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Endocrinology Vol. 138, No. 8 3276-3282
Copyright © 1997 by The Endocrine Society

ARTICLES

Lipoprotein Metabolism in the Fat Zucker Rat: Reduced Basal Expression but Normal Regulation of Hepatic Low Density Lipoprotein Receptors1

Wei Liao, Bo Angelin and Mats Rudling

Molecular Nutrition Unit, Center for Nutrition and Toxicology, NOVUM, and Metabolism Unit, Center for Metabolism and Endocrinology, Department of Medicine, Karolinska Institute at Huddinge University Hospital, Huddinge, Sweden

Address all correspondence and requests for reprints to: Wei Liao, M.D., Ph.D., Department of Cell Biology, Baylor College of Medicine, 1 Baylor Plaza, Houston, Texas 77030. E-mail: wliao{at}bcm.tmc.edu


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Hyperlipoproteinemia is one of the phenotypic characteristics of the fat Zucker rat that carries a mutation in the leptin receptor gene. In the present study, we studied the regulation of hepatic low density lipoprotein (LDL) receptor expression in lean and fat Zucker rats. Compared with lean rats, the fat ones had a pronounced (~60%) reduction in hepatic LDL receptor expression, whereas the levels of receptor messenger RNA (mRNA) were not reduced. Fat rats had increased levels of very low density lipoproteins and high density lipoproteins, but their plasma apo B100 within LDL was reduced. Challenge with 2% dietary cholesterol for 8 days suppressed hepatic LDL receptor expression in lean animals to similar levels as seen in fat ones, whereas the reduction in mRNA levels was much less pronounced. Treatment with ethynylestradiol (5 mg/kg BW·day) for 4 days strongly stimulated hepatic LDL receptor expression in both lean and fat rats; this treatment also increased LDL receptor mRNA levels, but to a lesser extent. In conclusion, the basal expression of hepatic LDL receptors is reduced in fat Zucker rats, but the capacity for the regulation of the receptors remains intact.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
ZUCKER RATS inherit obesity as an autosomal recessive trait (1). The fat rats (Rattus norvegicus) originally appeared as a spontaneous mutant in a cross between the Merck Stock M and Sherman rats (2). The homozygotes are obese, hyperphagic, insulin resistant and hyperlipidemic. Lean Zucker rats have lipid and lipoprotein patterns similar to rats of the Sprague-Dawley (3, 4) and Wistar strains (5), whereas a marked increase in plasma lipids and lipoproteins is one of the earliest abnormalities in fat Zucker rats (3, 4, 5, 6, 7, 8). In particular, very low density lipoproteins (VLDL) and high density lipoproteins (HDL) are increased (4). It has been shown that hepatic overproduction of lipoproteins (3, 8, 9, 10), rather than impaired lipolysis (4, 10), contributes to the development of hyperlipidemia in these animals.

The fat Zucker rat has recently been shown to have a defective receptor for leptin (11, 12, 13), which is the molecular basis for its typical phenotype. Fat Zucker rats also have pronounced hormonal changes. Thus, in addition to increased plasma insulin levels (14, 15, 16), fat Zucker rats have impaired metabolism of glucagon (17), GH (14, 15, 18), and thyroid hormones (14, 19). These hormones have been shown to play a role in the regulation of low density lipoprotein (LDL) receptors (20, 21, 22, 23, 24, 25, 26, 27), an important structure in the control of plasma lipoprotein metabolism. We therefore explored if the expression and regulation of hepatic LDL receptors are altered in fat Zucker rats. Our data show that the basal expression of hepatic LDL receptors is suppressed in fat rats. Similar levels of suppression could be achieved by the feeding of dietary cholesterol to lean Zucker rats, whereas both lean and fat animals showed a normal stimulation of the hepatic LDL receptors in response to estrogen treatment.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Material
Cholesterol and 17-{alpha} ethynylestradiol were purchased from Sigma (St. Louis, MO). Cholesterol-enriched diets (2%) were made by mixing ground rodent chow with hot Mazola corn oil (CPC Foods AB, Kristianstad, Sweden), 9:1, into which cholesterol had been dissolved.

Animals and experimental procedure
Male lean and fat Zucker rats were purchased from Harlan Sprague-Dawley, Inc. (Indianapolis, IN) and maintained under standardized conditions with free access to chow and water. The light cycle hours were between 0600 h and 1800 h. Animals were allowed to adapt to the environment for at least 2 weeks before starting the experiments. The experiments were approved by the institutional Animal Care and Use Committee.

For studying the effect of dietary cholesterol on hepatic LDL receptor expression, lean and fat rats (~ 8 weeks old, five animals per group) were fed standard rodent chow or 2% cholesterol-enriched chow for 8 days. On day 8 at approximately 1100 h, blood was taken and the livers were removed as described (28). Body weight was recorded the day before starting the food regime, and body and liver weights were recorded at sacrifice (Table 1Go).


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  		Table 1. Body and liver weights for the cholesterol feeding experiment

 
For studying the effect of estrogen treatment on hepatic LDL receptor expression, lean and fat rats (~10 weeks old, six animals per group) received ethynylestradiol (5 mg/kg BW, dissolved in propylene glycol) or the same amount of vehicle by daily sc injections (at 1000 h with the exception that the last injection was at 0800 h) for 4 days. On day 4, at approximately 1100 h, blood and liver samples were obtained as described above. Body weight was recorded the day before the ethynylestradiol treatment, and body and liver weights were recorded at sacrifice (Table 2Go).


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  		Table 2. Body and liver weights for the ethynylestradiol treatment experiment

 
Lipid determinations and size-fractionation of lipoproteins
Hepatic lipids were extracted from approximately 0.2 g samples according to the method of Folch et al. (29). Total cholesterol and triglycerides in plasma and liver extracts were assayed individually using commercial kits (Boehringer Mannheim, Mannheim, Germany). Size-fractionation of lipoproteins was performed on the pooled plasma samples of each group by fast protein liquid chromatograph (FPLC) as described (28, 30, 31). For this purpose, equal volumes of plasma from every rat in each group were pooled (5 ml), and the density was adjusted to 1.21 g/ml with solid KBr. After ultracentrifugation at 100 x 103 g for 48 h, the supernatant (lipoprotein fraction) was removed and adjusted to 2 ml by adding FPLC elution solution (0.15 M NaCl, 0.01% EDTA, 0.02% sodium acid, pH 7.3). After filtration through a 0.45 µm filter, 1 ml (corresponding to 2.5 ml plasma) was injected on a 54 x 1.8 cm Superose 6B column. Fractions of 2 ml were collected, and total cholesterol and triglycerides were measured.

Separation of apolipoproteins by SDS-PAGE
For separation of plasma apolipoproteins, 1.1 ml from every other FPLC fraction (fraction numbers 21 through 43) was precipitated with trichloroacetic acid (15%), washed twice with acetone, and solubilized in 120 µl of loading buffer as described elsewhere (22, 28). Samples were boiled for 5 min in the presence of 5% 2-mercaptoethanol, and 80 µl were loaded on 4–20% gadient SDS-polyacrylamide gels for electrophoresis separation of apolipoproteins (4 h, 45 mA/gel). Gels were stained with Coomassie blue. For reference, human LDL (apo B100), high (Bio-Rad Laboratories, Richmond, CA) and low (Pharmacia Fine Chemicals, Piscataway, NJ) molecular weight standards were used.

Ligand blot assay of LDL receptors
Hepatic membranes were prepared from the pooled liver samples (0.5 g) of each group as detailed previously (21, 28). The protein in the membrane preparation was assayed (32), using reagents from Bio-Rad. The membrane preparation was mixed with loading buffer (10% glycerol, 0.5% SDS, 2 mM CaCl2, 0.5% Triton X-100, 0.05% bromophenol blue, and 50 mM Tris-HCl, pH 6.8) and the proteins were separated by SDS-PAGE (6% polyacrylamide gels containing 0.1% SDS). No sulfhydryl reducing agent was added, and no heating was performed. The separated proteins were electrotransferred onto 0.45-µm nitrocellulose filters. The nitrocellulose filters were incubated for 1 h in the buffer (5% BSA, 2 mM CaCl2, 1 mM KI, 50 mM Tris.HCl, pH 8.0). 125I-labeled rabbit ß-VLDL (5 µg/ml) was then added. After an additional 1 h of incubation, filters were washed with 0.5% BSA, 2 mM CaCl2, 50 mM Tris. HCl, pH 8.0, and thereafter with the washing buffer without albumin. Filters were exposed to Kodak XAR-film. LDL receptor expression in blots was quantitated by using a Bio-Imaging Analyzer (Fujix, BAS 2000, Fuji Photo Film Co., Tokyo, Japan). Background levels measured in irrelevant filter areas of the same size have been subtracted from the data presented. It has been well established that LDL receptor after SDS-PAGE and transferring to nitrocellulose under nonreduced conditions retains its ligand binding activity (33). We used ß-VLDL as the ligand because ß-VLDL gives better signal to background ratio than LDL in visualization of LDL receptor (34). For quantitation of hepatic LDL receptor, ß-VLDL ligand blot assay is well correlated with RIA (34) and immunoblot assay (22) by using antibody.

Total nucleic acid (TNA) preparation and analysis of LDL receptor messenger RNA (mRNA)
TNA was prepared according to Durnam and Palmiter (35). The liver samples of each individual were homogenized with a Polytron (Kinematica, type PT 10/35, Kriens, Lucerne, Switzerland) in 4 ml of buffer (1% SDS, 10 mM EDTA, 20 mM Tris-HCl, pH 7.5) and digested for 45 min at 45 C with proteinase K (200 µg/ml). TNA was precipitated by adding 2 volumes of pure ethanol after phenol-chloroform extraction, and the pellet was suspended in the buffer. Quantitation of LDL receptor mRNA was done by a solution hybridization titration assay using a mouse [35S]UTP-complementary RNA probe (31). The slopes of the linear hybridization signals were calculated by the method of least squares and compared with the slope generated from a synthetic mouse LDL receptor mRNA standard. Data are expressed as attomoles (amol) per microgram TNA.

Statistics
Data are presented as means ± SEM and analyzed using Statistica software (StatSoft, Tulsa, OK). One-way ANOVA was used to evaluate the presence of significant differences between groups, followed by post hoc comparisons of the group means according to the method of Tukey.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
First, fat and lean rats fed the standard diet were compared. Hepatic LDL receptor expression was reduced by approximately 60% in fat rats, but the LDL receptor mRNA level was not reduced (Fig. 1AGo and B). There was no significant difference in hepatic total cholesterol between lean and fat rats, whereas hepatic triglycerides tended to be increased in fat rats (by 180%, Fig. 1CGo). Plasma total cholesterol was increased by 58% (P < 0.001) and plasma triglycerides were increased by approximately 4.5-fold (P < 0.001) in fat rats (Fig. 1DGo). FPLC analysis of plasma lipoproteins showed that the increased plasma cholesterol was within HDL and VLDL (Fig. 1EGo), and the increased plasma triglycerides were within VLDL (Fig. 1FGo). Separation of apolipoproteins by SDS-PAGE showed that LDL apoB100 was markedly decreased in fat rats (Fig. 2Go).



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  		Figure 1. Effects of dietary cholesterol on hepatic LDL receptors and plasma lipoproteins. Fat and lean Zucker rats received either standard food or 2% cholesterol-enriched food for 8 days. Blood was then drawn for analyses of plasma total cholesterol and triglycerides and for FPLC analysis of plasma lipoproteins; the livers were obtained for determination of LDL receptor expression, LDL receptor mRNA levels, total cholesterol and triglycerides. Cholesterol, triglycerides, and LDL receptor mRNA were determined individually. Ligand blot assay of LDL receptor and FPLC analysis of plasma lipoproteins were performed on the pooled samples of each group. A, Ligand blot assay of LDL receptor. For each group, 50, 100, and 200 µg of membrane protein (from left to right) were loaded onto the gel. The molecular mass is indicated in kDa on the left side of the figure. B, Quantitation of LDL receptor expression and LDL receptor mRNA. LDL receptor expression of lean rats receiving standard food was set to 100% as control. C, Hepatic total cholesterol and triglycerides. D, Plasma total cholesterol and triglycerides. E, FPLC lipoprotein cholesterol pattern. F, FPLC lipoprotein triglyceride pattern. *, P < 0.001 compared with lean rats fed standard chow. {dagger}, P < 0.001 compared with fat rats fed standard chow. {ddagger}, P < 0.005 compared with fat rats fed standard chow.

 


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  		Figure 2. Effects of cholesterol feeding on apolipoproteins. See Fig. 1Go for experimental procedure. Apolipoproteins in every other FPLC fraction of lipoproteins (fractions number 21 through 43) were separated by SDS-PAGE, as described in Materials and Methods.

 
To determine the response of hepatic LDL receptors to dietary cholesterol in Zucker rats, fat and lean rats were fed standard or 2% cholesterol-enriched diet for 8 days. Following dietary cholesterol, the expression of hepatic LDL receptors in lean and fat rats was reduced by 75% and 37%, respectively (Fig. 1Go, A and B). This suppression was much less pronounced at the mRNA level, where 36% (P = 0.08) and 8% reductions were seen in lean and fat rats, respectively (Fig. 1BGo). The cholesterol-enriched diet markedly increased hepatic total cholesterol and triglycerides in both lean and fat rats (P < 0.001; Fig. 1CGo). In lean rats, dietary cholesterol had minimal effects on plasma total cholesterol and triglycerides and on plasma lipoprotein pattern (Fig. 1Go, D–F). In fat rats, dietary cholesterol caused a more than 2-fold increase in VLDL and IDL cholesterol, whereas HDL cholesterol was reduced (Fig. 1EGo), so that plasma total cholesterol was only slightly increased (Fig. 1DGo). The cholesterol-enriched diet increased plasma triglycerides by 53% in fat rats (P < 0.005; Fig. 1DGo). However, this increase was not seen in the lipoprotein triglyceride patterns (Fig. 1FGo). This may be due to the loss of large chylomicrons and VLDL particles during the filtration procedure. SDS-PAGE analysis of plasma apolipoproteins showed that feeding lean rats the cholesterol-enriched diet did not cause major changes in apolipoproteins, whereas feeding fat rats the same diet increased apo B48 and apoB100 in VLDL and IDL fractions, while markedly reducing apo E in HDL (Fig. 2Go).

To determine if the regulatory responsiveness of hepatic LDL receptors is altered in fat Zucker rats, lean and fat rats were treated with ethynylestradiol or vehicle by daily sc injections for 4 days. This treatment is known to result in a pronounced increase in hepatic LDL receptors in normal rats (36, 37, 38).

In the rats that received vehicle only, the basal hepatic LDL receptor expression was again reduced by approximately 60% in fat rats as compared with lean ones (Fig. 3Go, A and B). Plasma total cholesterol and triglycerides were markedly increased in fat rats (P < 0.001; Fig. 3CGo), due to increased VLDL and HDL (Fig. 3Go, D and E). Again, apoB100 in the LDL fractions was clearly decreased in fat rats (Fig. 4Go). These findings thus confirmed the basal differences between fat and lean rats described above.



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  		Figure 3. Effects of estrogen treatment on hepatic LDL receptors and plasma lipoproteins. Fat and lean Zucker rats received either ethynylestradiol (5 mg/kg BW, using propylene glycol as vehicle) or equal amounts of vehicle by daily sc injections for 4 days. Blood was drawn for analyses of plasma total cholesterol and triglycerides and for FPLC analysis of plasma lipoproteins; the livers were obtained for determination of LDL receptor expression and LDL receptor mRNA levels. Cholesterol, triglycerides, and LDL receptor mRNA were determined individually. Ligand blot assay of LDL receptor and FPLC analysis of plasma lipoproteins were performed on the pooled samples of each group. A, Ligand blot assay of LDL receptor. For each group, 50, 100, and 200 µg of membrane protein (from left to right) were loaded onto the gel. Arrow indicates the precursor of LDL receptor. The molecular mass is indicated in kDa on the left side of the Figure. B, Quantitation of LDL receptor expression and LDL receptor mRNA. LDL receptor of lean rats receiving vehicle only was set to 100% as control. The LDL receptor precursor was not considered in the quantitation of LDL receptor expression shown in the figure. C, Plasma cholesterol and triglycerides. D, FPLC lipoprotein cholesterol pattern. E, FPLC lipoprotein triglyceride pattern. *, P < 0.001 compared with lean rats without estrogen treatment. {ddagger}, P < 0.0025 compared with lean rats without estrogen treatment. {dagger}, P < 0.001 compared with fat rats without estrogen treatment.

 


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  		Figure 4. Effects of estrogen treatment on apolipoproteins. See Fig. 3Go for experimental procedure. Apolipoproteins in every other FPLC fraction of lipoproteins (fractions number 21 through 43) were separated by SDS-PAGE, as described in Materials and Methods.

 
We then evaluated the effect of ethynylestradiol administration. This treatment stimulated the hepatic LDL receptor expression in lean and fat rats by approximately 5- and 17-fold (if the LDL receptor precursor band was also considered, the corresponding values were 8- and 24-fold), respectively (Fig. 3Go, A and B), whereas ethynylestradiol only increased the LDL receptor mRNA levels by approximately 1.5- (P < 0.001) and 3-fold (P < 0.001) in lean and fat rats, respectively (Fig. 3BGo). Following treatment, plasma total cholesterol was reduced by 66% in lean rats (P < 0.0025) and by 42% in fat rats (P < 0.001); plasma triglycerides were reduced by 81% (P < 0.0025) and 61% (P < 0.001), respectively (Fig. 3CGo). In lean rats, ethynylestradiol reduced cholesterol in all lipoproteins (Fig. 3DGo). In fat rats, the treatment markedly reduced HDL cholesterol, whereas cholesterol in IDL and VLDL fractions was increased (Fig. 3DGo). The reduction in plasma triglycerides following ethynylestradiol treatment was due to reduced VLDL triglycerides in both lean and fat rats (Fig. 3EGo). Analysis of plasma apolipoproteins in ethynylestradiol-treated lean rats revealed that most apolipoproteins disappeared almost completely (Fig. 4Go). Ethynylestradiol treatment caused a similar change in fat rats, but HDL apo AI was reduced moderately. Apo B48, apo B100, and Apo E remained in VLDL and IDL fractions, and apo AI and apo AIV bands appeared in VLDL and IDL fractions (Fig. 4Go).


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
One of the major new findings of the present study was that the basal expression of hepatic LDL receptors in fat Zucker rats was markedly lower than in lean ones. Although fat rats had a markedly reduced expression of hepatic LDL receptors, they had no clear accumulation of plasma LDL cholesterol. Actually, LDL apo B100 levels in fat rats were lower than in lean rats, a finding consistent with a previous report by Witztum and Schonfeld (8). Lipoprotein analysis by FPLC clearly demonstrated that the increased concentration of plasma triglycerides in fat rats was accounted for by plasma VLDL accumulation, and that the increased plasma concentration of cholesterol was due to increased cholesterol in VLDL and HDL fractions. These data are essentially consistent with previous observations (4). Thus, these findings indicate that the reduced LDL receptor expression in fat Zucker rat does not contribute in a major way to the development of hyperlipidemia in this animal.

The mechanism(s) responsible for this decrease in hepatic LDL receptors is unclear and may only be speculated upon. The decreased expression of hepatic LDL receptor in the fat Zucker rats may be explained by an increased turnover of the receptor. However, the fact that hepatic cholesterol was not increased in fat animals, and the lack of decrease in receptor mRNA levels, would clearly argue against a suppressed LDL receptor expression as the consequence of an increased influx of lipoprotein cholesterol to the liver. The changes in LDL receptor expression may be related to the hormonal abnormalities known to be present in fat Zucker rat. Thus, these rats are insulin-resistant and develop hyperinsulinemia (14, 15, 16). Furthermore, fat Zucker rats also have an impaired metabolism of glucagon (17), GH (14, 15, 18) and thyroid hormone (14, 19), all known to regulate LDL receptors (20, 21, 22, 23, 24, 25, 26, 27). A further intriguing possibility relates to the fact that Zucker rats have a defective leptin receptor (11, 12, 13) and therefore high circulating levels of plasma leptin (39). It is not known if this hormone may still exert some biological effects in tissues such as the liver via unaffected leptin receptor subtypes. Against this concept speaks the fact that obese humans, who have high levels of plasma leptin (40, 41, 42), in general express high levels of LDL receptor mRNA in their livers (Ståhlberg, D., M. Rudling, B. Angelin, P. Forsell, K. Nilsell, and K. Enarsson, unpublished data).

A second novel finding of the present study was that dietary cholesterol had a suppressive effect on hepatic LDL receptors in both lean and fat Zucker rats. This was somewhat surprising because rats have been reported to be resistant to the suppressive effect of dietary cholesterol on hepatic LDL receptor expression. Thus, dietary cholesterol stimulates rather than suppresses the expression of hepatic LDL receptors in rats of the Wistar and Sprague-Dawley strains (23, 43). In terms of hepatic LDL receptor expression, however, the response in both lean and fat Zucker rats to dietary cholesterol was more similar to that observed in rabbits and hamsters (44, 45). It is known that lean Zucker rats have lipid and lipoprotein patterns similar to rats of the Sprague-Dawley (3, 4) and Wistar strains (5), whereas fat Zucker rats develop marked hyperlipidemia (3, 4, 5, 6, 7, 8). In rats of the Wistar and Sprague-Dawley strains, dietary cholesterol reduces HDL cholesterol and increases VLDL and IDL cholesterol (23, 43). In the present study, dietary cholesterol caused similar changes in fat animals. However, lean Zucker rats were resistant to dietary cholesterol in terms of plasma lipoproteins, in spite of the fact that this diet markedly reduced the expression of hepatic LDL receptors. Again, this argues against the reduced LDL receptor expression as a major explanation of the hyperlipidemia in the fat Zucker rat.

A third finding of interest was that the hepatic LDL receptor expression could be stimulated in the fat Zucker rat by administration of pharmacological doses of estrogen. This indicates that the capacity for a maximal stimulation of hepatic LDL receptors in the fat Zucker rats is not defective. Furthermore, it implies that the relative deficiency in GH release known to be present in the fat Zucker rat (14, 15, 18, 46, 47, 48) does not affect the responsiveness to estrogen in this animal model of obesity, in contrast to the known requirement for GH in maintaining the response to estrogen in hypophysecomized rats (21). Estrogen may increase VLDL production in rats and rabbits, but also induces strong hypolipidemic effects (49, 50, 51). The hypolipidemic effect of estrogen treatment in lean Zucker rats found in the present study is similar to that shown in rats of other strains, but the estrogen treatment caused accumulation of VLDL and IDL particles in the fat Zucker rats. This may be the consequence of the stimulatory effects of estrogen on VLDL production; the accumulation of apolipoproteins of intestinal origin, such as apo AIV, in VLDL in this situation may actually indicate a state of saturation of lipoprotein clearance in these animals.

Our study also provides further evidence of the existence of a posttranscriptional regulation of hepatic LDL receptors in vivo. First, although fat Zucker rats had a clear reduction in hepatic LDL receptor expression, their LDL receptor mRNA levels were not reduced. Second, dietary cholesterol caused 75% and 37% reductions of hepatic LDL receptor expression in lean and fat rats, whereas the LDL receptor mRNA levels were only decreased by 36% and 8%, respectively. Third, estrogen caused 1.5- and 3-fold stimulations of LDL receptor mRNA in lean and fat rats, respectively, whereas it increased the LDL receptor expression by 5- and 17-fold. Thus, regulation of LDL receptor mRNA by dietary cholesterol and estrogen could not simply explain the change in LDL receptor expression in Zucker rats. Evidence of posttranscriptional regulation of LDL receptors has been presented in both in vitro (52) and in vivo (22) studies. It has also been shown previously that, in rats of the Wistar and Sprague-Dawley strains, dietary cholesterol stimulates hepatic LDL receptors but does not increase the LDL receptor mRNA (23, 43).

In summary, our study has demonstrated that the basal expression of hepatic LDL receptors is deficient in fat Zucker rats, that dietary cholesterol suppresses hepatic LDL receptors in both lean and fat Zucker rats, and that the response of hepatic LDL receptors to stimulation with estrogen remains intact in fat Zucker rats. Thus, the capacity for regulation of hepatic LDL receptors in the fat Zucker rat is normal. The observed suppression of basal hepatic LDL receptor binding activity in fat Zucker rats is presumably due to the altered hormonal balance in these animals.


   Acknowledgments
 
We thank Lilian Larsson for the preparation and labeling of rabbit ß-VLDL.


   Footnotes
 
1 This study was supported by grants from the Karolinska Institute, the Swedish Medical Research Council (03X-7137), from the Thuring, Widengren, Jeansson, and Lundström Foundations and Swedish Heart-Lung Foundation, and from the Ruth and Richard Julin, the Old Female Servants, and the Ax:son Johnson and the Nordic Insulin Funds. Back

Received January 28, 1997.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

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