Alterations of lipids and apolipoprotein CIII in very low density lipoprotein subspecies in type 2 diabetes

Aims/hypothesis Very low density lipoprotein (VLDL) particles are heterogeneous, comprising two main subspecies, VLDL 1 (Sf 60-400) and VLDL 2 (Sf 20-60). The aim of the study was to examine the distribution and composition of VLDL subspecies in type 2 diabetes.Subjects, materials and methods We studied the composition and concentration of triglyceride-rich lipoproteins (TRLs) in 217 type 2 diabetic patients and 93 control subjects between 50 and 75 years of age. Lipoprotein subspecies were separated by density-gradient ultracentrifugation. Apolipoprotein (apo) CIII and apo E in plasma and apo CIII in TRL subspecies were measured by nephelometry and apo CII in serum by a commercial kit using a single radial immunodiffusion method.Results The concentrations of VLDL 1, VLDL 2 and intermediate density lipoprotein were significantly increased in type 2 diabetes subjects, the change being most marked for VLDL 1. There was a strong linear correlation between VLDL 1 triglycerides and plasma triglycerides in both groups (r=0.879, p<0.001 and r=0.899, p<0.001). Diabetic subjects had markedly higher plasma ratios of apo CII:apo CIII and apo CIII:apo E. Despite elevated plasma apo CIII, type 2 diabetic subjects had a relative deficiency of apo CIII in all TRL subspecies, suggesting profound disturbances of apo CIII metabolism.Conclusions/interpretation The elevation of VLDL 1 triglycerides is the major determinant of plasma triglyceride concentration in normal subjects and in type 2 diabetic individuals. Both apo CIII and apo E metabolism are disturbed in type 2 diabetes.     

Differences in low density lipoprotein subfractions and apolipoproteins in premenopausal and postmenopausal women

Postmenopausal or oophorectomized women are at higher risk for the development of coronary artery disease than are premenopausal women. These differences in risk may be due to alternations in plasma lipoproteins modulated by hormonal changes. Plasma cholesterol, triglyceride, lipoprotein cholesterol, and apolipoprotein A-I and B (apoB) concentrations, as well as low density lipoprotein (LDL) particle size (LDL 1-7), as assessed by 2-16% polyacrylamide-agarose gradient gel electrophoresis, were determined in 87 premenopausal and 43 postmenopausal women. All were participants in the Framingham Offspring Study, were gynecologically normal, and were not taking any hormones. The postmenopausal women had significantly (P less than 0.05) higher plasma LDL cholesterol concentrations than did the premenopausal women. Plasma triglyceride, total cholesterol, very low density lipoprotein cholesterol, and apoB levels were higher, and apoA-I and high density lipoprotein cholesterol were lower in the postmenopausal group, but these differences were not significant at P less than 0.05. The postmenopausal women were likely to have small LDL particles compared to premenopausal women. Controlling for age and body mass index effects significantly reduced the differences in total cholesterol, LDL cholesterol, apoB, and LDL particle size and broadened the differences in apoA-I and high density lipoprotein cholesterol. These data indicate that menopause is positively correlated with LDL cholesterol (P less than 0.05) and decreased LDL particle size (P less than 0.05) after adjusting for significant covariates.     

Transfers of esterified cholesterol and triglyceride between high density and very low density lipoproteins: In vitro studies of rabbits and humans

Mixtures of rabbit very low density lipoproteins (VLDL) and high density lipoproteins (HDL), one of which was endogenously labeled with ³H in the free and esterified cholesterol moieties and with ¹⁴C in the triglyceride (TG) moiety, were incubated at 37°C in vitro in the presence of rabbit lipoprotein-free serum. There was a net mass transfer of esterified cholesterol from HDL to VLDL and of TG from VLDL to HDL. Both esterified ³H-cholesterol and ¹⁴C-TG, however, transferred in each direction between the two fractions. In the presence of parachlormercuryphenyl sulfonate (PCMPS), a thiol group blocker that inhibits lecithin:cholesterol acyltransferase (LCAT), the net mass transfers of esterified cholesterol and TG were reduced, as were the bidirectional transfers of ¹⁴C-TG, but the bidirectional transfers of esterified ³H-cholesterol were much less affected. Qualitatively similar results were obtained with incubations of human VLDL and HDL containing tracer amounts of either VLDL or HDL labeled with ³H in the esterified cholesterol moiety. In incubations of unlabeled human VLDL, HDL, and lipoprotein-free plasma, the molar ratio of the net mass transfers of TG:esterified cholesterol was 0.55 ± 0.10 (mean ± SE, n = 8) in the absence of PCMPS, and 0.25 ± 0.04 in the presence of PCMPS. The difference between these ratios was statistically significant (p < 0.02). Therefore, it has been concluded that the net mass transfers of esterified cholesterol and TG between VLDL and HDL may be independent processes.     

Metabolism of esterified cholesterol in the plasma very low density lipoproteins of the rabbit.

The in vivo metabolism of esterified cholesterol and triglyceride in plasma very low density lipoproteins (VLDL) has been studied in postabsorptive rabbits injected with endogenously 3H-labelled preparations of VLDL. The rates at which the injected esterified [3H]cholesterol and [3H]triglyceride were removed from the recipient VLDL fraction were remarkably similar. But, whereas most of the [3H]triglyceride was rapidly lost from the plasma compartment, more than 60% of the esterified [3H]cholesterol removed from VLDL was recovered in the plasma low density lipoprotein (LDL) and high density lipoprotein (HDL) fractions, an observation in direct contrast to previous findings in rats and guinea pigs in which hepatic uptake accounted for the major proportion of esterified cholesterol removed from VLDL. The appearance of VLDL-esterified [3H]cholesterol in rabbit LDL was quite compatible with the well-documented catabolic conversion of VLDL to LDL. The appearance in HDL, on the other hand, could not be so explained and was further investigated in studies performed in vitro. It was found that, concurrent with a net mass transfer of esterified cholesterol from HDL to VLDL, there was a much more rapid transfer of esterified [3H]cholesterol in the reverse direction, implying the existence of a process of molecular exchange of esterified cholesterol between the two fractions, analogous to that recently described between VLDL and LDL.     

Cloning and expression of apolipoprotein B, the major protein of low and very low density lipoproteins.

We report the cloning of cDNAs for rat liver apolipoprotein B (apo B) and the use of the cloned sequences to examine apo B expression at the level of mRNA in rat tissues. Fifteen putative apo B clones were identified by antibody screening of a rat liver cDNA library in the lambda gt11 expression vector. The identity of the clones was confirmed by immunological studies of the fusion protein products. All clones appear to contain sequences found only in apo B PI, the high molecular weight form of rat liver apo B. Blotting studies show that the clones hybridize to a single 20-kilobase liver mRNA species, sufficiently large to encode the entire apo B PI peptide, which is estimated to be 400 kDa in size. Apo B PI mRNA is abundant in liver and present in lower amounts in intestine but is absent in a variety of other tissues examined. This tissue distribution is consistent with that expected from studies on the in vivo synthesis of apo B. One clone, corresponding to a 240-base segment of the apo B PI mRNA, was sequenced and found to exhibit homology with a short region of rat apo E mRNA. Analysis of the secondary structure of the corresponding peptide did not show the preponderance of amphipathic alpha-helical structures characteristic of other apolipoproteins examined thus far.     

Serum lipoproteins, apolipoproteins and very low density lipoprotein subfractions during 6-month fibrate treatment in primary hypertriglyceridaemia

Serum lipoproteins and apolipoproteins were studied in 14 hypertriglyceridaemic (HTG) patients during a 24-week period of treatment with gemfibrozil, and after a 6-week washout period. A marked decrease in very low density lipoprotein (VLDL) cholesterol and triglyceride was observed. There was an increase in high density lipoprotein (HDL) cholesterol, particularly the HDL3 component. A slight increase in low density lipoprotein (LDL) cholesterol was observed after 12 weeks, but this had almost disappeared after 24 weeks. The treatment resulted in an increase in serum apolipoprotein A-II levels and a reduction in serum apo C-III and apo E. VLDL subfractionation by density gradient centrifugation in four subfractions of decreasing size (A, B, C and D) showed a predominant reduction of the large subfractions A, B and C, while the decrease in VLDL-D was less marked. Percentage changes from the baseline level of VLDL-A and VLDL-D cholesterol were found to be inversely correlated with percentage changes in HDL and LDL cholesterol, respectively. This might reflect a transfer of cholesterol from VLDL-A to HDL, and from VLDL-D to LDL. The above data suggest fibrate-induced stimulation of lipoprotein lipase, and indicate that the enhanced transfer of cholesterol from VLDL to LDL, induced by fibrates in HTG patients, is less pronounced after a prolonged period of treatment.     

Effects of estrogen replacement on plasma lipoproteins and apolipoproteins in postmenopausal, dyslipidemic women.

The effects of oral estrogen replacement (ethinyl estradiol 0.02 mg/d) on plasma triglyceride, total cholesterol, very-low-density lipoprotein (VLDL) cholesterol, low-density lipoprotein (LDL) cholesterol, high-density lipoprotein (HDL) cholesterol, and apolipoprotein (apo) A-I and B levels and LDL particle size were assessed in 20 postmenopausal women with a previous hysterectomy and various forms of dyslipidemia (LDL cholesterol > or = 4.14 mmol/L [160 mg/dL] and/or HDL cholesterol < or = 1.03 mmol/L [40 mg/dL]). All subjects were studied while on a standard cholesterol-lowering diet, and were sampled in the fasting state before beginning estrogen therapy and after a mean of 13 weeks of estrogen therapy. Lipids were measured by standardized enzymatic techniques, apos were measured by enzyme-linked immunoassays, and LDL particle size was measured by gradient gel electrophoresis. Mean values for plasma lipid parameters (mmol/L) at baseline and during estrogen replacement were as follows: triglyceride, 2.11 and 2.75 (30% increase); total cholesterol, 7.45 and 6.52 (13% decrease); VLDL cholesterol, 1.09 and 1.22 (12% increase); LDL cholesterol, 5.09 and 3.70 (27% decrease); and HDL cholesterol, 1.27 and 1.58 (24% increase). Mean values for apo A-I were 163 and 254 mg/dL (56% increase), and for apo B they were 170 and 148 mg/dL (13% decrease). The LDL particle score was 4.09 and 4.52 (11% smaller). Changes in all parameters were statistically significant (P = .05) except for VLDL cholesterol. These data indicate that estrogen replacement is effective in decreasing LDL cholesterol and apo B concentrations and increasing HDL cholesterol and apo A-I concentrations in dyslipidemic postmenopausal women, but it should not be used in patients with baseline fasting triglyceride levels higher than 2.82 mmol/L (250 mg/dL) unless it is accompanied by a progestin. Our data indicate that this form of estrogen replacement could lower the risk of coronary artery disease (CAD) by more than 50% in these women, based on favorable alterations in plasma lipoproteins.     

Effect of high density lipoproteins on permeability of rabbit aorta to low density lipoproteins

A study was made on the effect of high density lipoproteins (HDL) on the permeability of rabbit aorta to low density lipoproteins (LDL) after intravenous administration of human HDL and human [125I]LDL to normal and hypercholesterolemic rabbits. Evaluation of radioactivity in plasma and aorta has shown that the administration of a large dose of HDL decreased the aorta permeability rate for [125I]LDL on an average by 19% in normal rabbits, and by 45% in rabbits with moderate hypercholesterolemia. A historadiographic study showed that HDL also decreased the vessel wall permeability to [125I]LDL in normal and particularly in hypercholesterolemic animals. The suggestion was made that HDL at very high molar concentration can hamper LDL transportation through the intact endothelial layer into the intima due to the ability of HDL to compete with LDL in sites of low affinity on the surface of endothelial cells.     

Effect of pravastatin on intermediate-density and low-density lipoproteins containing apolipoprotein CIII in patients with diabetes mellitus

The apolipoprotein (apo) B lipoproteins, intermediate-density lipoproteins (IDL) and low-density lipoproteins (LDL) that contain apo-CIII are associated with coronary heart disease in patients with diabetes mellitus. Apo-CIII is prominent in diabetic dyslipidemia. We studied whether these apo-B lipoprotein types containing apo-CIII in diabetics are reduced by 1 year of pravastatin treatment. We randomly selected 45 age- and gender-matched placebo/pravastatin pairs from diabetic patients in the Cholesterol and Recurrent Events trial, a randomized, double-blinded trial of pravastatin 40 mg monotherapy. Very-low-density lipoproteins (VLDL) and IDL + LDL particles were subdivided based on the presence of apo-E and apo-CIII to yield 3 particle types: E+CIII+, E-CIII+, and E-CIII-. Compared with placebo, pravastatin reduced IDL + LDL apo-B concentrations for E+CIII+, E-CIII+, and E-CIII- by 42% (p = 0.02), 17% (p = 0.7), and 29% (p = 0.002), respectively, commensurate with IDL + LDL cholesterol concentration reductions in the particle types of 29% (p = 0.002), 25% (p = 0.2), and 36% (p <0.0001), respectively. These IDL + LDL CIII+ particles are rich in triglycerides and cholesterol and are likely to be remnant particles of VLDL. Thus, pravastatin reduced potentially atherogenic remnant particles, a prominent component of diabetic dyslipidemia associated with coronary events; these results may contribute to its demonstrated effectiveness in reducing coronary heart disease in diabetics.     

Rate of exchange of esterified cholesterol between human plasma low and high density lipoproteins.

In vitro incubations of human low density lipoproteins (LDL), high density lipoproteins (HDL) and lipoprotein-free plasma at 37 degrees for 6 h revealed no significant net mass transfers of esterified cholesterol from either lipoprotein fraction to the other. Observed transfers of esterified [3H]cholesterol from LDL to HDL must, therefore, have represented a process of molecular exchange between the two fractions. The time course of the esterified [3H]cholesterol transfers indicated that, from the point of view of the exchange process, the pools of esterified cholesterol in LDL and HDL were (mathematically) homogeneous, thus permitting a precise calculation of the rate of the exchange between the two fractions. In incubations of lipoproteins isolated from the plasma of 9 clinically normal human subjects, there was an hourly exchange of 8-12% and 7-31% of the pools of esterified cholesterol in LDL and HDL respectively, which, in molar terms, represented an exchange of 108-301 mumol/l plasma/h between the two fractions. The exchange of esterified cholesterol between human LDL and HDL is, therefore, rapid when considered in relation to other parameters of esterified cholesterol and lipoprotein metabolism.     

Changes in very low and low density lipoproteins with dietary fat modification

Since VLDL and LDL are involved in atherogenesis, their response to dietary modification was studied in 15 normal male prisoners. A 3-month reference diet (P/S ratio 0.3, daily cholesterol intake 370 mg) was compared with a modified fat diet (P/S 1.0, 250 mg) given for further 3 months. The decrement in serum cholesterol by 32 mg/dl reflected a decrease in VLDL and LDL. It was associated with a decrease in serum apolipoprotein B by 16 mg/dl and in serum apolipoprotein E by 1.2 mg/dl. The decrement in VLDL cholesterol was paralleled by a lowered VLDL apolipoprotein E content. Serum and VLDL triglycerides, HDL cholesterol and the serum apolipoproteins A-I and A-II did not change significantly. One beneficial result of a conventional dietary regimen is lowered LDL with unaffected HDL. Another effect is the apparent modification of VLDL with a decrement of cholesterol and apolipoprotein E-enriched particles.     

Targeting cardiovascular risk associated with both low density and high density lipoproteins using statin-niacin combination therapy

BACKGROUND: Cardiovascular risk might be reduced by targeted changes in both low density and high density lipoprotein cholesterol (LDL-C and HDL-C). This dual strategy will require a well tolerated, effective regimen, as well as a better understanding of how HDL-C may be targeted. DESIGN: An open-label, uncontrolled, retrospective cohort study of combined statin-niacin therapy. METHODS: We reviewed all patients ( n= 132) started on this combination in a referral lipid clinic over a 6.5-year period for tolerability, safety and effectiveness. RESULTS: Combined therapy was tolerated by 77% of patients. No serious adverse events attributable to medication were encountered. In drug-naive patients (n = 37), moderate doses of statin and niacin (mean 1180 mg/day) reduced LDL-C 31% and increased HDL-C 29% ( P< 0.002, both comparisons). At niacin doses >or= 1000 mg/day (mean 1480) added to a constant statin regimen (n=29), HDL-C increased 20% ( P< 0.001). Even at niacin doses < 1000 mg/day (mean 580, n= 23), HDL-C increased 13% ( P< 0.05). Although mean HDL-C increased, the initial and final HDL-C distributions were broad and largely overlapping. Any chosen cutpoint for HDL-C goal would apply to only a minority of patients. The total/HDL cholesterol ratio had narrower distributions, as the percentage of patients with ratio < 5.0 increased from 17% to 67%. CONCLUSION: Combined statin-niacin therapy lowers LDL-C and raises HDL-C with acceptable tolerance and safety. If treating LDL-C is the primary goal, consistent with current guidelines, then a strategy of targeting the total/HDL cholesterol ratio as a secondary goal is applicable to more patients than targeting HDL-C itself.     

Effect of hypochlorite (HOCl)-modified low density lipoproteins and high density lipoproteins on platelet function

As hypochlorous acid (HOCl) might be a potential candidate for generation of modified (lipo)proteins in vivo , the present study was aimed at investigating the effects of HOCl-modified lipoproteins on platelet function in vitro. Lipoproteins modified with HOCl at concentrations that occur physiologically did not induce spontaneous platelet aggregation. However, low density lipoproteins (LDL; 100 to 500 microg protein/ml) increased platelet aggregation and fluorescence anisotropy in response to ADP (1 and 10 microM) and thrombin (0.1 and 0.5 U/ml) as well as malondialdehyde (MDA) formation as a function of increasing HOCl-concentrations (0.2 to 1.6 mM). HOCl-modified high density lipoprotein subfraction 3 (HDL3) had no effect on platelet function.     

Effect of estrogen replacement plus low-dose alendronate treatment on bone density in surgically postmenopausal women with osteoporosis

This prospective randomized, double-blind, placebo-controlled, clinical trial was performed to evaluate the effectiveness of estrogens plus low-dose alendronate on bone metabolism. A total of 150 surgically postmenopausal women with osteoporosis were randomized in three groups: group A, micronized E2 (2 mg/d) plus standard-dose alendronate (10 mg/d); group B, micronized E2 plus low-dose alendronate (5 mg/d); and group C, micronized E2 plus placebo (one tablet per day). In all women, bone mineral density (BMD) and serum bone metabolism markers were assessed at admission and every 6 months for 2 yr. After 2 yr, BMD significantly increased compared with baseline in all groups. The percentage BMD change was significantly higher in groups A and B than in group C. The differences in BMD detected between groups A and B were not statistically significant. Since the 6-month follow-up and throughout the study, serum osteocalcin and bone alkaline phosphatase levels and urinary deoxypyridinoline and pyrilinks-D excretion were significantly reduced in all groups. Serum bone alkaline phosphatase levels significantly decreased in groups A and B, without difference between them, in comparison with group C. In conclusion, in surgically postmenopausal osteoporotic women treated with estrogen replacement, the addition of alendronate at a low dose of 5 mg daily induces a gain of bone mass not significantly different in comparison with that obtained using a standard dose of 10 mg daily.     

Lipids, lipoproteins, apolipoproteins, and other risk factors in Chinese men and women with and without myocardial infarction

One hundred and fifty-four male and 69 female Chinese patients, aged between 40 and 60 years, who had suffered myocardial infarction (MI) were investigated and compared with 216 men and 219 women who had no history or ECG evidence of coronary heart disease. The male MI patients had significantly raised levels of triglycerides (160 mg/dl), cholesterol (194 mg/dl), VLDL-CH (31 mg/dl), apolipoprotein B (122 mg/dl) and apolipoprotein E (4.7 mg/dl) and a lower apolipoprotein A-I level (126 mg/dl) than the control group (triglycerides 131, cholesterol 179, VLDL-CH 26, apo B 102, apo E4.2, and apo A-I 138 mg/dl). The women with MI also had higher values for the atherogenic lipids than the control group (triglycerides 175 vs. 134 mg/dl, cholesterol 218 vs. 186 mg/dl, LDL-CH 128 vs. 104 mg/dl, VLDL-CH 32 vs. 26 mg/dl, apo B 121 vs. 103 mg/dl and apo E 5.4 vs. 4.3 mg/dl), as well as lowered apolipoprotein A-I (128 vs. 144 mg/dl). The Lp(a) levels (men and women considered together) were significantly higher for the MI patients (34.3 mg/dl vs. 26.2 mg/dl). Anti-atherogenic lipoproteins such as HDL-cholesterol, HDL2-CH, HDL3-CH, phospholipids and apolipoprotein A-II, C-II and C-III showed no difference between the groups.     

Metabolism of apolipoproteins CII,CIII1, CIII2 and VLDL-B in human subjects consuming high carbohydrate diets

Simultaneous kinetics of apoproteins CII, CIII₁ and CIII₂ and VLDL-B were determined in six normal subjects initially while consuming a control diet and subsequently while consuming a high carbohydrate diet. Mean triglyceride levels in plasma and VLDL were significantly increased, from 110 to 173 mg/dl and 40 to 114 mg/dl respectively, by the high carbohydrate diet. Specific radioactivity of each C peptide was determined, following injection of ¹²⁵I-VLDL, in both VLDL and HDL from plasma samples taken over 48 hr using a technique involving analytical isoelectric focusing. Specific activity vs time curves for VLDL and HDL were almost superimposable on either diet, suggesting that CII, CIII₁ and CIII₂ were catabolized as a group. The high carbohydrate diet resulted in a significant increase in the mass of each C peptide which was brought about by an increase in production since no change in fractional removal rate (FRR) was observed. The pool size of VLDL apo B was also significantly increased and was a function of both increased production as well as decreased FRR. The proportion of CIII₂ mass relative to those of CII and CIII₁ was lower in both VLDL and HDL with the high carbohydrate diet. These observations indicate that the metabolism of the B and C apoproteins are independent of each other and do not respond in the same manner to changes in VLDL triglyceride and that increased triglyceride synthesis and secretion stimulates C apoprotein production.     

Effect of ketoconazole on plasma sex hormones, lipids, lipoproteins, and apolipoproteins in hyperandrogenic women

The aim of the current study of 18 hyperandrogenic women was to determine the affects of ketoconazole (KTZ), an oral synthetic antifungal imidazole derivative that inhibits gonadal and adrenal steroidogenesis, on lipids, lipoprotein cholesterols, apolipoproteins, endogenous sex steroid hormones, and their interactions. Eighteen hyperandrogenic women, ages 18 to 35, with a history of severe acne and/or hirsutism, were randomly divided into two groups of nine, both receiving KTZ (group 1, 400 mg/d; group 2,800 mg/d) for 10 days. In groups 1 and 2, KTZ therapy reduced cholesterol (10%, P less than or equal to .01; 19%, P less than or equal to .05) and low-density lipoprotein (LDL)-cholesterol (13%, P less than or equal to .05; 33%, P less than or equal to .025), and increased apolipoprotein (apo) A1 (7%, P less than or equal to .005; 13%, P less than or equal to .01). KTZ, 800 mg/d, decreased apo B (21%, P less than or equal to .005), and lowered the ratio of LDL-cholesterol to high-density lipoprotein (HDL)-cholesterol (40%, P less than or equal to .01). KTZ therapy more than doubled the levels of estradiol (E2) in both groups (136%, P less than or equal to .01; 171%, P less than or equal to .01) and, in the high-dose group, decreased the levels of free testosterone (FT) (48%, P less than or equal to .05) and dehydroepiandrosterone-sulfate (DHEA-S) (36%, P less than or equal to .005). The reductions of total and LDL-cholesterol appear to be attributable to the increases in E2 and possibly to the decrease in FT. KTZ therapy may have beneficial effects on atherogenic lipid and lipoprotein patterns in women with hyperandrogenicity.     

Effects of apolipoprotein C-II (apoC-II) on the lipolysis of very low density lipoproteins from apoC-II deficient patients

Lipoprotein lipase requires apolipoprotein C-II (apoC-II) from plasma very low density lipoproteins (VLDL) and high density lipoproteins (HDL) for maximal activity. To understand the mechanism by which apoC-II enhances the activity of the enzyme, the kinetic parameters for the hydrolysis of VLDL-triglycerides and phospholipids by purified bovine milk lipoprotein lipase have been determined in two patients with apoC-II deficiency. The absence of apoC-II in these patients was demonstrated by a specific radioimmunoassay for apoC-II (<0.05 mg apoC-II/dl plasma; normals ?5.0 mg/dl) and by isoelectric focusing of the isolated apoVLDL. The plasma levels of apoC-III, another apoprotein of VLDL, in the two subjects were 18.8 and 22.0 mg/dl (normals 11.1 ± 0.9 mg/dl). The kinetics of lipolysis of VLDL in either the presence or absence of apoC-II were monitored by using the patients' VLDL which were labeled in vitro with tri[1-¹⁴C]oleoyl glycerol and dansyl phosphatidylethanolamine (DPE). The release of [¹⁴C]oleic acid and the rate and extent of the DPE fluorescence increase were dependent on lipoprotein lipase and apoC-II concentration. Maximal hydrolysis of VLDL-triglycerides by lipoprotein lipase occurred at 2.5 μg apoC-II/mg triglyceride. The value of the Michaelis-Menten constant (K_m) of lipoprotein lipase for apoC-II deficient VLDL-triglycerides decreased from 7.8 mM in the absence of apoC-II to 1.0 mM at 2.5 μg of apoC-II; there was only a slight change in V_max. When normal HDL were used as the source of apoC-II, the rate of lipolysis of apoC-II deficient VLDL also increased and the value of K_m decreased to 1.0 mM. These results suggest that the effects of apoC-II on the rate of lipolysis of VLDL result from an apoC-II induced decrease in the apparent K_m of the enzyme for the substrate. One possible explanation for this decrease in K_m is that apoC-II enhances the interaction between lipoprotein lipase and triglyceride within the surface monolayer of the lipoprotein particle.     

Formation of high density lipoprotein2-like particles during lipolysis of very low density lipoproteins in vitro.

The effects of lipolysis of human plasma very low density lipoprotein (VLDL) on the structure and composition of high density lipoproteins (HDL) have been investigated. Lipolysis was performed in a controlled system in vitro containing VLDL (d less than 1.006 g/ml) and HDL3 (d = 1.125-1.210 g/ml) from human plasma and lipoprotein lipase (EC 3.1.1.34) purified from bovine milk. Lipolysis of VLDL caused profound changes in HDL3. Protein, phospholipid, and cholesterol liberated from VLDL during its lipolysis were transferred to the HDL3 particles. As a consequence of this in vitro transfer, the chemical composition and biophysical properties of HDL3 were substantially altered. The newly formed particles exhibited a flotation rate (F01.20) of 6.7 and a hydrated density of 1.110 g/ml. The chemical composition closely resembled that of native HDL2, and their size was slightly larger than that of the precursor HDL3. When HDL3 and postlipolysis HDL2 were subjected to ultracentrifugation under flotation velocity and equilibrium conditions, both proved to be stable particles. These results, when extrapolated to in vivo conditions, suggest an important metabolic relationship between the levels of circulating VLDL and HDL2 in plasma. This relationship now permits a reasonable explanation for numerous in vivo observations in which the levels of VLDL and HDL2 change reciprocally.     

Delayed clearance of very low density and intermediate density lipoproteins with enhanced conversion to low density lipoprotein in WHHL rabbits.

Rabbit livers express two genetically distinct receptors for plasma lipoproteins: (i) the low density lipoprotein (LDL) receptor and (ii) the chylomicron remnant receptor. In homozygous Watanabe-heritable hyperlipidemic (WHHL) rabbits, an animal model for human familial hypercholesterolemia, LDL receptors are genetically deficient, but chylomicron remnant receptors are normal. Hence, WHHL rabbits clear LDL from the circulation at an abnormally slow rate, but they clear chylomicron remnants at a normal rate. The current studies show that WHHL rabbits clear 125I-labeled very low density lipoprotein (VLDL) and its metabolic product, intermediate density lipoprotein (IDL), from plasma at a markedly decreased rate. The impaired clearance is due to a profound decrease in the rate of uptake of 125I-labeled VLDL and 125I-labeled IDL by the liver. Because of its rapid clearance in normal rabbits, only a fraction of the 125I-labeled apoprotein B component of VLDL is converted to LDL. In WHHL rabbits, the impaired clearance of VLDL leads to a markedly increased conversion of 125I-labeled apoprotein B from VLDL to LDL. These results indicate that: (i) in rabbits, the LDL receptor mediates the rapid removal of VLDL and IDL from plasma, and (ii), a deficiency of LDL receptors leads to an enhanced conversion of VLDL to LDL. The combination of overproduction and impaired plasma clearance of LDL, both resulting from a single gene mutation in the LDL receptor, leads to a massive increase of plasma LDL levels in homozygous WHHL rabbits.