipoprotein B (apoB) lipidation. Studies were performed to define the direct effect of TNF-alpha on VLDL secretion rate and composition in rat hepatocytes cultured in conditions resembling the fed situation. Increases of 17-24% in the number of VLDL particles secreted and of 44-88% in the cellular levels of apoB mRNA were caused by 5, 20, or 100 ng/mL TNF-alpha in 8 h. Lipoprotein secretion returned to baseline levels in 16 h, whereas TNF-alpha-treated cells continued to exhibit higher apoB transcript levels. The mass of each lipid class in secreted VLDL and of MTP mRNA in cells was not affected by any of the tested TNF-alpha doses or treatment periods. These findings indicate that over a wide range of concentrations, TNF-alpha was capable of inducing sustained upregulation of apoB mRNA expression and transient increase in secretion of its protein, but, apparently, VLDL triglyceride secretion was not a TNF-alpha target under conditions in which fatty acids were not extracellularly provided.

Stimulation of Lipogenesis by Pharmacological Activation of the Liver X Receptor Leads to Production of Large, Triglyceride-rich Very Low Density Lipoprotein Particles

The oxysterol-activated liver X receptor (LXR) provides a link between sterol and fatty acid metabolism; activation of LXR induces transcription of lipogenic genes. This study shows that induction of the lipogenic genes Srebp-1c, Fas, and Acc1 upon administration of the synthetic LXR agonist T0901317 to C57BL/6J mice (10 mg/kg/day, 4 days) is associated with massive hepatic steatosis along the entire liver lobule and a 2.5-fold increase in very low density lipoprotein-triglyceride (VLDL-TG) secretion. The increased VLDL-TG secretion was fully accounted for by formation of larger (129 +/- 9 nm versus 94 +/- 12 nm, a 2.5-fold increase of particle volume) TG-rich particles. Stimulation of VLDL-TG secretion did not lead to elevated plasma TG levels in C57BL/6J mice, indicating efficient particle metabolism and clearance. However, T0901317 treatment did lead to severe hypertriglyceridemia in mouse models of defective TG-rich lipoprotein clearance, i.e. APOE*3-Leiden transgenic mice (3.2-fold increase) and apoE-/- LDLr-/- double knockouts (12-fold increase). Incubation of rat hepatoma McA-RH7777 cells with T0901317 also resulted in intracellular TG accumulation and enhanced TG secretion. We conclude that, in addition to raising high density lipoprotein cholesterol concentrations, pharmacological LXR activation in mice leads to development of hepatic steatosis and secretion of atherogenic, large TG-rich VLDL particles.

ANGPTL3 Decreases Very Low Density Lipoprotein Triglyceride Clearance by Inhibition of Lipoprotein Lipase

KK/San is a mutant mouse strain established in our laboratory from KK obese mice. KK/San mice show low plasma lipid levels compared with wild-type KK mice despite showing signs of hyperglycemia and hyperinsulinemia. Recently, we identified a mutation in the gene encoding angiopoietin-like protein 3 (Angptl3) in KK/San mice, and injection of adenoviruses encoding Angptl3 or recombinant ANGPTL3 protein to mutant KK/San mice raised plasma lipid levels. To elucidate the regulatory mechanism of ANGPTL3 on lipid metabolism, we focused on the metabolic pathways of triglyceride in the present study. Overexpression of Angptl3 in KK/San mice resulted in a marked increase of triglyceride-enriched very low density lipoprotein (VLDL). In vivo studies using Triton WR1339 revealed that there is no significant difference between mutant and wild-type KK mice in the hepatic VLDL triglyceride secretion rate. However, turnover studies using radiolabeled VLDL revealed that the clearance of (3)H-triglyceride-labeled VLDL was significantly enhanced in KK/San mice, whereas the clearance of (125)I-labeled VLDL was only slightly enhanced. In vitro analysis of recombinant protein revealed that ANGPTL3 directly inhibits LPL activity. These data strongly support the hypothesis that ANGPTL3 is a new class of lipid metabolism modulator, which regulates VLDL triglyceride levels through the inhibition of LPL activity.

The role of transhepatic bile salt flux in the control of hepatic secretion of triacylglycerol-rich lipoproteins in vivo in rodents

Bile salts (BS) have been shown to suppress the secretion of very-low-density lipoprotein-triglyceride (VLDL-TG) in rat and human hepatocytes in vitro. In the present study, we investigated whether the transhepatic BS flux affects VLDL-TG concentration and hepatic VLDL-TG secretion in vivo. In rats, the transhepatic BS flux was quantitatively manipulated by 1-week chronic bile diversion (BD), followed by intraduodenal infusion with taurocholate (TC) or saline for 6 h. In mice, the transhepatic BS flux was manipulated by a 3-week dietary supplementation with TC (0.5 wt.%) or cholestyramine (2 wt.%). In rats, BD followed by saline or TC infusion did not affect plasma triacylglycerol (TG) concentration, hepatic TG production rate or VLDL lipid composition, compared to control rats. In mice supplemented for 3 weeks with TC or cholestyramine, the transhepatic BS flux was increased by 335% and decreased by 48%, respectively, compared to controls. Among the three experimental groups of mice, an inverse relationship between transhepatic BS flux and either plasma TG concentration ( R 2=0.89) or VLDL-TG production rate ( R 2=0.87) was observed, but differences were relatively small. Present data support the concept that BS can reduce VLDL-TG concentration and inhibit hepatic TG secretion in vivo; however, this occurs only at supraphysiological transhepatic BS fluxes in mice.