Very low density lipoprotein (VLDL) receptor-deficient mice have reduced lipoprotein lipase activity. Possible causes of hypertriglyceridemia and reduced body mass with VLDL receptor deficiency.

Although very low density lipoprotein (VLDL) receptor (VLDLr) knockout mice have been reported to have no lipoprotein abnormalities, they develop less adipose tissue than control mice when fed a high calorie diet. Mice that are deficient in adipose tissue expression of lipoprotein lipase (LpL) also have less fat, but only when crossed with ob/ob mice. We hypothesized that the VLDLr, a protein that will bind and transport LpL, is required for optimal LpL actions in vivo and that hypertriglyceridemia due to VLDLr deficiency is exacerbated by either LpL deficiency or VLDL overproduction. Fasted VLDLr knockout (VLDLr0) mice were more hypertriglyceridemic than controls (2-fold greater triglyceride levels). The hypertriglyceridemia due to VLDLr0 was even more evident when VLDLr0 mice were crossed with heterozygous LpL-deficient (LpL1) and human apolipoprotein B (apoB) transgenic mice. This was due to an increase in apoB48-containing VLDL. [(3)H]VLDL turnover studies showed that VLDL-triglyceride clearance in VLDLr0/LpL1 mice was impaired by 50% compared with LpL1 mice. VLDLr0/LpL1 mice had less LpL activity in postheparin plasma, heart, and skeletal muscle. Infection of mice with an adenovirus-expressing receptor-associated protein, an inhibitor of the VLDLr, reduced LpL activity in wild type but not VLDLr0 mice. Therefore, the VLDLr is required for normal LpL regulation in vivo, and the disruption of VLDLr results in hypertriglyceridemia associated with decreased LpL activity.     

Role of lipoprotein lipase in plasma triglyceride removal.

Intravenous injections of anti-lipoprotein lipase serunis quantitatively block the catabolism of very low density lipoprotein (VLDL) and portomicron triglyceride and specifically inhibit triglyceride transport into ovarian follicles. The immunological studies presented provide information on the site of action of lipoprotein lipase (LPL). In the anti-LPL serum-treated animals initial plasma triglyceride accumulation occurs at the time of antiserum injection. This instantaneous inhibition of triglyceride removal provides direct evidence that the functional LPL responsible for VLDL and portomicron triglyceride hydrolysis is located in sites within the plasma compartment readily accessible to immunoglobulins. In vitro immunological studies show that the adipose, heart, ovarian, and liver LPL share common immunological determinants. Biochemical studies on highly purified heart and adipose LPL suggest that these enzymes have identical protein moieties.     

Insulin-mediated modifications of myocardial lipoprotein lipase and lipoprotein metabolism

Recirculating organ perfusion in vitro was conducted with hearts from control rats, animals given a single dose of streptozotocin (65 mg/kg) 48 h earlier, and streptozotocin-treated rats administered insulin (5 units), 2 h prior to organ perfusion. During 45-min perfusions, the lipolysis of very low density lipoprotein (VLDL) triglyceride was significantly less in hearts from diabetics than in controls (41.9 +/- 7.3% of control). This was associated with significant reductions in heparin-releasable (functional) lipoprotein lipase and tissue lipoprotein lipase of perfused hearts. The decreases in VLDL triglyceride metabolism and the levels of myocardial lipoprotein lipase were completely reversed by treatment of diabetic rats with insulin 2 h prior to study. Similar improvement of VLDL triglyceride metabolism and increases in myocardial lipoprotein lipase activity were observed in hearts from diabetic rats by direct addition of 100 milliunits/ml of insulin to the recirculating perfusion media. Under these conditions, the increase in both fractions of lipoprotein lipase in response to insulin was completely inhibited, and utilization of VLDL triglyceride was partially inhibited by pre-perfusion with cycloheximide for 10 min. The data derived from either VLDL triglyceride lipolysis in organ perfusion or direct measurement of myocardial lipoprotein lipase demonstrate a direct effect of insulin on myocardial lipoprotein lipase activity, and suggest that the response to insulin may be due in part to effects on protein synthesis.     

Effect of an anti-lipoprotein lipase serum on plasma triglyceride removal.

Anti-lipoprotein lipase sera injected intravenously in roosters blocked quantitatively the catabolism of very low density lipoprotein (VLDL) triglyceride. Antibodies were produced in rabbits immunized with highly purified lipoprotein lipase (LPL, glycerol ester hydrolase, E C 3.1.1.3) prepared from chicken adipose tissue. Following anti-LPL serum injection there was a linear increase in plasma triglyceride concentration. The rate of entry of triglyceride in plasma was estimated from the rate of triglyceride accumulation in the plasma of animals injected with anti-LPL serum, or from the disappearance curve of biologically labelled VLDL. In instances where both measurements were conducted in the same animals there was very close agreement between the two procedures. Inhibition of VLDL triglyceride catabolism of anti-LPL serum provided a way to characterize newly secreted VLDL that exhibited a broad spectrum of particle sizes with a median of 625 A degrees. They contained 76.2 +/- 1.2% triglyceride and had a high ratio of free to ester cholesterol (2.46 +/- 0.45). In control VLDL samples there was 46.1% triglyceride, and the ratio of free to ester cholesterol was 1.19. The complete inhibition of triglyceride removal by an antiserum prepared against adipose tissue LPL demonstrates that the NaCl-inhibited, serum-activated lipase prepared by affinity chromatography on heparin-Sepharose and concanavalin A-Sepharose columns is the enzyme responsible in vivo for the catabolism of VLDL triglyceride. Further, the kinetics of triglyceride accumulation in the plasma provide evidence that the site of degradation of VLDL triglyceride is within the plasma compartment.     

Resistance of a very low density lipoprotein subpopulation from familial dysbetalipoproteinemia to in vitro lipolytic conversion to the low density lipoprotein density fraction

In vitro lipolysis of very low density lipoprotein (VLDL) from normolipidemic and familial dysbetalipoproteinemic plasma by purified bovine milk lipoprotein lipase was studied using the combined single vertical spin and vertical autoprofile method of lipoprotein analysis. Lipolysis of normolipidemic plasma supplemented with autologous VLDL resulted in the progressive transformation of VLDL to low density lipoprotein (LDL) via intermediate density lipoprotein (IDL) with the transfer of the excess cholesterol to high density lipoprotein (HDL). At the end of 60 min lipolysis, 92-96% of VLDL triglyceride was hydrolyzed, and, with this process, greater than 95% of the VLDL cholesterol and 125-I-labeled VLDL protein was transferred from the VLDL to the LDL and HDL density region. When VLDL from the plasma of an individual with familial dysbetalipoproteinemia was substituted for VLDL from normolipidemic plasma, less than 50% of the VLDL cholesterol and 65% of 125I-labeled protein was removed from the VLDL density region, although 84-86% of VLDL triglyceride was lipolyzed. Analysis of familial dysbetalipoproteinemic VLDL fractions from pre- and post-lipolyzed plasma showed that the VLDL remaining in the postlipolyzed plasma (lipoprotein lipase-resistant VLDL) was richer in cholesteryl ester and tetramethylurea-insoluble proteins than that from prelipolysis plasma; the major apolipoproteins in the lipoprotein lipase-resistant VLDL were apoB and apoE. During lipolysis of normolipidemic VLDL containing trace amounts of 125I-labeled familial dysbetalipoproteinemic VLDL, removal of VLDL cholesterol was nearly complete from the VLDL density region, while removal of 125I-labeled protein was only partial. A competition study for lipoprotein lipase, comparing normolipidemic and familial dysbetalipoproteinemic VLDL to an artificial substrate ([3H]triolein), revealed that normolipidemic VLDL is clearly better than familial dysbetalipoproteinemic VLDL in competing for the release of 3H-labeled free fatty acids. The results of this study suggest that, in familial dysbetalipoproteinemic individuals, a subpopulation of VLDL rich in cholesteryl ester, apoB, and apoE is resistant to in vitro conversion by lipoprotein lipase to particles having LDL-like density. The presence of this lipoprotein lipase-resistant VLDL in familial dysbetalipoproteinemic subjects likely contributes to the increased level of cholesteryl ester-rich VLDL and IDL in the plasma of these subjects.     

Lipoprotein composition as a component in the lipoprotein clearance defect in experimental diabetes

The hypertriglyceridemia associated with streptozotocin-induced diabetes in rats is largely reflected in the plasma lipoproteins of density less than 1.006 g/ml. Analysis of the plasma apolipoproteins of these rats indicated marked alterations in both the total levels and in the lipoprotein distribution of the major apolipoproteins. In whole plasma, diabetes was associated with significant increases in apolipoprotein (apo)-AIV, apo-AI, and apo-B (mainly in the intestinally derived apo-B240) and a marked decrease in apo-E. In the d less than 1.006 g/ml lipoprotein fraction (very-low-density lipoproteins (VLDL], there were significant increases in apo-B240, apo-AI, and apo-AIV and decreased levels of apo-E and the C apolipoproteins. The decrease in apo-C was primarily due to lower levels of apo-CII, and the ratio of the lipoprotein lipase inhibitor, apo-CIII, to the lipoprotein lipase activator, apo CII, was significantly increased over that in controls. The comparative clearance of triglycerides of VLDL particles from control and diabetic rat plasma was tested in recirculating heart perfusion in vitro. During 45-min perfusions of hearts from control donor rats, lipolysis of triglycerides of VLDL from diabetic rats was only 63-64% of that using plasma VLDL from control rats. Perfusion of hearts from diabetic rats with VLDL from control rats gave lipolysis values of only 53% of that obtained with normal hearts. Where both the VLDL and hearts were obtained from diabetic rats, lipolysis was 23% of that observed when both the lipoprotein and the organ were from control rats. The data suggest that in addition to depressed lipoprotein lipase activity in the tissue from diabetic rats, there are also major compositional changes in circulating lipoproteins which may contribute to defective triglyceride clearance from the circulation.     

Tamoxifen inhibits lipoprotein activity: in vivo and in vitro studies

Tamoxifen, a nonsteroidal antiestrogenic antitumor agent, has weak estrogen-like effects on lipid metabolism, however, the mechanism remains unknown. We previously reported that tamoxifen decreases the activity of lipoprotein lipase (LPL), a key enzyme in triglyceride metabolism, in patients with breast cancer. This study evaluated the effect of tamoxifen on LPL activity in vitro and in vivo. In experiment 1, total cholesterol, triglyceride, adipose tissue weight, and LPL activity of post-heparin plasma were measured in ovariectomized female rats with and without tamoxifen treatment. In experiment 2, purified very-low-density lipoprotein (VLDL) and purified LPL were incubated with and without tamoxifen or estrogen, and the triglycerides in VLDL were measured using an enzymatic method. In experiment 1, total cholesterol and adipose tissue weight decreased significantly in tamoxifen-treated rats (p < 0.001 and p < 0.01, respectively). Triglyceride measurements were not significantly different between the two groups, however, the LPL activity was lower in tamoxifen-treated rats (p < 0.005). In experiment 2, triglycerides in VLDL were significantly higher after VLDL and LPL were incubated with tamoxifen and estrogen (p < 0.005). We concluded that tamoxifen inhibits the hydrolytic activity of LPL in vivo and in vitro. This mechanism may explain the elevated serum triglyceride levels in some patients treated with tamoxifen.     

Differential binding of triglyceride-rich lipoproteins to lipoprotein lipase.

In comparison to very low density lipoprotein (VLDL), chylomicrons are cleared quickly from plasma. However, small changes in fasting plasma VLDL concentration substantially delay postprandial chylomicron triglyceride clearance. We hypothesized that differential binding to lipoprotein lipase (LPL), the first step in the lipolytic pathway, might explain these otherwise paradoxical relationships. Competition binding assays of different lipoproteins were performed in a solid phase assay with purified bovine LPL at 4 degrees C. The results showed that chylomicrons, VLDL, and low density lipoprotein (LDL) were able to inhibit specific binding of (125)I-labeled VLDL to the same extent (85.1% +/- 13.1, 100% +/- 6.8, 90.7% +/- 23.2% inhibition, P = NS), but with markedly different efficiencies. The rank order of inhibition (K(i)) was chylomicrons (0.27 +/- 0.02 nm apoB) > VLDL (12.6 +/- 3.11 nm apoB) > LDL (34.8 +/- 11.1 nm apoB). By contrast, neither triglyceride (TG) liposomes, high density lipoprotein (HDL), nor LDL from patients with familial hypercholesterolemia were efficient at displacing the specific binding of (125)I-labeled VLDL to LPL (30%, 39%, and no displacement, respectively). Importantly, smaller hydrolyzed chylomicrons had less affinity than the larger chylomicrons (K(i) = 2.34 +/- 0.85 nm vs. 0.27 +/- 0.02 nm apoB respectively, P < 0.01). This was also true for hydrolyzed VLDL, although to a lesser extent. Chylomicrons from patients with LPL deficiency and VLDL from hypertriglyceridemic subjects were also studied. Taken together, our results indicate an inverse linear relationship between chylomicron size and K(i) whereas none was present for VLDL. We hypothesize that the differences in binding affinity demonstrated in vitro when considered with the differences in particle number observed in vivo may largely explain the paradoxes we set out to study.     

Overexpression of human apolipoprotein A-II in mice induces hypertriglyceridemia due to defective very low density lipoprotein hydrolysis

Two lines of transgenic mice, hAIItg-delta and hAIItg-lambda, expressing human apolipoprotein (apo)A-II at 2 and 4 times the normal concentration, respectively, displayed on standard chow postprandial chylomicronemia, large quantities of very low density lipoprotein (VLDL) and low density lipoprotein (LDL) but greatly reduced high density lipoprotein (HDL). Hypertriglyceridemia may result from increased VLDL production, decreased VLDL catabolism, or both. Post-Triton VLDL production was comparable in transgenic and control mice. Postheparin lipoprotein lipase (LPL) and hepatic lipase activities decreased at most by 30% in transgenic mice, whereas adipose tissue and muscle LPL activities were unaffected, indicating normal LPL synthesis. However, VLDL-triglyceride hydrolysis by exogenous LPL was considerably slower in transgenic compared with control mice, with the apparent Vmax of the reaction decreasing proportionately to human apoA-II expression. Human apoA-II was present in appreciable amounts in the VLDL of transgenic mice, which also carried apoC-II. The addition of purified apoA-II in postheparin plasma from control mice induced a dose-dependent decrease in LPL and hepatic lipase activities. In conclusion, overexpression of human apoA-II in transgenic mice induced the proatherogenic lipoprotein profile of low plasma HDL and postprandial hypertriglyceridemia because of decreased VLDL catabolism by LPL.     

Metabolism of very low density lipoproteins in genetically lean or fat lines of chicken

Metabolism of very low density lipoproteins (VLDL) has been compared in fat (FL) and lean (LL) lines of chicken. When refed after fasting, plasma triglyceride concentration reached a significantly higher plateau in FL, although their feed consumption was lower than in LL. Newly synthesized VLDL were studied using anti-lipoprotein lipase antibodies. VLDL triglyceride (TG) concentrations were increased by antibody injection and reached a higher concentration in FL plasma than in LL. Newly synthesized VLDL exhibited a similar lipid composition. Fatty acid profiles were also similar when birds ingested a very low fat diet. Comparison of in vitro affinity of lipoprotein lipase and VLDL from both genotypes did not reveal any difference in Km and Vmax. [14C]labelled VLDL from fat or lean donors were prepared and were injected into chickens from both genotypes. Fractional rate constants did not differ between lines. However, as plasma VLDL-TG pools were very different, plasma turnover was higher in FL than in LL. About 3-fold more VLDL-TG were incorporated in abdominal fat of FL than in LL. Difference in fattening between both genotypes seem to be due to both increased VLDL secretion and VLDL removal from plasma without difference in VLDL characteristics.     

Synthesis of recombinant high density lipoprotein with apolipoprotein A-I and apolipoprotein A-V

It has been shown that apolipoprotein A-V (apoA-V) over-expression significantly lowers plasma triglyceride levels and decreases atherosclerotic lesion development. To assess the feasibility of recombinant high density lipoprotein (rHDL) reconstituted with apoA-V and apolipoprotein A-I (apoA-I) as a therapeutic agent for hyperlipidemic disorder and atherosclerosis, a series of rHDL were synthesized in vitro with various mass ratios of recombinant apoA-I and apoA-V. It is interesting to find that apoA-V of rHDL had no effect on lipoprotein lipase (LPL) activation in vitro and very low density lipoprotein (VLDL) clearance in HepG2 cells and in vivo. By contrast, LPL activation and VLDL clearance were inhibited by the addition of apoA-V to rHDL. Furthermore, the apoA-V of rHDL could not redistribute from rHDL to VLDL after incubation at 37°C for 30 min. These findings suggest that an increase of apoA-V in rHDL could not play a role in VLDL clearance in vitro and in vivo, which could, at least in part, attribute to the lost redistribution of apoA-V from rHDL to VLDL and LPL binding ability of apoA-V in rHDL. The therapeutic application of rHDL reconstituted with apoA-V and apoA-I might need the construction of rHDL from which apoA-V could freely redistribute to VLDL.     

Mechanisms of triglyceride-lowering effect of an HMG-CoA reductase inhibitor in a hypertriglyceridemic animal model, the Zucker obese rat.

Inhibitors of 3-hydroxy-3-methyl glutaryl coenzyme A (HMG-CoA) reductase have been approved for treatment of hypercholesterolemia in humans. This class of therapeutic agents, in addition to lowering plasma cholesterol, reduces plasma triglyceride levels. We have investigated the mechanism of triglyceride-lowering effect of lovastatin in the hypertriglyceridemic state by using a rodent model of hypertriglyceridemia and obesity, the Zucker obese (fa/fa) rat. Lovastatin treatment (4 mg/kg), as compared to placebo, caused a 338% reduction in plasma triglyceride (146 +/- 5 vs. 494 +/- 76 mg/dl), a 58% decrease in total cholesterol (99 +/- 13 vs. 156 +/- 18 mg/dl), and a 67% reduction in high density lipoprotein (HDL)-cholesterol (69 +/- 8 vs. 115 +/- 15 mg/dl). The fall seen in plasma triglyceride was due to a decrease in hepatic secretion of very low density lipoproteins (VLDL), determined after blocking the clearance of triglyceride-rich lipoproteins with Triton WR-1339. Lovastatin treatment did not affect either the activities of hepatic lipogenic enzymes, glucose-6-phosphate dehydrogenase, or malic enzyme, or the activities of the lipolytic enzymes of adipose tissue, lipoprotein lipase, or liver, hepatic triglyceride lipase. Supplementation of mevalonolactone in the diet partially reversed the changes in plasma triglyceride (265 +/- 37 vs. 146 +/- 5 mg/dl), but not in total or HDL-cholesterol. These data demonstrate that, in the hypertriglyceridemic Zucker rat model, HMG-CoA reductase inhibitors reduce the rate of secretion of VLDL and this effect can be partially reversed by administration of mevalonolactone.     

Relationship of lipoprotein lipase activity to triglyceride uptake in adipose tissue

Fasted rats injected with actinomycin or fed glucose show increased lipoprotein lipase activity of epididymal adipose tissue. Data from the actinomycin-treated animals showed a direct correlation between the lipoprotein lipase activity and the uptake of lipoprotein triglyceride by the epididymal fat pad in vitro and in vivo. Data from the animals fed glucose confirmed these findings in vitro. These data strongly suggest that lipoprotein lipase plays a major role in triglyceride deposition in adipose tissue.     

Glucose tolerance, plasma lipoproteins and tissue lipoprotein lipase activities in body builders

Oral glucose tolerance, insulin binding to erythrocyte receptors, serum lipids, and lipoproteins, and lipoprotein lipase activities of adipose tissue and skeletal muscle were measured in nine body builders (relative body weight (RBW) 118 +/- 4%), eight weight-matched (RBW 120 +/- 5%) and seven normal-weight controls (RBW 111 +/- 3%). The body builders had 50% higher relative muscle mass of body weight (% muscle) and 50% smaller relative body fat content (% fat) than the two other groups (P less than 0.005). Maximal aerobic power was comparable in the three groups. In the oral glucose tolerance test (OGTT), blood glucose levels, and plasma insulin levels were lower (P less than 0.05) in the body builders than in weight-matched controls. Insulin binding to erythrocytes was similar in each group. On the basis of multiple linear regression analysis, 87% of the variation in plasma insulin response could be explained by body composition (% muscle and % fat) and VO2max. Plasma total cholesterol, low-density lipoprotein (LDL) cholesterol, and very low-density lipoprotein (VLDL) triglyceride concentrations were significantly lower in the body builders than in weight-matched controls. In comparison with the normal-weight group, the body builders had a lower total cholesterol level. High density lipoprotein (HDL) cholesterol, its subfractions (HDL2 and HDL3 cholesterol) and lipoprotein lipase (LPL) activities of adipose tissue and skeletal muscle were comparable in all three groups. Partial correlation analysis showed a positive relationship between plasma total triglyceride, total cholesterol and LDL cholesterol on the other hand and the % fat on the other.(ABSTRACT TRUNCATED AT 250 WORDS)     

Function of hepatic triglyceride lipase in lipoprotein metabolism

Rat hepatic triglyceride lipase (H-TGL) was purified from liver tissue extracts by affinity chromatography on Sepharose 4B with covalently linked heparin. The purified rat H-TGL exhibited the properties previously described for this enzyme. Enzyme protein was injected into rabbits for anti-H-TGL antibody production. Antirat-H-TGL did not react against rat lipoprotein lipase (LPL) but inhibited H-TGL-activity both in vitro and in vivo greater than 90%. These antibodies were injected into rats and lipoprotein analyses were performed over a 36-hr period. It could be shown that inactivation of H-TGL by anti-H-TGL gamma-globulins in vivo led to an increase in total triglyceride concentration from 70 mg/dl to 230 mg/dl due to an increase in very low density lipoprotein (VLDL) and low density lipoprotein (LDL) triglycerides 4 hr after antibody injection; a marked increase in high density lipoprotein (HDL) phospholipid concentration was observed with almost no change in HDL-cholesterol and HDL-triglycerides. This study documents the ability of antirat-H-TGL-gamma-globulins to inhibit H-TGL in vitro and in vivo. Furthermore, the inhibition of triglyceride removal in vivo demonstrated that this enzyme together with LPL is responsible for the catabolism of VLDL-triglyceride.     

Inhibition of lipoprotein lipolysis by polyarginine and evaluation of the mechanism of its interaction with lipoprotein lipase

It was found that polyarginine (Mr 40 000-60 000) is a strong inhibitor of the lipoprotein lipase activity in vivo and in vitro. The inhibitory effect in vivo was observed after a single intravenous injection of 0.85-3.5 mg/kg to rabbits, that in vitro at the polypeptide concentration of greater than or equal to 2.5 micrograms/ml. Within the first few hours after intravenous injection of polyarginine hyperlipidemia occurred with an obvious increase in the plasma triglyceride and VLDL fractions and a slight decrease of the LDL and HDL fractions. These changes typical for reduced lipoprotein lipolysis were due to the formation of a polyarginine-heparin complex, on the one hand, and to the formation of a polyarginine-enzyme complex devoid of the lipolytic properties, on the other. The inhibitory effect of polyarginine on lipoprotein lipase is related to the whole polypeptide molecule or its large fragment, since arginine and metformine (bi-guanidine compound) have no effect on the enzyme activity.     

Role of lipoprotein lipase separated from VLDL during VLDL catabolism

Lipoprotein lipase (LPL) which is associated with very low density lipoprotein (VLDL) separated from the VLDL-LPL complex during hydrolysis of triglyceride in the presence of HDL in vitro. When further VLDL was added to the mixture, the separated LPL became associated with the freshly added VLDL and hydrolyzed its triglyceride. These results suggest that LPL separated from the substrate during catabolism of VLDL may act on other VLDL particles in vivo.     

Postabsorptive VLDL‐TG Fatty Acid Storage in Adipose Tissue in Lean and Obese Women

Adipose tissue lipoprotein lipase (LPL) is a necessary enzyme for storage of very‐low‐density lipoprotein–triglyceride (VLDL‐TG), but whether it is a rate‐determining step is unknown. To test this hypothesis we included 10 upper‐body obese (UBO), 11 lower‐body obese (LBO), and 8 lean women. We infused ex vivo‐labeled VLDL‐¹⁴C‐TG and then performed adipose tissue biopsies to understand the relationship between VLDL‐TG storage and LPL activity in femoral and upper‐body subcutaneous fat. Both fractional tracer storage and rate of storage of the VLDL‐TG tracer were evaluated. VLDL‐TG storage was also examined as a function of regional adipose tissue blood flow (ATBF), insulin, VLDL‐TG turnover, regional fat mass, fat‐free mass (FFM), and fat cell size. LPL activity per adipocyte was significantly greater in obese than lean women but not significantly different per gram lipid. Both VLDL‐TG fractional tracer storage per kg lipid and VLDL‐TG storage rate per kg lipid were similar in abdominal and femoral fat in all three groups and were not significantly different between groups. Multiple regression analysis identified FFM and femoral fat mass as significant independent predictors of VLDL‐TG fractional tracer storage and insulin as a significant predictor of VLDL‐TG fatty acid storage rate. LPL activity, ATBF, and VLDL‐TG turnover did not predict VLDL‐TG storage. We conclude that lower FFM and greater plasma insulin are associated with greater VLDL‐TG deposition in abdominal subcutaneous and femoral fat. Greater femoral fat mass signals greater femoral VLDL‐TG storage. We suggest that the differences in VLDL‐TG storage in abdominal and femoral fat that occur with progressive obesity are regulated through mechanisms other than LPL activity.     

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.     

Very low density lipoprotein. Removal of Apolipoproteins C-II and C-III-1 during lipolysis in vitro.

In this study we have investigated the effects of very low density lipoprotein (VLDL) lipolysis on the removal of radiolabeled apolipoprotein C-II and apolipoprotein C-III-1 from in vitro lipolyzed lipoproteins. Lipolysis was carried out in vitro using lipoprotein lipase purified from bovine milk, and mixtures with or without plasma. Lipoproteins were isolated by ultracentrifugation and by gel filtration. Labeled apo-C-II and apo-C-III-1 distributed among plasma lipoproteins, predominantly VLDL and high density lipoprotein (HDL). Lipolysis induced transfer of apo-C-II and apo-C-III-1 from VLDL to HDL. The transfer was proportional to the extent of triglyceride hydrolysis, and similar for the two apoproteins. The apo-C-II/apo-C-III-1 radioactivity ratio did not change in either VLDL or the fraction of d greater than 1.006 g/ml during the progression of the lipolytic process. Similar observations were recorded while using plasma-devoid lipolytic systems. Gel filtration of incubation mixtures, on 6% agarose, revealed that the removal of labeled apo-C molecules from VLDL is not a consequence of either centrifugation or high salt concentration. These results suggest that there is no preferential removal of apo-C-II or apo-C-III-1 from lipolyzed VLDL particles. They further indicate that the ratio of apo-C-II to apo-C-III-1 does not regulate the extent of lipolysis of different VLDL particles, at least in VLDL isolated from normolipidemic humans.